Program Book - American Society of Andrology
Transcription
Program Book - American Society of Andrology
SUMMARY OF 1987 ANNUAL MEETING OF THE AMERICAN SOCIETY OF ANDROLOGY ALL ACTIVITIES ARE ON THE LOWER LEVEL 2 (LL2) IN THE COLORADO OR DENVER BALLROOMS I Friday March 6 Postgraduate Course Saturday Annual March 7 Meeting Sunday March 8 Annual Meeting Monday March 9 Annual Meeting Time Event Time Event Time Event Time Event NEUROENDOCRINOLOGY 0800 Neuroanatomy and Animal Studies; A. Plant 0800 Session A, Opening R. P. Amann W.D.Odell 0800 Session F, Clinical Andrology; Papers 62 to 69 0800 Session K, Epldldymis; Papers 117 to 124 0850 Pituitary Tumors; P. Snyder 0810 The Serano Lecture R. V. Short The Androgens Regulation of Gonadotropin in Man; W. Bremner 0900 1000 BREAK Session G, Toxicology and Ageing; Papers 70 to 76 Therapy with GnRH and Analogs: W. Crowley 1015 Session B, Endocrinology and Sertoli Cell; Papers 1 to 7 1200 LUNCH 1200 LUNCH ON YOUR OWN 1200 LUNCH ON YOUR OWN 1330 Session C, Sperm Evaluation; Papers 8 to 14 1330 Session H, State of the Art Lecture, E. 0. Price Evolutionary Aspects of Male Behavior 1430 Session I, Posters coffee and soda; Papers 77 to 116 1630 Session J, Workshop on Animal Rights, B. Rollin 1330 Donor Selection; N.Alexander 1400 Sexually Transmitted Disease Screening; M. E. Guinan BREAK 1020 Awards Ceremony 1050 Annual Business Meeting 1145 Session L, The Serano Lecture, R. V. Short; The Size of the Testes 1245 Adjourn BREAK BREAK BREAK 1505 Legal and Liability Aspects of AID; D.Young 1550 Screening Couples; K. Jones 1620 Sperm Freezing; R. L. Urry 1650 Insemination and Results; K. Jones 1745 Exhibitor Tutorials, 1730 Pennrose & Pomeroy Rooms (on L3) Exhibitor Tutorials, I & J Colorado Ballroom 1800 Student Mixer, Matchless Room (on LL 1) BREAK 1900 Cocktails in Denver Ballroom 1900 Opening Reception, Denver Ballroom Session E, Posters, Papers 22 to 61, wine and cheese 1930 Banquet In Denver Ballroom 1530 Session D, Infertility; Papers 15 to 21 1730 Student Colloquium, G/H Colorado Ballroom; Speaker: Stephen W. Byers : �- 1015 1050 DONOR INSEMINATION 1315 Overview of AID; R. L. Urry ,__ State of the Art Lecture R. P.Michael Neuroendocrine and Behavioral Aspects BREAK 1900 PRESIDENTIAL MESSAGE On behalf of the American Society of Andrology, welcome to our Twelfth Annual Meeting. This year's meeting, held in beautiful Denver, Colorado, commences with a Postgraduate course addressing two important topics: . one physiologic, the neuroendocrinology of male reproduction; the other, socio-legal, the collection and use of reproductive samples. The following three days comprise presentation of 124 scientific papers, four lectures by three distinguished andrologists, and a workshop on animal welfare and animal use in biologic research. This year's Serano lecturer is Roger Short, Professor of Physiology at Monash University, and the State of the Art lecturers are Richard Michael, Professor of Psychiatry at Emory University and Edward Price, Professor of Animal Science at the University of Califor nia at Davis. We hope all will enjoy, learn from, and profit by this broad program. I encourage all attendees to actively participate in the exchange of data and ideas. William D. Odell President, American Society of Andrology AMERICAN SOCIETY OF ANDROLOGY OFFICERS President Vice President Secretary Treasurer Past President William D. Odell Larry L. Ewing Joel L. Marmar Brian H.Vickery Anna Steinberger C. Wayne Bardin Arnold M. Belker Don F. Cameron Anita P. Hoffer Richard Horton William B. Neaves EXECUTIVE COUNCIL James W. Overstreet B.Jane Rogers Lonnie D.Russell Sherman J.Silber Keith D. Smith Rebecca Z. Sokol COMMITTEE CHAIRPERSONS Awards ...................................Terry T. Turner Constitution and Bylaws ....................Anthony V. Boccabella Finance ...................................Arnold M. Belker Future Meetings ...........................Luis J. Rodriguez-Rigau International Liaison ........................Philip Troen Liaison ....................................Balwant Ahluwalia Local Arrangements ........................Rupert P. Amann Membership ...............................Larry Johnson Nominating ..................... , ......., .Claude J.Migeon Postgraduate ..............................Richard Horton Program ..................................Rupert P.Amann Publication ...., ...........................David W. Hamilton Student Affairs ............................Barry T. Hinton Editor, Journal of Andrology .................Marie-Claire Orgebin-Crist P-1 PROGRAM CHAIRMAN Welcome to the 1987 Annual Meeting of the American Society of Andrology. It is a pleasure to have you join us in Colorado. I hope you will take this opportunity to visit with clinicians and scientists located along the Front Range. Your Local Committee has worked diligently to insure that our meeting is unique and rewarding. The Program Committee has strived to develop an outstanding program and assemble the submitted abstracts in logical groups. Hopefully, they will elicit discussion and improve our under standing of both basic and clinical aspects of andrology of man and animals of economic importance, as well as the principles of reproductive biology established using appropriate animal models. The strength of the American Society of Andrology is in bringing together clinicians and scientists with diverse backgrounds so that each may learn from others. I challenge you to approach our Twelfth Annual Meeting with this inquisitive and comparative approach. Only in this way will andrology best serve society. I am thankful to the members of both the Program and Local Committees, the many individuals who helped make the meeting a success, our corporate sponsors and exhibitors, and for the support of the College of Veterinary Medicine and Biomedical Sciences of Colorado State University. Rupert P. Amann, Ph.D. Local and Program Chairman I Animal Reproduction Laboratory Colorado State University Fort Collins, Colorado Program Committee Local Committee Robert D. Atherton Bruce D. Schanbacher Francis R. Tekpetey Rupert P. Amann William Bremner William D. Odell Bruce D. Schanbacher Ronald L. Urry Rupert P. Amann AMERICAN SOCIETY OF ANDROLOGY PAST PRESIDENTS 1975-1977 .................................Emil Steinberger 1977-1978 .... .............................Don W. Fawcett 1978-1979 C. Alvin Paulsen .. . . .. . . 1979-1980 ..............................Nancy).Alexander 1980-1981 . .............................Philip Troen 1981-1982 . .............................Richard M. Harrison 1982-1983 . .............................Richard). Sherins 1983-1984 Andrzej Bartke . . . .. . . .. 1984-1985 Rudi Ansbacher 1985-1986 Anna Steinberger . . . . • . . 2-P . . . . . . . . . . . . . . . . . . . . . . . . . . • . . . . . . . . . . . . • . . . . . . . . . . . . . . . . . . . . . . . . . . . . . • . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . . GENERAL INFORMATION Transportation Taxi (about $8) and Limousine Service (about $5) from Denver's Stapleton Airport to the hotel are available outside the baggage area. The Denver Marriott DOES NOT provide pick-up service. Headquarters Marriott City Center 18th St. and California St. Denver, CO Tel. (303) 297-1300 On-site Registration Conference Center (Lobby, Lower Level Two) Thursday, March 5 4:00 to 7:00 P.M. Friday, March 6 7:15 AM. to 5:00 P.M. Saturday, March 7 Registration Fees Postgraduate Course With CME credit $125.00 Without CME credit $125.00 Students (w/o lunch) $ 30.00 Annual Meeting 7:15 AM. to 5:00 P.M. Sunday, March 8 7:30 AM. to 5:00 P.M. Monday, March 9 7:30 to 10:30 AM. Annual Business Meeting Monday, March 9, 10:50 A.M. Any motions should be sent to the society Secretary prior to the meeting. Non-member $95.00* Regular member $80.00* Student member $45.00* "Includes the $15.00 late registration fee for individuals who did not preregister by mail prior to February 14, 1987. The Press Room is located in the Silverton Room (lower level 1) The Slide Preview Room is the Homestake Room (lower level 1) Sponsors and Supporters of the 1987 Annual Meeting Animal Reproduction Laboratory, College of Veterinary Medicine & Biomedical Sciences, Colorado State University Serono Laboratories, Inc. Beverages during the morning and afternoon breaks are provided through the courtesy of the commerical exhibitors. Please express your thanks to them. Sustaining Members of the Society The following organizations are sustaining members of the American Society of Andrology. The Society is very grateful for their support. Buckeye Urological Associates Syntex Company Knoll Pharmaceutical Company Syva Company National Medical Enterprises, Inc. TAP Pharmaceuticals Ortho Pharmaceutical Corporation The Upjohn Company Serano Laboratories, Inc. West Michigan Reproductive Institute P-3 SERONO AWARD LECTURESHIP DISTINGUISHED ANDROLOGIST AWARD Dr. Roger V. Short, Pro fessor of Reproductive Biol The American Society of ogy at Monash University, Andrology presents its 1987 Clayton, Australia, has a Distinguished Andrologist distinguished career in repro Award to Emil Steinberger, ductive biology. He attended M.D., in recognition of his the University of Bristol vision and deep commitment School of Veterinary Medi to the development of an cine and received his B.V.Sc. and his M.R.C.V.S. in 1954, drology. He was a founder his Ph.D. degree from the of the Am.erican Society of Andrology and served as its .\ University of Cambridge in 1958, and his Sc.D. in 1969. He first president. His contri was an honorary member of the Agricultural Research butions in basic and clinical research have left an indelible Council Unit of Reproductive Physiology and Biochemistry at Cambridge until 1972, \vhen he was appointed Director mark on the discipline and have given direction to the \vork of others. Dr. Steinberger, President of the Texas Institute for Reproductive Medicine and Endocrinology in Houston, and Clinical Professor of Endocrinology, Department of Medicine, at the University of Texas Medical School at Houston, was born in Germany and received his early of the Medical Research Council Unit of Reproductive Biology in Edinburgh, a position he held until leaving for 1982. Dr. Short has published approximately 235 manuscripts in the area of reproductive biology and is Australia in widely respected as a teacher. Together with Professor education in Europe. He emigrated to the United States in C.R. Austin, he edited a series of texts entitled "Reproduc 1948 and obtained an M.Sc. in Anatomy and Endocrinology and an M.D. from the State University of Iowa in 1955. Dr. tion in Mammals," which have been reprinted and trans Steinberger's research has encompassed all cellular com lated into several languages. As Serono Lecturer, Dr. Short will present two lectures on comparative andrology. ponents of the seminiferous tubule, the kinetics of sperma togenesis, the function and interaction of the Leydig and Sertoli cell, genetic, hormonal and metabolic factors influ encing testicular development and function, as \veil as the endocrinology of the testis and the development of male contraceptives. Additionally, he is a dedicated teacher and superb clinician. He has championed two important causes: that treatment of infertility must involve understanding the fertility potential of the couple, and the integration of various disciplines into the area of reproductive medicine. YOUNG ANDROLOGISTS 1982 1983 1984 1985 1986 1987 L.J.D. Zaneveld, D.V.M., Ph.D. William B. Neaves, Ph.D. Lonnie D. Russell, Ph.D. Bruce D. Schanbacher, Ph.D. Stephen J. Winters, M.D. Ilpo T. Huhtaniemi, M.D., Ph.D. YOUNG ANDROLOGIST AWARD Dr. Ilpo T. Huhtaniemi, DISTINGUISHED ANDROLOGISTS 1976 1977 1978 1979 1980 1981 1982 1983 1984 1985 1986 1987 4-P Roy 0. Greep, Ph.D., M.D., Sc.D. M.C. Chang, M.D., Ph.D. Roberto E. Mancini, M.D. Robert J. Hotchkiss, M.D. Thaddeus Mann, M.D., Sc .D., Ph.D. John Macleod, Ph.D. Alexander Albert, Ph.D., M.D. Eugenia Rosemberg, M.D. Kristen B.D. Eik-Nes, M.D. Mortimer B. Lipsett, M.D. Robert H. Foote, Ph.D. Alfred D. Jost, D.Sc. Emil Steinberger, M.D. M.D., Ph.D., was educated in Finland and currently is Professor and Chairman of the Department of Physiol ogy at the University of Turku. His research has cen tered on the endocrine reg ulation of development and function of the fetal, neona""" tal a n d adult testis. This research has resulted in more than 125 papers in refereed journals. He also is an outstanding teacher. In recognition of his elucidation of testicular endocrinology, the American Society of Andrology is pleased to present Ilpo T. Huhta niemi its 1987 Yoyng Andrologist Award. NEW INVESTIGATOR AWARD (Prior to 1986 called the Student Presentation Award) Donated by the West Michigan Reproductive Institute This winner of this award will be announced at the Awards Ceremony, Monday, March 9, 10:20 a.m. Previous Recipients 1983 Thomas T. Tarter 1984 Peter S. Albertson Randall S. Zane 1985 none 1986 Mark A. Hadley STUDENT INFORMATION Student Mixer .., ...., .........Friday, March 6, 6:00 p.m. to 7:00 p.m. in the Matchless Room, lower level 1. Student Colloquium . ......... ..Saturday, March 7, 5:30 p.m. to 6:45 p.m. in G/H Colorado . Ballroom Speaker ... ................Stephen W. Byers . Topic . ...................fo Vitro Studies of the Male Reproductive System (Organized . by the Student Affairs Committee) Student Travel Awards . ........ Five awards of $100 each, authorized for payment from the . general fund by the Exe culive Council, will be presented to five students at the Awards ceremony. Winners were selected by the Awards Committee. Placement Service ............ .Available throughout the meeting at a board near the registra . tion desk. Joanne Killinger will answer questions. SPECIAL EVENTS Opening Reception Friday, March 6, 7:00 p.m. Denver Ballroom on the conference level (LL2) This event is free to all registrants at the Annual Meeting and their guests. Hot and cold treats will be provided and extra beverages (after the first hour) will be available for purchase. Banquet Sunday, March 8, 7:30 p.m. The banquet will be preceded by cocktails, beginning at 7:00 p.m., in the Denver Ballroom (left end). Entertainment will be provided. Special banquet tickets are required for both registrants and their guests. Awards Ceremony Monday, March 9, 10:20 a.m. A brief introduction will be given by President William Odell. Five student travel awards will be made and awards will be presented to the New Investigator, the Young Andrologist and the Distinguished Andrologist. P-5 SPECIAL EVENTS SATURDAY 8:10 A.M. THE SERONO AWARD LECTURE I. The Androgens: Nature, Red in Tooth and Claw Roger V. Short Department of Reproductive Biology Monash University, Clayton, Australia 9:00 AM. STATE OF THE ART LECTURE Neuroendocrine and Behavorial Aspects of Male Reproduction Richard P. Michael Departments of Psychiatry and Anatomy Emory University SUNDAY 1:30 P.M. STATE OF THE ART LECTURE Evolutionary Aspects of Male Sexual Behavior in R. P. Michael Domestic Ungulates, Laboratory Rodents and Primates Edward 0. Price Department of Animal Science University of California at Davis 4:30 P.M. WORKSHOP ON ANIMAL RIGHTS Speaker and Moderator: Bernard E. Rollin Departments of Philosophy and Physiology Director, Bioethical Planning Colorado State University E. 0. Price MONDAY 11:45 A.M. THE SERONO AWARD LECTURE II. The Size of the Testes Roger V. Short Department of Reproductive Biology Monash University, Clayton, Australia B. E. Rollin COMMERCIAL EXHIBITORS As this program goes to press, a complete list of commercial exhibitors is not available. However, an Exhibitors' Program and Buyers Guide will be included in the material distributed to all registrants at the Postgraduate Course or the Annual Meeting. The commercial exhibits are in I/II Denver Ballroom, and will be open Friday through Sunday. 6-P TWELFTH ANNUAL MEETING POSTGRADUATE COURSE IN ANDROLOGY Friday, March 6, 1987 Conference (lower) Level, Marriott City Center, Denver, CO Richard Horton, M.D., Course Director NEUROENDOCRINOLOGY OF MALE REPRODUCTION-Richard Horton, Chairperson, LAC/USC Medical Center, Los Angeles, CA 8:00 Neuroanatomy and Animal Studies-Anthony Plant, University of Pittsburgh Medical School, Pittsburgh, PA 8:50 Pituitary Tumors-Peter Snyder, University of Pennsylvania School of Medicine, Philadelphia, PA 9:40 Break 10:00 Regulation of Gonadotropin in Man Seattle, WA 10:50 Therapy with GnRH and Analogs-William Crowley, Jr., Massachusetts General Hospital, Boston, MA 11:40 General Discussion 12:00 Lunch William Bremner, VA Medical Center, - CURRENT ISSUES WITH DONOR INSEMINATION-Ronald L. Urry, Chairperson, University of Utah School of Medicine, Salt Lake City, UT 1:15 Overview of AID-Ronald L. Urry, University of Utah School of Medicine, Salt Lake City, UT 1:30 Donor Selection-History, Physical and Genetic Screening, and Population Pools Nancy Alexander, Eastern Virginia Medical School, Norfolk, VA 2:00 Sexually Transmitted Disease Screening for AID-Mary E. Guinan, Center for Disease Control, Atlanta, GA 2:45 Break 3:05 Legal and Liability Aspects of AID-David Young, Attorney-at-Law, Salt Lake City, UT 3:50 Screening Couples and Psychological Aspect s-K irtly Jones, University of Utah School of Medicine, Salt Lake City, UT 4:20 Sperm Freezing, Thawing and Preparation for Insemination-Ronald L. Urry, University of Utah School of Medicine, Salt Lake City, UT 4:50 Insemination Techniques, Results of AID, and Other Facets to Consider-Kirtly Jones, University of Utah School of Medicine, Salt Lake City, UT 5:20 General Discussion The School of Continuing Medical Education, University of Utah School of Medicine, has designated this Postgraduate course for 8 credit hours in Category 1 of the Physicians Recogni tion Award of the American Medical Association. P-7 TWELFTH ANNUAL MEETING FRIDAY evening, March 6 6:00 7:00-9:00 ------- Student Mixer in the Matchless Room on lower level ILLI). Opening Reception in the Denver Ballroom. SATURDAY, March 7 8:00 Scientific Session A, E/F Colorado Ballroom Opening and Welcome -Rupert P. Amann, Local/Program Chairperson; William D. Odell, 8:10 The Serono Lecture I. 9:00 THE ANDROGENS: NATURE, RED IN TOOTH AND CLAW State of the Art Lecture President Roger V. Short, Monash University, Clayton, Australia- Richard P. Michael, Emory University, Atlanta, GA NEUROENDOCRINE AND BEHAVIORAL ASPECTS OF MALE REPRODUCTION 9:50 Break-coffee and soda in the central lobby and exhibition area Scientific Presentation Session B, ENDOCRINOLOGY AND SERTOLI CELLS Chairpersons: S.J. Winters and L.D. Russell 10:15 1 10:30 2 10:45 3 11:00 4 11:15 5 11:30 6 11:45 7 Estradiol (E,), but not testosterone (T), allows for the development ofLHRH self-priming in the adult male rat. P.M. Grosser,* Z. Xue' and B. Robaire. Role of testosterone metabolites in the maintenance of the self-priming effect of LHRH in pubertal male rats. S.J. Nazian. Photoperiodic maintenance of the ram testis during exposure to contrasting stimulatory and inhibitory daylengths. Bruce Schanbacher. Purification and comparative structural analysis of a FSH and testosterone responsive glycoprotein from rat and mouse primary Sertoli cell culture media. C. Yen Cheng, Josephine Grima, Kenneth S .K. Tung and C. Wayne Bardin. Germ cells bind testicular "Fe-transferrin following the transferrin mediated transport of ••Fe across Sertoli cells. Daniel Djakiew and Martin Dym. Germ cells influence the polarized secretion of androgen binding protein (ABP), but not transferrin (TRF) by Sertoli cells in vitro. Andrzej Janecki,* Andrzej Jakubowiak' and Anna Steinberger. 12:00-1:30 Effect of age on composition of human seminiferous tubule boundary tissue and on the volume of each component. J.G. Abdo,* L. Johnson and W.B. Neaves. Lunch-on your own Scientific Presentation Session C, SPERM EVALUATION Chairpersons: A.M. Belker and R.H. Hammerstedt 1:30 8 Relationship of human sperm DNA stability and fertility potential. J.E. Sokoloski, M.M. Quigley' and A.J. Thomas 1:45 9 2:00 10 Studies on the function and regulation of sulfhydryls in caput epididymal sperm induced to acquire progressive motility in vitro. G. Cornwall, S. Tillman' and T.S.K. Chang. Automated semen analysis: reproducibility studies. Kenneth A. Ginsburg' and Ernest L. 2:15 11 Comparison of fully automated and traditional methods of semen analysis. D. Vantman, M. 2:30 12 Analysis of prevasectomy ejaculates by the hypoosmotic swelling (HOS) test. L.J.D. Zaneveld, Abel.' Zinaman and R. Sherins. R.S. Jeyendran, M. P.P. de Castro and P.J.M. Silveira.' . ty of Andrology. *Not member of the American Socie 8 P - SATURDAY, March 7 2:45 13 3:00 14 Relationship between the hypoosmotic swelling (HOS) test and the sperm penetration assay (SPA). C. Bastias,' T. Thompson,* A. Buck,' M. Hinson' and B.J. Rogers. Human sperm acrosin as a fertility marker. H.H. Van der Ven,' W.P. Kennedy,* J.M. Kaminski,' R.S. Jeyendran and L.J.D. Zaneveld. Break-coffee and soda in the centrnl lobby and exhibition area 3:15 Scientific Presentation Session D, INFERTILITY Chairpersons: N.J. Alexander and K.D. Smith 3:30 3:45 15 16 Lymphopenia in male partners of infertile couples. Allan R. Glass. Effects of soluble products of activated lymphocytes and macrophages (lymphokines and monokines) on human sperm motility. Joseph A. Hill,* Florina Haimovici' and Deborah J. 4:00 17 Evidence for antibody binding specificity to donor sperm. Stephen E. Howe and Dona M. 4:15 18 Inability of sperm washing to remove antisperm antibodies. Gilbert G. Haas, Jr., Osmond 4:30 19 4:45 20 A comparison of !VF cycles with failed versus successful fertilization: significance of cycle, semen and patient variables. S.P.Boyers, J.B. Russell,• J.N. Stronk,• L. Fino,' E. Jones' and 5:00 21 Gamete intra-fallopian tube transfer (G.I.F.T.). Influence of the male factor on the success rate. Luis J. Rodriguez-Rigau, George M. Grunert, Rell M. Woodward, Eberhard C. Lotze, Anderson. Lynch. D'Cruz' and Fayez Nahhas.• Clomiphene and tamoxifen in oligospermic infertility. W.S. Maxson, B.J. Rogers, C.M. Herbert, C. Markuson and F.K. Kirchner, Jr. A.H. DeCherney. Joseph R. Feste, William Gibbons, Keith D. Smith and Emil Steinberger. 5:30 Student Colloquium-G/H Colorado Ballroom Speaker Stephen W. Byers, IN VITRO STUDIES OF THE MALE REPRODUCTIVE SYSTEM - Scientific Presentation Session E, Denver Ballroom 7:00-9:30 Papers 22 to 61 and wine and cheese. Presenters of posters should ensure that their poster is in place by 1:30 p.m. to allow casual viewing during the afternoon and should be at their poster during the evening session. Posters must be removed by 10:00 p.m. 22 Effects of prenatal irradiation on testicular LH receptors in adult Syrian hamsters. A.G. Amador, A. Bartke, V. Chandrashekar' and H.G. Klemcke.' 23 24 25 Changes in serum testosterone and luteinizing hormone levels in male rats with adjuvant-in duced arthritis. Jeffrey W. Clemens' and Brent C. Bruot.' The effects of castration and testosterone on plasminogen activator activities in rat ventral prostate. M. Wilson, C. Ludowese, A. Sinha and R. Estensen. Effect of seasonal changes in Leydig cell (LC) number on the volume of LC smooth endoplas mic reticulum (SER) and intratesticular testosterone content (IT). Larry Johnson and D.L. Thompson, Jr. 26 27 28 Influence of 35°C and melatonin (MEL) on testosterone (T), testis and accessory sex organs (ASO) after HCG in juvenile deer mice. Pat Fail. Computerized analysis of sperm motion: effect of stage temperature and storage at room temperature and 4°C. AB. Stockwell,• P.J. Burns and D. Douglas-Hamilton.' Validation of a computerized system for evaluating motility of bull sperm. P.R. Budworth,• R.P. Amann and P.L. Chapman.' 29 Correlations between fertility and computer-determined swimming parameters of bull sperm. P.R. Budworth' and R.P. Amann. *Not member of the American Society of Andrology. P-9 SATURDAY, March 7 30 Automated semen analysis in large epidemiologic studies. Frank DeStefano,' Joseph L. 31 32 Assessing human sperm velocity in semen and during the capacitation process using auto mated semen analysis. G. Koukoulis,' D. Vantman, M. Zinaman and R. Sherins. Dissociation of hypo-osmotic tail swelling patterns in sperm from fertile and infertile men. 33 Differential evaluation of HOS test minimizes false positive fertility results. R. Basuray, S. 34 35 Employment of the hypoosmotic swelling (HOS) test to assess the functional integrity of equine sperm membrane. Panayiotis M. Zavos and Gary W. Gregory.* The P and G pattern in patients with a normal sperm morphology< 14%. T.F. Kruger,• K.F. 36 A comparison of morphometry and volume of human spermatozoa. T. Turner, S. Schrader, Annest,' Marcie-jo Kresnow,' Melinda L. Flock and Steven M. Schrader. D. Vantman, M. Zinaman and R. Sherins. Tarchala, H. Van der Ven,' M. Perez-Pelaez and R. Jeyendran. Simmons* and A.A. Acosta. 37 38 R.S. Jeyendran., M. Perez-Pelaez, R.F. Karuhn and H.H. Van der Ven.* A new method of evaluating sperm morphology with predictive value for human IVF. T.F. Kruger,' A.A. Acosta, K.F. Simmons,' R.J. Swanson, J.F. Matta,' LL. Veeck,' M. Morshedi* and S. Brugo. Usefulness of scanning and transmission electron microscopy in clinical andrology. Paul W. Musselman, Anthony J. Thomas, Jr.,' James T. McMahon and Howard S. Levin.* 39 40 Paper withdrawn The role of sperm morphology evaluated by strict criteria in the hamster ova/human sperm penetration assay (SPA). R. James Swanson, Thinus F. Kruger,' Mary Hamilton,' Kathy F. 41 The relationship between sperm fertilizing capacity and the penetration of zona-free ham ster ova by frozen-thawed boar spermatozoa. R.N. Clarke, T. Almlid,* V.G. Pursel* and L.A. Simmons* and Anibal A. Acosta. Johnson.* 42 43 Penetration of hamster ova by non-human primate spermatozoa. Barbara S. Durrant. Potential utility of rabbit salt-stored zona penetration assay. R.A. Fayrer-Hosken and B.G. 44 Value of ATP assay in predicting the outcome of in vitro fertilization (IVF) trials and sperm hamster ova penetration assay (SPA). M. Morshedi,* M. Hamilton,' and P. Pleban,' A. Acosta, 45 Use of Penetrak to study sperm penetration patterns. Richard M. Harrison and Ronald W. 46 Effects of cervical mucus shearing on the biomechanics of sperm penetration. D. Katz, R. Brackett. T. Kruger' and R.J. Swanson. Lewis. 47 Happ,' A. Yudin,* J. Overstreet, F. Hanson* and J. Chang.* Predictive value of quantitative and qualitative bacteriologic studies in zona-free hamster egg penetration test failure. Laszlo Sogor, Hunter Hammill,' Daina Glavan* and Arlette Coulter.* 49 Comparison of spermatid and epididymal sperm analyses with testis and epididymal histo pathology in compound evaluation. S.L. Wilczynski, L.D. Russell and J.M. Killinger. New observations on the morphological features of the guinea pig sperm acrosome reaction. 50 Analysis of sperm fertilizing ability in the stallion. E. Bustos-Obregon, Hector Rodriguez* 51 Ultrastructural-physiologic correlates of human sperm egg penetrating ability. R. A. Bronson, 48 S.P. Flaherty,* V.P. Winfrey* and G.E. Olson. and Sylvia Leiva.* G . W. Cooper• and D. Phillips. 52 Regional differences in androgen binding by ram epididymal tissue. F .R. Tekpetey and R.P. Amann. 53 The effect of varying FSH pulses on transferrin secretion in superfused Sertoli cell cultures. Andrzej Jakubowiak, * Andrzej Janecki' and Anna Steinberger. *Not member of the American Society of Andrology. 10-P SATURDAY, March 7 54 55 Neutral amino acid absorption by rat epididymis. B.T. Hinton and H. Hernandez. Direct and indirect immunologic response following acute testicular torsion. Johnny B. Roy, 56 57 Incorporation of ("C)gossypol and formation of ("C)gossypol-conjugates in mouse trans formed Sertoli, TM4, cells. Nongnuj Tanphaichitr' and Lisa Fitzgerald.• Spectral analysis of short-term fluctuations in salivary testosterone concentrations. Norma 58 59 Doppler evaluation of varicocele. C.B. Dhabuwala and Anil Kumar.• Is sexual dysfunction of hormonal origin in chronic alcoholic males? J.P. Wincze,' M. James Mays,' Gilbert G. Haas, Sharon Einfeldt,' Brian Lansford' and Fayez Nahhas.' F. Besch, James M. Dabbs, Jr.• and Charles H. Hopper.• Engle-Friedman,' T. Nirenberg' and M.R. Leipman.' 60 Effects of cyclosporine on testicular structure and function. L. Seethalakshmi, D.A. Diamond 61 The clinical features of spermatogenic failure. Anne M. jequier and S.C. Holmes. and M. Menon. SUNDAY, March 8 -------- Scientific Presentation Session F, CLINICAL ANDROLOGY Chairpersons: A.R. Glass and R.Z. Sokol 8:00 62 Physiological attributes of episodic follicle stimulating hormone secretion in normal men. J.D. Veldhuis, J.C. King,' R.J. Urban,• AD. Rogol and M.L. Johnson.' 8:15 63 8:30 64 8:45 65 Effects of chronic endurance exercise training on male reproductive hormonal profiles. A.C. 9:00 66 9:15 67 The role of sexual stimulation and ejaculate characteristics during the clinical use of a seminal collection device (SCD). Panayiotis M. Zavos. Experience with needle biopsy of the testis as an office procedure. R.A. Appell, j.S. Deutsch,' 9:45 68 9:45 69 Effect of altering sex steroid levels with human chorionic gonadotropin or testolactone on sex hormone binding globulin level. Richard V. Clark, James F. Dunn' and Richard). Sherins. Subnormal prolactin (PRL) response to thyrotropin-releasing-hormone (TRH) does not necessarily support the diagnosis of hypogonadotropic hypogonadism. Duarte Pignatelli, Davide Carvalho' and Manuel Hargreaves.' Hackney' and W.E. Sinning.' j.R. Goodman' and j.N. Macaluso.' Induction of acrosome reactions by the human zona pellucida. N.L. Cross,' P. Morales,• j.W. Overstreet and F.W. Hanson.' Factors affecting microinjection of mammalian spermatozoa into hamster eggs. R.N. Clarke and L.A. Johnson.' Break-coffee and soda in the central lobby and exhibition area 10:00 Scientific Presentation Session G, TOXICOLOGY AND AGEING Chairpersons: B. Robaire and j.L. Fourcroy 10:15 70 Semen characteristics of "unexposed workers." A comparison of five worker populations. S.M. Schrader, T.W. Turner, j.R. Burg and J.M. Ratcliffe. 10:30 71 10:45 72 11:00 73 11:15 74 Increased degeneration of germ cells late in meiosis and no loss during spermiogenesis in aged men. Ali Bagheri' and Larry Johnson. Effects of ethanol on the acrosome reaction in human ejaculated spermatozoa. Juan G. Alvarez,' J.C. Touchstone,' Isabel M. Lopez' and Bayard T. Storey. Effect of liver damage, alcohol, and recreational drugs on seminiferous tubules and Leydig cells in men. Tho Q. Nguyen,• L. Johnson, Robert E. Wolf,• Ali Bagheri,• Michael E. Bailey' and W.B. Neaves. CNS and testicular sensitivity to ethanol as a function of pubertal development. R.A. Anderson, J.F. Phillips' and S.H. Berryman.' •Not member of the American Society of Andrology, P-11 SUNDAY, March 8 11:30 75 11:45 76 The effect of dose, duration and age of exposure on the expression of lead toxicity on the hypothalamic-pituitary-testicular axis. R.Z. Sokol. Potentiation of gonadotoxicity of cyclophosphamide in the dog by adjuvant treatment with an LHRH agonist. J.C. Goodpasture, K. Bergstrom•and B.H. Vickery. 12:00-1:30 Lunch-on your own 1:30 Scientific Session H, E/F Colorado Ballroom State of the Art Lecture Edward 0. Price, University of California, Davis, CA EVOLUTIONARY ASPECTS OF MALE SEXUAL BEHAVIOR IN DOMESTIC UNGU LATES, LABORATORY RODENTS, AND PRIMATES Scientific Poster Presentation Session I, Denver Ballroom Papers 77 to 116 and coffee and soda 2:30-4:30 Presentors of posters should ensure that their poster is in place by 8:00 a.m. to allow casual viewing during the morning and mid-day and should be at their poster during the afternoon session. Posters must be removed promptly at 4:30 p.m. 77 The effectiveness of an insulated jar in protecting semen quality from temperature extremes. 78 Is nocturnal penile tumescence (NPT) a test of pituitary-testicular function? F.T. Murray, M. 79 80 Hydroflex penile prosthesis: early experience and recommendations. H. David Mitcheson. Penile plethysmography on impotent men using vacuum-constrictor devices. Joel L. Marmar, 81 Testicular biopsy in the evaluation of male infertility. Miguel Diaz, Fernando Rivas, Lidia G.E. Peterson, S.M. Schrader and T.W. Turner. Sciadini• and H. Wyss.' Thomas J. DeBenedictis and Donald E. Praiss. Garcia, Horacio Rivera and Jose Maria Cantu. 82 Characterization and quantitation of white blood cell subsets in human semen. Hans Wolff,• Heidi M.P. Faris: Joseph A. Hill: Joseph A. Politch•and Deborah J. Anderson. 83 Influence of macroagglutinating (MA), microagglutinating (MI) and immobilizing (I) anti sperm antibodies on the hamster ova/human sperm penetration assay (SPA). Mary Hamil 84 85 ton: Anne Bogeart: Mahmood Morshedi,' Steven Ackerman and R. James Swanson. An in vivo technique for screening immunologic factors in the etiology of the unexplained poor post-coital test. Jerome H. Check and Chung H. Wu. Association of sperm agglutination and immobilization tests with immunobead (!MB) assay. M. Morshedi,' S. Ackerman, A. Bogeart,' A. Acosta and R.J. Swanson. 86 Comparison of the sperm penetration assay (SPA) with objective motility parameters. LR. Kossoy,' T. Thompson: A. Buck,' M. Hinson,' B. Brodie: W. Vaughn,' A.C. Wentz•and B.J. Rogers. 87 Intracornual semen deposition at different depths and subsequent fertilization and embry onic development in superovulated cows. John Fernandez-VanCleve: Panayiotis M. Zavos 88 Experience with intrauterine insemination in infertile couples. Christine L. Cook,•Leigh T. 89 Effect of neuraminidase on sperm separation for sex selection. Hugh C. Hensleigh• and 90 Increased rate of human sperm fertilization ability by Percoll-gradient centrifugation. N. 91 Ability of glass wool filtration to isolate fertile sperm fractions. W.J. Holmgren, R.S. Jeyen and Roger W. Hemken.' Price•and Arnold M. Belker. Denise Truzinski. * Tanphaichitr,•A. Agulnick, D. Diaz, J. Hill, L. Fitzgerald' and D. Anderson. dran, M.R. Neff, M. Perez-Pelaez. 92 Sperm preparation for artificial insemination using rayon filters. Hugh C. Hensleigh' and Denise Truzinski.* �Not member of the American Society of Andrology. 12-P SUNDAY, March 8 93 --- ATP decline of infertile patient semen after swim-up separation method (SSM). M. Mor shedi,' R. Acosta,* P. Pleban,* A.A. Acosta, J. Yuan' and R.J. Swanson. 94 Polylactosamine on the mouse sperm surface during maturation and capacitation. Charles H. Muller. 95 Capacitation of bovine sperm by removal of cholesterol. Eduardo Ehrenwald,' John E. Parks• and Robert H. Foote. 96 Interactions between selected fatty acids and the activation of palmitic acid in spermatozoa. Robert E. Jones and Stephen R. Plymale. 97 98 Changes in cytosolic protein and phospholipase activity during in vitro capacitation of sperm from fertile and subfertile bulls. Margaret Henault, Gary Killian and David Chapman.' Glycosidase activities in bovine sperm during in vitro capacitation and the acrosome reaction. 99 A new method for obtaining a high percentage of viable, capacitated sperm. Christopher De 100 101 Immunological comparisons between the boar and human proacrosin-acrosin proteinase systems. Mark S. Siegel,' Dana S. Bechtold,' Janet L. Willand' and Ke.�neth L. Polakoski.' Inhibitory effect of zinc on protein phosphorylation in the sperm head membranes in spisufa 102 Oxygen consumption rate varies with the concentration of bull spermatozoa. Carole 103 Movement characteristics of cynomolgus monkey sperm recovered from the cervix and uterus. E. Behboodi,' D. Katz, J. Overstreet and A. Hendrickx.' Pulsatile LHRH-treatment in male patients with severe oligospermia and selectively ele vated FSH, W. Aulitzky,' J. Frick,' F. Hadziselimovic. Identification of a progesterone receptor in human testis. Stephen J. Winters, Hugh S. Tamara McNutt,' Gary Killian and David Chapman.• Jonge, S.R. Mack and L.J.D. Zaneveld. sodidissima. Balwant Ahluwalia and George Nolan.' Wegner,* Gary Killian and David Chapman.' 104 105 Keeping and Philip Troen. 106 107 108 Effects of diethylstilbestrol (DES) on testicular LH receptors in Fisher 344 rats. A.G. Ama dor, A.I. Esquifino,' A. Bartke, V. Chandrashekar,' J.J. Fernandez-Ruiz' and R.W. Steger.' Leydig cell function in the cryptorchid rat. Tom 0. Abney. 109 Ethinyl estradiol inhibition of testosterone production in the LH treated hypophysectomized rat. R.B. Myers' and T.O. Abney. GnRH increases specific precopulatory responses in testosterone treated pony geldings. 110 Microsurgical accessory gland ablation: The effects on fertility potential in the rat. William 111 Protein contributions of the accessory organs to the composition of human seminal plasma as dete.rmined by high resolution two-dimensional electrophoresis. Edward E. Gaunt, Anibal A. 112 114 Characteristics of the accessory glands of the male reproductive tract of some native Califor nia rodents. B. Peitz and S. Membreno.• A rat model of fetal epididymal and gonadal development and its similarity to clinical abnormalities, J.L. Fourcroy, M.C. Shen, G.P. Riordan and D.W. Warren. Cloning and sequence analysis of human testis-specific lactate dehydrogenase C., Erwin 11.9 Evidence for polymorphic sperm antigen expression in inbred mice. Chong Xu' and Deborah Nancy Diehl,' Sue McDonnell and Marolo Garcia.' Blank, Eric Yousha' and Marc Goldstein. Acosta, Steven B. Ackerman, Patricia A. Pleban, • John F. Stecker and James H. Yuan.• 113 Goldberg, Catherine E. Driscoll' and Jose L. Millan.• J. Anderson. 116 Chronic silicone implants in the rat vas deferens. Donald P. Waller, Micha·el Szarley,' Annamarie Martin,' Cyrus L. Fakroddin,' Glen K. Adaniya,* Jeanne M. Quigg,* Andrejs R. Nikurs and Lourens J.D. Zaneveld. �Not member of the American Society of Andrology. P-13 4:30-6:15 Scientific Session J, E/F Colorado Ballroom WORKSHOP ON ANIMAL RIGHTS Bernard E. Rollin, Colorado State University, Ft. Collins, CO COCKTAILS AND BANQUET, Denver Ballroom 7:00-7:30 7:30-9:30 Cocktails (cash bar) Banque/ MONDAY, March 9 ------- Scientific Presentation Session K, EPIDIDYMIS Chairpersons: J.L. Marmar and M.-C. Orgebin-Crist. Androgen movement into the rat seminiferous and epididymal tubules perifused in vivo. T.T. 8:00 117 8:15 118 Protein secretion by rat epididymal epithelial cells growing in bicameral culture chambers. 8:30 119 Maturation-related glycoproteins in the mouse epididymis. Hwan-Wun Liu* and Charles H. 8:45 120 Correlations between changes in rat sperm membrane lipid composition and the tempera ture-dependence of the membrane physical state during epididymal maturation. Joseph C. 9:00 121 The role of superoxide dismutase in preventing lipid peroxidation and loss of motility in human ejaculated spermatozoa. Juan G. Alvarez,• J.C. Touchstone,* Luis Blasco and Bayard 9:15 122 9:30 9:45 123 124 Patency and pregnancy rates following microsurgical epididymovasostomy for epididymov asal obstruction. Charles 0. Turk' and Lawrence S. Ross. Results of specific tubule microsurgical vasoepididymostomy. Sherman J. Silber. Pregnancy after vasovasostomy. Sherman J. Silber. Turner. S.W. Byers. Muller. Hall' and Laura M. Edgerly.• T. Storey. 10:00 Break 10:20 Special Session, E/F Colorado Ballroom Awards Ceremony William D. Odell, presiding - The New Investigator Award Student Travel Avvards The Young Andrologist Award Ilpo T. Huhtanie:mi, M.D., Ph.D., Helsinki, Finland The Distinguished Andrologist Award Emil Steinberger, M.D., Houston, TX 10:50 Annual Business Meeting of the American Society of Andrology William D. Odell, 11:45 presiding Scientific Session L, E/F Colorado Ballroom The Serono Lecture II. Roger V. Short, Monash University, Clayton, Australia THE SIZE OF THE TESTES 12:45 Adjourn Meeting •Not member of the American Society of Andrology. 14-P f INDEX TO AUTHORS O F ABSTRACTS A Abdo,).G., 7 Abel,Ernest L., 10 Abney, Tom 0., 107, 108 Ackerman, Steven B., 83, 85,111 Acosta, Anibal A., 35, 37, 40, 44, 85, 93, 111 Acosta, R., 93 Adaniya,Glen K., 116 Agu !nick, A., 90 Ahluwalia, Balwant, 101 Almlid, T ., 41 Alvarez, Juan G., 72, 121 Amador, A.G., 22, 106 Amann, R.P., 28, 29, 52 Anderson, Deborah J., 16, 82, 90, 115 Anderson, R. A., 74 Annest, Joseph L., 30 Appel\, R. A., 67 Aulitzky, W., 104 B Bagheri, Ali, 71, 73 Bailey, MichaelE., 73 Bardin, C. Wayne, 4 Bartke, A., 22, 106 Bastias, C., 13 Basuray, R., 33 Bechtold, Dana S., 100 Behboodi, E., 103 Helker, Arnold M., 88 Bergstrom, K., 76 Berryman, S. H., 74 Besch, NormaF., 57 Blank, William, 110 Blasco, Luis, 121 Bogeart, Anne, 83, 85 Boyers, S. P., 20 Brackett, B. G., 43 Brodie, B., 86 Bronson, R. A., Sl Brugo, S., 37 Bruot, Brent C., 23 Buck, A., 13, 86 Budworth,P. R., 28, 29 Burg, J. R., 70 Burns,P. J., 27 Bustos-Obreg6n,E., SO Byers, S. W., 118 c Cantll, Jose Marfa, 81 Carvalho, Davide, 64 Chandrashekar, V., 22, 106 Chang, J., 46 Chang, T. S. K., 9 Chapman, David, 97, 98, 102 Chapman,P. L, 28 Check, Jerome H., 84 Cheng, C. Yan, 4 Clark, Richard V., 63 Clarke, R. N., 41, 69 Clemens, Jeffrey W., 23 Cook, Christine L, 88 Cooper,G. W., Sl Cornwall,G., 9 Coulter, Arlette, 47 Cross, N. L., 68 D D'Cruz,Osmond, 1 8 Dabbs, James M., Jr., S7 De Castro, M.P. P., 12 De Jonge, Christopher, 99 DeBenedictis, Thomas J., 80 DeCherney, A. H., 20 DeStefano,Frank, 30 Deutsch, J. S., 67 Dhabuv.'ala, C. B., S8 Diakiew, Daniel, S Diamond, D. A., 60 Diaz, D., 90 oraz, Miguel, 81 Diehl, Nancy, 109 Douglas- Hamilton, D., 27 Driscoll, CatherineE., 114 Dunn, JamesF., 63 Durrant, Barbara S., 42 Dym, Martin, S E Edgerly, Laura M., 120 Ehrenw·a!d,Eduardo, 9S Einfeldt, Sharon, SS Engle-Friedman, M., S9 Esquifino, A. 1., 106 Estensen, R., 24 f Fail,Pat, 26 Fakroddin, Cyrus L., 116 Faris, Heidi M.P., 82 Fayrer- Hosken, R. A., 43 Fernandez- Ruiz, J. J., 106 Fernandez- VanCleve, John, 87 Feste, Joseph R., 21 Fino, L., 20 Fitzgerald, Lisa, S6, 90 Flaherty, S.P., 49 Flock, Melinda L., 30 Foote, Robert H., 9S Fourcroy, J. L., 113 Frick, J., 104 G Garcia, Lidia, 81 Garcfa, Marolo, 109 Gaunt,EdwardE., 111 Gibbons, William, 21 Ginsburg, Kenneth A., 10 Glass, Allan R., lS Glavan, Doina, 47 Goldberg, Envin, 114 Goldstein, Marc, 110 Goodman, J. R., 67 Goodpasture, J. C., 76 Gregory, Gary W., 34 Grima, Josephine, 4 Grosser, P. M., 1 Grunert,George M., 21 H Haas,Gilbert G., Jr., 18, SS Hackney, A. C., 6S Hadziselimovic,F., 104 Haimovici,Florina, 16 Hall, Joseph C., 120 Hamilton, Mary, 40, 44, 83 Hammill, Hunter, 47 Hanson,F. W., 46, 68 Happ, R., 46 Hargreaves, Manuel, 64 Harrison, Richard M., 4S Hemken, Roger W ., 87 Henault, Margaret, 97 Hendrickx, A., 103 Hensleigh, Hugh C., 89, 92 Herbert, C. M., 19 Hernandez, H., S4 Hill, Joseph A., 16, 82, 90 Hinson, M., 13, 86 Hinton, B. T., S4 Holmes, S. C., 61 Holmgren, W. J., 91 Hopper, Charles H., S7 Howe, StephenE., 17 Jakubowiak, Andrzej, 6, S3 Janecki, Andrzej, 6, S3 Jequier, Anne M., 61 Jeyendran,R. S., 12,14, 33,36,91 Johnson, L. A., 41, 69 Johnson, Larry, 7, 25, 71, 73 Johnson, M. L., 62 Jones, E., 2 0 Jones, RobertE . , 9 6 K Kaminski, J. M., 14 Karuhn, R.F., 36 Katz, D., 46, 103 Keeping, Hugh S., 105 Kennedy, W.P., 14 Killian,Gary, 97, 98, 102 Killinger, J. M., 48 King, J. C., 62 Kirchner,F. K., Jr., 19 Klemcke, H.G., 22 Kossoy, L. R., 86 Koukoulis,G., 31 Kresnow, Marde·jo, 30 Kruger, ThinusF., 3S, 37, 40, 44 Kumar, Anil, 58 L Lansford, Brian, SS Leipmqn, M. R., 59 Leiva, Sylvia, 50 Levin, Howard S., 38 Lewis, Ronald W., 45 Liu, Hwan-Wun, 119 Lopez, Isabel M., 72 Lotze, Eberhard C., 21 Ludowese, C., 24 Lynch, Dona M., 17 M Macaluso, J. N., Jr., 67 Mack, S. R., 99 Markuson, C., 19 Marmar, Joel L., 80 Martin, Annamarie, 116 Matta, J.F., 37 Maxson, W. S., 19 Mays, James, SS McDonnell, Sue, 109 McMahon, James T., 38 McNutt, Tamara, 98 Membreno, S., 112 Menon, M., 60 Millan, Jose L., 114 Mitcheson, H. David, 79 Morales,P., 68 Morshedi, Mahmood, 37, 44, 83, 85, 93 Muller, Charles H., 94, 119 Murray,F. T., 78 Musselman,Paul W., 38 Myers, R. B., 108 N Nahhas,Fayez, 18, S5 Nazian, S. J., 2 Neaves, W. B., 7, 73 Neff, M. R., 91 Nguyen, Tho Q., 73 Nikurs, Andrejs R., 116 Nirenberg, T ., 59 Nolan, George, 101 0 Olson, G. E., 49 Overstreet, J. W., 46, 68, 103 p Parks, JohnE., 9S Peitz, B., 112 Perez-Pelaez, M., 33, 36, 91 Peterson,G.E., 77 Phillips, D., 51 Phillips, J.F., 74 Pignatelli, Duarte, 64 Pleban,Patricia A., 44, 93, 111 Plymale, Stephen R., 96 Polakoski, Kenneth L., 100 Politch, Joseph A., 82 Praiss, Donald E., 80 Price, Leigh T., 88 Pursel, V. G., 4 1 Q Quigg, Jeanne M., 116 Quigley, M. M., 8 R Ratcliffe, J. M., 70 Riordan, G. P., 113 Rivas, Fernando, 81 Rivera, Horacio, 81 Robaire, B., 1 Rodr(guez, Hector, 50 Rodriguez- Rigau, Luis J., 21 Rogers, B. J., 13, 19, 86 Rogol, A. D., 62 Ross, Lawrence S., 122 Roy, Johnny B., SS Russell, J. B., 20 Russell, L. D., 48 s Schanbacher, Bruce, 3 Schrader, Steven M., 30, 36, 70, 77 Sciadini, M., 7 8 Seethalakshmi, L., 6 0 Shen, M . C., 113 Sherins, Richard}., 11, 31,32,63 Siegel, Mark S., 100 Silber, Sherman J., 123, 124 Silveira,P. J. M., 12 Simmons, KathyF., JS, 37, 40 Sinha, A., 24 Sinning, W. E., 65 Smith, Keith D., 21 Sogor, Laszlo, 47 Sokol, R. Z., 75 P-15 Sokoloski, J.E., 8 Stecker, John F., 111 Steger, R. W., 106 Steinberger, Anna, 6, 53 Steinberger,Emil, 21 Stockwell, A. B., 27 Storey, Bayard T., 72, 121 Stronk, J. N., 20 S\vanson, R. James, 37, 40, 44, Tillman, S., 9 Touchstone, J.C., 72, 121 Troen, Philip, 105 Truzinski, Denise, 89, 92 Tung, Kenneth S. K., 4 Turk, Charles 0., 122 Turner., T. T., 117 Turner, T. W., 36, 70, 77 83, 85, 93 Szarley, Michael, 116 Urban, R. J., 62 u T Tanphaichitr, Nongnuj, 56, 9 0 T archala, S., 33 Tekpetey, F. R., 52 Thomas, Anthony J., Jr., 8, 38 Thompson, D. L., Jr., 25 Thompson, T., 13, 86 16-P v Van der Yen, H. H., 14, 33, 36 Vantman, D., 11, 31, 32 Vaughn, W ., 86 Veeck, L. L, 37 Veldhuis, J. D., 62 Vickery, B. H., 76 w Waller, DonaldP., 116 Warren, D. W., 113 Wegner, Carole, 102 Wentz, A.C., 86 Wilczynski, S. L., 48 Willand, Janet L., 100 Wilson, M., 24 Wincze, J.P., 59 Winfrey, V.P., 49 Winters, Stephen J., 105 Wolf, Robert E., 73 Wolff, Hans, 82 Woodward, Rell M., 21 Wu,Chung H., 84 Wyss, H., 78 x Xu,Chong., 115 Xue, Z., 1 y Yousha,Eric, 110 Yuan, James H., 93, 111 Yudin, A., 46 z Zaneveld, Lourens J. D., 12, 14, 99, 116 Zavos, Panayiotis M., 34, 66, 87 Zinaman, M., 11, 31, 32 ABSTRACTS ES'TRADIOL {Et ) , BlJl' 001' IESltsIEBl'./E ( T ) , m,a.'S FOO RAT. nm DEVELOFMElIT OF UffiH SELF-PRIMING IN nm AOOLT HALE P.M. Grosser" , Z . Xueu and B . Roba.ire3 • Centre for the Sti.rly of Reprcd.uction and Depts. of PhaTill/Therap and Ob/Gyn , !'�ill University, Montrea l , Canada.I• l and Dept. of Urology, University of Beiging , Beiging, Peoples Republic of China2 • T and E2 1 administered simultaneously to adult uale rats, synergize to suppress spermatogenesi s . Although exploited for the developnent of a male contraceptive, the mechanism responsible for this steroid interaction at the hypothala.mo pituitary complex is not known , 'Ihe present sti.rlies examined the effects of T- ( 2 . 5 cm) and E2-filled ( 0 . 1 cm) subdermal Silastic implants , alone or in combination, on pituitary lll responsiveness to UIRH and LHRl-I self-priming . Male SD rats (300-325 g) 1•ere equipped with jugular catheters . Bloo:l ssmples for RIA of plasma LH were obtained every 10 min for 7 hrs, 1 day before and 1 , 3 , 7 and 14 days after giving T, LHRH {600 ngjkg ) l<aS given at 90 min (pituitary LHRH responsiveness ) and at 240 and 270 min (LllRH self-priming) after initiation of sampling. Within 14 days , T treatment reduced LI-I response to LHRH by more than 70%; LHRH self-priming (ratio of 3rd to 2nd LI-I peak) was reduced from 133% i n controls to 108%. E2 alone reduced pituitary responsiveness by less than 20%. HQl..oever , LHRH self-priming was increased markedly from 120% in controls to 201% by day 14, The combination of T and E1 \.-ere additive with respect to pituitary responsiveness. In contrast , the self-priming E1 or T+E2 implants. response \.� 'Ihus T and Et greater than with E1 alone (338% by day 14), act differently on the hypothalwno-pituitary complex; T suppresses pituitary responsiveness while E1 primarily sensitizes the pituitary to IlffiH self-priming . 'These complementary actions ll'aY account, in pa.rt, for the synergistic action of T and E2 , Supported by MRC and FCAC , 2 ROLE OF TESTOSTERONE METABOLITES Ill THE HAINTEllANCE OF THE SELF-PRIMING EFFECT OF LHRH Ill PUBERTAL MALE RATS. S . J . Nazian, Department o f Physiology & Biophysics, College of Medicine, University of South Florida, Tampa, FL 33612. Pubertal male rats release roor e LH in response to LHRU if priced with sc.all doses of LHRH than if pretreated with saline. Castration eliminates this self-priming effec t . Treatment of castrated rats with Silastic capsules contain ing testosterone ( T ) , dihydrotestosterone (DHT) or estradiol (E) will aaintain the effect. It has been suggested that the E receptor plays a role in this process because the DHT �etabolite So. androstane-36-178-diol (B-DIOL) is kno'Jll to bind to i t . To examine this, pubertal male rats were cas trated and implanted sc with one o f two sizes of capsule filled with B-DIOL or its 3o. isome r , 4 days later these animals were anesthetized and primed with lOµg LHRH/lOOg BW or saline iv at � hr interva l s . 30 nin after the 3rd injec tion a blood sample was collected by heart puncture and all animals received a 50 ng LHRH/lOOg BW challenge injection. A final blood sample was collected 10 min later. Neither isomer was capable of maintaining a self-priming effect . B-DIOL seemed to reverse the effect; saline primed rats re leased n:-ore LH in response to LHRH challenge than did LllRH pretreated animals. Additional studies indicated that neither isooer had any effect on pituitary sensitivity to a When adninistered to intact rats for single LHRll injection. 4 days neither a Su reductase inhibitor (4-MA) nor an arona tase inhibitor (A.TD) was capable of blocking the self-pric ing effect. When given to castrated rats bearing T capsules capable of maintaining the effect, ATD did not block it . 1hese data suggest that both aromatization and Su reduction of T will maintain the self-priming effect and that in the Supported by absence of one, the other is sufficient. 3 PHOTOPERIODIC ltAillTE!IANCE OF THE RNI TESTIS DURING EXPOSURE TO COIHRASTi llG STIHULATORY AND HlHlBJTORY OAY LE!IGTHS . Bruce Schanbacher, RLHUSMARC, Agr. Re s. Serv . , U .S . Departnent of Agricu l ture , Clay Center, t/E 68933. The Suffo l k ram i s characterized by a cyclic reproductive response to changes i n day length. Seasonal responses of the ovfne testis can be experiirenta l l y driven by a l ternat i ng exposure to contrasting short (SL: 160) and long ( 16L:SD) days (D'Occhio e t al . , B i o l . Reprod . 30 : 1039-1054, Twelve nature Suffolk rams were assi gned to one o f 1984 ) . two groups and exposed for a period of o n e year to a 56-d (hexamestral ) pho tope rfod cycle of e i ther SL: 16D-')16L:80 ( G p i l or BL: 160--+7 l : 9 D : l l : 7 0 (GpI I ) . Al l rams entered the experiment late i n the breeding season (Nov) when mean testis diameters (6.4 cm) and mean serum testosterone levels ( 2 , 6 ng/ml ) were decrea s i n g . T e sti s diameters a n d serum testosterone for G p s I a n d I I were s i m i l a r a nd para l l e l ed each other over the course o f the study with m i n i mum average values ( 5 . 8 e n and 1 .0 ng/ml ) recorded during the first two cycles (Dec-f�arch ) , Thereafter, both testis diaf'.eters and serum testosterone i ncreased abruptly and were maintained near 7 . 5 crri and bet'Wee n 5 and 10 ng/ml over the next four cycles . Twenty -four hour secretory patterns of melatonin in these rams accurately reflected daylength exposure w i th nighttime values being three- to In view o f the four-fold higher than those during the day. evi dence that tes ticular regres s i o n requires weeks of exposure to f nhf bf tory long days, these fi nd1 ngs suggest that 56-d cyclic stimulation o f the pineal-pituf taryg-Onadal continuum is effective at preventi ng normal p h o tof nd u ce d , testicular regression in rams. Pre l i minary evid ence al so i n d i cates that these rams produce good semen and d i s p lay sexual aggres s i veness suitable for year-round mating. 4 PURlfICATION AND COMPJJUTIVESTRUCJ'lJRAL ANALYSIS OF A FSD AND TISTOSTERONERESPONSIVE GLYCOPROIEIN ffiOM RAT AND MOUSE PRIMARY SERTOLI CELL CULTURE MEDIA. C. Yan Cheng, Josephine Grima, Kenneth S.K. Tung• , and. C. Yayne B&tdln. The PopulaUon Council. 1230 York Avenue, Nev York, NY 10021; and •Department of Pathology, School of Medicine, University of Nev Meiico, Albuquerque, NM 87131. During a search for hormonally responsive proteins from primary Sert.oil cell-enriched cultures established from 20-day-old rats, ve noted aFSH· and testosterone-res9onsive protein designated C!rfB.-21 (Cheng et al. Endocrinology ll8:"80-"88, 1986). CMB-21 bas been purified to apparent homogeneity using sequentlal anion-eichange, chromatofocusing, gel permeation. and hydrophobic HPLC from batches of 2 litres of Sertoli cell culture medium. Also a monospecific an.ti�rum bu been ra,iS(ld ag&inrt CMB·ZI. CMB-ZI is a heterogeneous gtycoproteln of Mr 90,000 consistine oftvo subunits vith identical electrophor-etic mobility of Mr 4l,OOO. It is reactive to vheat germ 141glutinin but not to concanavalin A on lectin-blots. Using Serloli cell cuHures established from prepubertal mice at 20 days of age, it vas noted that it consisted of a CMB--21 immunoNactive macromolecule demonstrated by immunoblots. We have therefore partially purified lhis immunoreactive CM.B-21 9rotein from the mouse Sertoli cell culwres by HPLC and shovn that the mouse im.munoreaclive CMB-Zl is a glycoprotein of Mr 84,000 consisting of two subunits of Mr 42,000. Peptide maps vere generated for these tvo proteins and visualized by either immuno- or Jectin·staining. These analyses indicate that even though these proteins have some distinctive fealures, they have similar peptide maps and share common immunodet.erminants. In summary, ve have Cl) identified a FSH· and testosterone-responsive protein in rat primary Sertoli cell cultures; and {2) this protein shares es>itopes vith a similar macromolecule in the mouse ptima_ry �ertoli cell cultures. llD163ll. P-17 Journal of Andrology 5 GERM CELLS BIND TESTICULAR 59Fe·TRANSFERRIN THE TRANSFERRIN MEDIATED TRANSPORT OF If.FeLLOWING ACROSS SERTOLI CELLS. I&nill � & Martin Ihm, Dept. of Anal. & Cell Biol., Georgetown Univ., Washington D.C. 6 Using confluent epithelial sheets of Sertoli cells (3. xlO cells/chamber) grown in bicameral culture chambers (0. cm ) we have shown the transfcrrin (TF) mediated transport of Fe across Scrtoli cells (l). We now report the effect of coculturing germ cells on Sertoli cells during this process. Two populations of germ cells (primary spcrmatocytes, round spermatids) were isolated fro adult T rats by unit gravity sedimentation and cocultured (Sx!O germ cells/chamber) overnigh with epithelial sheets of Sertoli cells. � When 520 p�f of human 9Fe-TF were placed in media bathing the sal aspect of Sertoli cells alone for 6 hours, 3.7 pM of testicular � Fe-TF was immunoprecipitated from apical media. However, coculture of spermatocy l( or spermatids on Sertoli cells reduced � the immunoprecipitable Fe·TF i n apical media by 50% and 20% � respectively. Since spermatocytes and spermatids contain 31.5xl0 and I0.7x!04 TF receptors, resp �tively, on their cell surface, the % reduced immunoprecipitable Fe· in apical media of cocultures indicates that some of the Fe-TF secreted apically by Sertoli cells is subsequently bound to germ �ells. From the 5 percentage reduction in i mu noprecipitabl e·TF in the � presence of genn lls, 2.5xl0 spermatids or l.Ox l O spermatocytcs fg ence, will bind all the Fe-TF secreted apically by Sertoli cells. 5 less than one germ cell per Sertoli cell will bind all the Fe·TF secreted apically by Sertoli cells. Since in. tlY2. there are a number of germ cells per Sertoli cell this indicates there arc a number of unoccupied TF receptors which may act as a sink for the passage of Fe down a concentration gradient from blood (high concentration of Fe) to germ cells (lower concentration of Fe) during the transferrin mediated transcellular transport of Fe across Sertoli cells. (I) Djakiew et al, ( 1986). J. Androl. 7:355-366. �3 � ?: f J� e g §!, 6 GERM CELLS INFLUENCE THE POLARIZE.D SECRETIOfl OF AllDROGEtl B I NOIJIG PROTEIN (ASP), BUT HOT TRAtlSFERRill ( TRF) BY SERTOLI CELLS IN VITRO. Andrzej Janecki*, Andrzej Jakubowiak* and Aiina----ste"1nberger. Department of OB/GYN and Reproductive S c i ences, University of Texas Medical School at Houston, 6431 Fannin, Houston, TX 77030 The polarity of ASP secretion i n the rat testes changes w i t h age, possibly due to varying germ cel l populations. I n the present study, we investigated the infl uence of germ cells (Ge) on the polarized secretion of ASP and Trf by rat Serto l i cel l s (Sc) maintained in two-compartment cul ture chambers (Janecki and Stei nberge r , J. Androl . 7 , 1986). Sc from 14-day-old rats secreted ASP b i - d i rectiona l l y , the outer : i nner compartment ratio (OC/IC) being highest ('•·2.0) at Days 3-4 of culture when the monolayers became essential l y free of contami nating G e . This ratio was not infl uenced by FSH {200 ng/ml ) . Addition of Ge (> 85% pachytene spermatocytes) reduced the ratio to ....- 0 . 7 , but only in the presence of FSH. Co-culture with Ge did not infl uence the Trf OC/IC ratio. In cul tures of Sc from 32-day-old rats� intentionally containing many original Ge, the ASP OC/IC ratio was ..,,. 0 . 6 . However, i f these cul tures were treated w1th hypoton i c buffer, to remove the residual contaminating Ge, the ASP OC/lC ratio changed to � 1 . 3 , i ndependent of FSH. The hypoton i c treatment d i d not infl uence the Trf OC/!C ratio (.... 0 . 7 ) . These data suggest that Ge may influence the polarity of some Sc secretions i!! Y.U!:.Q, the effect being FSH-dependent i n Sc obtained from 14-day-old rats but not from 32-day-old anima l s . 18-P · January/February 1 98 7 Vol. B 7 EFFECT OF AGE Otl COMPOSITION OF HUMAN SEMillIFEROUS TUBULE BOUtlOARY T I SSUE ANO Otl VOLUME OF EACH COMPOtlEtlT. J . G . A b d o * , L. John s o n , and W . B . tl e a v e s . U n i v er s i ty of Texas Health Science Center at D a l l a s , D a l l a s , TX 75235 . T u b u l a r boundary t i s su e t h i c k e n s with age. I n a sma l l group of men, t h e percentage of myo i d c e l l s i n t u bu l ar boundary t i ssue was not infl uenced by age, and the compo s i t i o n of extrace l l u l a r components r e m a i n s unknown. The o b j e c t i v e w a s t o characterize the compos ition o f bound ary tissue i n 16 younger (20 to 29y) and 18 o l d e r { 51 to 84y) adult men. T e s t e s were perfused w i t h g l u t a r a l d e h y d e , p l aced in osmium, and err�edded in Epon. Volume of boundary t i s s ue/man was c a l c u l ated from the percentage boundary tissue determined by point counting 0 . 5JJ111 sections and the p a i r e d parenchymal v o l u m e . Percentage o f myoid c e l l s , col l agen, microfi br l l s , or other extrac e l l u l ar component in boundary ti ssue was determined from electron m i crographs taken randomly on predetermined sites of EM gr i d s . V o l ume of each component was calculated from the volume of bound ary t i s sue/man and the percentage of boundary t i s s u e occu p i ed by each compo n en t . There was no d ifference (P>0.05) between testes from younger and older men in the percentage of boundary t i s s u e occ u p i e d by myo l d c e l l s { 3 6 . 2 ± 1 . 6 vs 3 1 . 8 ± 1 . 8 % ) , c o l l agen (27 .0±2 .0 vs 2 9 . 2 ± 2 . 5%) , m l c r o f l b r i l s {22 . 1±1 .3 v s 2 4 . 1 ± 1 . 5 % ) , and other ( 1 4 . 8 ± 1 . 8 v s 1 5 . 0 ± 1 . 9 % ) . T h e volume o f boundary t i s sue/man {3 , 3 ± 0 . 2 v s 3 . 3 ± 0 . 3 m l ) , m y o i d c e l l v o l ume/man ( 1 .2±0 . l vs l . l tO . lm l ) , c o l l agen vol ume/man ( 0 . 87±0.08 vs 0 . 97±0.14ml ) , vol ume of m i c ro f i b r i ls/man ( 0 . 74 ± 0 . 0 8 vs 0 , 80±0 . lOm l ) , and volume of other ( 0 . 48±0.06 vs 0 . 50±0.0Bml ) a l s o were s i m i l ar ( P > 0 . 0 5 ) between a g e groups. T h i s study confirms that age-related thickening of boundary t i ssue i s not due to the depos i t i on of col l agen or other boundary tissue components. Supported by NIH grant AG2260. 8 RELATIONSHIP OF HUMAN SPERM O..'iA STABILITY' AND FERTIL ITY roTE:n'I'IAL. Sokoloski* ,J.E . , Qtligley* ,M.M. and Thomas1 A.J. Depts of Gynecology and Urology, Cleveland clinic Foundation, Cleveland, OH144106. Male fertility potential cannot be predicted by microscopic semen analysis alone. To augment the conventional analysis various in vitro tests have been developed to better delin eate the possible male factors involved in an infertile union. One of these tests takes advantage of the fluoro chromatic properties of Acridine Orange (AO ) . A01 when bound to native O..'iA, fluoresces green and when bound to de natured DNA glows red. This report profiles a population of fertile men and infertile patients whose sperm has been subjected to AO analysis. 72 patients were evaluated i n this preliminary study. The AO score ( % green) displayed a bimc<lal histogram with an arithmetic ranqe of 2 to 100% (x=79.8 + 29.7 sd ) . outliers (>2sd) were rem::ived from the set and a new, unimc<lal range of 54.9-100% was generated ( x=89.2+1 7 . l ) and was taken as the hypothetical �normal" range. Ten of the 72 sarrples fell into the "abnorrral"range. There was no significant difference bet��en the two groups in terms of: total sperm count/ejaculate and % motility. The sperm of the 10 abnor!TB.l pts and 23 of the normal AO group Yas subjected to the Hamster Egg Assay with no sig nificant differences in the % eqgs penetrated ( 3 7 . 4 vs 37.l p=ns ) . Seventeen pts were in the IVF-ER program, 6 had abnor!TB.l AO and fertilized only 13%(4/30) of their wives eggs Yhile the not:Tr'al AO partners fertilized 47%( 39/83) (p<0.05 ) . These data suggest that AO analysis may provide yet one rrore parameter by which male fertility potential can be evaluated . No. 1 TWELFTH ANNUAL MEETING 9 STUDIES ON THE FUNCTION AND Rl';GUI.ATION OF SULFHYDRYLS IN CAPUT EPIDIDYHAL SPERM INDUCED TO ACQUIRE PROGRESSIVE MOTILITY l!:f VITRO. G . Cornwal l , S . T i l l�an* and T . S . K . Chang, Dep t . of Urology, 'Ihe Johns Hopkins Hospital, Baltimore, MD. To study the function and regulation of sulfhydryls (SH) in epididyraal sperm naturation, imi:nature caput epididymal sperf.l. were induced to acquire progressive motility by incubation in induction mediU111 {IM) containing 15% seruinal plasma and 501" .M theophylline. Howev.e r , 95% of these sperm exhibited a flagellar angularity characterized by a 9 0 - 1 800 bend at the neck or midpiece. Addition to IM of diamide, a sulfhydryl oxidant which forms disulfide ( S O S ) bonds , completely prevented the flagellar bending in gotile- induced caput sperm. The shape-maintaining effect of diamide however, was transient; 180 min. after addition of diamide 85% of the sperm again exhibited flagellar angularity. Subsequent studies suggested that sperm glutathione reductase (GR) activity may be involved in the recurrence of flagellar bending by increasing the levels of SH in diamide In these studies addition o f nitrofurantoin treated sper�. (NF) , a GR inhibitor, to IK reduced caput sperm GR activity by 83% and prevented sperm flagellar bending. Flow cyto· metric studies using a SH fluorescent stain were used to m�nitor sperm SH levels in IM containing diamide and NF. These studies revealed that: 1) l!.o ttle- tnduced sperm with angular flagella contained high levels of SH; 2 ) these SH levels were significantly reduced by dtsmide; 3) sperm SH levels in diamide - treated sperm tncreased over time paral· le ling the increase in flagellar angulation; and 4) the presence of NF prevented increases in spern SH as well as flagellar bending. Taken together these studies suggest that the interaction between intracellular sperm sulfhydryls and glutathione reductase may play an important role in the maturation of sperm in the epididyais. EPA CR-812765-02·0 AUTOlV.TEO SEMEll ANALYS I S : REPRODUC I B I L I TY STL'°IES. Kenneth A. Ginsburg* and Ernest L. Abel*, Wayne State University ' C . S . Kott Center, Detroit, M l , �8201 . C:C.•put�r-assisted v ide°'"icrographic syste;is pernit analysis of spern "ova· r;ent with unpreced�n!ed speed and accuracy. We investigated the variabi l · ity and reproduc i b 1 l 1 t y o f such reasure-"'enh o n fresh ejaculated hu•.an sper""3tozoa in se-,inal p l as.,-a using the C e l l S<lft Auto,.-ated Se;oen Analyzer (CASA, Cry�Resources, lft) under g Hakler cha,..her. 1 0 specit'ens nith spern concentrations of 31 t':' �37 X 10 y,�re studied in 3 di fferent protocol s , . and concentration, ,,--0 t 1 l 1 ty, 1lnear1ty, velocity, head d i s p l ace,..ent {Al.HJ and head beat frequency (&Fl nare deternined: \ ) Three rethods "ere used to exa11ine d i f ferences bet>leen individual and pooled analy5es nith 5 repe· titions per "ethod. In �.ethod A a single field .. as evaluated in l{ethod B, i;ol t i p l e fields nere e�anined and data pooled, and in Method C, 4 pre· � frned fields Mre exanined in t r i p l i cate and average r-easurerents deter· mned. 2 ) Effects of nu,,,ber of c e l l s analyzed on the stabi l i ty of reasure· rents. 3) Reproduci b i l i t y of Feasureo-..ents "ithin subjects exa"lined by recording on v i deotape and repeated analysis. �rrelati�n betneen �.e thods A and B for concentration, rnti l i ty and veloc· 1ty nas high ( r000 . 85 t , r�.835, r�.899, p < 0 . 0 1 ) ilhether only one or five fields »ere e�anined. Linearity and BCF "ere not 1>el 1 correlated, l n d i · eating significant vari ation f o r SO'<e c e l l r:.:ive--:->ent reasure.•entscan occur over d i f ferent areas o f the s l ide. The 1011est variabi l i ty for a l l •easure r.ents nas obtained nith th<! averaging r-<=thod. After collection of at least 200 cells, reasured values d i d not vary a s rore c e l l s 11ere studied, HHhlne reproduc i b i l i t y using V ideotaped ir,,oges nas high nith average nithln-sa"f>le stand� r d erro � s of 1 . 7 \ , 2.7\, 3 . 2 \ , 3 , 3 \ , 4 . 9 \ , 8.2!;, and . 4.4\ for concentrat1�n, r.:it1 l 1 ty, velocity, l i nearity, Nxir:u--. ALH, t'ean ALH and BCF, respect1ve\y. Conclusions: 1) Spern d i s t r i bution on nicroscope s l ides ray be a s i g n i f i cant source of varia tion for linearity and & F •easure.-.ent b u t n o t for c<.mcen�ration, c.:i t i l i � y , �elocity and AUt, This variation ray need to be taken 1 �to cons1der� t1on rn developing a r<:1ve--ent prof i l e o f tl"oe spern popuht10� fro., a srngle sec,;,n analysis. 2 ) A sa--p l i n g 1'etITT!d 11ith 4 predeternrn� d areas done l n tr �_ p l icate, calculating ,-,;,an values, gives _ the least l'>1thin·sa,..p l e variation. 3) !lo change in results occurs after analysis of 200 c e l l s , �) The CASA gives highly reproducible results nith an uvera l l average standard error of 4.1\ for a l l parreters reasured, and stwuld thus be suitable for detecting ��all changes i n spern r-0ve,-,;,nt character i s t i c s i n both a research and c l i n i c a l setting, 10 11 CCtu'AlllSON OF FULLY AUTU1ATW ANO TRAD ITIONAL HETHUDS Of SL'"l EN ANALY SIS . o . Vantnan, H . Zinanan and R . 91erins . Developnental Flldocrinology Branch, N ICHO, Bethesda, HD. Recent technology fo r fully auto�ated conputer-as sisted analysis of spern count and t:lOtion characteristics ( C e l l So f t ) potent ially offers an objective nethod for senen analysis , We tested the accuracy of this nev sperra count a11d approach by cooparing est inates of notility in seminal plasma obtained by two cethods ; Cell Soft and traditional senen analysis using hellilcyto� . for cel l density and subjective assessnent for �otility Semen fron 63 spero positive and 7 and spero velocity. azoospermic �en were provided after )b-48 hrs abstinence F.stii:1ates o f Cell Soft precision (coef .of var ,nean±SE) � count and Xnotil_� are shown using U ,7 sec exposure Concentration k aotility 3 flds 9 flos c e l ls/field 3 flds 9 tlos 57 t 1 7 40 t 1 2 (n"'5) 40 t 0 5J t 5 3U t 4 34 ! ) l l-2U tn,,.12) £ 5 t 4 £5 ± L 2l ± lJ t ) 2 1 - 4 U ( n =l 9) 21 t 3 18 t 1 IL t 2 11 t _ ) 41 (n<>l7) 12 t 4 16 i.J...___ To assess l.el l &:lft ac.curacy ,9 randon flds of -obServ8.tion were usi;,o to estitiate sperc-. count ( r:i ill ion/ml ,tiean ± S E) cells/tield 0 1 - 10 11 20 )20 Henocytoiwter U 3 .7tl .3 15 .4tl .9 o"/ .9t9 ,3 CellSoft 3 .btU , 3 12 ,6±3.U 21 .9±2 ,3 90 ,9±10 ,3 0 °0 1 NS NS p<Q ,05 Corre lat ion <:oe f fie ien t for cone ent ration :O . 1:! 7 , Xnoti l i ty U .71:! and velocity 0 .58 were significant p((J .01 , We conclud�rl that Cell5oft provides acceptable accuracy and precision when analyzing core th�n 10 c el l s /fiel d ( I S nillion/ml ) . HOW<!Ve r , spen1 velocity nay require aore detailed analysis , l lU 12 AllALYSIS OF PREVASECTOMY EJACULATES BY THE HYPOOSH.OTIC SWELLING (HOS) TEST. L , J , D . Zaneveld, R, S , Jeyendran, H.P.P. d e Castro and p,J,M. Silveira*, Dept. of Ob/Gyn, Rush University, Chicago, IL, USA and Propater Institute, Sao Paulo, Brasil. The hypoosmotic swelling (HOS) test assesses the biochenical integrity o f the sperm aembrane by determining the influx of water under hypoosgotic conditions, resulting in the expansion o f the spern. membrane and the The HOS test was shown to be curl ing of the tail fibers, nore reliable in predicting the outcoae o f human in vitro fertilization ( IVF) than the more standard semen � �� Based on these and paraneters ( J . Andro l . 7 : 190, 1986). other in vitro dat.a, cut-offs for .. nomal� (fertile) and ,.abnortDal .. (infertile) were tentatively assigned; .60% o r more reactive sperm being "nornal. . , l ess than 50% being "abnormal"" and between 50 and 60% representing a ""grey" In order area where it is difficult to reach a diagnosis. to assess the validity o f these values in vivo, ejaculates fron 1,890 men who had fathered childre"il""at one or more times in their life and requested a vasectomy, were analyzed by the HOS test. 96% o f these men gave a positive HOS test ()60% reactive spern), 2% gave a negative test {(50%-reactive spern.) and 2% fell into the By coaparison, 13% o f the ejaculates had "grey" area. less than SO% motile sperm and 68% possessed 60% or more motile sperm, AsstCTJ.ing that the large naj ority of these men were still fertile at the tlae o f evaluation, the results suggest that the HOS test correlates well with the fertility status o f the aale and should be a worthwhile addition to the senen analysis, Supported by NIH HD 19555 and support froB. Rush-Presby terian- St, Luke' s Hedical Center • .. ., P-19 Journal of Andrology 13 RELAT IONSHIP BETWEEN THE HYPOOSMOTIC SWELLING (HOS) TEST AND THE SPERM PENETRAT ION ASSAY (SPA) C. Bastias,* T. Thompson� A. Buckt H . Hinsort. and B . J , Rogers, Dept. OB /GYU, C-fARR, Vanderbi l t University School of Medicine, Nashvi l le , Tn. 37232 The (HOS) test has been proposed as a useful assay in the diagnosis of the i nfert i l e male. A good correlation between the HOS and SPA in ferti l e and normal semen samples was i ni t i a l ly presented but subsequently no s i gnificant correlation was demonstrated with ferti l e and infertile patients. To val idate the potential usefulness of the HOS test c l i ni c a l l y we have eval uated 68 patients using both the HOS and SPA as well as traditional se.11en parameters. The methodology for the HOS test was as originally described by Jeyendran et a l . The SPA was performed by the original procedure uti l i zing an 18 hour preincubation period. Values of �60% for the HOS and �10% for the SPA were considered positive and <50% for HOS and 0% for SPA w�re considered negative. The gray area (50 59%) for HOS and ( 1 - 9) for SPA were excluded. The sens1tivity of the HOS test was 74% and the specific ity was 85% for predicting the SPA resu l t s . The false positive rate i n the HOS test was 22% and the false negative rate was 18%. The HOS had an 80% success for correct prediction of SPA resu l t s . Correlation coeffi c i ents were: HOS vs SPA - 0.44 ( P < 0 . 0 2 ) ; v s mot i l i ty 0 . 5 0 (P< 0.01 ) ; vs count - 0.41 (P<0,05 ) . Due to the high degree of success of the HOS test i n predicting the SPA results and the ease of performance of the HOS test we bel i eve that it i s a useful adjunct to evaluation of standard semen parameters as part of a comprehensive male fert i l i ty assessment. 14 HUHAN SPERM ACRO\I H AS A FERT!L �TY MARKER . H . H . Van der Ven*, W . P . Kennedy, J . M . Kaminski , R . S . Jeyendran and L . J . D , Zanevel d , Oept. of Ob/Gyn , Univ. of Bonn, FRG and Dept. of Ob/Gyn, Rush University, Chicago, IL. T h e acrosin activity of 36 ejaculates obtatned from men i n an IVF program was compared with the fert i l i z i ng capacity of their spennatozoa in order t o ascertain whether acrosin could be used as a ferti l ity marker. Total acrosin was determined using a recently developed s i m p l e assay system ( Kennedy et a l , American Fert i l i ty Soci ety Abstracts , 87 , l g8 5 ) , Briefly, spermatozoa from each ejaculate were washed free of seminal p l a sma by mi l d centrifugation over ficol l . The washed spennatozoa were then i ncubated with the substrate, 11-a-benzoyl-dl-arginine p - n f t roani l i de ( BAPHA; lmg/ml ) and 0 . 1% Triton X-100 detergent ( which causes dispersal of the sperm acros011al membranes and releases acrosin) at pH 8 . 0 . After 3 hrs of i ncubation, benzamidfne, a potent acros i n inhibitor was added to quench the reaction and the absorbance of the supernatant was read at 410 nm after m i l d centrifugation. T h i s simple assay was reported t o be l f near in response to sperm numbers i n the range of 2-10 m i l l i o n and a l s o compared w e l l with the standard acrosin assay, A s i gn i ficant di fference in the total acros i n activity was noted between the 26 fert i l e {mean ± S . £ . ; 0 . 3 4 ± 0.03 mIU) and 10 infert i l e { 0 . 1 9 ± 0.03 mlll) ejacul ates. Moreover, the acrosin levels observed in the individual ejaculates significantly correlated with their ferti l i z ing capacity ( r=0.475; p<0,01 ) . The data demonstrates the c l i nical applicabi l i ty of the hLJ'llan acros i n assay for determining the fert i l izing capacity of spermatozoa. Supported i n part by tHH HO 19555, 20-P · January/February 1 9 8 7 Vol. 8 15 LYHPHOPENIA IN HALE PARTNERS OF HIFERTI LE COUPLE'S. Allan R. Glass, Walter Reed Hospital, Washington, DC 20307 The etiology of oligospernia in nost infertile nen ls unknown. To explore whether the process that disrupts speroatogenesis in infertile aen might also affect another rapidly proliferating cell population, nanely blood cells, �e conducted a retrospective study of blood counts in 72 unselected nale partners of infertile couples attending our clinic and in 119 healthy controls undergoing routine peri odic health examination s . There was no difference between infertility clinic patients and controls in nean levels of hemoglobin, he�..atocrlt, or red cell volune, or in percent age of subjects with abnoroal values for these paraneters. Platelet counts were also sinilar in both groups. However, the nean total leukocyte count was lower in the infertility clinic group (S818±17 9(SEM) vs 6397±174 per vL; p < . 05) . The infertility clinic group also had lower total lymphocyte count than controls (1697±69 vs 2206±80 per vL; p <,001) but did not have lower neutrophil counts (3331±144 per µL; con trol=3678±139) . In the male partners of infertile couples, there was no correlation between sperm density and either l)'Ill phocyte count or total leukocyte count and no difference in nean values of these parameters between those with spern density above or below 20 aillion per c c . Preliminary re sults fron an ongoing prospective study of infer tile, oll gospernic oen (currently n=6) and fertile controls (n=24) show no difference between the groups in counts of total T lynphocytes, helper lyophocytes (T4), or suppressor lynpho cytes(T8) . We conclude that nale partners of infertile cou ples have significant lynphopenia without other changes in blood count, and this lynphopenia is not clearly related to reduction in sperm density, Whether lymphopenia in this populat ion reflects an underlying imaune defect which pre disposes to infertility re!l'.ll ins to be deternined. 16 EFFECTS OF SOLUBLE PRODUCTS OF ACIIVATED LYKPHOC'iTES AND MACROPHAGES (LYMPHOKINES AND HONOKINES) ON HUMAN SPERM MOTILITY, Joseph A. Hill*, Florina Haimovici*, and Deborah J. Anderson, Harvard Medical School, Boston, HA 0 2 1 1 5 . The male and feaale reproductive tracts are immunologically dynaaic tissues containing imsune response cells (lymphocytes and macrophages) that elaborate soluble factors upon activation ( lyaphokines and 1tOnokines, respectively). The aajority of these soluble imnune factors provide intercellular signals that potentiate and regulate the i!!fllune response; in addition •any lymphokines and aonokines have been shown to be cytotoxic to nonillllune l. cells that are �argets of Experiments were conducted to evaluate itlllu l ne attack. the effects of supernatants from activated leukocyte cultures , and purified lymphokine and monokine preparat ions on sperm motion, An automated semen analyzer was used to obtain accurate measurements of Supernatants from several paraaeters of spere l!,Otion. activated peripheral blood leukocyte cultures significantly decreased sperm motility. In studies of effects of individual lymphokines and monokines, significant antilil<Otility effects were observed when spermatozoa were incubated with the lymphokine gamr:aa-interferon, and the monokine tumor necrosis factor. Subnoraal fertility aay result froa defective sperm function caused by lynphokines and monokines elaborated by activated lymphocytes and macrophages residing in the reproductive tracts of infertile nen and women. No. 1 TWELFTH ANNUAL MEETING 17 EVIDE!ICE FOR ANTIBODY BINDING SPECIFICITY T O DONOR SPERM. Stephen E . Hove and Dona M . Lynch, Department of Pathology, Rose Medical Center , Denve r , D i f f erences Colorado 80220, i n antisperm antibody binding have been 19 CLOMIPHENE AND TAMOXIFEN I N OLIGOSPEAMIC INFERTILITY. W S. Maxson, B.J. Rogers. C.M Herbe1!, C. Markuson·. F. K. Kirchner. Jr., Vanderbilt Univ. Med. Ctr., Nashville, TN 37232 and Northwest Center !or Infertility and Reproductive Endocrinology. Margate, Fl demonstrated betveen husband sperm and donor sperm in s o me 33063 antibody binding specificity may p l a y in noncorrelations To clarity the role of antiestrogen therapy for o!igospermic infertility, indirect ELISA method, sera from i n f e r t i l e patients with p o s i t i v e imno b i l i z a t ion assays were investigated. The immobilization assay was chosen for comparison since it i s clomiphene citrate 25 mg BID (CC) and tamoxifen 1 0 mg BID (TX) on i n f e r t i l e wonen. To investigate the contribution antisperrn between a t r a d i t i o n a l indirect functional assay and an b e l i e v e d t o be m o r e specific than agglutination assays in the detection of antisperm antibody. Sera from 15 infertile patients with positive immob i l i z a t i o n assays (SIV > 2 , 0 ) were evaluated a s follows: ( I ) ELISA assays with target sperm froM 5 i n d i v i d u a l sperm donors, ( 2 ) SIT and ELISA assays performed specimens, and in p a r a l l e l - u s i n g the same sperm donor ( 3 ) ELISA assays u s i n g pooled sperm from four d i f f e r e n t d o n o r s , When the 1 5 sera were tested using the SIT against 5 individual random sperm donors, there was a 43% (31/72) positive correlation. When these 15 sera were tested in parallel using the same 5 individual random sperm donors, the correlation was improved to 80% (36/45), These data were comparable to our findings in a larger group of 292 i n f e r t i l e p a t i e n t s , In t h i s g r o u p , 64 p o s i t i v e immo b i lization assays were found and when compared with an ELISA using pooled target s p e r m , 80% (52/64) correlated posi t i v e l y . These data suggest that there are significant differences in antibody b i n d i n g between i n d i v i d u a l sperm samples, probably due to antibody specificity, which may e f f e c t correlations between sperm antibody t e s t s w h e n d i f f e r e n t sperm donors a r e used, These differences nay minimized by utilizing appropriately with significant antigen e cess, x 18 semen parameters, bovine mucus penetration (BMP). and the zone lree hamster oocyte penetration !est (SPA) i n 19 normogonadotropic r--...m men with oligospermia. negative antisperm antibodies and normal physical exams. M1dcycle artifical cup insemination with husband's sperm was performed twice per cycle during the 3 placebo. 6 drug, and 3 post-therapy months_ The results (mean: N:CC""7, TX�12): ,,., ,_ .. � Baselme Placebo Drug Drug Washou1 �� 4 143 102 168 17 5 , ;; 148 112 117 14 8 '".. � I cc35 4;x 51 41 35 37 30 37 34 37 cc49 5 �x 59 55 f>6 54 48 53 49 53 No d1f!erence was round between CC, TX, c; 0 ""....., I '; 32 86 0 5 0 0 0 0 c;, ;.x 32 25 29 27 30 24 20 27 or placebo on any parameter evaluated. Despi!e no obvious enhancement in semen parameters. eight pregnancies occurred (42%) ( 1 CC, 5 TX, 2 placebo). These data suggest that: 1) these semen parameters are inadequate to predict fenility before or alter therapy: 2) continued study is necessary to compare the relative el!icacy of CC and TX versus placebo. be selected sperm pools INABILITY OF SPERM WASHING TO REKOVE ANTISPERH ANTIBODIES. Gilbert G. Haas , Jr . , Osll!ond D ' Cruz*, and Fayez Nahhas*, University of Oklahoma Health Sciences Cen t e r , Oklahoma C i t y , OK 73190. Attempts have been made to reoove antibodies attached to the human spern surface by repeated washing. Although such therapeutic maneuvers have resulted in pregnancy, most men treated in this manner were assayed for serum antibodies by agglutination a ss a y s , and antibodies on the men's spera sur· face were not identified. Using a radiolabeled antiglobulin assay, we showed that washing up to 18 tines could not sig· nificantly diainish ant ibody placed on the spern surface . Hunan sperm from multiple donors were pooled and incubated with a l • l dilution of plasna previously shown to be posi tive for IgG antispern antibodies . After 3 washes, tripli cate samples of 2 m i llio n spera were incubated with 1251labeled antihucan lgG, the saaples washed 4 tiaes, and the residual spern-associated radioactivity deteraine d . These steps were repeated after 6, 9, 1 2 , 1 5 , and 18 washe s . The results were expressed as the percentage of the total radioactivity added to each spera a l i quot that remained associated with the sperm surface. The percentage o f radio activity for the 6 experimental conditions was 1 6 . 8 7. , 1 8 . 76'1., 1 4 . 7 8 % , 1 5 . 79'!., 1 6 . 26'1. , and 1 3 . 8%, respectively. Repeat assays have shown sinilar r e s u l t s . There does not appear to be a significant removal of spera-bound IgG from the human sperm surface even after extensive washing with isotonic PBS. Exposure of sp er to acidic (pH 4 ) or alka line (pll 1 1 ) failed t o renove sperm-associated antibody. a this double-blind. placebo-controlled study evaluated the effect or A COMPARISON Cf IVF· CYCLES WITH FAILED VERSUS SUCCESSFUL FERTILIZATION: SIGNIF ICANCE Cf CYCLE, SEMEN MO PATIENT VARIABLE S, S,P .Boyers , J .B .Russel l *, J . N . Stronk*, L .F1no*, £,Jones*, A.H.OeCherney, Dept , rtJ/Gyn , Yale Univ. Sch. Med . , New Haven, CT 06510-8063. From 5/83 to 1/86 we completed 631 l aparoscopies for IVF, 3081 oocytes were recovered (4.9/cyc l e ) and 1924 (62.4%) fert i l i zed . There were 53 cycles with at least 1 mature oocyte but no fert i l i zation (8,4% · The purpose of this study was to compare 53 cycles wi h no fert i l i zation to 40 control cycles with regard to cyc l e , semen and patient var i ab l e s . For each failed fert i l i zation cyc l e , a control was randoml y selected as the cycle closest in time (same day or 1-2 days before or after) fn 'n'hich the fert i l i zation rate was > 60%. Hean fert i l i zation rate in cont rols was 79.6%. The fiurrb e r of eggs was significantly lower i n cycles with zero fert i l i zation (3.8+2,7 vs 5.9+2.7) (p<0.001 ) . There was no s i g n i f i cant di f ference i n day of laparoscopy ( 12 .1±_ 1 .2 vs 1 1 .8+ 1 .2 ) (p=0.25) or E 2 level on day of hCG ( 7 69.:!:_ +446 vs 84D+229 pg/ml ) (p=0.391) for failed and control Cycles , resPectively. Peak E 2 level was higher i n the con trol group ( 1 193+577pg/ml ) compared to cycles with no fer t i l i zation ( 972+513pg/ml ) , (p=0.059), After semen washing and migration, Spe rm dens1ty ( x lo6 /ml ) 1 n the migrated sample was not di fferent in failed ( 39 . 5 + 38 . 9 ) vs control { 4 3 . 2 + 3 9 , l ) cycles {p=0.64), However, moti l i ty of the migrafed sample was s i g n i f i cantly greater in the control group { 6 3 . 8 + 1 7 . 0% vs 56.1+17.3%) (p=0,035). There was no s i gn i f i cant-d i f ference in-patient age (34 .0.:!:_4 .2 vs 3 4 , 3 + 4 . 2 years) { p = 0 , 6 4 ) . Tuba l infert i l ity was present i n 82.5'.4 v s 54.7'.4 o f control and failed cycles, respect i vely. Failed cycles were rrore often associated with unexplai ned or rr.3. l e fnfert i l ity { 49 . 1% vs 1 7 . 5%) and nul l i gravity (69.2% vs 47.53). 20 / P-21 Journal of Andrology 21 GAMETE INTRA-FALLOPIAN TUBE INFLUENCE OF THE MALE FACTOR ON THE TRA?ISFER (G, I . F . T . ) , SUCCESS RATE, Rell M . Luis J. Rodriguez-Rigaul , George M, Grunert2 , Woodvat:d 2 , Eberhard C . Lotze 2 , Joseph R, Feste2 , William Gibbons3, Keith O. Smith l , and Emil Steinberger l . l rexas Institute for Reproductive Medicine and 3 saylor 2woman1s Hospital, Endocrinology, College of Medicine, Houston, TX, In .Y..!..U2 fertilization ( I . V . F , ) and G . I . F . T , have been proposed as therapeutic approaches in infertile couples where a significant male factor is present. To date1 no data have been published relating the success rate of G, I , F . T , to the severity of the male factor. In this report ve analyzed the results of our first 100 G , 1 . F , T , The overa l l pregnancy rate was 21%. We examined cases. the relat ionship between occurrence of pregnancy and spern count (S,C, 1 mil lion/ml) , total sperm count (T. S , C . 1 mill/ejaculate), % ootility, motile sperm count ( H . S , C . 1 lll i ll/ml) and total motile sperm count (T.H, S . c . , mill/ejaculate). Signi ficant relationships were observed bet<Jeen pregnancy rate and S , C , , T , S , C . 1 M , S . C , 1 and T,H,S.C, The best correlat ions were noted with H , S , c . Pregnancy rates over 30% occurred i n groups and T . H , S , C , with H . S , C. over 40 mil lion/ml or T , H . S , C , over 100 mil lion/ejaculate, With H , S , C . below 10 and T , H . S . C , below 25 the pregnancy rates were 1 2 . 5 % and 9 . 5 % , respectively. N o correlation was found between % motility and pregnancy rate. In a subpopu lation o f 24 patients results of sperm penetration assay ( S , P . A . ) using zona free hams ter eggs were available. No correlation between S , P , A . and pregnancy rate could be deconstrated. 2 2 EFFECTS OF PREtlATAL IRRAOIATIOII ON TESTICULAR LH RECEPTORS 111 ADULT SYRIA/'/ HAMSTERS. A . G . Amado r , A . Bartke, V . Chandrashekar* and H . G . Klemcke*l , Dept. Physiology, SIU School of ��d i c i n e , Carbonda l e , IL 62901-4531 ; 1USOA, Roman l. Hruska JI.ea t Animal Research Center, Clay Center, NE 68933. M a l e Syrian hamsters were prenatal l y i rradiated with a 1 37-cesium source for 1 , 2, 3 or 4 m i n . {128 rads/mi n ) , and were k i l l e d after reaching adulthood. Irradiation caused a dose-related decrease i n body , testes and seminal vesicle 'fl'eights. LH receptor (LH-R) content was reduced i n hamsters i rradiated for 3 or 4 m i n . However , LH-R con tent was significantly h i gher in those i rradiated for 4 m i n . Significant correlations were observed between the changes in PRL leve l s and LH-R content ( P< 0 . 0 5 ) , and the changes in LH levels and LH-R concentrat i o n (<O.Ot ) , a l though the variation 1n the levels o f these hormones was Similarly to LH-R concentrati o n , plasma not significant. FSH levels 'fl<ere signi ficantly higher i n animals irradiated for 4 min than in those from any other group. Furthenrore, these two parameters were correlated ( P < 0 . 00 5 ) . These results i ndicate that exposure to a radiation dose capable of pennanently denaturing semlni fe�·ous tubules (4 mi n ) , can increase LH-R concentration, through a tubular damage rel ated elevation in FSH l evel s , or perhaps by direct effects on Leydig cel l s . . January/February 1 98 7 Vol. 2 3 CllA!!GF.S IN SERU!I TESTOSTERO�r Atl!l LUTEINIZING HOfil'.ONE LEVELS IN ?'ALE RATS HIT\\ ADJUVANT-INDUCED ARTHRITIS, Jeffrey W . Clemens* and Brent C . Bruot*, Department of Biological Sciences, Kent State University, Kent, OH 44242 It has been suggested that androgens may protect aales froa autoirr::iune diseases such as rheur)l.toid arthritis. This study was designed to examine the effects o f adjuvant induced arthritis (AA) on s eruo testos terone (T) and lutein izing horr:.-0ne (Lii) levels. Male Le-n'is rats were made arthri tic by injection of 1 ng Uycobacterium butyr iCU!!l i n Freund's adjuvant into the r i ght hind footpad. Groups of arthritic and non-inj ected control rats ( n=lO/group) were s ac ri f iced on days 18, 21, 24 and 27 post-injection. Body and thymus weigh t s , left and right hind paw vo lume , and serun T and Lil were deterained. Adjuvant injection resulted in significant ly {p<0.05) l arge r right and left hind paw volume and siB nificantly reduced body and t hymus ueights, Serum T leve ls were also significantly (p<0,05) reduced i n AA anireals . Testosterone levels were 0 . 37 ± 0 . 0 7 , 0 . 61±0 . 1 4 and 0 . 39 ± 0 . 09 ng/ m l in the AA animals compared to 4 , 79±0.43, 2, 77±0.38 and 3 . 19 ± 0 . 4 8 ng/nl in the control aniF-als on days 1 8 , 21 and 24, res p ec t ive ly . In contra s t , serua LH levels were sign i f i cant ly (p<O, 05) e levated in the arthritic rats. LH levels in the AA an i�4ls were 5 . 3 2 ± 0 , 4 4 , 4 . 99 ± 0 , 3 9 and 4 , 75 ± 0 . 2 9 ng /ml compared to 3 . 6 7 ± 0 . 2 8 , 3 . 3 2 ± 0 . 4 2 , and 2 . 70± 0 . 28 ng / ol on days 18, 21 and 27, re s pec tively . These re sults denonstrate that serum levels of testosterone are reduce d in the AA rat . The elevated levels of Lii suggest that this reduction is probably not the result of impaired gonadotropin secretion. (This work was supported in part by BRSG 307 RR07208 and the Northeastern Ohio Chapter of the Arthritis Foundation.) 24 T HE EFFECTS OF CASTRATIOll AUD TESTOSTEROllE Oii PLASHINOGEll ACTIVATOR ACTIVITIES Ill RAT VENTRAL PROSTATE H Wilson , 1 , 2 C Ludowes e , 1 A Sinha , 1 , 3 R Estensen, 2 VA Medical Center 1 and DeptS Lab Hed and Path2 and Genetics and Cell B i o l , 3 Univ Hn, Minneapoli s , HJI 551.J 1 7 . Plasminogen activators (PA) are highly speci f i c , argi nine-type proteases which are thought to mediate controlled localized proteolysis in regulation of tissue remodeling. Hence, increased PA activities could be expected during regression or the prostate following castration. However, PA are also secreted by the prostate. Therefore, the effects of castration and testosterone (T) replacement on PA activities in ventral prostate of the rat were studied. Castration resulted in a r i s e in days, PA activity: 60-80% at 2 days, 21.Jo-280% at l.J 85% at 7 days, and no difference at 10 days castration as compared With intact controls (activities expressed per unit protein or DllA respect i v e l y ) , T treatment of 1 1.J day castrates ror l.J days increased PA activity 150% above oil treated controls . SOS-PAGE zymography showed 3 major forms of PA (67, 60, and 38 KDa) which d i d not appear to change with castration. minor band of acti vator at 35 KDa was absent in day castrates but present i n l.J T treated castrates. A & 10 Three bands of low mol wt proteolytic activity ( 2 1 .J , 2 2 , & 1 7 KDa) cleaved gelatin in control gels. These bands were more intense in lanes of 2 & containing plasminogen. 4 day castrates in gelatin gels Thus, increased PA activity measured at l.J days castration does not appear to reflect changes in urokinase (38 KDa) or tissue-ty pe (67 & 60 i t reflects either Rather, KDa) activators. uncharacterized low rool wt PA activities which comigrate w i t h these proteases or these proteases are capable or activating plasruinogen to plasmin. 22-P B No. l TWELFTH ANNUAL MEETING 25 EFFECT OF SEASONAL CHANGES IN LEYO I G CELL ( L C ) NUMBER O N THE VOLUME OF L C SMOOTH EtlDOPLASMIC R E T I C U L U M {SER ) AND INTRATESTICULAR TESTOSTERONE CONTEllT ( I T ) . Larry Johnson and O . L . Thompson , Jr . , University of Texas Health S c i e n c e Center at D a l l a s , D a l l a s , TX 75235 and Louisiana State U n i versity, Baton Rou 9 e, LA 70803. T e s t e s from 47 adu l t ( 4 - 2 0 y ) stal l i o n s o b t a i n e d in Nov.-Jan. ( nonbreeding) and 41 adult s t a l l ions obtafned in May-July (breed i n g ) were perfused with g l ut a r a l dehyde, p l aced in osmium and embedded in Epon 8 1 2 . P e r c e n t a g e L C cytop l asm o r L C nu c l e i I n the tes t i s was determined by point counting 0. 5µm sect i o n s under b r i g h t f i e l d m i c r o s c o p y . T e s t e s from 6 r a n d o m l y selected horses per season were processed for el ectron microscopy. The volume ( m l ) o f SER/te s t i s was c a l cu l ated from the percentage SER i n the cytopl asm (measured by point counting of 12 to 20 e l ectron m i c r o g r a p h s per st a l l i o n ) , percentage LC cyto p l asm, and parenchymal volume. Number of LC was c a l cu l ated from t h e p e r c e n t a g e L C n u c l e i , parenchymal volume, h i s t o l o g i c cor rection factor, and volume of s i n g l e nucleus. IT was deter m i n e d from the contr a l a t e r a l testis by radloirrrn u noassay. SER/g ( 89±4 vs 102±7µ1 ) , lT/g ( 0 . 58±0 .08 v s 0 . 7 0 ± 0 . 0Bmg ) , L C number/g ( 2 1 ± 1 vs 24±1 x 1 0 6 , P <0 . 05 ) , SER/test i s 1 1 . 1 ±0 . 7 v s 1 7 . 0±l . 4ml , P<0 . 01 ) , IT/test i s ( 77±12 vs 1 1 2 ± 1 2 m g , P <0 . 05 ) , and number of LC/test i s ( 2 .6±0 . 2 vs 4 . 0±0 . 2 x l 09 , P<0 . 0 1) were greater in the breeding season . SER/testis (r::0.81, P<0.01) and IT/testis ( r = 0 . 4 6 , P<0 . 0 1 ) were r e l ated t o the number o f LC/test i s . A l s o , SER/test i s was r e l ated t o IT/te s t i s ( r =0 . 4 1 , P <0 . 01 ) . These d a t a emph a s i ze the importance o f seasonal changes In the nurrber of Leydlg cel l s on the amount of SER av a i l ab l e to produce testosterone and on I T /test i s . Supported by IHH grant HO 16773. 26 INFLUENCE OF 35°c AND MELAiONIN (HEL) ON TESTOSTE RONE (T) , TESTIS AND ACCESSORY SEX ORGANS (ASO) AFTER HCG IN JUVENILE DEER MICE: Pat Fai l , Laboratory of Reproductive Endo c r i nol o gy , Center for Life S c i ences and Toxicology, Research T r i angle Institute, P . O . Box 12194, RTP, NC 27709, Melatonin ( a e l ) capsules suppressed testicular and ASO develop•ent in juvenile deer mice (Whitsett et a l . , J. Reprod. Fer t i l . deter•ine i f : 72: 287-293, presses T , and ( 2 ) HCG. the Ninety 3-wk-old Objectives were to suppressed testis responds to nale l i g h t : 8 h dark (LO) , were treataents 1984 ) . ( 1 ) •el or high anblent teaperature sup (TR) . TRS LO+sha11+350C; 3) 27!20C or 35!2oc. were assigned to 1 of 3 l) LD+shaa+270C; Teaperatures body weight) in 1/2 the •ales, 1/2 Blood seru� collected before and 1 h after HCG was stored at -2ooc unt i l RIA for t. JICG, 2) were Silastic capsules w i t h 5 ag eel or inplanted. At 7 wks, HCG was injected ( 2 5 IU/lOg after in 16h n i c e , housed LD+me l + 3 5 0 C . without (shan) were received s a l i n e . deer randomly trunk blood and organ O ne wk weights were collected. RESULTS: At 7 wks serun T was highest in LD controls [Tr 1 ) 4425±605, Tr 2 ) 1654±235 and Tr 3 ) 1250!235 pg/n l ; {P< . 001 ) ] . One h after HCG, T was higher { P < . 0 0 1 ) in LO controls ( 3 3 , 000±3383 pg/n l ; n"15 ) , than after 35oc ( 1 5 , 123!3313 pg/n l ) o r 1".1.el Tr ( 1 0 , 042±2755 pg/ml ) . T was lowest i n nales pretreated with •el (P<.001 ) . also suppressed by Testicular 35°C weight (P<.001) and ASO were but only ael testis weight. Thus , the suppressive suppressed influence of nel and 35oc was reversed by HCG for the ASO ( P < . 0 1 ) and T (P<.03) but no t for the t e s t i s . ( E P A ICR809428801 0 ) . COMPUTERIZED ANALYSIS OF SPERM MOTI O N : EFFECT 27 OF STAGE TEMPERATURE AND STORAGE AT ROOM TEMPER ATURE AND 4 ° C . A . B . Stockwe l l * , P . J . Burns, and D , Douglas-Hami l t o n * . Hamilton-Thorn Research, 30A Cherryh i l l Park, Danve r s , M a . 0 1 9 2 3 . I n the present study a new computerized sperm motion analysis instrument , developed in our lab oratory, was used to investigate the effect of stage temperature on f r e s h ( F ) and stored semen ( 2 6 to 30 hrs . ) , Storage conditions were s low cool to 4 ° C 1 SC : - 0 . JC/min. at 3 7 ° C ) and room temperatu r e ( R T : 2 5 ° C ) . Two extended ejaculates from each o f 4 stallions were us e d . Parameters studied were: Moti l i t y , Progressive Motility, Velocity, and Progressive V e l o c i t y . The rer. u l t s were: Sperm Motion Parameters (mean ± S E ) v Vp SEMEN TEMP M Mp 74±1.1 36±0 . 6 F 57±1 . 8 2 1 ±0 . 9 6 7 ± 1 . 2 3 1 ±0.8 37 ± 1 . 8 SC 12±1 . 0 18"C 25±0 . 6 63!1 . 1 RT 21 ±1 . 1 4±0 . 3 28"C F SC RT 61 ± 1 , 9 55±1 . 7 32!1 . 7 28±0 . 8 21±1 . 5 10±0 . 7 97±0 . 8 95±1 . 3 80±2 . 4 51±1 . 0 44±1.6 36±1 . 2 F 58±1 . 3 29±1 . 0 1 1 6± 1 . 5 65±1 . 4 SC 57 ± 1 . 3 22±1 . 3 119±1 . 6 56±1 . 7 RT 42±1 . 4 14±0 , 6 100± 1 . 9 43±1 . 1 our results indicate that stage temperature has a marked effect o n several parameters studied, and therefore should be c l o s e l y controlled. A l s o , S C semen parameters appeared superior to R T . 38°C 28 VALIDATION OF A COMPUTERIZED SYSTEM FOR EVALUATING MOTILITY OF BULL SPERM. P.R. Budworth•, R.P. Amann and P.L. Chapman• Colorado State University, ir rt Collins, Colorado 80523. We evaluated the Cel!Soft system (CRYO Resources, Ltd.) for measuring the percentage of progressively motile sperm (>20 um/sec; mot) and velocity (vel) of motile sperm. Measures of linearity, lateral head amplitude and beat·cross frequency were not validated. Video tapes of sperm in filtered (0.2 um) egg·yolk citrate or egg·yolk Iris extenders (8 x 106 sperm/ml) were analyzed at 30 frames/sec. With a IOX objective and a 6.7X projection ocular, optimum minimum and maximum settings for :irc:i of .1 hull �rcrrn hcnrl were '.:'.0 '.Hid 70 pixels. Virtunlli all motile sperm were detected and debris rarely were classified as immotilc sperm if the extender had been filtered. With nonfiilered extender, debris similar in size to a sperm head were classified as immotile sperm. For mot, variation about the mean was greater when only 10 or 20 fields (about 8 sperm/field) were analyzed than 25 fields (CV "' 34, 32 and 26%; data for 8 bulls). Variation was similar when 12, 20 or 30 frames/field of view were analyzed (CV = 16, 16 and 18%). To evaluate precision, proportional mixtures of Hve and killed sperm were evaluated (4 bulls). Mot was correlated (r,,Q.97) with the ratio of live:killed sperm and the slope was similar to the theoretical slope. }.fean vel of motile sperm was similar for each mixture (P>0.05). To estimate accuracy, mot determined by computer and by "track motility" were compared for 20 samples (0 · 63% motile sperm); the values were correlated (r,.,0.95). The Cell-Soft system is precise and reasonably accurate for evaluating swimming parameters of bull sperm if the extender has been filtered and 30 random fields (about 240 sperm) are evaluated using 30 frames/field. Supported by the National Association of Animal Breede.rs. P-23 Journal of Andrology • January/February 1 98 7 29 CORRELATIONS BETWEEN FERTILITY AND COMPUTER DETERMINED S\\'IMMING PARAMETERS OF BULL SPERM. P.R. Budworth* and R.P. Amann. Colorado State University, Fort Collins, CO 80523 We determined if any parameter evaluated by the CellSoft system (CRYO Resources, Ltd.) might be useful for predicting the fertility of a semen sample. Trial I: Semen from 10 bulls was extended in egg-yolk citrate containing I of 2 antibiotics (7 to JO x 106 sperm/AI dose) and cryopreserved. Mean ( 1 000 cows/sample) 7S·day nonreturn rates ranged from 58.8 to 76.4% for the 20 samples. The percentages of progressively motile (mot) sperm or linearity (lin), linear velocity (Ivel), straight-line velocity (svel), btrnl hc�d �rnl' t i 1 t 1 . J c llh 1\. 311'1 h"ll·,·rn\� frC(]UCOC} {11_-f) uf mo1ile sperm at 0 or 1.5 h post·thaw were not significantly correlated with nonreturn rate (r.. ·0.01 to 0.34). Multiple correlations with fertility based on all data for 0 or 1.5 h were 0.39 and 0.64. Trial 2: In a competi1ive fertilization trial, frozen·thawed sperm from pairs of bulls were mixed in equal numbers to provide 4 or 5 samples per bull of mixed sperm for insemination involving each of 9 bulls. The proportion of calves sired by each bull (260 calves total) was used 10 calculate a competitive fertility index (CI) which ranged from ·46 to +24. Evaluations of mot, Ivel, and svel at 0 h post·thaw were correlated (P<0.05) with the Cl (r,,0.86, 0.68, 0.79) as were evaluations of mot, Ivel, svel, and !in at 1.5 h (r ..0.78, 0.77, 0.73, 0.66). l'.tuiliple correlations with CI based on these da1a for 0 or 1 . 5 h were (R=0.94 and 0.99). Based on the latter daia, the Cel!Soft system might enable prediction of fertility (R2>0.80). (American Breeders Service and Dr. R.G. Saacke kindly provided samples and fertility data.) Supported by the National Association of Animal Breeders. 31 HUHAll SPERH ASS!::Ssnr; CAPACITATION G, Developnental Frank Destefano*, Joseph L. Annes t * , Marcie-jo Kresnow*, Melinda L . Flock* and Steven H. Disease Control, Atlanta, Schrad e r , Centers Georgia for 30333 .cooputer-assis ted In addition, conventional methods o f senen analysis have inspection and have been subject to considerable observer variabi lity. Recently developed automated methods for senen analysis such cethod, We have used one the Computer Assisted Senen Analysis (CASA) in a large cross-sectional study status of Vietnam-era veterans, of the health The CASA system uses digitized image analysis to oeasure sperm concentration and notility (percent cotile cells, velocity, trajectory, lateral head displacem�nt amplitude and frequenc y ) , i n c ludes a and t!Orphology module for categorizing sperm head shape and ceasuring individual sperm head dimensions ( a rea,- perine t e r , length /wi d t h ratio, roundness, and major axis length ) , We present the results of semen measureeents on over 500 study participants and evaluate the influence of various denographic and lifestyle factors, such a s age, cigarette S!?loking, u s e , on the different aeasures. alcohol and drug Since the participants were generally healthy men, cos t of whoa had no reported fertility problems, the study data should be useful for e stablishing guidelines on the expected values for most of the above ceasures i n the general population. 24-P recent Bethesda, HD . correlated availab l l i t y analysis (CellSoft) provides Sherins of an with of fully s pe m i:1otlon opport11nity to i n 111 e dia d u r i ng the capacitation process . Se�en froo 1 6 t1en 11 i t h proven f e r t l l l t y �-ere provided after 36-48 and hrs abstinence . o f l i near velo c i t y (LV) (u/sec) EsticJates �-e re obtained u t i l i z i � varying nunbers of tracking points dur ing a 0 .7 sec interval o f observation ( Table I ,oeant SE) (15 Tracking points I These data 15-19 39.3tl.6 4 6 . 1± 1 , l linear velocity (u/sec) the ioportance o f the nU1'1ber P"lints of 0 ,7 during 3 .5% vi th ( Ha,r FIO analyzing rao t i o n 50 ( sec Table \.'!'!re provided LV (u/sec) 30.4±1.3 LV u t i l i zed , durif18' show the that under hu•Hln of II ) , linear 32 DISSOCIATION Of' SPDU1 FRCM t t . Zinai;an and R. did by 4hr not froni in vary that also CellSoft ced i a neasured i n sugges t that t'l!ay require even , ANIJ 5'/ELLI/'3 PATTnt!IS IN lNff.RTILE D. Vantna n . HEN. 9\ e r i n s . Develop�ntal Endocrinology Branch , NICHD. Bethesda , ttO. oedium fertile the and of d i s t ribution fron 15 lOtal a C'\<lrker Accordingly , S\l"elling we as between analyzed well as patterns of irnrninal spern and Se�n and 12 i n f e r t i l e r..en llt:!fl'! collected by fertile tor of s1.1elling pattern incubation in capacitation e<;<d i u n . nasturbation after were taken as d i s t i nguish t&il of swe l l i ng during serves would i n f t! r t i l e oe n . percent spent ( HOS) 1./'e quest ioned 1.1hetht!r capacit<1tion 3b-48 hrs abstinence and aliquots routine analysis and H(J�. The r..•M lnder was centrifuged , washed w i t h cap.icitation nedia ( Haa-FlO 111th J .5% to hunan s11il'.l up st!run a l bu(lin) and i n t o mediuco incubating lhl! Spera allowed a t J 1 ° i n 5% C Ol/954 air , Aliquots were taken <1t 'l ,4 , and b hrs for HOS which involved incubation of the final aixture at 37° for 3U n i n , fixation of the cells w i t h fornalin and quantitation of swe l l l 11�'. patterns. In contrast to recent l l t erat ure % total s11elllng in sec::e n and ._,.,e diun in both groups sio.Uar(NS) but %ope n swelling (type g) was and % res t r ic ted ��tterns were significantly d i fferent (neantSE) semen fertile infertile ( type g} fertile infertile � 2 hr l l .5t3 . 1 % 25 .St4 .4i: 2 1 . lt 5 . 3 35 . l t l . � 42 .3±3 . 2 i: 25. . 8.±-2 .ui: p( 0 .0 1 3 9 .3.:t3 .6 2 0 . 7±-2 .6 p< O .U I p<O .(J) N.S 9 8 hr 3 1 .9!2 . l HYPO- OSl-IOTlC TAIL FERTILE fro� abstinence . conditions process velocity analysi s Se11en hrs by trac ki ng 32.4±1,9 sefl!inal plas'l!a (!IS) , Ho\.'l'!v e r , our d a t a detailed 20 th:> experinental s pe rn capacitation alb1.E1in ) using after 36-48 2 hr 33.Stl.8 seroen These d a t a i n capaci t a t i o n se n n hu'llan cells/subject f e r t i l e donors of h:cordl ng l y , we evalua ted LV in sea.en arr! a f t e r prolonged incubation nedia , 20 3 1 .3±0.8 p<0.01 p<0.01 de"1onstrate tr�cking points in assess ing LV . during it has been impractical to perform semen have been labor-intensive and tine-consuming, s ystea1 been TH£ ANALYSIS R. Recent data suggest that t a i l s11elling on exposure to analysis in large population studies because the methods substantially overcom� these problems. The SEMEN and NICIID, Branch, prev iously auto>Jated spern function. occupational exposures may adversely affect male relied on subjective visual has ANO DURI/'.'G SEHEN Zinocan assess spem velocity obj e c t i v e l y und e r experiBental conditions. 1./'e decided to exanine the accuracy o f Cell Soft <Jhen e s t l n a t i re h1.1;an spern notion both in seraen hypo-ost'!O t i c Despite concerns that various environ�ental and fertility, H. potent ial . character i s t i c s IN AUIOMATEO usu� Vantnan, Endoc rinology tiotil i ty assessing 30 D. fert i l i t y cnore AUTOMATED SEMEN ANALYSIS IN LARGE EPIDEHIOLOGIC STUDIES VELOCITY PROCESS Koukoulis• , S pe m Vol. B 4 i1.r 3 0 . fl t 3 . 3 37 .9t3 .9 /LS 8 hr 23 .Stl . 7 47 ,9.;t2 .b p< u . 0 1 36 :1:1;1.:3 .3 32 .3;t2 .3 14 .9,tl .U 16 .lt2 .3 p( Q , 0 1 ____� ____rS conclude that our nod i f i c a t i o n ot HOS "'.ay be useful We in d i s l lnguishing f e r t i l e fron infi.rtlli. nen. No. 1 TWELFTH ANNUAL MEETING DIFFERENTIAL EVALUATION OF HOS TEST MIHlMIZES FALSE POSITIVE FERTILITY RESULTS. R. Basuray, S. Tarchala, H. Van der Ven*, M.Perez-Pelaez, R, Jeyendran. Insti tute of Reproductive Med i c i n e , Chicago, IL, Dept. Ob/Gyn , Univ. of Bonn, FRG. The hypoosmot i c swel l i ng (HOS) test, which measures the functional i ntegrity of spel1!l membranes, has been shown to be more reliable in predicting the outcome of IVF than the standard semen parameters (J. Androl , 7 : 190,1986 ) . Based on these data, 60% or more reactive spenn i n the HOS test was tentatively considered n ferti l e" . Although false negative fert i l ity results were minimized by this 60% cut-off level , false posi t i v e results were st i l l preserit. To minimize t h i s , a differential evaluation of HOS test was performed on 31 ejaculates before and after IVF "swim-up" procedure. Though there was a significant di fference {p<0.01) on the HOS test between 24 fert i l e (mean±SEM; 85.4±1.8) and 7 inferti l e (72.1±3.6) ejaculates, both groups had more than 60% HOS-reactive sperm. There was no significant di fference on the di fferent ial evaluation of HOS between fert i l e or i nfert i l e groups. However, on differential evaluation, percent sperm with tai l - t i p swel l i ng was significantly decreased (p<0.05) after swim-up (10±2 . 4 ; before: 17 ,6±2 . 1 ) fn the i n fe rt i l e group. I n the fert i l e group, an i ncrease after swim-up (35.5±3,0; before: 30.7±3.0) was not s i g n i ficant, but an i nverse response was observed for whole-tail swel l i ng (fert i l e : before/after: 45.6±3.7/4 1 . 6±2, g ; Infert i l e : before/after 40/6±3.3/50.9±3 . 8 ) , These results i n<licate that differential evaluation of HOS test before and after swim-up procedure of sperm may miminize false positive fert i l ity resul t s . 33 34 EMPI.OYMENI' CF 'IHE H't'P(XS."[1l'IC SWELLIOO ( IDS ) TEST 'IO ASSESS 'IlIB FW:TIONAL Itmr;RIT'l CF s:;JJINE SPERM MEM8AANE . Panayiotis M. zavos and Gary w. Gregory*, University of Kentucky, Aninal Sciences Department, Lexington, KY 40546. "Previous stuiies indicated that hura.n spenratozoo reacted � exp:>sed to the IDS test oorrelated highly with the in vivo fertilizing ability of ejaculate:i spematozoo . The obje::tive of this stuiy was to determine the effectiveness of the IDS test as a means of evaluating the integrity of the men'brane of equine sperrratozoo . ·Fquine ejaculates fran six stallions of kf'IC1.ffi fertility Cn=72) .,;ere analysed for starrlard sperm pararreters and then exposed to the IDS test. (J, Reprod. Fert . 70:219, 1984 ) To assess the swelling pattern of the spenmtozoo, three different dilutions of the IDS diluent �e used originally i . e . 115, 135 arrl 150 irOsrri/l . Fran these , the 115 rrl>sn/l dilution yielded the best results ard was dlosen for further stOOies . Fran the ejaculates assessed stallions represented) the i:ercentage SWJllen (6 spernatozoa w:is 37.8+1 . 3 (range 29. 5-59 . 2 ) a.rd the rrajority of spernatozOa were rraxinally s-..ullen {coil type ) . 'Ihese firrlings suggest that equine sperrratozoo may react to hflX)OSrrotic swelling arrl that the rrost efficient swelling occurred \roilen the IDS diluent was adjusted to the 115 nOsni/l level. Further stu:lies will be carried out to establish the significance of the swelling patterns observed with equine sperrratozca. 35 THE P AND G PATTERN IN PATIENTS WITH A NORMAL SPERM MORPHOLOGY < 14%. Kruger TF*, Simmons KF•,Acosta AA. EVMS, Dept. OB/GYN, Norfolk, Va. In patients with a sperm concentration> 20 million and motility of ::-30%, but with a low normal morphology (<14%), the fer tilization rate is markedly impaired. In previous studies the fertilization rate In the group <14.% ranged between 37% and 49% and in the group >14% between 85 and 88%, (p < .0001) The specific morphological pattern was studied in 25 patients with a normal sperm morphology of <14%, 7 of 25 patients did not fertilize any oocyte at all, 9 of 25 fertilized less than 50% of oocytes and 9 of 25 patients fertilized >50% of oocytes. In 6 of the 7 patients who did not fertilize there was an identical morphology pattern, indicating a poor prognosis ( p pattern). In these patients, the average normal morphology was 2%, the severe head abnormalities (severe amorphous) 10% and slight amorphous forms 19%, In 11 of 18 patients who did fertilize at least one oocyte the average normal morphology was 10%, severe amorphous Corms 33% and slight amorphous forms 30%. (G pattern or good prognosis pattern) We propose that by evaluating the normal sperm morphology by strict criteria patients can be classified In two groups <14% and >1-4%. In the group <14% there are two subgroups with the P and G pattern which can assist the clinican to indicate prognosis and to give answers to no fertilization in an !VF Program . 36 A COMPARISOU O F NORPHOMETRY AND VOLUME O F HUMAN SPERMATO ZOA, T. TURNER, S . SCHRADER, NIOSH, CINCINNATI, OH 45226 , R . S . JEY.ENDRAN , M. PEREZ-PELAEZ, INSTITUTE OF REPRODUCTIVE MEDICINE, CHICAGO, IL, R. F, KARUHN , PARTICLE DATA LABS, "' ELHHURSI, IL, H . H . VAN DER VEU, UNIV OF BOilll, BONN, FRWG l'lor-phcmetric lll'iUUrEJrienls and sperwtozoan YOlmes were performed on 29 hlmln ejaculates which were previously analyzed for fer t i l i t y (IYF), s�n voltme, spem concentration, mrphology and mtility. These par<rneters wre analyzed to detemlne whether or not they could be used solely to pl'Edict fertility. S l i des used for routine mrphology classification were a11alyzed for rrorphcm?try which indudes sper1111 head area, perimeter, length and width. These mrphaMtrlc measurEtrenh were obtained by tracing the ou t l i ne of 200 spem heads fran each s""(>le using the Zeiss Videoplan ccnputer interfaced with a digitizing board, a Zeiss microscope and video ll'IOflitor. The enlarged imge of each spem head was projected onto the ronitor and traced with a digitizing pen, using hand and eye coordination. Sperwtoioan Yoltmes were obtained using an £1zooe l'todel 112 electronlc particle analyzer, and a General Electric T P 50 hlgh s� terminal interfaced with a Digital Oata SystO!l 1103 c�ter and ronitor. S�hs previously frozen fl'Clll the !VF proceduN! were tMWl'd and diluted for YOllf!l!'tric analysh. VolU!'i!s on ten thousand cells from each ejaculate IH!re analyzed for population ll'Ode, median and irean. Pearson ' s l i near regression correlation coefficients, weN! used to detemine the degree of correl a tion between mrpt.omtry al'!d spem Yoh.me. The best relationships existed between the geanetric n>an sperm Yohme and mrphcm:!trlc area {r:0.304, p=0 . 1 1 6 ) , perimeter (r=0.316, p:0. 101), ar.d length (r:0.314, p�.104). fven though the population size was YM l l (n;29), these paraneters were not closely reJatro al'ld prob<lbly naluated different s�nn cell characteristics. P-25 Journal of Andrology 37 A NEW METHOD OF EVALUATING SPERM MORPHOLOGY WITH PREDICTIVf VALUE FOR HUMAN*IVF. Kruger TF•, Acost� AA1 Simmons KF, Swanson RJ, Matta JF, Veeck LL: Morshedi M, Brugo S. EVMS, Dept. 08/GYN, Norfolk, Va. A prospective study was planned to evaluate the role of sperm morphology as a predicting parameter In an IVF Program. The male patients had a sperm concentration of> 20 million/ml and a motility of >30% and based on new strict criteria for evaluating normal sperm morphology patients wer-e divided prospectively into 2 groups. In group I (25 pts.) normal sp er m morphology was214% and in group 11 (71 pts,) normal sperm morphology was>l4%1 using a thereshold that proved to be of value in a previous publication. Multiple regression was used to evaluate different parameters; concentration, motility and morphology against the dependent variables fertilization and cleavage. 'Ille only factor which was significantly correlated with fertilization and cleavage was normal sperm morphology (P <0,0001). 'Ille fertilization rate (per oocyte) and the cleavage rate were 49.4% and 47.6% respectively in group I and 88.3% and 87% in group JI, (P <tl,0001) The ongoing pregnancy rate/1aparoscopy and embryo transfer was 4% and 5.5% respectively in group I and 18.3% and 18.5% in group IL This study demonstrates the value or evaluating normal sperm morphology using the strict criteria reccomended, Patients can be better counseled and the probability of fertilization or no fertilization can be more accurately predicted, 38 AND TRANSMISSION ELECTRON MICROSCOPY AllDROLOGY. Paul w. Musselman, M . D . ' Anthony J . Jr . , H.D. � James T . Kc1"'.ahon, Ph . D . , Howard S . Levin, USEFULNESS OF SCANNING IN CLINICAL Thomas , H.D.� Cleveland Clinic Foundation, Cleveland, OH, 44106 'f'ne spern of twenty men was evaluated using scanning and/or transr.rlssion electron nicroscopy (TEH) . (SEM) Eighteen men had severe asthenospermia, the average rr�tility being 12\. Two men exhibited abnornal spern head morphology. SEM and TEH findings included: ( 1 1 ) , sperm autolysis abnorma l tail morphology ( 4 ) , head dissociation axonerr.e ( 1 ) , absent dynein arms These findings were correlated with the and physical examination. ( 1 ) , absent ( 1 ) , absent acroso:ne ( 2 ) . patient ' s h i s tory Of sixteen men with no prior urologic surgery and a noroal physical examination, 4 5 \ h a d previously undergone extensive and unsuccessful trials of drug therapy . None of the 20 patients has improved or achieved a pregnancy. This data sugges t s that, scanning and/or transmission e l ectron nicroscopy selected patients with unexplained, and abnoroal spem head morphology, is of benefit in severe asthenospermia 'Ihe of a n irreversible �natomic abnoroal i t y early discovery as described in these patients should prevent costly, t�me consuming and ultimately useless therapy and allow those couples to ir<0re quickly choose other a lternative methods o f parenting. · January/February 1 9 8 7 Vol. 39 PAPER WITHDRAWN 40 THE ROLE OF SPERM MORPHOLOGY EVALUATED RY STRICT CRITERIA Ill THE HAHSTE.ll OVA/HUMAN SPERM PENETRATION ASSAY (SPA) , R, James Swanson, Thinus F, Kruger,*Hary Hamilton t Kathy F . S i=.-0mf and Anibal A, Acosta, Andrology Laboratory, Departnent of Obstetrics an<l Gynecology, Eastern Virginia Medical School and Department of University, Norfolk, Riological Sciences, Old Dominion VA 23507. A nev strict nethod for evaluation o f sperm corphology is currently used in this laboratory . This parameter proved highly predictive for fertilization rate and cleavage rate (p� . 000 1 ) in another !VF prograrn. l A prospective study evaluated the following seoen parameters in samples of all patients referred to the Andrology Laboratory over a set period of tine: sperm morphology . SPA. concentration, Eotility and normal These factors vere correlated with the The concentration and �otile spera fraction were evaluated with the aid of computer analysis (Cellsoft Sei-:,en Analysis Systen, Lab-soft Division of Cryo Resources Ltd . , New York) . Evaluation of concentration ranging from ranging froa 7% to ranging from lh 63 patients yielded a 2 to 219 million/ml, motility 87X, and normal spern morphology to 39%. T�e haaster test was performed according to a protocol described in previous publications. The SAS general linear model was used to evaluate the different variables, There was a significant regression relationship between the percent nornal sperm morphology �nd penetration rate in the SPA lKruger TF, lfenkveld R, Stander Herve JP, Van Zyl JA, Soith K. (p=.0025) . FSH, Lombard CJ, Van der Sperm oorphology as a prognostic factor in in vitro fertilization. Steril (in pre s s ) . 26-P Fertil B No. 1 TWELFTH ANNUAL MEETING 41 ll\E RELAT!OKSHIP BElllEEM SPERM FER'IILIZlllG CAPACITY IJHI THE PEHETRATIOIC OF ZOKA·FREE HAMSTER OVA BT FROZEM· TltAWED BOAR SPERMA.TOZOA. R, H. Cl arke , T. Almlid,* V. G, Pursel ,* and L.A. Johnson,* U . S . Department o f Agr\culture, Agr\cultural Research Serv\ce, Reproduction laboratory, Seltsvi l l e, I-[), 20705, Sperm fro:n two boars t hat had sho""1 wide di fferences 1 n senen quality characteri stics after freezi ng and thawing were tested for fert i l i t y and for their ability to Thi rteeri penetrate zona-free hamster {ZFH) ova in vitro. ejaculates from Boar A {h\gh post-thaw-qlia"Tity'}' and soar B { l ow post-thaw quality) were frozen by the straw method 111 a l actose-yolk extender. Eggs were recovered froro 52 sow� and g11ts 36-96 hr after i n semi nati on ' and evaluated for ferti l i zation and sperm b\nding to the zona pe l l ucida, For the sperm penetration assay {SPA). boar spennatozoa were ·acrosome-reacted i n vitro, placed ...0 th ZFH ova al 39°C, and evaluated forspenn penetrati on. sperm binding, Soar A and degree of polyspermy after 3 hr i ncubation, exhibited a signifi cantly {P<.05) higher l evel of sperm ferti lity and sperm binding {77'1. and 93- sperm/egg, respectively) than d\d Boar B {421 .. and 10 spenn/egg, respectively} . With the SPA, sperm from Boor A penetrated a higher {P<.05) n1.111ber of ZFH ova than did Boar B {431 However, .sperm binding and degre� and 241. respectively} . Based on of polyspermy did not differ bet'rlE!en boars. t hese l im-tted dat a , there may be a rel ati onshi p between sperm penetration and spem ferti l i zi ng capac\-ty, whi ch would make the SPA useful in predicting potential boar di fferences 1 n ferti 1 1 zi rig · capacity of deep frozen spenn. 42 PE!IETRATION OF HAMSTER OVA B'i !ION-HUMAN PRIMATE SPERMATOZOA. Barbara S , Durrant, Ph . D . , Zoological Society of San Diego, P . O . Box 551 , SanDiego, CA 9 2 1 1 2 Many zooa vorldwide are currently collect ing and freezing the secen of exotic ani-mals, Freezing methods employed are similar to those proven successful for the semen of domestic animals, These semen freezing techniques remain virtually untested in exotic anieals due to the lack of large nunbers of females available for artificial insemination studies. It is, therefore, icperative that laboratory analysis of semen be accurate in predicting the potential fertility of valuable sanples. The sperc. penetration assay (SPA) is being used as part of a CoDprehensive semen evaluation program, Senen of six non-human prit!.Ste species, collected by electro-ejaculation or post-aorten epididymal extraction was frozen for periods of one oonth to .five years. Frozen human sperm was used as an assay control. After a slow thaw to 0°C all seoen, regardless of freezing extender, was diluted 1 : 1 with TEST-'i and incubated a t 0°C for 1 8 hr. Sperm was heat-shocked by the rapid addition of 12-fold volume of 37°C BWW. Capacitation continued a � 37°C for l� hr. Hamster ova were co-incubated With 2 x 10 spermatozoa for 2-3 hr. Penetration was achieved with the spera of the pygcy chimpanzee, patas monkey and lion-tailed macaques. No penetration was achieved by the spern of the gorilla, uakari or lemur. Poor post-thaw motility in a l l species was correlated with failure to penetrate hamster ova. The SPA nay provide ic.portant information in addition to traditional senen analysis to deten:1ine potential fertility of spern of exotic species as well as facilitating the development of opticua cryopreservation techniques , 4 3 FO'l'ENTIAL UTILITY OF RABBIT SALT-STORED ZOHA PENETRATION ASSAY. R . A . Fayrer-Hosken and B . G . Brackett , Dept . of Physiol. and Pharmacol. College of vet. Medicine, Athens , GA, 30602. Study of sperm interaction with salt-stored zonae involved 1 . bindinq of epididymal sperm 2 . the rate of capacitated sperm penetration into the perivitelline space ( PVS) 3. the effect of uridine diphosphate (UDP) sugars on sperm-zona bindinq and 4 , the requirement of calcium for sperm-zona binding. Hean numbers of sperm attached to ssz ( n > 4 0 ) for sperm from proximal and distal oaput, proximal and distal corpus, cauda and vas deferens were O . 00, O . 1 3 , O . 14, 18 . 4 2 1 5 0 . 4 1 1 and 5 9 , 2 7 , respect ively , In vivo capaoitated sperm were attached by 10 min and increasing proportions of ssz were penetrated up to lOOt by 7 0 min. Increasing numbers of sperm reached the PVS of inseminated ssz by 270 min. UDPMannose did not affect the attachment of sperm. UDPGalactose effected a highly significant increase in attachment (P < 0 . 0001) , UDPGlucose highly significantly decreased attachment (P < 0 , 0001) . UDPH-AcetylGalactosamine decreased attachment (P < 0 , 05 ) when compared to both controls and UDPll-AcetylGlucosamine. In vivo capacitated sperm-ssz binding was partially inhibited in a calcium-free medium and totally inhibited when EGTA ( 1 mg/ml) was added. Results indicate significant influences of UDP sugars and of calcium on sperm-zona interaction and emphasize potential use o f ssz. (supported by NICHO Grant HD 1 9 2 8 8 , USDA Grant 85-CRCR-1-1-1707 and UGA VMES Project #86-2 2 5 . ) 4 4 VALUE OF ATP ASSAY IN PREDICTil->G 'IHE OUTCOME OF IN VITRO FERTILIZATION (lVF) TRIALS AND SPERM HAMSTER OVA PENETRATION ASSA'i (SPA) , M. Morshedi, * M. Hamilton , * P. Pleban, * A. Acosta, T . Kruger ,* and R. J , Swanson, Andrology Laboratory, Department of Obstetrics and Gynecology, Eastern Virginia Medical School and Department of Biological Sciences, Old Dominion University, Norfolk, VA 23508. Semen samples fron 20 !VF patients were used for ATP measurements. ATP levels (both in M/ml semen and in picoM/106 sperm) were compared to human egg penetration, cleavage and embryo transfer rates as well as to basic semen parameters. Using t-test ( t = l , 8 3 ) , a p<0.06 was observed for ATP values compared to penetration rate alone. A good correlation between sperm mean velocity and percent penetration (r=0 . 6 , p<0.01) was also observed. In a separate study of 30 patients, ATP levels of spermatozoa subjected to a swim-up separation method (SSH) at the time of inseaination, were compared to SPA results. An overall positive predictive value of 90% was calculated. Almost all patients with low ATP had abnormal SPA results. Based on a study of 22 infertile patients, ATP levels drop an average of 20-30% between reeasurements in a fresh versus SSH treated, but otherwise identical, specimen. Studies are underway to evaluate a two-srep ATP v.easurement (fresh and SSH treated spermatozoa at time of insemination) in patients with low or no penetration and cleavage in IVF and SPA procedures to determine if their poor performance is due to low ATP levels at time o f insemination, P-27 Journal of Andrology 45 USE OF PENETRAK TO STUDY SPERM PENETRATION Richard H. Harrison and Ronald W , Lewi s , PATTERN S , Delta Regional Primate Research Center, Covington, LA 70433 The standard PeneTrak test determines the distance traveled (in mo) in the cervical �ucus in 90 minutes by the vanguard spern (VA?I) . More than 50 semen sa�ples were evaluated using the PeneTrak test vith the following determinations: the average distances traveled by the lead 2 0 spertt (M-20) , lead 50 sperm (M- 5 0 ) , and lead 100 sperm(H-100 ) ; the dis tance traveled by the lOOth sperm (TA I L ) ; and the percent Samples of that distance to the vanguard distance (%VAN). were evaluated for sperm concentration (CONC ) , % norsal forns (NORM), and % not ile (KOT) . A motility index (HI) was calculated. From these data regression analysis equat ions showing the fit of VAN to M-20, M-50, H-100, and %VAN were calculated . The data were further analyzed by ranking the se11:en samples according to VAN values and determining mean %VAN values for various categories of VAN . All the regression equations had coefficients of determination The n:ean %VAN value for all specimens greater than 90%. was 6 6 . 6 % . Significant differences w e r e found when the mean %VAN values were compared for samples with VAN values o f 30 or less (54 . 2 5 ± 3 .0 1 ) with those having VAN values o f 31-40 ( 7 2 , 9 7 ± 2.89) or VAN values greater than 40 There were no significant differences in ( 7 4 . 2 5 ± 2 . 29 ) . HOT values between these groups b u t the mean HI values These studies indicate that were significantly different . the distance traveled by a single vanguard sperm is usually mean indicative of the distance traveled by the following sperm, that oany sperm parameters correlate well with the vanguard value, that grade of uotility is important in penetrating cervical nucus, and that the standard PeneTrak kit can be a research tool used for studying characteristics of sperm. This research supported by RR00 164 from N . I . H . to D . R . P . R . C , 46 EFFECTS O F CERVICAL l{UCUS SHEARING ON T H E B IOHECHAIHCS OF SPERM PENETRAT ION. 0. Katz, R. Happ*, A . Yudin*, J , Overstreet , F . Hanson*, J , Chang*, University o f Cal ifornia Medical School , Dav i s , CA 95616. Cervical mucus i n vitro i s subject t o shear forces due to contractions of the fema l e viscera. Given the hetero geneity of the mucus secretion, the res u l t is a mucus c o l ulllll consisting of a mosaic of microdoma i n s whose mate r i a l properti e s r e s i s t sperm to different extents. This heterogeneity may effectively select which sperm succeed in penetrating the mucus. We are studying the mechanics of t h i s phenomenon , characteri z i n g mucus properties on the s c a l e of individual sperm. Our "sperm probe" technique a n alyzes the relation between flagel l a r beat k i nematics and swirrming velocity to infer mucus r e s i s t i v e propert i e s . W e have found that mucus shearing does reduce microscale r e s i stance to sperm, and that this may be manifest in changes in both viscous and elastic mucus proper t i e s . The effect of the latter upon sperm mo t i o n , which has not pre v i o u s l y been addressed, may be particul a rl y important. We have also employed transmi ssion e l ectron m i c roscopy, using a new freeze -substi tution technique , to v i s ua l ize how shearing effects the mucus microstructure and i t s inter act ion with sperm. The micrographs suggest a s i g n i f i cant e l a s t i c response to sperm, and a "poreQ s t ructure which i s a l tered by shea r i n g . ( S upported by N I H grant HD1297 1 . ) 28-P · ]an11ary/Febr11ary 1 9 8 7 47 PREDICTIVE DACTERIOLO::ac VALUE STUDIES Vol. B OF QUANTITATIVE AND Ill ZONA-FREE QUALITATIVE HAMSTER EGG PENETRATION TEST FAILURE. Laszlo Sogor, Hunter Hamraill*, of Doina Glavan* and Arlette Coulter*. Department Reproductive Biology, Case Western Reserve University, Cleveland, OH, 44106. Tlo'enty-four patients had sperm which was donated for evaluation in zona-free hamster egg penetration assay and for microbiologic qualitative colonization in fashion. agar plates and anaerobic a quantitative Fresh specimens plated on and brain heart infusion broth (BHI) and blood were The grown under aerobic and anaerobic co�ditions. specimens had serial dilutions done to 10 colony forming units (CFU) per ml, and they were incubated under aerobic negative conditions. cultures for B_. All patients gonorrhea and had prior Chlamydia trachomatous sera types E through K , Eleven patients had penetration test failure based on less than lO hamster penetration. j Evaluation of the microbiology revealed 1 0 CFUs present in tests, However, had and controls that had successful hamster o the eleven failures, three of them greater than 10 ; CFU of Staphylococcus 5 epid rmidis only one patient with counts (greater than 10 CFU) of Strepviridans, Enterococcus and Staph had successful penetration with the hamster t e s t . With failures i t is possible that quantitative evaluation of bacteria acting synergistically could explain one component in certain I n this limited series cases of the failed hamster t e s t . this '-'Ould give an 80% predictive value v en the quantitation forming culture Further � of the bacteria i s greater than 10 colony units and there is a combination for mixed and Enterococcus, involving Staph epidermidis studies will and potential therapy. be needed to further clarify this 48 COMPARISOll OF SPERHAT IO AND EPIOI DYHAL SPERM ANALYSES WITH TESTIS AtlD EPIOIDYMAL H I STOPATHOLOGY Ill C{),'-IPOUND EVALUAT!Otl. S. L. Wilczynski, L . O . Russell + and J . H . Ki l l i nger, Stauffer Chemi �a l Co. , Envi ronmental Health Center, Farmington, CT, University of Southern I l l i no i s , Carbonda l e , I L Caput and cauda � i d i dymal s perm a n d t e s t i c u l a r spermatid analyses and reproductive hormone concent rations were com pared t o epididymal and testicular h i stopathol ogy during coffllound evaluat i o n . H a l e Sprague-Dawl ey rats were fed 0 and 1000 ppm of R -40244, a m a l e rat s p e c i f i c antifert i l ity agent, for I O week s . Animals were sacrifi ced after 24, 48 and 96 hours and 2 , 4 , 6 , 8 and 10 week s . Sperm analysis was performed on a caput and cauda ep i d i dymis and spermat i d analysis was performed on a homogenized t e s t i s from one s i d e of the animal . The rema i n i n g t e s t i s and e p i d i dymis were perfused and prepared for histopathology. General toxicity was indicated by decreased body weights and food consurrpt i o n . Histopathologic changes i n t h e t e s t i s due t o Sertol i cel l damage were indi cated by germ c e l l degenera t i o n , f a i l u r e of spermatid release and abnormal sperm mor phology. Test i c u l a r spermatid analyses showed simi l ar i n creases i n abnormal morphology and decreased cel l concen tration. Changes i n the testis resulted in germ cel l sloughing into the epididymis and depletion of normal epi didymal sperm. Corresponding results were seen in the caput and cauda ep ididymis with increases in abnormal sperm and decreases i n cel l concent ration and mot i l ity. Increases in FSH and LH occurred l at e r and coincided with changes i n testis and ep i d i dymal wei ght s , h 1 s t opathology and sperm parameter analyses. The r i s e in FSH and L H was secondary t o the testicular changes. No. 1 TWELFTH ANNUAL MEETING 49 NEil OBSERVATIO!IS ON THE MORPHOLOGICAL FEATURES OF THE GUINEA PIG SPERM ACROSOME REACTION . S . P . Flaherty* , V . P . Winfrey* and G . E . Olson, Department of Cell Biology, Vanderbilt University, Nashville, TN 372 32 . The Rembrane fusion events of the guinea pig spena acrosome reaction (AR) have been examined using routine thln sectioning and negative staining, Spena were induced to acrosome react e i ther vith ionophore A2 3187 and ca2+ , or by preincubation in Ca2+.free �edium with lysolecithin followed by the addition of ca2+ . Fusion of the outer acrosomal membrane (OAK) and plasma membrane (PM) results ln hybrid vesicles, and also membrane tubules which are often oriented in parallel arrays . In rouleaux, the PK over parts of the apical segcent are adherent between adjoining sperm; in these zone s , the PKs of adjacent sperm are linked by periodic cross-bridges, the PK and OAK are linked by periodic bridging elements , and the luminal face of the OAM exhibits a filamentous substructure. Fusion between the OAK and PK does not occur in these regions, and they remain as sheets after completion of the AR. Henc e , in guinea p i g sperm there appears to be a degree of directionality in the membrane fusion events of the AR, and stable non-fusigenic domains may be present in both the OAK and PK. Ongoing studies of hybrid membranes isolated on Percoll gradients aim to characterize further their s tructural and chemical properties . Supported by HD-20419 and HD-05797. 50 ANAL Y S I S OF SPERM FERT I L I Z I N G A B I L I T Y IH THE STAL L I ON . E . B u s t o s - O b r eg 6 n , Hector R o d r i g u e z * a n d Syl v i a L e i v a * , U n i v e r s i t y of Ch i l e M e d i c a l S c h o o l , S a n t i ag o , C h i l e . P r e d i c t i o n of t h e s p e rm f e r t i l i z i n g a b i l i ty based on cl assical semi n a l parameters is l i m i t e d . T h e r e f o r e , s p erm v e l o c i t y , o b j e c t i v e mo t i l i ty and A T P c o n c e n t r a t i o n w e r e a n a l y z e d f o r n c t i v e a n d f r o z e n s t a l l i o n semen d u r i n g a n d after the breeding period to broaden our u n d e r s t a n d i n g o f s p e r m a b i l i t i e s . S p e rl7! v e l oc i ty r a ng e s from 1 0 . 4 t o 1 2 . 2 u .l s e g ; A T P c o n c e n 1 t r a t i on from 0 . 8 7 t o 3 . 4 x 1 0 - M a n d t h e o b j e c t i v e m o t i l i t y f r o m 3 3 t o 38% . T h e s e t h r e e p a r a me t e r s w e r e s i g n i f i c a n t l y a n d p o s i t i v e l y c o r r e l a t e d among them and t o t h e s U b J e c t i v e m o t 1 l 1 t y a n d s p erm v i t a l i t y . Ne g a t i v e c o r r e l a t i o n w a s f o u n d w i t h s p e rm a b n o r m a l i t i e s . H o s i g n i f i c a n t changes c o u l d b e found between n a t i v e a n d f r o z e n s a mp l e s , t h o u g h A T P d e c r e a s e d in 50% after freez i n g . The new p a r am e t e r s a n a l y z e d may const i tute v a l u a b l e t oo l s f o r f i n e a s s e s sment o f s t a l l i o n seme n . ( Supported by DIB - U. of Ch i l e 0 1 464- 8655 ) . 51 �PH'iSIOI.CGIC OORREI.ATES OF Hl.W.N SPmi r03 PEllEI'AATIN3 ABILITY. AA Bronson, G1 Ccoper/' Division of fum\an Fepro:iuction, North Shore University Jbspital, and D Phillips , Pop..tl.ation Council , center for Biare::lical Research Follc1'1in} incul:ation in capacitating rredia, only a ::rna.11 i:ereentage of spematozoa have teen foond acrosare reacte:i, as judged by stained preparations of fixe:l SfeIJil at the light microscope level. selecticn of a highly notile p:::p. .i lation of ultrastnlcturally unifonn spann has allc:1.-'8d us to categorize the appearance of the acroscma in large nu:rters of sperrratozoa by transnission electron microscopy . Fol100.ng incubation in a m:xlifie:l aw1 m;:di1.IT1 o::ntai.ning hlllBn serun albunin at o:::t1Ce1t 1 rations r,;mgi.ng :Eran l. to 30 rrq/ml, the prqxirtion of acrosure reacte:l sperm was de termined and o:::nparej with the nurter of spern:a.tozoa {:ene trating zona-free hanster eggs. An increased proportion of acrosare reacted Sfel:Yil was seen at higher concentrations of HSA and oon:elaterl with greater egg penetration freqUencies. As speim rrotility was similar between treabrent gro..ips , the vaxyin;J ability of p:::ipulations of spenn to fenetrate eggs � to te on the basis of the observed differences in acroscrral status. Nota-.urthy was the fin::ling that even at the highest concentration of aJJ.:unin utilizerl, only a minority of spematozoa �'ere acrosare reacted. 'Ihese find ings suggest that capacitating cortlitions ;!!! vit.ro are inefficient, -..nen carpared with trose within the farale repro::luctive tract. Alternatively, only a small oohort of Sfenratozro. may l:e capable of acrosare reacting within a larger p:p..Uation of rrotile, rroq:bologically nonnal spenn. 'Ihese results E!fl'Fhasize the nee:I for the establisl'Brent of starrlard OOlldi.ticns ur.der Wich the fertilizing potential of h\.film si;enratozoa is judged, 52 REGIONAL DIFFERENCES IN ANDROGEN BINDING BY RAM EPIDIDYMAL TISSUE. F.R. Tekpetey an d R.P. Amann, Colorado State University, Ft. Collins, CO 80523. We determined differences associated wilh season and epididyma\ region on binding of dihydrotestosterone (DHT) to tissue from the caput, corpus and cauda (study I) or 7 regions of the epididymis (study 2). Tissue obtained in Feb·hfay (non-breeding season; NBS} or Aug (breeding season; BS) was used to prepare total low salt ( 1 4 mhi) and nuclear high salt (414 mM) extracts; low-salt extract was stripped of endogenous free steroid with dextran coated charcoal (DCC). Scatchard analyses used 2·40 nM [ 3HJDHT + I OOX excess nonradioactive DHT in a DCC assay. Addition o f ;-nd rogc n binding protein to cxtr:i.ct before DCC stripping Jid not increase binding of DHT. Relative to DHT ( 100%). binding of testosterone was 12% and of androstenediooe, estradiol, or progesterone was <4%. Concentrations of binding sites in low salt and high-salt extracts were similar. Binding (fmol/mg wet tissue) of DHT was higher (P<0.05) in BS than NBS, but affinity of l 1 1 DHT binding was about 4.7 x 10 M" in all samples. In BS, there were regional differences (P<0.05) in concentration of DHT binding sites (low- & high-salt extracts) which averaged (N .. 6 rams) I I.I, 8.7, a n d 6.3 (±0.8) for the caput, corpus and cauda in study I. In study 2, maximum DHT·binding was by tissue from the distal caput epididymidis; tissue from the proximal, central, or distal caput; proximal or distal corpus; an d proximal or distal cauda averaged (N = 3 rams) 9.4, 10.8, 13.3, 10.3, 9.1, 7.2 and 4.9 (±.0.8). For NBS, regional differences were not significant in either study and respective means were 8 . 1, 5.6 and 5.6 (±0.8) for study I and 5.8, 9.6, 7.9, 7.7, 7.5. 7.0 and 7.l (±.1.2) for study 2. These data might be interpreted as evidence that the central caput through proximal corpus regions are most dependent on androgenic stimulation (Supported by EPA CR·8127525-0I). P-29 journal of Andrology 53 THE EFFECT OF VARYJtlG FSH PULSES ON TRAllSFERRIN SECRETION Ill SUPERFUSEO SERTOLI CELL CULTURES. Andrzej Jakubowiak.*, Andrzej Janeck1 * and Anna Steinberger. Department of 08/GYN and Reproductive Sciences, University of Texas Medical School at Houston, 6431 Fann i n , Houston, TX 77030 have recently demonstrated that pulsat1 l e FSH We stimulation of rat Serto l i cel l s ( S c ) i n superfused cul tures leads to a pulsat l l e secretion pattern of transferrin (Trf) which differs dramatically from that resulting from continuous superfusion with the same hormone concentration {Jakubowiak et al , 1986 ) . In the current study we investigated the secretion of Trf over a 24�hour period in response to FSH pulses which differed i n frequency, duration and ampl itude. Confluent monol ayers of Sc from 18-day-old rats cul tured on RSM matr i x were superfused with serum-free, defined cul ture media and were exposed for 20 m 1 n , or longer, intervals to various concentrations of FSH (llIADOK oFSH-Sl6 ) . Sequent i a l samples of effluent media were c o l l ected and radioirmlunoassayed for Trf. The total amount and the secretory pattern of Trf varied depend i n g on the frequency and duration of the FSH pulses as w e l l as on the hormone concentration. The response to FSH pu l ses appeared to be dose - related and biphasic with a rap i d increase i n Trf secretion fo 1 1 owed by a drop, then a sma1 1 er more to continuous FSH prolonged stimulation. Trf response superfusion showed a di fferent dose - r e l ated and biphasic These results clearly demonstrate that Sc pattern. i n f l uenced not only by the presence of FSH secretion is but also by the mode of its d e l i very. Supported in part by UIH grant HO 17802 . 54 NEUTRAL AHillO ACID .ABSORPTial BY RAT EPIOIOYHIS. B . T . Hi nton & H. Hernandez, Vepartirent of Anatomy & rell Biology, Univers i ty of Virginia School of 1-'.ed i c f n e , Charlottesv i l l e , V i r g i n i a 22908 . For several years there has been cons i derable empha s i s upon the role of secretion by t he e p i d i dyml s , there have been relatively few studies directed towards u n de rstanding its absorptive role. Uti l izing the technique of stopped-flow spli t-drop m i croperfus i o n , the absorption of the neutral amino acid, ci-aminoisobutyric acid (AIB) from the luir.en of different epidfdymal regions was studied. Absorption from the caput, corpus and cauda regions was found to be via a time dependent and saturable system for each region w i t h s i m i l a r appKm values (6!r.M) but di fferent Vmax value s ; ?O , 26 and 68 pmoles AIB absorbed/min/rrm3 tubular vol ume for caput, corpus and cauda respectively, The absorption of AIB from the caput lumen was 'Ja+-dependent and inhibftable by 2-methyl AIB ( 2 Me A I B ) , absorption from the corpus luiren was also ua+-oopendent but not inhibi table by 2MeAIB. of AIB from the cauda luiren was Absorption Na+-fndependent and not i nhibftable by 2f'ie .AIB. Results that neutral amino acid suggest absorption along di fferent regions of the epfdidymis may occur via di fferent transport pathways; that it i s also l i nked to ou tward Ha+ movement i n proximal e p i d f dymal regions. These findings further support our prev i ous observations that there exists a selective permeabi l ity barrier from lumen to blood along the epfdidymal duct. Supported by H . I . H . grant HD18257 . 30-P · January/February 1 9 8 7 55 Vol. B . DIRECT AND INDIRECT IMMUNOLOGIC RESPOllSE FOLLOWING ACUTE TESTICULAR TORSIOU. Johnny B . Roy, Janes Hays*, Gilbert G . Haas, Sharon Einfeldt*, Brian Lansford*, and Fayaz Nahhas*. University o f Okla. Health Sciences Center, Okla. City, OK. Current reports remain contravers i a l in regard to the delet erious effect o f a unilateral torsion o f one testicle t o its contralateral counterpar t . llistologic as well as iln!llu nologic alterations have been demonstrated in several of the studies. In this report the authors investigated the i1m1unologic eff ect o f unilateral torsion both directly on the sperm and in directly in the plasma. Unlike nost o f the previous studies which based their ic. �unologic response upon visual examin ation of fluorescein labelled antibody, we employed quanti tive RIA neasurenent o f anti-spern antibody, Additionally we obtained the ejacualte by electroej aculation while in virt ually all past experiments the animals were sacrificed t o obtain sperm. Of t h e 35 adult m a l e Sprague-Dawley rats, 5 had a shao operation for control . The renaining rats were subjected to 360° torsion and divided into 3 groups . Group I : After a 2 h r , torsion, half had the torsed testis detorsed, the other half orchiectomized. Groups II & I I I had sinular events done on the torsed testis save for the duration of torsion o f 4 and 6 hrs. respectively. Four and 8 wks. later, the immunoreactivity was neasured by an RIA technique devel oped by one o f the authors (GGll) u t i l i z ing goat anti-rat I 1 2 5 IgG. Electroejaculation was obtained through the u s e o f a rectal probe operating on a 60-cycle alternating current de livering 0 - 1 5 volts. Increase i�inunoreactivity was apparent in the detorsed group compared to the orchiectomy rats. This wasevident both directly in the peripheral circulation. ( P 05) , This study showed that a tarted testicle left in the scrotum stimulates an innunologic response apparent in 4 wks. (. even after only a 2 hr. torsion. Orchiectomy o f the torted testis aneliorates & supresses the response. 56 INCORPORATION OF [ 1 4 c ] GOSSYPOL AND FORMATION OF [ 1 4 c ] GOSSYPOL-CONJUGATES IN MOUSE TRANSFORMED SERTOL I , TM4 CELLS. Nongnuj Tanphaichitr* and Lisa Fitzgerald*. Dept o f Obstetrics and Gynecology, B e t h I s r a e l Hospital, Harvard Medical School, Boston, MA 022 1 5 Gossypol preferentially causes adverse effects to mouse tran sfor�ed Sertoli TM4 c e l l s . To see if the selective effects to TH4 cells is due to their preferential drug incorporation, [ 1 4 c ] gossypol was, therefore, used for this study. TH4 c e l l s exposed t o 2 5 ug/al of [ 1 4 c ] gossypol ( 1 . 6 7 aCi/mno l e ) f o r one hr incorporated the radioactivity at 6 0 , 000 cpa/mg protein. This level was twice and ten times as high as that incorporated by marsupial kidney epithelial cells (PtK2) and nouse BALB/c JTJ fibroblasts, respectively. The incorporation kinetics o f [ 1 4 c ] gossypol at 2 . 5-25 ug/al into TH4 cells w a s biphasic. The level incorporated [ 1 4 c j gossypol reached the first plateau within 60 llli n, then rose again and attained the second plateau within 60 llli n. The level at the second plateau was twice that o f the first one, When [ 1 4 c ] gossypol-labelled TH4 c e l l lysates were dialyzed extensively against H 2 0 , half of the radioacti vity s t i l l reaained in the tubing. The results suggest that gossypol forms conjugates with TM4 c e l l s ' macromolecules. To further characterize the nature o f gossypol-protein conjugates TM4 cells labelled with [ 1 4 c ] gossypol at 10 ug/ml for l hr were solubilized in SDS-sao.ple buffer and elec trophoresed on an SDS-polyacrylamide ge l , [ 1 4 c ] gossypol-protein conjugates detected by fluorography appeared as proninant bands o f Mr 's These proteins may 6 3 , 5 9 , 5 2 . 5 , SO, 4 1 . 5 , 38 and 3 4 , 5 kd. serve as gossypol receptors accumulating the drug from the mediua into the c e l l s . Supported b y the World Health Organization grant 185057 and the Rockefeller Foundation grant #82014. No . 1 57 TWELFTH ANNUAL MEETING SPECTRAL. ANALYSIS OF SHORT-TERM FLUCTUATIONS IN SALIVARY TESTOSTERONE CONCENTRATIONS. Norma F. * * Besch, Ja.mes M. Dabbs, Jr. and Charles H. Hopper, Reproductive Research laboratory, St, Luke's Episcopal Hospita l and Department Ob/Gyn, Baylor College of Medicine, Houston, TX 77030 and Department Psychology, Georgia State Unlverslty, Atlanta1 GA )0303. Ser�m free . testosterone is believed to represent the biologically active fractlon of the total testosterone concentration and ls thus probably the more useful determination for clinical purposes. In the male, salivary testosterone concentrations are highly corre lated and concordant with serum free testosterone concentra tions. Given the non-invasive and stress-free nature of saliva c?llection, measurement in saliva is particularly suitable to . s1tuat1ons where multiple samples are required, In order to determine the degree to which testosterone concentrations in saliva show short-term episodic or cyclic variation, a study was conducted in which 1 .5 men (median age = 19) collected saliva at 2-minute intervals for one hour. Testosterone was measured in 0.2.50-ml sample volumes using an RIA with an 1-12.5 tracer and a PEG assisted second antibody separation, Each set of 30 deter minations was analyzed with the BMDPIT Spectral Analysis Computer program following removal of any linear or first quadratic components by a regression analysis. Cyclicity was demonstrated which was unique for each subjecti however' the majority showed cycles in the range of 9-1.5 per hour, or pulses every 4-7 minutes. The pulses, however, were of low amplitude suggesting that multiple sampling would not significantly increase the diagnostic reliability of the testosterone determination over that obtained for a single sample, 58 C. B. Dhabuwala and DOPPLER EVALUATION O F VARICOCELE. Anil Kumar*, Wayne State University, Hutzel Hosp i ta l, Detroit, MI 48201. Varicocele is a CO!!li!Lon surgically correctable cause of male infertility. Various oodalities such as ultra sonography, thermography, and venography have been used for the evaluation of varicocele. Doppler evaluation o f the panpiniform plexus offers a simple noninvasive oethod of assessing reflux in the venous systen of the speniatic 174 patients (348 spert!latic cords) were evaluated cord. using a dual frequency direct ional doppler with a paper recorder. The diagnosis of varicocele was based on shift of baseline during deep breathing (DB), valsalva �aneuver (VM), or manual abdominal coopression (MC) . An audible bruit during these oaneuvers helped to confirm the presence of varicocele. Shift of baseline or the presence of bruit lasting two seconds or less were not considered as indicative of varicocele. Of the 174 patients, 74 (42.5%) had left-sided varicocele, 37 patients ( 2 1 . 2%) had bilateral varicoceles , and 4 ( 2 , 2%) had right-sided varicocele. No varicocele was detected in 59 ( 3 3 . 9%) patients. The 4 right-sided varicocele patients had left orchiectooy in the past due to reasons such as testicular torsion or tu�or. During VM, closure of the inguinal canal, moveoent of the testis, and other sioilar factors may result in a false shifting of the baseline. Various pat terns of varicocele records and oethods of preventing In conclusion, misinterpretations will be discussed. doppler exanination affords a dyna�ic, inexpensive, non invasive and sensitive oethod for the denonstration of duration and intensity of reflux in the pampiniform venous plexus. 59 I S SEXUAL DYSFUNCTION OF HORMONAL ORIGIN 111 CHRONIC ALCOH OLIC MALES? Wincze, J . P . *, Engle-Fr iedaan, M.* , ?lirenberg, T . * , Leipcan, M , R . * , Brovn University Progran in Medicine, Providence VA Medical Center, Providence, R. I . 02908. Inpatients from an alcohol rehabilitation unit volunteered to participate in a prospective study to evaluate effects o f alcohol intake on sexual and horconal function on entry and a t 3, 6 and 12 conths after entry. Data is reported at baseline i n 1 8 of these patients. The patients vere 22-54 years old and had a drinking history of 12-31 years and had scores of 22-106 on the Michigan Alcoholism Screen ing Tes t , At entry patients had a drink free period of 2-44 days and had little evidence of liver damage. Ten, (56%) patients reported sexual difficulties and 6 attribut ed then to alcohol directly. Patients with sexual complaints reported significantly lower subject ive arousal while viewing an erotic Eilo (P< . 0 1 ) lJhen conpared to subjects with no cooplaints; the two groups had similar changes i n penile c ircumfrence during the erotic s t imulus and sleep. Total and free Testosterone, Estradiol, Pro lactin, LH and FSH levels lJere nornal i n the group as a In conclusion assessment whole and in the two sub groups. of 18 nale alcoholics showed a high incidence of sexual complaints i n a nornal horoonal oileu and a decreased congitive responsiveness to erotic st imulus but intact penile erectile responses. Sex Conplaints (10) No Conplaints (8) p value Free T 2 1 . 5! 1 . 0 2 4 . 8 .t l . 0 NS (pg/ol) Estradiol 29.2!5.8 30,0!7 . 2 !IS (pg/1!1.1) LH mIU/ml 10,1!3.2 7 , 5! 2 . 9 !IS 60 EFFECTS OF CYCLOSPORINE ON TESTICULAR STRUCTURE AND FUNCTION. Seethalakshmi , L . , Diamond , D . A . and Menon, M . , Uivision o f Urological and Transplantation Surgery, tr.-las s Medical Center, Worcester, MA 01605 We examined the effects of the i m�unosupressive drug, cyclosporine (CsA) on the structure and function of testis in sexually nature rats. The drug vas administered sub cutaneously in three doses ( 10 , 20 and qo mg/kg/d) for 14 days. Cyclosporine caused a dose dependent decline i n body and reproductive organ weight s . Histology o f the testis showed definite degenerative changes and sperrn counts declined. A reduction in circulating testosterone and an increase in gonadotropins and creatinine were seen. Determination of CsA levels revealed that both tubular and interstitial compartments of testis take u p CsA. Cyclosporine i s kno"�n to induce a decline in renal function (Whitirg et . al . , 1985 ) . Renal failure has been reported to affect spermatogenesis and Leydig cell function (Lim A comparison was therefore carried out and Fang, 1975). between the qo ng/kg/d CsA group and a group subjected to subtotal nephrectomy, Renal function in both groups was cooparable in terms o f seruo creatinine levels, but neph rectomy did not induce the changes in the reproductive system seen with CsA. This suggests that the testicular dysfunction seen in CsA treated rats is due to the direct action of CsA on the testis. P-31 Journal of Andrology 61 ' rrcn a g<oop �u·.i.tcq!nk failure Is a CO"ro1l e>use of Nle infertility. of 749 rnn frcn OOtrei ""rciaqe!i, 115 � C23ll '-'Ire O'!e:-1 to be infert ile pi:e;e!\Ce of spirmt()9E'nle failure "'35 to spermt09<nic failure. l « b'/ its presen:.e en a identified 'r.r/ either a rais.ed !X'l"U'l fSli l test!C1Jlar biopsy. � e'o'e Of tho� 175 r:en "ith Spef>"dtogenk failure, 65 OHl wre azoospernic Mil 110 pe r nll in at least or.e specia!n of �r e fS3'J oHgozoospernic sEO?n, Th! aqe and length of infertility at pt...Sentation -.ere sbilar in tt.e aetiOIO'JY of the b.lth n,.., azoospero;ic orrl ollgozoo5{'er"1c o::'n. spermtcqenic failure ""'-5 also very si,,.Uu in b:>th grOJP:!! , Ml�t of the testes. touion anl tr� being the mst cccron le causes. f ic 3i'rl in 80\ of the pe 7est!C11l.or site >.es rWJced in 92\ of th! sen.n �Sll "'3S ele'P.lted in 95' of the noosper>tlc oli90zoospef"1'1ic '""'° ' and 001 c.f tm 0H90ioospe mic potimts. In SH Qf the aro::.spemie Md 13\ n.e senn so ulsOO . of the oligozoospemic �n, the SetU'l Ul �\1.5 pe r c a."ld 'JoJ !82\) Prolactin le','<!ls 1.>e•e �Hur<'d in 48 nn> of too of the oligoioosper"ic non Mil ""re "°'""'l in b::lth gcoops. f<Wdc.5 & l.rub uoos m � al .uoos "1 �<\'Otypic MK>'.'"dlies ""1re pre-sent in H ¢f the aroospemic �. all of "otlich Test!<:\.!lU histology ""'"' . pe r ic Wri! 47X:(i, Md in l of ti">! ol 90 avilable frcn 12 of th!! azoospemic W frm 18 of too oli90toospereic <>?fl n- testicular lesions wre rore se•-ere in tr" azoosper,.lc tten in t graticule, �Jdlg c-eU nr.> 'i!/ ;;wtyil'll tt"<! I..en:r oligoroospe rrn lc group. trfP'!rplasia roJld be dexnstrated. .. i toos rn Vol. B EFFECT OF ALTERING SEX STEROID LEVELS WITH HUMAN CHOR IONIC GONADOTROPill OR TESTOLACTOllE 011 SEX HORMONE BillDING s.c. IDIJ'.ES, Dept. Of Cl;st�rics s G:,m�IO'J"f, lhh'ersity of lt;.tt �ha..<>, lhh'<'rslty flospltal, !btti�.,, 1C7 ?\Ai. �oo January/February 1 9 8 7 63 The C)!n!raJ feat11res of fn>•Mt""f(llc E,1!lurg N�E )I. JOOU! tfl · ...-n � A total Qf 52 of tt.e oligo?oospunic patients uere &.bjected either to Th:! resp:n� to treatnont ..,,5 v:incoo;ele tie or to "'ldical tteraP'/· ertr�ly p:.x arid no pregiuocles re,,,.,lted. This st<ld'/ �CO'\Strates ti'>!t sporrutoqenlc !allu£e r-.ly ptesent with either n.e siroilarlty in aetlol�y aru n::Qe of azoospernia or oli90toospemia. The lesion My affeo:;t pcesentatim 1oOJld suggoest the/ are the s.>.""' disease. t .e intra Nrl also the extra tutular CO'"f'Ht."E!nts Qf the te-stis. It Cd/\ be diagiosa> sl.rl>lY ard easily in <m infertility clinic and fails to respxrl to � tceatr:ent. GLOBULIN LEVEL, Richard V Clark, Janes F, J. Sherins, Depts of Medicine, Dunn* and Richard Eoory Univers i t y , A t l a n t a , GA 3 0 3 2 2 , University of Texas , San Antonio, TX 78284, Develop nental Endocrinology Branch, NICHD, Bethesda, MD 20892. The level of sex horoone binding globulin (SHBG) i s af fected by sex steroids: SHBG tends to be increased by estro gens and decreased by androgens, but their r e l a t i v e poten cies are uncertain. Indirect potent than testosterone. data suggest e s t r a d i o l i s nore in nen by in We evaluated this ducing elevations of both e s t r a d i o l and testosterone with huaan chorionic gonadotropin (hCG) . Testolactone, t a s e inhibitor, was used t o reduce estradio l . severe oligosperaia but noroal sized an arooa Five nen with and nornal sex testes s t e r o i d and gonadotropin l e v e l s , were treated with hCG (2000 IU, I M tiw) for 16-34 weeks. Testolactone a f t e r 4 weeks of hCG therapy and continued (2 g / d ) was begun throughout . SHBG was neasured by a s o l i d phase nethod using concanava l i n A Sepharose. Steroids were neasured by RIA. l\CG treatoent in creased t e s t o s t e rone 2.507, and e s t r a d i o l 4407., l e v e l s increased 3 20i. and 4607. respectively. Free horoone SflBG decreased by 237. d e s p i t e the greater increase in e s t r a d i o l . Addition o f testolactone reduced estradiol by 337, and t e s t o s terone by 2.57. though both renained ouch higher than basal values; SHBG was reduced a further 43i., to less and than h a l f the basal Two value. The ratio of free hornones changed very l i t t l e . weeks after discontinuing both hCG and testolactone, of SHBG, testosterone, levels and estradiol had returned to basal v a l u e s . A control group o f nen had stable levels o f SHBG and sex steroids during a sinilar tine period. We conclude that t e s t o s terone nay be a stronger modulator of SHBG l e v e l s than estradiol, and that t e s t o l actone oay have a direct effect in lowering SHBG. 62 PHYSIOLOGICAL ATTRIBUTES OF EPISODIC FO....LICLE STIMULAT ING � SECRETION IN �AL !-'EN, J.D. Veldhuis, A.O. Rogol, and M . L . J.C. K:lp;I, * R , J , Urban, Jotmson, University o f Virginia School o f Ner1icine ' Charlottesville, VA 22908. Although the pulsatile attributes of LH release have been defined in considerable detail there are no detailed characterizations of physiological properties of episodic FSH secretion. Accordingly we withdrew blood at 10-roin intervals For 36 hr in 5 healthy men and subjected the resulting irrmunoactive FSH time ser� ies to 4 mathematically independent methodologies for detecting underlying endocrine rhythms. By Cluster an alysis, the mean (+SfM) FSH interpulse interval deter mined From a totar of 101 significant FSH peaks was 103.:!._5 , l min, with a corresponding FSH pulse frequency of 20+ 0.73 peaks/36 hr. FSH pulse amplitudes reached maximi"l value� of 5. 3.:!.0.08 mIU/ml, represent!� a O. n ,tO. 04 mIU/ml increase above preceding nadir, Interven ing regions of nonpulsatility ("valleysu) had a durat ion of 37.:!.l . 3 min, and mean nadir concentrations of 4.4+ .26 mIU/ml , The Detect program yielded a similar estlinate of FSH pulse frequency: viz. , 22.6+3,0 puls �s/36 hr. Fourier transfoI'ination teVe"aled s1gnificant Partial intrinsic FSH periodicities every 107 min. autocorrelation analyses disclosed significant positive autocorrelations at a lag of k 100 min in each man. We corclude that the present application of multip le independent analytical rr.ethods to extensive FSH time-series can delineate pulsatile FSH release occur ring at a mean interval of 103 min in normal men. Such observations should provide a ne"# basis for investigat ing episodic FSH secretion in man in various conditions of health and disease. 1 = 32-P SUBNORKAL PROLACT Ill (PRL) RESPOllSE TO THYROTROP 111-RELEAS l tlG-HORHONE (TRH) DOES I/OT NECESSARILY SUPPORT THE D IAGtlDS I S OF HYPOGO!IAOOTROP IC HYPOGO!IADI S H , 64 . . Duarte Pignate l l i , Davide Carva l ho and Hnnuel Hargreaves Dept. of Endoc r i nology - Hosp i ta l de S8o Jo8o, Porto, Portuga I . �e studied 1 7 p a t i e n t s , 9 w i t h Hypogonadotroplc Hypogona d i sm (HH) and 8 w i th Constitutional Delay on Onset of Puber ty (COOP) , by p e r forming a TRH IV 1' njectlon (� O iig) andana l ys i n g the peak PRL response d i f ferences between the 2 groupS. The mean age of the HH group was 27 . 1 ±4 . 4 (SE) yrs and they were fol lo1-i-ed-up for 3 . 0±0.73 (SE) y r s . The CPOO group had a rriean age of 1 6 . 3±0.7 (SE) yrs and were f o l l owed-up for 2 , 9 ± D . 8 ( S E ) y r s . The mean peak PRL responses t o the IV-adml n l s tered TRH, I n the HH p a t i e n t s was 23.60 ng/ml {i6.24 SE) wh i l e i n the COOP patients i s was 3 0 . 96 ng/ml ( ±6 . 35 S E ) . The d i fference between the two groups1 response was not sta t i s t i ta c 1 l y s i g n i f i cant ( S tudent 1 s t test) . On the other hand , our HH patients cou l dn ' t a l s o be d i s t i n g u i shed fr�� the patients w i t h rr� re l y a COOP on the ba s i s of a IO'n�r se rum PRL response to TRH. In fact , in the HH group we found 4 patients (out of 9) w i t h a response greater than 25 ng/ml (peak) , and a l so , i n the COOP group, there have been 3 (out of 8) w i th a lower than 20 ng/ml response to TRH IV Injec tion . �e conclude that the d i agnos i s of HH must contlnue to rely essen t i a l l y on prolonged observation (unless a Ka l l man syndrome can be demonstrated) un t i l other t e s t s , currently under investigation w i l l actua l l y prove to be a solution to this preplexing problem. No. 1 65 TWELFTH ANNUAL MEETING EFFECTS OF I NG O N HALE Hackney• Laboratory, This study CHRON I C �.E. Kent of exercise t r a ined < LH > , at by RIA co l l e c t e d 0800 h), ( m e a n�SE) wh i l e the cu l a r the The UT and 60 y, of and were s i g n i f i c an t l y 23 . 6 !.,0 . 6 in both not the g r o up s . i n d i ca t e T and 66 and that f r ee - T t e s t i cu l a r h <B h ( s tart ing for Ne i th e r 4 group in height, h mean T and C p < 0 . 05 > 7 . 25!.,0 . 67 for The LH vs. PRL l ow e r C ET and UT of the ET 1 1 . 7!.1 . 2 l e ve l s groups. a l U/m l , and The were This l e ve l s w e r e n o t f r ee - T ma l e s l ev e l s . the the groups. C in and l ev e l s between and age, f r ee - T ng/m l , f r equency chron i c in were 45�20 m i n / d , pu l s e between PRL a n d T 4 h y p o t ha l am i c - p i t u i t a r y di ffer l ow e r LH d i f f er norma l e l evated d/wk , pg / m l , ( 1 5 . 3 �1 . 9 howev e r , tude d i d not did vs. r e s p e c t i ve l y > . e l evated suggest o v e ra l l ( 4 , 99 !.,0 . 46 p = 0 . 06 ) ; tor simi lar were were hormone <C> hypotha l am i c - p i t u i t a r y - t e s t i levels vs. T e s t o s t e rone samp l e s g roup w a s s e d e n t a r y . The 1 7 . Z:t,.1 . 4 min 6�1 fat. g r o ups , hormonal ET group had been a c t i v e d i so r d e r s group 44242. l u t e i n l z ing b l oo d A.C, endurance cor t i s o l resting every OH, and males. <El , <PRL > , in Ken t , n=1 i > nc i l > % body ET TRA I N Phy s i o l o gy reproduc t i v e <UTJ <ET; 1 2 . 4 �6 . 7 h i s to r i e s had EXERCISE A pp l i e d Uni v . , estradlol p r o l ac t i n tastJ State u n t r a i ned f ree-T, measured Sinnl n g • , compa r e s prof i l es <T> , ENDURANCE REPRODUCT I VE HORMONAL P R O F I L E S . and The a mp l i LH data f u nc t i on norma l and suggests the causes of f i nd i n g s endurance training po s s l b l y by l owe r s impai r i n g function. IBE OOLE OF SEXUAL srlf.«JIATION AID amAACl'ER.Isrrcs OORIN:; 'lliE CLINICAL USE OF =tATE A SEMINAL CDLLECI'ION DEVICE {SCD) . Panayiotis M. zavcs, University of Kentucky arrl Ardrology Institute of Lex , , Lexington , KY 40546. Recent stOOies have sh<:f,m that ejaculates collected at intercourse with the use of a Silastic seminal fluid collection device { I-S.::O l were significantly improved �n axr.p::i.red to ejaculates oollected via rrasturbation {Fertil. Steril. 4 3 : 491 arrl 4 4 : 517, 1985 ) , TOO routioo use of the I-SCD rrethod in our arrlrol03y facilities has provided ad::l itional data concerning this device. Of a group of 50 patients that produced specimens via the tWJ rrethcds , 45 patierits (90,0%) preferred the I-SCD rrethod for collection of ejao.ilates rather than rrasturbation (Mast ) , Also the same p:::pulation of patients rated sexual stinlllation elicited during production of the ejaculates (0-10) higher when the 1-s:D was used ( 7 . l l versus Mast ( 2 . 3 ) . 'Itere was a EQSSitive correlation (r=0.83) between the sexual stirulation rating provided by the patients and the TFSF values of all seminal ejaculates collected . Other features of noted irnp:>rtance �e the ease by t.tiich the I-s:D rrethod may be used at the privacy of the patient's heme , and the wife or husban:l may bring the specimen for analysis or sperm wash for artificial insemination purposes (A!Hl without tirre loss, The data suggest that increases in seminal characteristics ooted �en the SCD was used may be aS5CX'iated with increases in sexual stitrUlation elicited during the production of these ejaculates. FUrthemore, the I-SCD rretho:i as awlied, may not only irr.prove specirren characteristics b.lt also may provide other attractive practical aspects rraking the SCD quite favorable for use during treatrrent of infertility. by the clinician ar.::1 the infertile oouple . 67 EXPERIENCE WITH NEEDLE BIOPSY OF THE TESTIS AS All OFFICE PROCEDURE. R . A . Appel l , MO, J . S . Deutsch*, MD, J . R. Goodman*� MO, J . N . Macal uso*, Jr . , MD. Meadowcrest Fert i l i ty Center, Gretna , Louisiana 70056 The i n d i cations for testicular bi opsy i n the evaluation of male subfert i l i ty have been well categorized. F u l l scrotal exploration with open biopsy of the testis is a hospital based, expensive, and painful procedure with some morbidity that necessitates a substantial delay in the p a t i e n t ' s abi l i ty t o return t o work. Testicular needle biopsies were performed on 65 testes in 35 patients (at the time of submission of this abstract) in the office under local anesthesia u s i ng a new a l l - inclusive biopsy kit that con tains a modified Tru-Cut needle for biopsy purpos e s . The procedure has resulted in no episodes of bleeding or swel l i ng and minimal patient di scomfort requ i r i n g only acetominophen for analgesia. Suffi cient tissue is obtained for uncompromi sed diagnostic accuracy, including the num ber of tubules necessary in quantitative biopsy for spermatid counts . The negative aspects include a l o s s in the preservation of the interstitial tissue integrity (which is not of major import i n the evaluation of sub fertil ity) and the fact that the procedure requires an untrained assistant to stabil ize the testis during the biopsy. The technique is simpl e , rapid, inexpensive and the patient does not lose employment time w h i l e an appro priate diagnosis is obta ined. I n addition, this has a l l owed specific phar�acologic management based upon biopsy findings. Results of therapeut i c i ntervention done in this manner wi l l be presented with reference to efficacy of pharmacolog1c i ntervention with respect to specific biopsy findings. 68 IllOUCTlON OF ACROSOME REACTIONS BY THE HUMAN ZONA PELLUCIDA. 11 . L . Cross*, P. Morales*, J . W . Overstreet, and F. W. Hanson*, Dept. of Obstet. and Gynecol . , Univ. of C a l i f . Davi s . We have tested the abi l i ty of the human zona pel l ucida ( Z P ) to induce acrosorne reactions i n human sperm. Oocytes were di ssected from ovarian tissue and stored at -70°C; they are non-vi abl e . P.o t i l e sperm were selected by the swim-up technique, resuspended in 35 mg/ml huma serum albumin in modified BWW medium at 30 to 50 x 10 sperm/ml , and i ncubated at 37°C in 5% C02 0 95% a i r for 5 hr. Acro some reactions (AR) were detected as described by Cross et a l . J Androl . 7 : 28P, 1986. Oocytes were incubated 1 min with 50 x ro 6 sperm/ml (4 ± 0 . 4% AR, mean ± SEM, n = 6 ) , t o a l l ow b i n d i n g of a n average 1 7 6 sperm/ZP. One group of oocytes was fixed i rrrned iately and another was fixed after a further l hr i ncubation without sperm. Sperm on the ZP were 5 ± 2% AR at 1 min and 44 ± 5% reacted at 1 hr. The increase i n AR could be due to induction of AR by the ZP, or to preferential binding by a subset of sperm destined to complete the AR in the next hr. To distinguish these two possibi l i t i e s , we tested the ability of solubi l i zed ZP to induce AR in a sperm suspension. ZP were removed with a sma l l bore mouth pipette, and 6 ZP were solubi l i zed in I µ1 of 5 mM !1aH 2P04 , pH 2 . 5 . One µ1 of 2x medium was added , and then 1 µ1 of sperm (50 x 106/ml ) . Contro l s were a c i d i c buffer alone, or a zona-free v i t e l l u s treated with a c i d i c buffer. After 1 hr the sperm were fixed for the AR assay. The results were solubilized ZP: 24 ± 3% AR, n=3; buffer control : 3 ± 1%, vitellus control : 5 ± 1%. We conclude that the human Z P , or mater i a l intimately associ ated with i t , can i nduce AR. Supported by NIH grants HD19587 and H015149. g P-33 Journal of Andrology 89 FACTORS AFFECTING HICROINJECTION OF MAMMALIAN R.U. Clarke and L . A . SPERMATOZOA INTO HAMSTER EGGS. Johnson, U . S . D e p t . of Agriculture, Agricultural Research Service, Reproduction Laboratory, Beltsville, HD, 202705 Sperm microinjection i s a valuable tool for obtafni ng k aryotypes of. sperm that are other'ri\ se unable to fert 1 1 1 ze eggs i n vitro. Hicroi nject1on i s typi cally characterized by a lllg'f\Te9ree of egg mortality and a low rate of egg activation, which reduces the quantity and quality of metaphase spreads obtai ned frOOl mtcrot njected eggs, A study was conducted to tdenti. fy cr1t1ca1 factor{s) that tnight improve egg viability and activation aner f!licro i njection o f spenn nuclei into hamster eggs, In preliminary experiments, spenTI nuclei fr'om several mamnal s [ch1 nchi l l a, ran, hamster, and the creeping vol e (Hicrotus oregoni ) ] were microi njected i n t o 863 hamster eggswrt'lla g radually tapered needl e , After 6 hr i ncubation at 37°C, 58% of the i njected eggs· were viable and, of these, 22% were activated. In subsequent experiment s , the construction of an i njection need l e with a longer, more uni fonn barrel (5-8 µ __OD) substanti ally i ncreased both egg 11 i a bi l i ty and egg activation (92% and 66%, respectively; Mditionally, wh i l e Holmes defined mediLm was i1=1 29 ) . superior to Hams F-10 i n mai ntaining egg viabi lity and activation after microinjection, pretreatment of eggs w1 th 15-10% ethanol did not alter viab11ity or acti vation of These data indicate that size and rn i c ro i njected eggs. shape of the i njection needle and condi t i ons of egg culture are critical factors i n mai ntaining egg viabi lity anti activation after microinject1 on. Optimi zing these factors m ay enhance the quantity and qual ity of spenn karyotypes obtai ned from microinjected e<,Jgs. 10 SEMEN CHARACTERISTICS OF "UNEXPOSED WORKERS". A COMPARISON OF FIVE WORKER POPULATION S . S .M . Schrader, T . W . Turner , J . R. Burg and J . M . Ratcliffe . National Institute for Occupational Safety and Health, Cincinnat i, OH 45226 The National Institute for Occupational Safety and Health (NIOSH) has conducted five occupational f ield studies a ssessing potential reproductive hazards of male workers, as measured by semen character istics. Each of these studies vas at a different geographic location and each evaluated a concurrent control population . The first study was conducted in May, 1983 in Dillon, CO (N = 19) , the second in Deceaber, 1983 in Hilo, HI (N = 4 6 ) , the third in June, 1984 in Portland, OR (II = 3 6 ) , the fourth in December, 1984 in Groton, CT (N = 40) , and the fifth in June , 1986 in Cincinnati, OH (N = 45). T h i s report cospares the five control populations for semen vo lune and pH, and spera notility, velocity, viability (vital stain and hypo osmotic stress assay), Eorphology and norphometry. The confounding effects of age, snoking and a lcohol consuaption also �ere evaluated. The priaary conclusion eaphasizes the need for evaluating a concurrent control with each study population and not to rely totally on historical controls. 34-P · January/February 1 9 8 7 Vol. 8 71 INCREASED OEGENERATION OF GERM CELLS LATE I N MEIOSIS AUD NO LOSS DURING SPERM I O G E N E S I S JN AGED MEN. A l i Bag heri* and Larry Johnson, University of Texas Health Science Center at Dallas, Dallas, TX 75235. The amount of germ c e l l loss late i n meiosis was similar between younger (20-48yr) and older ( 58�85yr) adult men ( B i o l . Reprod. 31:779 ) , and no degeneration occurred during spermlogenes l s i n younger adults ( B i o l . Reprod. 2 5 : 2 17 ) . The object l ve of t h i s study was to evaluate the impact of germ c e l l loss during late meiosis or during spermiogenesls on sperm production i n aged men. Pote n t i a l d a l l y sperm production ( POSP/g) based on pachytene primary spermatoytes and d a l l y sperm production (OSP/g) based on early (Golgi and cap�phase) or maturation-phase spermat i d s were determined by count i n g those c e l l s in the homogenates of g l utaral dehyde perfused testes. Testes from 14 men ( 69 to 90, 77±2y) were compared with testes from 13 men (24 to 5 1 , 31±2y) , POSP/g o r OSP/g w a s the number o f c e l l s I n the homogenate times the theore t i c al yield of that cell type divided by the wei ght of t i ssue homogen i z ed and the l i fe span of that cell type. Percentage loss late in meios i s was calcul ated by d i v i ding POSP/g into the product of 100 t imes the d i fference between PDSP/g and OSP/g. No loss occurred during spermlogenes l s as noted by s i m i l ar ( P > 0 . 05 ) OSP/g based on early or maturation -phase spermat ids i n younger ( 4 . 9 ±0 . 2 vs 3 . 9±0 . 2 xl06 ) or aged men ( 1 . 5± 0 . 3 vs 2 . 2±0.5 x10 6 ) . DSP/g was lower (P<0.01) than POSP/g (10 .0±0.6 x106 for younger and 7 . 4± 1 . 1 x106 for aged ) . Degeneration late I n meiosis was greater (P<0 . 0 1 ) in aged ( 7 9 . 6± 3 . 0%) than younger men (54 .8±2 .4% ) . Aged men had an incre ased loss late in meiosis not previously detected i n older adult men. Increased germ cel l J o s s l ate i n meios i s contributes to reduced sperm production i n aged men. IHH grant AG2260. 72 EFFECTS OF ETHANOL OH tHE ACROSOME REACTION IN HUMAN EJACULATED SPERMATOZOA. Juan G . Alva r e z , * J . C . * Touchstone, * Isabel H , Lopez and Bayard t . Storey, Departaents of Physiology and Obstetrics and Gynecology, University of Pennsylvania, Phila, PA. 19104 Although some iaolated ethanol effects on ma�ma lian sperm have been documented, i t s effect on fertility in tercs of effects on human sperl!'atozoa is poorly understood. In this study the effects of ethanol on hul!l.8.n spenn acrosome reaction (AR) , as eeasured by chlortetracycline fluorescence patterns (CICF) and indirect immunofluorescence ( I I F ) , were examined. Aerobic incubation of human spena at 37°C in BVW medium ( 0 . 3% HSA) in the presence of 0 and 25 d{ ethanol ( 0 . 1 1 vol %) for 6 hr resulted in 4.±. 0 . 5 % and 23 .±. 3 % AR respectively (X + 5 . 0 , n = 1 2 ) , No effects were observed a t 0 h r . IncUbation for 0 and 6 h r i n the presence of 250 mH ethanol resulted in 25 .±. 3% and 46 .±. 5 . 5% AR, respectively (X + S .D . n = 12) , AR values for 0 mH ethanol were below 4%. Incubation of huaah sperm in the presence of the c a l c ium ionophore A23167 ( 1 0 pH) for 0 and 3 hr. resulted in 3 . 2 .±. 2 . 2% and 50 .±. 1 . 5 % AR respectively (X .±. S . D , n = 5). Interaction o f 250 EM ethanol with ionophore for 0 and 3 hr incubation r esulted in 35 .±. 6 . 4 % and 49 .±. 6 . 5% AR, respectively (X .±. S , D , n = 5 ) . No effects on motility were observed with concentrations of ethanol up to 500 irJI, These findings indicate that ethanol ingestion ll!-BY play a role in unexplained male infertility by inappropriately accel erating the acrosoae reaction in hua.an spent. Supported by March of Dimes Birth Defects Foundation, and NIH Grants HD-15842, HL-07027 and HL-19737, No. 1 TWELFTH ANNUAL MEETING 73 EFFECT OF LIVER DAMAGE , ALCOHOL , ANO RECREATIONAL DRUGS ON SEMIN IFEROUS TUBULES ANO LEYOIG CELLS IN M E N . Tho Q. Nguyen*, L . Johnson, Robert E. Wol f * , A l i Bagheri*, M ichael E. B a l l ey*, and W . B . /leaves . Un i ve r s i ty of Texas Health Science Center at Dal l as , Dallas, TX 75235. Testes from 15 control men (31±2y) , 9 men (35±3y) w i t h l i ver damage, 14 men {35±2y) known heavy al�ohol users, and 13 men (30±2y) known recreational drug users we�e perfused with g l ut araldehyde within 15 hours of death. T i ssues were pl aced In osmium and embedded l n E p o n . T h e percentage o f parenchyma o c c u p i e d b y s em i n l ferous tubu l e s and Leydig c e l l s were determined by point counting 0 . 5µm sections. For the contro l , liver, alcohol , and drug groups, respectively, the paired testicular weights ( 45±3 , 4 8 ± 5 , 4 6 ± 3 , 49±3 g ) , paired parenchymal weights (39±3, 41±4, 40±3, 42±3g ) , d a l l y spe production per man ( 1 64 ± 1 9 , 2 1 8 ± 3 4 , 196±24, 206±20 ir, XlO ) , sem l n i ferous tubu l e v o l u me/man (20±2 23±2, 22±2, 24t2m1 ) , semi n i ferous epi t h e l i um volume/man 1 5 ± 1 , 17±2, 16±1 , 18±lml ) , tubular d i ameter { 20 8 ± 4 , 225±3, 223±3, 223±5µm ) , tubular l e ngth/man { 1088±79, 969±86, 967±66, 102lt67m) , leyd i g c e l l v o l ume/man ( 1 . 4 ±0 . 1 , 1 . 2±0 . 1 , 1 . 5± 0 . 1 , 1 . 7 ±0. lml ) , volume o f a s i n g l e L e y d l g c e l l ( 6 1 7 1 ±1330 , 5070±492, 5049±489, 5464±461 11 ) , and number of l e y d i g eel ls per man { 2g6±45, 252±34, 327±37, 341±34 x106 ) reveal no signifi cant d i fference illOCln g means on most paraiie ers tested, The liver group had lower {P<0.05) L e y d i g eel l volume/man than d i d the drug group. Also, the d i <l'lleter of tubules in the control was l e s s (P<0 . 0 5 ) than the other groups. Hence, neither moderate l Iver damage, moderate use of alcoho l , nor moderate u s e of r e c r e a t i o n a l drugs i s damag ing to sem l n i ferous tubules o r leydig cel l s . NIH grant AG2260. { 74 CllS Al-ID T£STICULAA SENSITIVITY TO ETHANOL AS A RJNCTIO/l OF PUBERTAL DEVELOPMEITT. R.A. Anderson, J , F . Phi l l i ps* and S.H. Berr)man* , Rush 1".edical Center, Chicago, IL, 60612 Chronic ethanol ingestion duri09 adolescence delays sexual mat uration (Biol Reprod 32 S . l : 181, 1984). Repr-oductive impair ment is seen in pubertal males consUTiing ethanol diets 'ntlich result in minimal effects in adults, suggesting differential sensi tivity to ethano l , To excmine this central nervous systen (CtlS) sensitivity (rreasured by narcosis ! and steroi dogenic re sponse to ethanol was measured in Swiss-Webster mice at ages 23,26,30,40,50 and 60 days. "Sleep time" i ncreased with age (p< 0.05; taLAJ. 8 1 ) . ranging frcm 1.4 ± 0,4 to 111 ± 5 min at ages 23 and 60 days , respectively. Increased sleep tirre occur red at ages 35:45 days and 50-60 days (p<0.05). However, blood and brain ethanol levels at the time of awakening increased with age {p<0.05, tau=0 .78), with significant increases occur ring at 40-50 days . Blood ethanol ranged fran 278 ± 6.i to 402 ± 27 mgt fran ages 23-60 days. The data suggest both �reased ethanol metabolisn and decreased Cl\S sensitivity. Plasma andro gens {T + OHT) after 0.2 lU hCG increased with age (taw0.8, p<0.05 ) , levels ranging frcm 0.6 ± 0.4 to 29 ± 5 ng/ml at ages 23 and 60 days. There was no i ncrease in animals given 2 g/Kg ethanol (tatFO). Androgen levels were static {6.2 ± 0 , 4 ng/ml) in ethanol-treated animals fr-an ages 35-60 days, the peri od of highest steroidogenic response in control animals. Ethanol treatment prevented the increase in plasma OITT with age. In vitro steroidogenesis was not affected by 200 mg't ethanol at age 40 days, 'l'!tiile at 60 days, it was inhibited {60%; p<0.005 ) . Ethanol-i nduced delayed sexual maturation i s probably rrediated by CNS effects, due to relative insensitivity of prepubertal testes to ethanol , HCMever, ethanol suppression of plasma OHT may be partially responsible for delayed maturation, due to its possible rol e in the initiation of p..rbertal changes {Arch, AJidrol. 4: 283, 1980). Supported by NIH grant .M06604 75 THE EFFECT OF DOS£, OURATlOll AllD AGE OF EXPOSURE Oii THE EXPRESSION OF LEAD T O X I C I T Y 011 THE HYPOTHALAHIC P I T U I TARY-TESTICULAR AX I S . R.Z. Sokol , Harbor-UCLA Medical Center. Torrance, CA 90509 We have previously reported that exposure to lead (Pb) i nh i b i ts the H - P - T a x i s i n m a l e adu l t r a t s . In order to assess the s i g n i f i cance of dose, duration and age of exposure on the expression of P b toxci ty, 42d, 52d and 70d m a l e r a t s (8 rats/group) w e r e treated w i th water conta i n i ng o i , 0.1'.t o r 0 . 3 1 P b acetate. LH, T , sperm counts ( S P ) a n d P b l e ve l s w e r e measured a f t e r 30d or lOOd o f treatment. The r e s u l t s of the JOd e x p e r i m e n t were: Age Dose Pb(ug/dl) LH (ng/ml) T {ng/dl ) Sp x 106 42d 0 . ll <7 63 + 12 6 1 1 + 90 70 + 4 55 + 12 63 + - 3* 0 . 1'.t 25 + 2 577 + 77 27 + 12 318 + 64* 64 + 4* 0.31 36 + 3* 52d o.oi <7 607 + 109 103 + 8 53 + 1 1 3 1 1 + 52* 10 + 2* 43 + 9 o . 1 i 34 + 2 168 + 24* 48 + 2* 39 + 8 0 . 3 1 60 + 4* 1od o . o i <7 372 + 44 10 + 4 3o + 3 0 , li 37 + 2 29 + 7 186 + 30* 62 + 5* 83 + 17* 37 + 2* 57 + 8 0 .31 42 + 3* The results of the lOOd experiment fn 52d rats were: 0.0% <7 37 + 6 487 + 83 103 + 2 o . 1 i 30 + 5* 37 + 6 176 + 24* 10 + 2* {p <0.05) Conc l u s i on: 1) Serum T a n d spermatogene s i s are suppressed at a l l ages a n d a l l doses. 2 ) Lead exerts i ts toxi c i ty i n a dose response fashion. 3 ) I ncreased d u r a t i o n of exposure does n o t res u l t i n more severe suppression of the H-P-T axis. 76 POTEN1'IATION OF GONADOT'OXICITY OF CYCLOPHOSPHAMIDE IN THE: DOG BY ADJUVANT TREATMENT WITH AN LHRJl AGONIST. J.C. Goodpasture, K . Bergstrom* and B . H . Vickery, Syntex Research, Palo Alto, CA 94304. Relevance of the ��l e beagle dog as a r.lOdel for the gonado toxic effects of cyclophosphamide {Cl has been established p{; {J. Androl. 5 : P-28,1984) . The pur se of the present study was to determine if nafarelin (!!_) , an LHRJI agonist, by interrupting the hypothalamic-pituitary-gonadal axis prior to and during cherr.otherapy, rendering the testes "prepuber t a l " , could protect against gonadotoxicity and accelerate recovery. Four animals were dosed with N (2µg/kg/day s . c . ) 1 7 weeks after starting II, 2 animals additionally received C orally 3X/i.;eek for "-'45 weeks ('1.-6 45 mg/kg total C) . duration of ll was therefore "'52 weeks. The Plasma testosterone, spermatogenesis and ejaculation were completely suppressed by ti prior to starting C. By 2 weeks post-treatu,ent {PT} , plasna testosterone reached normal levels, and in a further 3 weeks e j aculation returned. Anirr�ls were heoicastrated at 5-9 ,,,-eeks PT. Those receiving U only began at 5 weeks PT to have spero in the ejaculate with normal motility, and normal sperm nu!Tlbers 1.;ere reached at 14 weeks PT. Animals receiving C+N had azoospermic ejaculates through 65 weeks PT, when the renaining testis was rerr.oved for histology. In contrast, C only animals (see ref) showed a rise in sperm numbers beginning at 10-11 weeks PT which reached 150xl06 sperm/ejaculate at 65 weeks PT. Testicular histology of C-+N animals at 5-9 weeks PT showed isolated tubules with apparently co!'l-plete spermatogenesis. However, In C only at 65 weeks PT no germinal cells were found. ani�a l s , spennatogenesis was nonJ>-al at this tirr.e . The addition of ti to C treatn:ent exacerbated the deleterious effects of C on the testes, and caution is therefore suggested for use of such a protocol clinically. P-35 Journal of Andrology 77 THE BFFBCTIVBNBSS OF AH IHSUJ.ATBD JAR IN PROTECTING G . E . Peterson, S . M . Schrader, T . W . Turner. Ni.tio_nal Institute for Occupational Safety and Health (DBBS) Cincinnati, OH. SKHEN QUALITY FROH TBHPBRATURB BXTRKMBS The Aladdin Thermo Jar (Aladdln •1100 Nashv i l l e , Tn) has been routinely used as a protective container for semen samples brought to field study s i tes after home collection. Three temperatures, 480 c. , 20 C . and -200 C. were used to assess the effectiveness of the containers in protecting semen quality from alterations For each temperature studied due lo temperature extremes. five ejaculates were pooled, vortexed and 3 llL were plac ed V i a b i l i t y estimates (vital Into each of three glass j a r s . stain, hypoosmo t l c stress (HOS) assay and percent motile sperm> and sperm veloc ity charac t e r i s t i c s (both pathlength end point to polnt velocities and progressive ratio} were determined before and after temperature exposures of 90 minutes, A thermocouple wes placed i n t o the senen of each glass j a r ; two jars were placed In the Insulated jars with one kept at roOtll temperature and the other placed In the controlled temperature environment along An A.HOYA with a LSD was With an unprotected glass j a r . conducted on the di fferences between pre- and poat treatment s . The i nsulated jar provided significant (p <0.0�) protec t i o n for the viability estimates at the temperature extremes but only provided partial protection These for the velocity parameters at these extremes. data Indicate that while the insul ated jar offers some protection against temperature effects on semen quality, men should be advised to avoid temperature extremes when transferring semen samples for analyses , particularly i f sperm veloc i t y measures are to be a s s e s s e d . 78 IS UOCTURNAL PENILE TUMESCENCE (?IPT) A TEST OF PITUITARY TESTICULAR FUNCTION? F. T. Kurray, M. Sciadini*, and H . Wyss*, Univ. of Florida, Gainesville, F L 32610. Nocturnal penile tumescence is a valuable tool for the HoYever, its determination of organic impotence in man. interaction with the pituitary-testicular axis reoains unclear, In an attempt to clarify the nature of such interaction we correlated age , serum testosterone and prolactin (pool of 9 samples over 24 hrs) , urinary FSH and urinary LH Yith parameters of nocturnal penile tumescence hypogonadotropic 1) in 56 iten representing five groups : hypogonadiso (n=7) ; 2) orchiectooized males with prostate cancer (na7); 3) prolactin pituitary tumors (n=9) : 4) diabetic males with nornal sexual function (n=9 ) ; 5) non"!al controls (n�24) . Data was analyzed utilizing both Pearson and Spearman correlation coefficients for the entire group of 56 subjects, S e rum testosterone correlated with all parameters of NPT including number of erections (E, r = . 5 8 , p < , 0 0 1 ) , maximal penile circumference change (MPCC, r = . 7 2 , p=<,0001) and total tumescence tiee (TIT , r=.62, p < . 0 0 0 1 ) , In control subjects there was a s i gnificant (r=-.48, p<. 02) inverse relationship between S e run prolactin was age and total tueescence tice only, inversely related to MPCC (r"- , 7 6 , p,.,02) in young controls ( 2 3 . 9 ! 1 . 2 y , n=9) but not older men (47 . 9 ! 2 . 3 y , n•l5) . We conclude that: l) A l l paraoeters o f NP! have a high direct correlation With serun testosterone, NP! correlates inversely with age in all controls and 2) seruo prolactin in only young males. 36-P · January/February 1 9 8 7 Vol. B 79 HYDROFLEX· PENILE PROSTHESI S : EARLY EXPERIENCE ANO RECO MMENDATIONS. H. David Hitcheson, New England Medical Center, Boston, MA 0 2 1 1 1 Fifteen impotence patients have been implanted with the Hydroflex (HFX) penile prosthesis since it became available one year ago, Follow up is necessarily short but useful CO!ll.!llents can already be made. The etiology of impotence was as follows: 7 patients had impaired penile arterial supply, 1 had severe diabetes mellitus, 1 haQ ideopathic hypogonadotrophic hypogonadism, 2 were para plegic, 2 were quadraplegic, 1 had empty seila syndrowe and had been previously irradiated for prostate cance= and one became impotent after radical prostatectony for prostate cancer. Noteworthy is that the man with empty sells syndrome received testos terone shortly before developing prostate cancer. Patient and partners con pleted questionaires before implanation and 2 and 12 weeks later. Penile measurements were nade in the flacid state, after erectorset tumescence and after HFX impl�n· tation. The HFX inplantation measurecents when compared to erectorset were excellent as were tunescence neasure· ments .. Deflation characteristics were good except in 1 . This Yas due to kinking o f the fluid passage way. Operative recott:Dendations will be made to show how best to determine the exact fit and to avoid complications. Most patients were comfortable after two weeks and learned to use the device in 2 sessions. Apart from the 1 deflation difficulty there were no early mechanical failures. 12 patients considered the device excellent, 2 good and 1 terrible. Partners were equally pleased. The HFX penile prosthesis provides excellent treat ment of impotence regardless of �tiology. 80 PENILE PLETHYSMOGRAPHY ON IMPOTENT HEN USING VACUUM CONSTRICTOR DEVICES. Joel L . Harmar, Thomas J . DeBenedictis, and Donald E . Praiss, Robert Wood Johnson School of Medicine at Camden, Camden, II. J. 08103. Vacuum-cons trictor devices ( V C ' s ) have been used success fully by over 6 , 000 impotent men. These devices create an erection-like state Yhen blood is drawn into the erectile tissues of the penis by a temporary vacuum, and tumescence is naintained by a constrictor ring placed at the base. Although these devices are non-invasive compared to penile implants and intra-corporal injections, they have been used less often because the early models were cumbersome and there Yere still some questions concerning penile blood flow during constriction. In this report we examined 31 patients with new V C models and rings. The patients were classified by diagnosis and penile/brachia} blood pressure index (PBI ) . Twenty-nine of 31 oen achieved satisfactory erections (buckling pressures) 450 go.) including 6 patients with PBI< 0 . 7 , 2 patients after implant reooval, 2 patients with poorly functioning inplants , 1 2 diabetics,and 7 with psychogenic problems. Penile plethys�ography yas recorded before, during, and after constriction. The amplitude (highest point on the pulse-pressure curve) declined by 70-75h during constriction but the blood flow was continuots and the crest tines (time from the start to highest point on the curve) reoained unchanged.After removal o f the rings the amplitude returned to baseline levels in all men, including those vith< 0 . 7 PB!. Thus, the new VC models and rings seem safe and effective for a variety of patients. No. l TWELFTH ANNUAL MEETING 81 TESTI CULAR B I OPSY ltl THE EVALUAT IO/I OF KALE l tlFERT l L I T Y . Miguel Dfaz, Fernando R i v a s , L i d i a Garcfa,. Horacio 83 Rivera and Jose Harfa CantU, Serv l c lo de Andrologfa, Hosp i tal de G l neco-Obstet r i c l a ; and D i v i s l6n de Gene t l c a , Unldad de lnvestlgac16n Bltmedica; Centro He'.dico de Occidente, I n s tl tuto Hexicano d e l Segura Soc i a l , Guadalajara, Ja l l sco, HEX I CO. - C l i n i ca l , hormon a l , h i s t o l og i ca l , and cytogenetlc data from 134 azoospermic men are presented. Med i c a l h i story, gen i ta l examination, serum gonadotroplns leve l , karyotype, and tes t i c u l a r biopsy for h i s t o l o g i c a l study were obtai ned from a l l patients, except In 8 i n d i v i d u a l s who n�r e not karyoty ped. The patients were d i v i ded in the f o l l ow i ng groups: only Sertoll ce l l s In tubules (SCO, n=66) ; 47 ,XXY K l i nefel ter syndrome (K, n,.26) ; obstruction (n:20) ; spermatogenes i s arrest ( n = 1 2 ) ; and orch l t l s sequelae (n:IO) , Besides group K , only two patients (both froo g roup SCO) had an abnormal karyotype: one a de novo del Yq and the other a fami l i a l t ( X ; J ) (q26 ; q 1 J . 2), -- Wi thout con s i d e r i n g the results of hi stological stud i e s , I t was observed that c l i n i c a l data, gonadotroplns leve l , and karyotype are suff i c i ent for a group d i agnos i s I n over 75% of patients. I t i s concluded that testicular b i opsy shou l d be performed only I n selected cases or for other spec i f i c studies or ves t i g a t i o n s . 82 CHARACTERIZATIOll AND QUANTITATION O F WHITE BLOOD in INFLUENCE OF HACROAGGLtrl'IllATING (HA) , MICROAGGLUTINATING (HI) AND IMMOBILIZING (I) ANTISPERH ANI'IBODIES ON THE HAMSTER OVA/HUHAN SPERM PENETRATION ASSAY (SPA) , Mary Haailton,* Anne Bogeart,* t:ahmood Morshedi , * Steven Ackerman and R. James Swanson , Andrology Laboratory, Department of Obstetrics & Gynecology , Eastern Virginia l!edical School and Department of Biological Sciences, Old Dottinion University, Norfolk, VA 23507, The results of the SPA and 3 !E!llunology assays in 156 infertile patients were compared to deten:iine the possible influence of antispern antibodies on the SPA outco�e. Positives were defined as an SPA rate o f 20% or above and a serum titer of 1/16 or above. Overall, 30% (47/156) of the patients had positive SPA results and 13,5% (21/156) dell!Onstrated positive antibodies i n their sera. Antispen!I antibodies did not appear to affect SPA, since positive SPA results were observed i n 29% (6/21) o f patients positive for antibodies, compared t o 30% (41/135) negative for antibodies, Examination of the individual itt!!lunological assays showed 31Z (4/13) o f HA-positive patients were SPA positive, In contrast, only 20% (2/10) of HI-positive patients and 0% (0/4) of I-positive patients were SPA positive,_ Although not statistically significant due to the small number of antispenn antibody positive patients tested i n this study, the observation that the presence of HI and/or I antibodies reduces positive SPA outcones warrants further investigation. 84 AN IN VIVO TECHNIQUE FOR SCREENING IMMUNOLOGIC FACTORS CELL SUBSETS IN HUMAN SEHEN, Hans Wolff*, Heidi H , P . Faris*, Joseph A, H i l l * , Joseph A. Politch*, and Jerome H. Check and Chung H. Wu, The Jefferson Medical Department of Obstetrics and Gynecology , Harvard Medical School, Boston, HA 0 2 1 1 5 , Occasionally, the physician is faced with the dileinna o f both the spermogram a n d the cervical nucus appearing normal Deborah J, Anderson, Fearing Research Laboratory, It i s known that white blood cells are present to varying degrees in the semen of both fertile and infertile men, even in the absence of apparent genitourinary tract infection, Little is known about the significance of these leucocytes, or whether certain white blood cell subsets are associated with characteristic abnormalities of semen. Recent studies by our group have provided evidence that some lynphokines and monokines in vitro cause significant impairment of spermatozoal--;totility. One aim of this study was to evaluate whether in vivo there are associations between the presei\Ce""""(i"f"certain white blood cells (as a potential source of the illtll!lunologic IN THE ETIOLOGY OF THE UNEXPLAINED POOR POST-COITAL TEST. College o f Thocas Jefferson University, Phil a . , Pa. 19107. and yet no sperm is seen with progressive forward motion (PFM} in the cervical nucus several hours after insemina tion or intercourse. By comparing the ability of donor spern versus husban d ' s sperm to denonstrate PFM in high dose estrogen (HDE) s t iculated mucus the couples could be separated into categories of either a nucus or a semen pro blem where there would be a strong possibility o f anti spermato�oa antibodies (ASA). Husband ' s sperm showing PFM with HDE alone would not suggest an icnunological problem and treatment with a HDE-human cenopausal gonadotropin the rapy would seem appropriate. Despite evaluating 4000 infer tility couples only 23 could be found to have a poor post aediators) and altered sperm motility patterns, coital (PK) test despite apparently norcal appearing sperm and nucus where an il'!l'llunological problem would be strongly helper/inducer, T cytotoxic/suppressor, and B classes tients and neither husband or wife had IgA antibodies (AB). Granulocytes , macrophage s , and lynphocytes of the T suspected. Improved PK with HDE alone occurred i n 7 pa were identified i n semen smears by applying a panel of cell-specific monoclonal antibodies i n an 3 of 7 conceived with HDE-hMG therapy. white blood cells are present in all semen samples to achieved a pregnancy. 2 nen achieved a pregnancy following clomiphene therapy and 1 had positive AB. No sperm with LPH was seen with either AID or AIH in 4 couples. Only 1 woman i1111unoperoxidase 11 assay, greatly varying degrees. Our results indicate that The lowes t numbers are recovered from semen of vasectoaized men, Studies on a possible relationship between white blood cell subsets and abnormal motility patterns are currently underway and these results will also be presented. Supported i n part by the Hax Kade Foundation and by the Fearing Research Laboratory Endovment, �2 couples showed good PK with AID but not with the husband. 7 men had posi tive AB and 6 were treated with corticosteroids and 5 had positive AB and she conceived following corticosteroid therapy. Thus, by using the HDE technique i n patients with an unexplained poor PK test those patients with ASA o f non ·clinical importance cay be separated from those where a positive test is of signif icance and the patient may re spond to corticosteroids. P-37 Journal of Andrology 85 ASSOCIATIO..'l OF SPERM AGGLUTINATION AND IMMOBILI ZATION TESTS WITa IMMUNOBEAD ( I Y.B ) man, A, Bogea r t , • A. Laboratory, ASSAY . M. Acosta, and R , J , Morshedi , • s . Swanson, Acker Andrology oeparto€nt of Obstetrics and Gynecology , Eastern Sera of 38 infer Virginia !-'.edical school, ?lorfolk, VA 23508. tile patients specifically sent for evaluation of antispern antibodies were tested to deterr:Une the association o f : standard tests micro (1 ) 0 (ST) for spern agglutination and iu.mobilization {1 1] test1 1 (2} �.aero (SA) above tests with SA observed in 6asic semen analysis and (3) centri fugation of IHB results. (1) (Kl , results o f the (bsa} 1 Fifty percent (19/38) o f sera were positive for at least one ST. All but one ST positive sera demonstrated Il'J3 attachments but only 30\ proved significant and incidence for ing was 85\, IgA, 69\ and 40\ respectively. with positive ST (K) IgG and IgM bind '!he one ��le serl.ll'l and no IMS attachment induced head to head binding of l!'IOtile donor spern in groups of 2 -S sperm. '!his phenomenon was observed in another 111ale serum. with pos itive K and strong IgG binding. Heither patient shon-ed SA in bsa. Sixty-one percent (14/23) of sett�n samples whose sera were tested showed no agglutination in their bsa. (9/14) had positive ST and I!-03, 64\ tive. Only 71\ (5/7) positive whereas all Of these, 3 being strongly posi o f samples with significant SA were ST (7/7) gave I!IB a t tachnent. ating the effect of centrifugation on IMB When evalu test results, 5 sera mixed and incubated with donor sperm were washed 4 x each and spun at either lOOOxg or 300xg. Four positive ST sera gave no IJ>!B attachnent at lOOOxg b u t gave p<Jsitive IMB binding after 300xg centrifugation. re!I'lained negative in the I!-03 test. (1) 'Ihe negative ST serum Results indicate that: 1113 test i s �-Ore specific and should be included in any i�i111unologic evaluation of infertile groups; (2) low centri fugation force i s essential to process saoples o f I�:B test1 and 86 (3) lack o f SA r:iay not prove (-) January/Febntary 1 9 8 7 Vol. s 87 IN'IRACORWAL SrnEN DEPCSIT!QN AT DIFFERENI' DfPlliS AN) SUBSE1;1JF.NI' Fm"l'ILIZM'ION AID EMBRYONIC IEVEl.DPMENI' IN SUPEROVU!ATED o:MS . Jctm Fernandez-VanCleve *, Panayiotis Zavce an:i Ro;jer W. Henken* , University of PUerto Rico-Maya.quez aOO. university of Kentucky, Dept. Animal "Sciences, Lexington , KY 40546. This stuiy was designed to assess the effects of intracornual ( IC) serrell dep:>sition at different depths or. fertilization rates (FR) an:i errbryonic retardation (ER) , �rovulated Holstein ct::/NS �e bred at 10.9+2.2 hr after the onset of estrus with a single dose of frozen serren from the same 0011. Animals �e grouped by site of A.I (TFS/group): l l right uterine horn (UH) 5 cm deep; 2 l left UH , 5 cm deep; 3 l right UH , 10 cm deep arrl 4) left c.rn, 10 an deep . The nurrber of unfertilized ova, norrral arrl atnorrral errbryos were recorded for each horn an::l depth at slaughter .. (5 d p::>St Al l . Entiryos, recovered from the horn \\here AI was i;;erforned. (ipsilateral ; !PS) were p:x:iled arrl carpared to arbryos recovered fran the oontralateral (CDNJ horn per depth. FRs -were higher in errbryos recovered from !PS than that of mN at 5 an (83.1% vs 43.4%; P<0.00 2 1 , respectively . FRs �re higher for errbryos recovered frcm the mN at 5 cm than those recovered at 10 cm CP<0 . 03 l . ER did not differ between IPS arrl OON at 5 an deep ( 5 . 1% vs 8.6%; P>0.101 or at 10 cm deep (8.3% vs 26.1%; P>0.05 ) , respectively . '!he results showed oo improvement in FR ....tien inseminating into the !PS horn deeper than 5 cm. In oontrast, IC-Al into the !PS horn deeper than 5 an nay decrease spenn migration to the CON horn arrl that rm.y result in lower CR (P<0.002) arrl/or possible increase in the erbryonic retardation ratio. M, in ST or I�:B test. COMPARISOtl OF THE SPERM PEflETRAT!Oll ASSAY (SPA) WITH OBJECTIVE !J.OT ILITY PARJIJ1.ETERS l . R . Kossoy*, T . Thompson*, A . Buck*, M. Hinson*, B. Bro<lie*, W. Vaughn*, A . C . Wentz*, B . J . Rogers, Dept. OB/GYrl, C-FARR, Vanderbilt University School of !J,edicine, tlashv i l l e , TfL 37232 The sperm penetration as say (SPA) has been used extensive ly to evaluate the fert i l i ty status of men. I t is a tedious and time consuming bioassay that requires ski l l ed ln the ongoing quest for a more per sonnel to perform. objective para'Tleter of sperm functional capacity we have eval uated so.11e rnot i l i ty parameters us i ng the Cel l Soft Automated Semen Analyzer and compared them to SPA results. Semen specimens from 29 patients were evaluated and the data analyzed using multiple l i near regres sion. The µarameters incl uded were moti l i ty { % ) , concentration, maximum aJll)l i tude of lateral head d i s p l acement (ALH max ) , mean ampl itude of l ate1·al head d i s p l acement (ALH mean), be at/cross frequency, velocity, l i nearity, penetration { % P ) in hamster eggs, and penetration index ( P I ) . tlot surprisingly a statistically significant correlation was found betn-een l i nearity and rr'3 ti l i ty ( P -" 0 . 0 2 6 ) . The P value for percent penetration vs ALH max was 0 . 058 and vs 1\LH rnean was 0.069 which failed to reach statistical The values for PI vs ALH max and ALH mean s i g n i ficance. were hon-ever s i g n i ficant (P�0.034 and 0. 039 respectively). A l l other correlations were not found to be significant. These results suggest the potenti a l usefulness of l ateral head di splacernent measurements in rna le ferti I ity eva l ua ti ons since there appears to be a s i gn i ficant correl ation with the penetration i ndex pararr.-eter of the SPA. The eval uation of l arger groups of individuals w i l l be " required to val idate the c·or.:!:!la: i9ns presented here, 38-P · 88 EXPERIENCE I/ITH IIHRAUTERINE INSEHINATION IN INFERTILE COuPLES. Arnold M. Christine L. Cook, Belker, M . D . , University of Louisville, H . D . * , Leigh T. Price"=, and Dept. of Obstetrics and Gynecology Kentucky 40292 Intrauterine insenination is suggested in cases of infer tility where cervical, nale, C03bined or unexplained factors are present . One hundred fifty-seven patients underwent intrauterine insemination (IUI} for a t least cycles (range 1-18) or until they conceived. Most patients had nultiple infertility factors. The overall pregnancy rate was 20%. The pregnancy rate in 41 factor infertility was 19%, spern antibodies, 0% in bodies, mucus, mucus, couples with oale 10% in 10 cases of male anti 8 cases of fenale antispero anti 35% in 20 cases of poor postcoital 42% in 12 cases of poor postcoital test with good test with poor 30% in 10 cases of cervical stenosis and 18% in 11 cases of unexplained infertility. None of the patients with mild ol igospermia as an isolated diagnosis or tubal or uterine factors as the primary diagnosis ( i n couples having multiple diagnoses) conceived. When couples with asthenospersia in addition to mild (10-20 x 106/m.l) or severe ( < 10 x 10 6/ml) oligospernia were evaluated separa tely, the pregnancy rates were 6% and 4 0 % , respectively. Spontaneous abortion occurred in 6 of the 31 pregnancies ( 1 9% ) , An ectopic pregnancy occurred in one case of poor postcoital t e s t with poor nucus {8% ) . The average number of cycles to conception was 3 . 6 ; no one conceived after the 9th insemination cycle. Intrauterine inseoination appears to b e particularly valuable in couples with poor postcoital tests and with asthenospernia associated with severe oligospernia, No. 1 TWELFTH ANNUAL MEETING EFFECT OF NEURAMINIDASE ON SPERM SEPARATION 89 FOR SEX SELECTION. Hugh C . Hensleigh* and Denise T r u z i n s k i * , D e p a r t m e n t of O b s t e t r i c s and Gynecology , University of Minnesota, Minneapolis, MN 5 5 4 5 5 . Human s e x select ion by sperm separation is being done cl inically by the albumen swim-down technique that selects for Y-bearing sperm (Ericsson, et al, N a t u r e 2 4 6 : 4 2 1 , 1 9 7 3 ) and the Sephadex column techn ique that selects for X-bear ing sperm ( S t e e n o , e t a l , Androlog i a 7 t 9 5 , 1 9 7 5 ) . Kaneko repor ted that s i alic acid residues on the surface of the sperm may account f o r the d i f ference in net charge observed on the surface of X - and Y-bearing s p e r m ( B i oc h e m . B i o p h y s . Re s . C o m . 1 2 4 : 9 5 0 9 5 5 , 1984) . To determine i f s ia l ic a c i d residues a r e important in the sperm separation techniques, s p e r m s ampl e s w e r e i n c u b a t e d w i t h 1 mg s i a l idase/ml (Type v, neuraminidase, Sigma) for lh a t 3 7 • c p r i o r to sperm selection. Samples were s t a i n e d with quinacrine mustard and scored blind f o r t h e p r e s e n c e of the Y-fluoresc ing body. No e f f e c t w a s s e e n u s i n g t h e S e p h ad e x c o l u m n t e c h n ique with a n average of 82 . 2± 4 . 1% X-bearing s p e r m f o l l o w i n g s i a l i d a s e i n c u b a t i o n {meant: Howeve r , t h e s . o . , c o n t r o l , 7 9 . J ± S . 5 5 % , N= 4 ) . s i al i d a s e i n c u b a t i o n n e g a t e d s e p a r a t ion in the albumen swim-down techniqu e : an average of 44. st8 . 3% Y-bearing sperm was observed (control, 7 1 . o:t s . 6% , N = 4 , P < 0 . 0 5) . These results suggest that a d i f f erential presence of s ialic acid r e s idues on sperm s u r f a c e m a y b e o f i m p o r t a n c e i n t h e a l b u m e n s w i m - d o wn t e c h n i q u e b u t n p t i n t h e Sephadex column techniqu e . 90 INCREASED RATE O P lMtAN SPERJi FERTILIZATION ABILITY BY PERCOLL-GRADIE.HT CEH'IilFUGATION. H. Tanphaichitrf A , d" and Agulnick, D , Diaz, J , H i l l , L . Fitzgeral o. Anderson. Dept. o f Obstetrics and Gynecology, Beth Israel and Brighaa and Women's Hospitals, Harvard Medical School, Soston, HA 02215 Human aperci can be capacitated i n Yitro to achieve fertilization ability, Sv1ia-up- anQP'ircoll-gradient centrifugation (PGC) procedures are routinely used for this purpoae clinically, Thi• rep<>rt is to illustrate that the PGC D4thod ia aore efficient than the svia-u p aethod in producing capacitated spera a a defined by ha.II.S t er egg penetration. In comparison to tbe svia-i.ip aethod, the PGC method yielded nonLll donors' and infertility patients� sperm that penetrated higher nuabera of zona-free haaater eggs vi.th 110re sperm/egg, Spena prepared by PGC did not ahov an increase in percent acroaoae reaction, nor did the percent · motility and velocity of motile spena differ significantly ' froa those capacitated by the avia-up aethod. However, the spera. .allbranea of PGC and svia-up sperm Ill.BY be disaiailar a.s evidenced by their different degrees of interaction with tvo aonoclonal antibodies, l lHS-63 and 8316 (gift• of or. Gregory Lee and Dr. Peter Johnson, respectively) , which recognize the the surface of spena acrosoaes. Noncapacitated and swim-up spera interacted aore with l lHS-63 than did PGC spet'll . In contrast , a higher percentage of PGC spen1 were recognized by H.3 1 6 . These results, therefore, suggest that the PGC ap•t'll 111.81 have altered surface properties that iocrea.ae their fertilization ability. 91 ABILITY OF GLASS WOOL FILTRATION TO ISOLATE FERTILE SPERM FRACTIONS, WJ Hologren, RS Jeyendran, MR Neff, M Perez-Pelaez. Mount Sinai Med Ctr, Univ of WI Hed School, Milwaukee, WI and Inst of Reprod Med, Chicago, IL. Semen filtered through a glass wool column yields an en hanced number of motile spern with chemically active and physically intact membranes (Fertil Steril 45 : 1 32 , 1986), As fertilizing -capacity is the u lticate test of sperm quality, the in vitro penetration of zona-free haoster oocyte (!VP) assay was performed to determine if filtered samples also possess greater penetrating capacity. Each sample was washed and divided into two aliquot s . One aliquot was filtered through a column containing 15 �g of precleaned glass wool microfiber packed to a depth of 3 . 5 mm. The concentration of the unfiltered control and filtered sample was adjusted to 10 x 106 /ml and incubated for 18-20 hr at 3 7 ' C , The lVP assay was performed using 1 x 106 sperm/ol for each aliquot coincubated with oocytes. Of the 10 ejaculates assayed, significantly higher (P<0 .01) !VP results were observed for the filtered sample (Medn ! S . D . ; 5 8 . 6 ! 2 6 . S ) than for the control ( 2 7 . 2 ! 20 . S ) . To determine whether the two fold increase in penetration ·:ate was due to enhanced sperl!I no tility, the IVP was repeated on five of the above sanples. However, progressive motility on the filtered fraction was adjusted prior to incubation with oocytes to the same level as in the control. Again, a higher penetration rate for filtered saoples (39.0 ! 2 4 . 3 ) was observed after adjusting motility to the same level as in the unfiltered control ( 2 4 . 2 ! 22 . 1) , Thus, the filtered saoples were controlled for concentration and/or motility to ainimize the effect of these paraoeters on the penetration rate. Therefore, it appears that glass wool filtration also effectively isolates a sperm population possessing a greater in vitro penetrat ing capacity. SPERM PREPARATION FOR ARTIFICIAL INSEMINATION 92 U S I N G RAYON F ILTERS, Hugh C . Hens l e ig h * and D e n i s e T r u z i n s k i * , Depa rtment of Obstetrics and Gynec ology, U n i v e r s i ty of Minnesota, Minneapol i s , MN 55455 Ar t i f i c i a l insemination w i t h t h e husband ' s sperm when there a r e high numb e r s of white blood cells ( W B C ) c o mb i n e d w i t h l o w or mod e r a t e c o u n t s presents spec i a l p r oblems f o r the l a b o r a t o r y prepar ing t h e s p e r m s am p l e , W a s h i n g by centrifugation g ives a h i g h recovery r a t e b u t does n o t r ed u c e t h e n u mb e r of we e s . G l a s s wool f i l t r a t i o n effectively removes WBCs b u t may leave s o m e g l a s s f ib e r s in the p r e p a r a t i on . Rayon f i l t e r s d o not shed f ib e r s , and a r e capable of remov i n g W B C s . The purpose of the present s tudy was to e s tab l i s h the p e r f o r m a n c e p a r ameters of Infer t i l i ty rayon f i l t e r s for sperm preparation. p a t i e n t s w i t h e l e v a t e d n u m b e r s of WBCs w e r e assigned t o a washed g r oup ( N "' l 3 , 3 1 cycles) o r a f i l t r a t i o n g r ou p ( N :z l 4 , 3 0 c y c l e s ) . Semen analysis and sperm preparations were d o n e followed by a c o u n t , motility estimate, and wec · c ount p r i o r to insemination. A h i g h recovery rate ( 87%) was ob s e r v e d w i t h the washed samples however the % motility and number of WBCs was not s i g n i f icantly d i f f e r en t . Rayon f i l t e r s reduced numbers of WBCs f r om an a v e r ag e number of 28 80/mm3 to less than 100/mmJ. A sign i f icant increase in motility from 56% t o 86% was observed in the f i ltered samples w i t h a 25\ recovery rate • Rayon f i l t e r s provided e f f e c t i v e removal of wees , increased m o t i l i t y , and mod e r a t e recovery r a t e s . P-39 Journal of Andrology 93 ATP DECLINE O F INFERTILE PATIENT SEMEN AFTER SWIM-UP SEPARATION ME!HOD (SSH) . H. Horshedi,* R. Acos t a , * P , Pleban,* A . A . Acosta, J . Yuan,* and R , J , Swanson, Andro logy Laboratory, Depart�nt of Obstetrics & Gynecology, Eastern Virg inia Medical School & DepartBent o f Biological Sciences, Old Dominion University, Norfolk, VA 23508-8503. In 22 infertile males, ATP content of 4 semen specimens de c l ined up to 95% when processed by SSH using Ham's FlO fetal cord serum medium, The SSH included a 10-minute 290xg cen t rifugat ion of sanple-mediun cixture followed by one-hour incubation at J7°c in 5% C02 /air. ATP values determined by cheoiluoinescence were calculated in picoM/106 sperm to ob The 4 draatic ATP declines were viate variations in count. not related to seiu?n count (oligozoospermia v s . normozoo spermia) , cotility or SSH recovery rate, All samples deconstrated an ATP decline averaging 20-30%. This finding may represent one possible explanation for the fertilization failure seen in some infertile individuals. Alternatively, low ATP values may represent a damage artifact inflicted upon sperm while processing for AIH, IVF or hamster proce dures. Recovery rates in non:.<>zoospennic saeples were consistently very low with SSH but very high with discon t inuous (40-60-90) percoll gradient separation (PGS) . In e va luating these 2 spern separation techniques in 6 patients, 4 of the 6 semen samples produced a smaller decline in ATP content with PGS. ATP values for the other 2 samples might have been compromised by elevated round cell counts (neutro phils and other types of round cells), which normally have much higher ATP content than spermatozoa. Some round cells and debris from the original sample normally remain after SSH. PGS eliminated round cells and debris. PGS is a method of choice in semen with elevated round cells or debris, or where drastic change in ATP content results fron SSH. ( 1 ) PGS replacement for Studies are underway to evaluate: SSH in low total notile recovery or low ATP content and ( 2 ) �-0difications in centrifugation/incubation protocol to aid recovery rate and stabilize ATP content. 94 POLYlACIOSANrnE 00 THE MJUSE SPERM SURFACE DURID; HAWRATIOO AND CAPACITATION Olarles H. Muller, Depts. CJ:,/Gyn and Biological Structure , lhiversity of Washingtoo , Seattle \.:A 98195 A polylactosamine-cootaining glycocoojugate (pLacNAc ) , recogiized by a m:noclonal a'ltiOOdy, first appears on the spenn surface in the mid-to-distal caput epididymidis of the rrouse. This is the sarre regioo in �ich a high nnlecular �ight, trypsin-sensitive sialylated pLacNAc appears in the apical cytoplasm of principal epididyrral cells and in the luninal fluid. Sial-placNAc in the epitheliun is independent of testicular fluid or spenn (as seen after efferent d\lct ligaticn), but ishighly dependent oo c irculating androgens. Poly l.acNac oo the spenn surface isdetected by inm.no flmrescence overlying the posterior acrosorre . Spenn from the proxirral caput are negati\e , and about 90% of sperm from the cauda are positive, During incubatioo mder in vitro fertilizaticn crnditicns,stn:face placNAc is slm.;ily lost such that 50% of sperm are positive by 60 min . In ccntrast, intracellular pLacNAc in the anterior acrosooe i s not accessable infreshly isolated sperm, but is e�sed progressively at 15,30 and 60 min of incubaticn. In nonrn.1 conditicns, no rrore than 50% of sperm are observed with the anterior acrosomal pattern. Ha"...e\er ,up to 100% of spenn are seen with this pattern after exposure t o calciun iooophore A23187, in a dose-dependent fashicn. S:inultaneously, the posterior acros0IT0l pattern is lost from iroophore-treated Intracellular placW..c is also exposed by washing spenn, or by extractioo with noo-icnic detergents. Intracellular pLacNAc appears in the forming acrosorre s of rornd spenmtids <md rray be a conpooent of the acrosomal rratrix. Spenn sm:face p Lac� rray sene as a stabilizaticn or zooa-recogiitioo (Supported by NIB Grants lID-16211 and HD-12629) factor. 40-P · January/February 1 9 8 7 95 Vol. B CAPACITATJOll Of 90VUE SPElM Bl REMOVAL Of CllOlESTEROl * * Ed\11rdo Ehremu\d , John E. Park• and Robert II. Foote Oep..a r t ,..tnt of Anfn!l sclen<:e, Cornell University I thaca, NY, 145S3-�6()1 hveral rtp-ort. hive lndlcued th1t c1p1eltuion of Memaltan spHl'l tru::ludn reduct t o n of the eholuterol (thol) to phosphollpld (Pl) r;itlo of th• phua Mebane. qu1nt l U t lvely reduce In the this study, Chot/PL rulo nethods of were developed bovine sperM and to the 1 f f e c t l ven111 of thh trntaent In c1p1ci t 1 t ln9 spern was duerrniMd. 8 (2X10 ) were lMubaied I n 1 . 0 nl of modified Tyrode's \luhed sptr!lll 1olutlon (TS) contalntna unllla.ellat l lp-osl).!;U of phosphaitdylchollne (PC), ·1th•nolu1tne (PE) ind 1 14cJ·Chot <35:35:30 111<1 l a r ratio, 300 rmol totll 3 t MJ · t r l o l e l n Pl), 11n Included n After 90 "'in iii 19°c, 1 ll:li net eAChange of C a noneuh1ngeable ioarker. H CJ ·Chol frtti1 l i poso�es to 1pern wu observtd (n•O 1nd sper111 Jl4t i l l t y was BOX. Spera were then 6 spern were tncub1ted as before with PC/PE l i poso�es wuhed ind SOX\0 contllnlna by no Chol. After 90 nln, sper111 were seperated frWI l i posoo:�s Musure1<1nt of 1 14c1-chol In the l lpose'!!-es centrtfug1tlon. (1upun1Unt) ind parallel gn chromatographic analysis of extracted, up-onlfled L lp-osonu (ns4J lndte1Ud th1t 30X of spern cholesterol was re«>ved by t h i s procedure. len thin 10X but no l l p-ost>Ms Chol efflux decrused sperm IW t i t f t y by red\iced sperm velocity by SOX. (Ml), with L i poso..es eonUlnlng Sperm Incubated with Chol {+Chol) and with Chol·free l i poSO!IU <-Chol) were washed end resuspended i n TS with o.n BSA ind 55 ua lyaophosphu ldylcholtne {lPC)/� SSA, Percent sperci IH'>der11otn11 the 1crosO!!le reaction {A�) upon ln<:ubitlon with TS·LPC was vud u a 11envre of aperll'I cap1cl u t l on. lPC, p.reent sperm n o t l l l t y 60.0+4,7 ind 57.0.!_6.& � All w re 14.0!_3 .4, Lon of Cho\ for NL, respectively. 20,3,t4.4 After 60 nin iii 39°c in TS· +Chol ind ·Chol WIS 57.0!_4.9, corresponding vatuu for percent •nd 39.7,!.l.2. These results suggest that fro• plu11-1 o:e11.br1nes of bovine spern "''Y be an iP-portant Hep in eapa � l t l l t o n In t h i s speci e s . Thh work was suppor!ed in part by NIM 9r1nt �o 1862&. 96 Ili!ERACtlOHS BETWEEN SELECTED FATTY AClDS AND THE ACTlVATION OF PALHITIC ACID IN SPERMATOZOA, Robert E . Jones and Stephen R . Plynate, Departments o f Medicine and Clinical Investigation, KAMC , Tacoaa , WA 98431-5454. Hunan sperm are capable of synthesizing scyl CoA thioestera from nascent fatty acids and coenzyae A. We have exten sively characterized the enzyne responsible for this pro cess, fatty acid: CoASH ligase (AMP) ( E . C . 6 . 2 . 3 . 1 ) , and have reported that, with the exception of docoaahexaenoic acid ( 2 2 : 6 ) , the saturated and unsaturated acyl substrates for ligase share a col!ICOn Michaelis constant (KB) of ap proxiaately 4uH. Because 2 2 : 6 coaprises the �jor fatty acid portion of aper� membrane phospholipids , we sought to investigate the effect of 2 2 : 6 on the synthesis of palmi In addition, we studied the influence of another toyl CoA, saturated fatty acid, stearic acid ( 1 8 : 0 ) , on palmitic acid Hul:lan sperm were isolated by centrifu ( 1 6 : 0 ) activation, gation and washing, and ligase was solubilized froa the sperm with 0 . 1 % Triton X-100 ( final concentration). The incubation aedis for the deteraination of ligaae activity consisted of increasing concentrations of tritiated 1 6 : 0 with varying levels of unlabelled 1 8 : 0 o r 2 2 : 6 plus satu rating levels of coreactanta. The unreacted 1 6 : 0 was extracted into heptane and the aqueous phase counted. Enzyaatic rates were calculated as lU!llO les palaitoyl CoA forned/ain/mg protein. 18 : 0 acted as a competitive inhibi tor but, aurprizingly, 2 2 : 6 was a noncompetitive inhibitor with an inhibition constant (Ki) of 9 . SuH. We conclude that in contrast to 1 8 : 0 (and presuoably other fatty acids) which appears to share the same active site as 16 : 0 , 2 2 : 6 has a different binding s i t e which i s distinct from the active s i t e , These data also imply that 2 2 : 6 cay require a separate enzyae for activation, and that 2 2 : 6 cay regulate palmitoyl CoA synthesis in sperm. No . 1 TWELFTH ANNUAL MEETING 9 7 CHANGES Ill CYTOSOLIC PROTEIN AllD PHOSPHOLIPASE ACTIV ITY DURING IN VITRO CAPACITATIOli OF SPERM FROM FERTILE AJID SUBFERTILE BUL�Hargaret Henault*, Gary Killian and 9 9 A NEW METHOD FOR OBTAIHillG A HIGH PERCENTAGE OF VIABLE, CAPACITATED SPERM, Christopher De Jonge, S , R , Mack*, L , J , D , Zaneveld, Dept , o f Physiology, Dept. o f David Chapman* , Dairy Breeding Research Center, University Park, PA 16802. Ob/Gyn, Rush University, Chicago, I l . 60612, Capacitation ( C ) i s the period o f time during which spera acquire the ability to undergo the acrosooe reaction subfertile (SF) bulls (non return rates 65- 741. and 59.96 1 . 6% respective l y ) . Washed ejaculated spern (50 x 106 vitro, but require extended periods of incubation. " BWW" (Biggers, Whitten and Whittinghaa, 1 9 7 1 ) nediWll is often Total protein and phospho lipase Az (PLA2 ) activity were deternined during in Vitro capacitation o f fertile (FT) and (AR), SpeTI!I. can be capacitated using various aedia in used for incubating spera, This study determined the effects o f incubation of huaan sperm in hypoosaotic saline cells/ml) were capacitated at 39° C , and St C02 /air in aodified Tyrodes mediun (Parrish, J . J. et al . , 1985 Sperm sonicates prepared at O , 3 , Theriogenology, 2 3 1 2 1 6 ) , o n C and AR and developed a system for capacltating ht.Gan cell cytosol. were washed in Ficoll and incubated in one of the fol low 6 , and 9 hr were centrifuged ( 12K x g , 1 5 ain) t o obtain a Cytosol protein decreased (P<.001) during capacitation and a significant ( P < . O S ) effect of fertility on total protein vas observed. Protein (µg/µ l ) for FT spera after short incubation periods, Ejaculated sperm ing solutions: Hypoosmotic ( 240mOsm) saline (0 or 3 . 5 % HSA), Blnl (0 o r 3 . 5 % HSA) or isosmotic saline lacking HSA AR, bulls vas 1 . 2 1 ± .04 at 0 h r decreasing to . 6 1 ± . 0 3 a t 9 hr. Protein for SF bulls began at . 8 2 ± .OS but �ecreased and calci\.llil , To synchronously induce the duced/ag protein) was deterMined in cytosols by measuring the decrease of fluorescently labeled L-u-phosphatidyl were added at O, 1, 3 , 4 , or 5 hr, tine periods and spern were incubated for an additional 15 mins. Sperm were then period , . 1 9 8 1 ) , Sperm were maximally capacitated by the number of only to . 70 ± . 10 . PLA2 activity (µg o f fatty acid pro choline and produc tion of labeled fatty acid for a 2 hr Lipids were extracted (Folch) and quantified by PLA2 activity for fluoronetry after separation by HPLC. all bulls increased frora 1 0 . 5 ± 2 at 0 hr to 2 3 . 3 ± 7 at 9 hr. Although PLA 2 activity in FT bulls increased frora 8 . 8 2 o f 3 SF bulls showed no change i n ac ± 2 t o 26 . 2 ± 1 3 , Percent tivity ( 1 5 , 2 ± 6 t o 1 2 . 7 ± 4 ) during capacitation. acrosooe reaction and PLA2 activity during capacitation were highly correlated for FT bulls ( r = . 9 7 ) but were nega tively correlated for the 2 SF bulls ( r= - , 49) shoving no change in PLA2 activity. These data suggest that cytosol protein, PLA2 activity and fertility may be related in bull spern. (Supported by Sire Power, Inc . ) 98 GLYCOSIDASE ACTIVITIES Ill BOVINE SPERM DURING Ill VITRO CAPACITATION AND THE ACROSOME REACTION . Taaara - McNutt*, Gary Killian and David Chapaan* , Dairy Breeding Research Center, University Park, PA 16802. Levels of 6-nannosidase (MAN), 6-galactosidase (GAL ) , 6glucosidase (GLU ) , 6-11-acetylgalactosaminidase (AGAL) and 6-11-acetylglucosaminidase (AGLU) activity associated vith spera during in � capacitation and the acrosome reac tion (AR) were determined fluorometrically (unit = ncoles 4-nethylumbelliferone liberated from substrate/hr•mg pro tein). Washed ejaculated sperm (50 x 10 6 cells/Ml) from 3 bulls vere incubated up to 7 hr at 39° C in 5% C02 /air in modified Tyrodes mediun (Parrish, J, J . et a l . , 1985, For whole c e l l sonicates prepared Theriogenology 2 3 : 2 1 6 ) . hourly, protein decreased over the 7 hr incubation ( l . 46 ± . 0 7 to 1 . 24 ± . 14 mg/ml ; r "' . 9 4 ) . MAii, GAL and GLU activi ties peaked early in the incuba tion. MAN activity vas high at 0 hr ( 1050 ± 836) but decreased gradually to 160 ± 131 at 7 hr. GAL and GLU increased from 100 ± 66 and 112 ± 62, respectively at 0 h r , t o 525 ± 206 and 589 ± 558 at 1 and 2 h r , respectively. Activities for both GAL and GLU then dropped rapidly to 100-200 units for the rest of the incub ation. HAN and GLU activities were negatively correlated ( r= - , 44 and - . 36) with the AR rate (ARR) , defined as the percent spern undergoing the AR each hourly interval of the incubation. AGAL showed peak activity (468 ± 290) at 3 hr, while AGLU alternately fluctuated between 50 and 230 units over the incubation . GAL, AGAL and AGLU lacked correlation with ARR (r=- . 0 7 , . 02 and - . 1 ) . We conclude that changes in glycosidase activities occur during capacitation. These may be important in modifying carbohydrate moieties o f the calciuri iono phore (A23187) was added to solutions containing calcitl!ll , C3lci� chloride was added to calciua-free aedia. processed for triple staining (J, Exp. Zool, Inducers 2 1 5 : 201-208, AR spern. After 3 hr. incubation )40X AR sperm were seen in hypoosmotic saline ( 3 , 5% HSA) and isosmotic BWW ( 3 . 5 % HSA). Longer incubations d i d n o t increase the nUGber o f AR spera. CalcilE'll-free, HSA-free saline proved to be ineffective in capacitating spen:1 ( 20% AR), l!ypooseo t i c saline ( 3 , 5% HSA) supported AR to the same extent a s isosmotic BWW ( 3 . 5 % !!SA) while yielding improved viability ()90% vs 80%). In conclusion, hypoosmotic saline ( 3 . 5 % HSA) capacitates a high percentage o f sperm i n a short period o f time and yields a highly viable population. Supported by NIH HD 19555. 100 DM.IIDu::GICAL C01PARIOO. 'IB BEIWEEN THE £0.l\R AID l?PDACROSIN-Jl.COOSlli PIDI'EIW\SE SYSTEMS . � Mark s. Siegel*' Dana s . Bechtold* , Janet L. Willand* and Kenneth L. Polakoski*, Washington University Medical Schcx:il, St, Louis, !>O 63110. Proacrosin is believe::l to be a critical o::rtp)flent of rm ture si:erm that is required for fertilization. Proacrosin derived frcrn boar spermatozoa has been utilized as a rrodel for the 11\3.jority of detaile::l studies involving this i.rrpor tant si:erm zyrro:e :J n. �;e have purifie::l l::oar and hman pro acrosin and have OCM developed i::clyclonal anti.lx:dies to these antigens. 'Ile present studies \*!re undertaken to o:upare the imnunological properties of the proacrosin fran these b\Q i:ipecies . FollMng west.em-blotting, it was sho'...10 that the antib::xlies produced against boar pro acrosin ;rere specific only to the kmwn o::uponents of the proacrosin-acrosin system. 'Ihese antib::x:lies reacted with boar proacrosin, alpha-acrosin and to a lesser extent beta acrosin. lbwever, the boar proacrosin antilxxlies did not cross-react with any antigens in the hurran sperm extracts . 'Ih:! antib::xli.es to hurran proacrosin reacted with htll'all pro acrosin and alpha-acrosin but apparently rot with the beta acrosin. Hman proacrosin antitodies also cross-reacted with the l:oar proacrosin and to a lesser extent with the boar alpha-acrosin but rot with the l::oar l:eta-acrosin. 'fuus, in spite of biochanical similarities (rrolecular ;reights and activation kinetics) between the hurran and boar proacrosin-acrosin systans distinct imnunological d.if fe� exist. (SUpp:r :l ted by NIH grant HD-128 6 3 . ) sperm and egg in preparation for fertilization. P-41 Journal of Andrology 101 INHIBITORY EFFECT OF ZINC Oll PROTEIN PHOSPHORYLATION IN THE SPER..'i HEAD MEMBRANES IN SPISULA SODIDISSIHA, Balwant Ahluwal i a and George Nolan*, Departcent o f Obste trics and Gynecology, Howard University College of Medicine Washington, D , C , 20059 zn 2+ when added in the media as low as one nicroc-0lar de creased protein phosphorylation from 70 to 75% in the iso lated sperm head membranes. Other divalent cations (calciuc, cobalt, barium and copper) in the similiar con centration were ineffective. Sodium fluoride, a phosphatase inhibitor, did not reverse zn2+ induced effect on protein phosphorylation,and when zn2+ was added to the nedia after the phosphorylation was started, the inhibitory effect was l o s t suggesting that phosphatase are not involved. The autoradiogram of 32p labelled sperm head membranes separa ted on SDS-polyacrylamide gel electrophoresis de�onstrated lack of protein phosphorylation in the presence of zn2+, The inhibitory effect of zn 2+ on protein phosphorylation in the sperm head menbranes suggest physiological role of t h i s cation in the nembrane functions. (Supported in part by Project grant -POHD-17104-0lAl) 102 OXYGEN COl/SUHPTION RATE VARIES WITH THE CONCENTRATION OF BULL SPERMATOZOA . Carole Wegner*, Gary Killian and David Chapman* , Dairy Breeding Research Center, University Park, PA 16802. The effect of spern concentration on the rate o f oxygen Suspen consunption (�l 02 /hr·l08 cells) was evaluated. sions of 2X washed ejaculated spern were prepared over a concentration range of 70-575 x 106 sperm/al by sequential d i lution o f the highest concentration w i t h 305 mOsm buffer (Brackett & Oliphant 1 9 7 5 , Biol. Reprod. 2 5 : 9 8 5 ) , 0 2 con sumption was neasured with a Clark elect rode at 34° C on 3 ml of suspension. Although initial values for 02 consunp tion were variable among ejaculates ( 1 0 . 6-22 . 0 ) , 02 con sumption and sperm concentration were h i ghly correlated ( r "' 0 , 8 3 ) for all 5 replicates. Sirailar effects o f sperl'! concentration on 0 2 consumption were o b s erved for rabbit and bull sperm incubated at 27° C. Another expericent was per forned to e l iminate the possibility t h a t the time re· quired to dilute spern, negatively a f f e c te d the 02 con sumption of sperm at low concentrations. When Spera oxygen consunption vas measured for sperm at lower concentrations f i r s t , 02 consumption increased with increasing spera con centration (r,,0 . 8 7 ) , as was observed in the original study. A third study vas conducted to deternine if a factor re leased into the supernatant by sperm at high concentra When supernatants of tion stiaulated oxygen consumption. sperm at high concentration were used t o dilute sperm to lover concentrations, oxygen consunption rates reoained high. For 6 replicates, oxygen consumpt ion and sperm con centration vere negatively or not correlated (r"'-0.64 to +O. 26). lhese studies suggest that sperm at high concen t r a t ion release a factor into the environment which st i mulates 0 2 consuaption. (Supported by HD20272) 42-P · January/February 1 9 8 7 Vol. s 103 MOVEHENT CHARACTERISTICS ,OF CYIWKOLGUS MONKEY SPERM RECOVERED FE«>M THE CERVIX AllO UTERUS. E . Behbood1 * , D. Katz, J, Overstreet , A. Hendrickx*, University of Ca l i fo r n i a , Davis, C A 95616. We are studying sperm transport i n Hacacca fasciculari s , using both surgica1 and non-surgical collection techniques. The menstrual cycle and mucus-laden cervix o f this species suggest its use in modeling events i n gamete biology i n the human. Sperm numbers and motion are quantitated using videomicrography. I n i t i a l l y , we compared sperm coll ected 5 hr and 30 h after mat i n g (pc ) . large numbers of vigorous, mot i l e sperm are retained by the cervix ( s i m i l a r to the human ) , a l though there are twice as many total and mot i l e sperm a t the e a r l i e r time. Sperm mo t i l i t y i s a l s o nearly twice as vigorous, and is more homogeneous at 5 hr pc. At 30 h pc over half the cervical sperm exhibit i ntermittant velocities, with i ncreased trajectory curvature and lateral motions of the sperm head. Over 90% of uterine sperm swim rapidly along straight trajectories at both time points. A small fraction exhibit alternating episodes of straight and tortuous , non-progressive swiPrning: this pattern resembl es the hyperactivated moti l i ty of other mall'flla l s which i s associated w i t h capac itation. Thus, capacitation i n the cynomolgus monkey may begin i n the lower fema l e (Supported by llIH grants H012971 and RR00169 . ) tract. 104 RJl.SATILE lJIRH-TREAn!ENT IN !>'ALE PATIENfS WI1H SEVERE (1.IGJSPERMIA AND saOCTIVELY ELEVA1liD FSH. W. Aulitzky� J. Fri ck � F. Hadziselimovic, *�partir::nt of Urology , General Hospital, Salzburg , Austria and Chil dren ' s Hospital , Basel, Switzerland. Evidence was presented, that severe oligospermia with high R'1I and normal lJl-levels may be caused by hypothalamic dysregulation. It was further demonstrated that restora tion of physiological UJ-pulse frequency by long tenn pulsatile LJ-!R!-1-application led to a marked decrease of elevated FSH-levels. The aim of this study was to confirm these preliminary data and to investigate the effect on the spermatogenesis. 15 male patients with severe oligo spennia, high FSH-levels and normal LR-levels were treated by p.:ilsatile UlRH for 6 coonths. All showed disturbed UJ pulse pattern (<10 peaks/24hrs. ). According to semen ana lysis and spermatogonia/tubule ratio (SPT-ratio) 3 groups of patients were created. Group I (n=S) sperm density <1 mill, SPT-ratio <0, 1 . Group I I (n=S , 1-10 ttrill sperms/ml , SPT-ratio 0,2S-0, 1 . Group I I I (n =S ) , 10-20 mill spenns/m l, SPY-ratio > 1 , 0. Normalisation of 1H/FSH secretion was seen in all pat. of group I I and I I I and in 3/5 pat. of group I during IRRH therapy. The transient effect on lli/FSH secretion supports the hypothesis that the hormonal dysregulation is of hypothalamic origin, Im provement of sperm density was observed in 9/10 pat. of groups I I and I l l , Wi.ereas no changes h�re found in group I . 1hese findings correlate with the SPT-ratio. Therefore h� conclude that SPT-ratio is a useful method for patients selection for �lsatile UlRH therapy and can be used as prognostic parameter . \ No. 1 TWELFTH ANNUAL MEETING 1 0 6 IDE!!l'IFICATIONCE A PROOESTERONE Rlm'nR IK llUJIAHTESTIS. Stephen J. Iinters, Buah S. l:oepina and Philip troen. DepUt.a.entof Medicine, Montcfiore Hospital, a.nd University of Piu.sburah School of Medicine, Pituburgh PA, 1'213. Testes from patients vith tc.sticula.r feminization a.nd chit.tons have bHn found t.o contain a cytosollc pro1uterone (Pa) biDdin1 protein. Hovever, a sim.Jlar protein has not boon found in norma.I huf!Wl U.SU.s. Further. s:t.udlo.s iD �of other e190rl.m.ontai a.nima.ls have produced confikt.ins results. Ye have ree1am.ined tc.sticular Pa bindln1 protei.111 using the syntheUc pro1ostin, 2JS-hydrol}' prome1estone, IS& rad.io l&bolled probe (Rouml-Uclaf, RU-27937). De1amothasono VIS used t.o block RU-27937 binding t.o glucocortlcoid receptors. Jlul.m.al bind ing at2Cva.s achieved by 8h, and rom&.i.o.ed stable for 92h. Bindinl YIS quantativoly similar in vhole to.sUs andsemlniforous tubules. Rel&t.ive binding aftiniUes vere RU-27937 > promegestono > methyltrienolone > miboJerone. Heither t&Stost.erone nor estrad.iol competed for binding of (3B)RU-27937. Sucrose density aradient centrifusation using a vertical lube rot.or allowed for the identific..Uon of a 8-95 bind.in& peat in lov salt buffer containing sodium molybdate, Saturation analysis revealed saturablo bindln1 of hiah affinity O::D.0.32.0nM). Binding capacity in tems from untroatcd men undera;oiD1 orchiect.omy for prostate cuclnoma 00.6tl.O fmol/ma protein) vu 1pprolimately 33� ofandro&en receptor (mlbo!erone) binding. RU27937 Mndlna was areater than normal ( 16.7 fmoJ/m1> in the ums of a patient vho had been treated with diethylstllbestrol. 111d 'V&S mutedly increased (233 fmol/mg) in the testes of a paUent •ith teslicular feminization. Ho bind.in& vas found in teslicular nuclei. Binding wu also detected in test.is from cynomolgus monkeys a.nd rams, but not froa Spraaue-Dawley ttts, Sviss-llebster mice or Syria.n hmsters. These results indicate that a Pa bind.in& protein is found in human tc.stis a.nd appears to be inhibited by 111dtoaen. AJthouah pre90ntin ma.n 111d in cert4in a.nimaJs, its physioloaic Nie is unknown. Studio.s are undervay in subhuman primates to further investi1&to its funcUont.l sitnifict.0.ct, 1 07 Laydig Call Function in the Cryptorchid Rat. Abney. To!ll 0 , D e p a r t n e n t o f Physiology and Endocrinology. Medical College of GA, Augu s ta , GA 30912-3395 Adult male rats vere rendered b i l a terally (bl) and unilaterally ( u n i ) post-surgery, populations gradients. 1 cryptorchid, A t 1 4 and 2 8 d a y a p u r i f i e d Leydig c e l l s ( LC ) , d e s ignated and 11 L C were i s o l a t e d from. aetrizaelde Gonadotropln binding a n d capacity were assessed in vitro, I n t e rs t i t i a l c eTi"n'Uiib e r s decreased a t 1 4 a n d 28 days (10 6 s teroidogenlc /testis) were i n the b i a n d u n i groups. compared to intact ind eutopic ( u n i - s c r o t a l ) contro l s , n u m b e r s ( 1 0 / t e s t i s } w i th i n each L C pop were Cell statistically equivalent for a l l groups at 14 days; LC pop II froa the 28 day uni group was decreased ( p ( 0 ,0 5 ) . Binding o f 125I-hCG (dpl!l/testls) was equivalent in pop I and II LC for all groups a t 14 days; LC pop II frOJ;1 the 28 day bi and uni groups exhibited decreased (p(0,05) binding, T e s t o s te r o n e ( T ) production in response to hCG (0-200 nlU) and dbcAHP(0-5 BH) was decreased (p(0,05) in bi and uni pop I and I I LC a t 14 d a y s , c o a p a r e d to A t 2 8 d ay s , u n i p o p I and 1 1 exhibited c o n tr o l s . decreases (p(0,05) in T production, however, eutopic and bi LC exhibited increased T production above controls , These data suggest that LC from. the uni group exhibited the m o s t dramatic c r y p t - i n d u c e d a l te r a t i o n s in c e l l func tion, particularly a t 28 d a y s , Supported b y N I U grant #16983, 1 0 6 EFFECTS OF DIETHYLSTILBESTROL (DES) 011 TESTICULAR LH RECEPTORS Itl FISHER 344 RATS. A . G . Ama:lor , A . I . Esquifino*, A . Bartke, V. Chandrashekar*, J . J . FernanC:ez-Ruiz* and R . W . Steger*, Dept. Phys iology, S I U School of Me d i c i n e , Carbonda l e , IL 62901-4531. 1 0 8 BTIIINYL BSTRADIOL INHIBITION OF TESTOSTEROUE PRO Treatment with DES causes the developrr:i!nt of prolactin ( PRL ) -producing adenomas, and hyperprolactinemia i s known to have dramatic effects on testicular function. How ever, there is a l s o evidence for independent effects o f estrogens o n Leydig c e l l function. Therefore , the present study was undertaken to analyze the di fferent effects of DES on the testes during and after treatment with the steroid. Fisher 344 rats (>B weeks o l d ) were implanted for 14 weeks with DES or empty s i l astic capsul e s , which were removed 7 weeks before sacrifice. A second group of rats received DES capsul e s 7 weeks before sacrifice. DES caused a dramatic decrease i n body, testes and semi Removal of DES capsules a l l owed a nal vesicle weights. partial recovery of these weights. Treatment with DES a l s o produced a great decrease in plasma L H . DES treat ment caused a decrease in LH receptor content without affecting LH receptor concentration. However , ren:-0val of DES capsules al lowed LH receptor concentration and content to increase well above the levels rr�asured i n The present results thus indicate that untreated rats. DES by regul ating LH receptors independently fr-Orn PRL and LH, can counteract the hyperprolactinemia-induced i ncrease in LH receptor level s . steroidogenesis while estrogens inhibit steroidogenesis. DUCTIO?l IN THE LU TREATED HYPOPHYSBCTOHIZIID RAT, R,B, Hyers* and T . O . Abney. Departaent of Physiology and Endocrinology, Medical College of GA, Augusta, GA 30912. Leydig cell function in the nature rat is controlled by various hon:iones. Luteinizing hornone (LU) aticulates This study is an attecpt to characterize Leydig cell function by exru;!ining the capacity of rat Leydig cells to produce testosterone (T) iE_ vitro after i:!! vivo treat�ent with various co::ibinationa of ethinyl estradiol (BB2) and LU. Hypophysecto�ized rats were injected with various doses of EE2 (25 or 10 ug/lOOg B .W . ) , LH (25, 10 or 5 ug/rat) or coabinations of the two. The rats were treated 4 days and killed on the fifth day. Testicular interstitial cells vere isolated and incubated for 3 hours in the presence or absence of hCG and dibutyryl cAMP. The incubation aedia were subsequently assayed for T. All doses of LR resulted in an increase in T production, while both doses of EE2 resulted in a 40-50% decrease in T production. The groups which received various doses of LU plus 10 ug of 882 responded with increased T production as c01'lpared to the group receiving EE2 alone. The groups receiving the two higher doses of LH coapletely overcame the inhibitory effect of 10 ug EE2, When 25 ug of EB2 was injected, treatoent with LR resulted in a dose dependent increase in T production coapared to the BE2 treataent alone. However, only the 25 ug dose of LH vas capable of overcoming the effect of 25 ug BE2. These data suggest that the testes of 8&2 treated rats are capable of responding t o LH and that high levels o f LH can overcooe the inhibitory effects of EB2, Supported by NSF#PCH-8109847. P-43 Journal of Andrology 10 9 GNRH INCRFASES SPECIFIC PRECOPULATORY RESPONSES TESTOSTERONETRFATED PONY GELDINGS. IN "2 2 1 NancyDiehl: Sue McDonndl, and Marolo Garcia 1 Pennsylvania State Uni</ersity, Department of Animal Sckna, Uni.,.ersity Park, 2 PA 16802 and Univenity of Pennsylvania School of Veterinary M:edkine, New Bolton Center, Kennell Square, PA 19348. The effects of exogenous GnRH \Vere studied in 12 long·term castrated male ponies (geldings) with and without androgen replacement. Ba.s.ed on petfor· marn:e in pre-treatment seAllal behavior tests, ponies were assigned as rank matched sets to four treatment groups: GnRH alone (G), 1estosterone and GnRH (TO), testosterone alon<: fO, and no treatment (C). Testosterone treat· ment consisted of 200 ug/kg testosterone propionate in sesame oil SQ c\'Cry 48 hr for three weeks. GnRH treatment consisted of 25 ug Cystorelin (CEVA Laboratories) SQ every 3 hr for three weeks. All animals recei'o'ed treatments or equivalent volume vehicle on the same injection schedule. Sexual behavior tests consisted of 4·minute exposure to an ovarieclomiud estrogen·treated pony mare restrained behind a teasing rail on one side of a small (3.05m X 3.05m) outdoor enclosure. Tests were conducted three times weekly between 0730 and 0930 for tvm weeks before the start of treatment, three weeks during treatment, and for two weeks after treatments had stopped. 1be following behavioral measures were recorded: sniff mare, sniff ground, flehmen response, lick, bite, strike, vocalization, and roll frequenc:ks: penis drop latency and duration; erection latency, frequency and duration; mount latency, frequency, and duration; and at· tention late1Ky and duration. During treatment TO ponies exhibited greater (p<.05) flehmen response and sniff ground frequencies. Attention duration and bite frequeocy were a!w greater (p<.05) among TO ponies. These findings are consistent with our dinka! obseivation offlehmen response in sta!lions following intra\'enous injection of GnRH. These rerults suggest a testosterone·dependent effect of OnRH on certain precopulatory investigatory responses in stallions. and support the hypothesis that OnRH is involved in the regulation of sexual be· havior. MICROSURGICAL ACCESSORY GLArm ABLAT IOU: THE EFFECTS ON FERTILITY POTENTIAL Ill THE RAT. W i l l iam B l a n k , Eric Yousha* and Hare Goldstein, The Population Counc i l , 1230 York Avenue, New York, /lew York 10021 . Hale Lewis rats were randomly divided into seven groups ( n = 5 ) . Using microdissection techniques, accessory glands were removed from each group as follows: seminal vesicles, coagulating glands, dorsal prostate, ventral prosta te , entire prostate, a l l accessory g l ands a n d sham-operated contro l s . After a minimum of four weeks, each m a l e was placed for o n e week in a separate cage with two fema l e s of proven fer ti l i ty. Tn� weeks after the mating study the females were ki. l l ed and the uterine implantation s i t e s were counted. Animals whose seminal ves i c l e s , dorsal prostate, entire prostate and a l l accessory glands were a b l ated failed to fertilize any ova i n every case. lkist o f those whose co agulating glands and ventral prostates were ablated d i d fertilize ova as d i d the shams. To prove this "selective inferti l i ty" was not secondary to impotence caused by the surgery, four rats from the seminal ves i c l e , dorsal prostate and ent i re prostate abla t i o n groups were placed with t1� fema l e s for one week. 1 Intravaginal washings were performed on each fema l e every day for seven days. Sperm were found in a t least one fe ma l e from each individual rra l e durin9 the study, thus proving that each ma l e was potent. It i s su9gested that each accessory gland makes a contribution to the fer t i l i ty Of the subject and that future studies a ttempt to identify these factors. 110 · January/February PROTE!tl CONTRIBUTIONS OF THE ACCESSORY ORGANS TO THE COMPOS ITION OF HUHAH SEHlllAL PLASHA AS OETERHIHED BY HIGH RESOLUTION Tl/0-DIHENSIOIIAL ELECTROPHOR E S I S . Edward E . Gaunt, A n i b a l A . Acosta , Steven B . Ackerman * Patricia A. Pleban!' John F. Stecker, and James H. Yuan. Department of Chemical Sciences, Old Dominion University, tlorfo l k, Va. 23508. 111 Human Seminal Plasma (HSP) was evaluated using Hi�h Resolution Two-Dimensional E l ectrophore s i s (HR2D E ) . HSP from healthy volunteers with recently proven ferti l i ty were characterized using this technique to establ i s h "normal11 prote i n distribution patterns i n the 2-D s l a b gel. T f me course studies were performed o n selected specimens to determine the effects of l i quifactlon on HSP protein composition wfthfn the first several hours after c o l l e c t i o n . A series of "coagulation� proteins were identified which di sappeared with t i me . few changes in protein compos i t i o n were seen after 60-90 minutes post ejaculation. Spl i t ejaculates were col lected ( u p to 6 fracti o n s ) t o identify w h i c h proteins i n t h e 2 - D gel pattern 11-ere contri buted by the various accessory glands. Initial fractions correlated well w i t h pure prostatic f l u i d . Terminal fractions .represented pure seminal ves i c l e f l u i d . HR20E of macroscopic g e l globules (globoid bod i e s ) separated from whole semen gave a protein pattern s i m i l a r t o terminal fractions of the s p l i t ejaculate. Pre ejacul atory fluids (urethral gland secretions) 11-ere evaluated by this technique as wel l . CHARACTERISTICS OF THE ACCESSORY GLANDS OF THE MALE REPRODUCTIVE TRACT OF SONE NATIVE CALIFORNIA RODENTS. B. 112 Peitz and S . Membreno*, Biology Department, California State University at Los Angeles, LA, CA 90032 The reproductive tracts o f males from the genus Peromyscus and Dipodonys were examined and compared to those of several laboratory rodents. Aninals were collected by overnight trapping at several field s i t e s , sexed in the field and males were brought to the laboratory. Weights of the accessory sex glands (ASG) , their fructose content (Foreoan, e t al . , 1973 Analyt. Biochen. 56: 584) , and ability of extracts of ASG to cross coagulate when mixed with extracts of other ASG (Peitz, et al . , 1979 J . Reprod. Fert. 5 7 : 183) were deteroined. Weights o f the ASG are I n both species higher in p; naniculatus than in P. truei. the coagul8'ting glands (CG) had the highest fructose con centration. E_. agilis had heavier ASG than E.· oerriarni, but they contained less fructose. I n E.· agilis, fructose concentration was highest in the CG, whereas in !?_. cerriami, the highest concentration was found in the arnpullary glands. When ASG extracts o f a single species were tested for coa gulation with each other the P . maniculatus extracts of all ASG coagulated to some extent-when mixed with extracts o f t h e CG, whereas, i n D . rnerriami S V extracts reacted with the CG much better than 8ny�other glands. Coagulation tests were performed between extracts of ASG of P . maniculatus and D . oerriarni and the rat and house mouse to determine which ilands are involved in the fornation of the vaginal plug. I n these tests between species the CG of P . maniculatus and D . nerriaoi coagulated more quickly than their SV. These-s tudies indicate that the ASG of Perooyscus species are oore similar to the Xurid rodents than Dipodornys species. 44-P Vol. B 1987 No. I TWELFTH ANNUAL MEETING 1 13 A RAT MODEL OF FETAL EPIDIDYMAL AND GONADAL DEVELOPMENT AND ITS SIHILARITY TO CLINICAL ABllORHALITIES, J . L . Fourcroy, UMDNJ-RWJMS, llew Brunswick, NJ 08903; H . C . Shen, Naval Hospital , Ports1nouth , VA 23708; G . P . Riorda!"!, Naval Hospi t a l , Bethseda, HD 20814 and D.W. Warren, University of Southern California, Los Angel es, �A 9 0 0 3 3 . Testicular cel l popula t ions are of multiple origins; normal gonadal developrr.ent and function depends on the interactions of these populations soree thought t o be o f mesonephric-ttesenchymal origin. The ACI rat (August x Copenhagen) provides an interesting model of the dependency on the �esonephri c anlage for normal development with a 1 0 - 1 5 % incidence of agenesis of the mesonephr ic duct and its derivatives. The agenesis i s noted predomi nantly on the right side with occasional contralateral renal abnormal i t ie s . The testes on the affected side are markedly smaller by puberty. We have reviewed the light and electron microscopy of normal and abnormal t i�e-ges tation fetuses. The defect is seen prior t o androgen secretion ( 1 5 days) and appears to coincide with vulnerable periods of definit ive u�b i l ical vascular reorganization. Altered intratesticular compo nents are observed in the a f fected gonad . C l i n i c a l l y , patients with agenesis of vas deferens display similarity to the rat model but unlike the r a t , maintain normal spermatogenesis . The at rophy i n t h e rat may reflect an earlier developmental insult blocking migration or mitosis of c r i t i c a l cel l ul ar element s . 114 CLONINCl AND SEQUENCE ANALYSIS O F HUMAN TESTIS-SPEC I F I C LACTATE DEllYDROClENASE c4 . Erwin G o l d b e r g , Catherine E. D r i s c o l I * and Jose L . H i 1 1 an•, Departnent of Biochemistry, H o I ecu lar B i o 1 ogy and Ce l l B i o l ogy, Northwestorn University, Evanston, IL 6020 1 : La Jo l l a Cancer Research Foundation, La Jo l l a , CA 92037. The t e s t i s s pe c i f i c l ac t a t e dehydrogenasc i s ozy®e of aaaaa l s constitutes one of the aost dranatic exa•ples of the te�poral and spatial specificity of gene function. Of special interest i s the close association of LDH-C4 with spermatogen e s i s . T h i s i s oz y p e f i r s t a p p e a r s i n a i dpachytene pri aary speri.atocytes and i n c r e a s e s i n concentration unt i l the sper11.atid stage. As a first st ep towards Investigating regulation of gene function in the gerninal epithelium we have constructed and probed a human t e s t e s cDNA expression l i brary, A c o a p l e a e n t a r y DNA (cDNA) c l o ne c o a p r i s i n g the c o n p l e t e c o d i n g region o f huaan testis-specific LDH-C4 (LDll-X) has been i s o l ated and sequenced. A f t e r n i c k trans l a t i o n the cDNA d e t e c t e d a s l ng l e a R N A s p e c i e s o f 1 . 5 kb f r o a t e s t e s b u t n o t p l a c e n t a . The cDNA begins with 6 6 b p o f 5 ' u n tran s l at e d region a n d e n d s w i t h 1 1 7 bp of 3 ' f l anking sequence. The c o d i n g region coaprises 987 bp that code for 329 am in o The co•plete anino acid sequence deduced from the acids. nuc l e ot i d e sequence has 74% honology w i th the s o a a t i c lactate dehydrogenases. The cDNA probe wi l l b e useful to i d e n t i fy a nd i s o l a t e t h e g e n o n i c DNA in o r d e r to investigate regu lation of the LDH-c4 gene. In view o f the r e l a t i on sh i p between LDH-c 4 and s p e r111 a togen e s i s , t h e s e s t u d i e s aay c o n t r i b u t e to our u n d e r s t a n d i n g o f t h i s process a t the 111o lecular l e v e l . Supported by srants Nill CA21961, NIH CA42595, Nill HD05863, and by the Prosrcnc for 115 EVIDENCE FOR POLYMORPHIC SPERM ANTIGEN EXPRESSION IN INBRED HICE, Chong Xu*, Deborah J , Anderson, Fearing Research Laboratory, DepartGent of Obstetrics and Gynecology, Harvard Medical Schoo l , Boston, KA 0 2 1 1 5 . I t has been known for some time that T/t complex gene products affect fertility in aale mice, Furthers-0re, it vas recently reported that allogeneic mouse spern is more il!J!lunogenic than syngeneic sper111 , and that certain infertile men have sperm that is l'lOre illl!rt.lnogenic and antigenic than sperA of fertile donors, These data suggest that there may 1>e genetic differences in sperm antigen expression that affect fertility. To nap sperm antigenic differences in mice of different inbred strains, 8-week old virgin female Balb/c mice were illl$!iu nized with epididymal sperm from the following inbred strains: Balb/c, DBA/2, C51 Bl/6 A/J and 1 29, l1£11unizations consisted of 2 x 10 sperm in Complete Freund's Adjuvant injected subcutaneously, fol }owed at 21 day intervals by injection of 2 x 10 sperm i n Inco�plete Adjuvant. Sera obtained seven days following final injection were applied to epidid}"lllal sperm extracts from the various inbred strains using the Western Blot procedure. Forty-seven major protein bands and no differences were revealed by Cooeassie Blue stain following electrophoretic separation of each of the sperm extracts. However , Western blot analysis indicated that certain major sperm membrane components show strain-dependent differences in i1m1unogenicity and antigenicity. These data provide evidence for the existence of polymorphic antigens on sperm, 116 CHRONIC SILICOHE IMPLANTS IH THE RAT VAS DEFEREN S . Donald p , Waller, Michael Szarley•, Glen K. Annamarie Martin• , Cyrus Fakroddin•, Adaniya • , Jeanne M . Quigg* , Andrej s R . Nikurs, and Lourens J, o. zaneveld, University of I l l inois at Chicago and Rush Presbyterian-st. Luke 1 s Medical Center, Chicago, I L 6 0 6 1 2 A new reversible male contraceptive device (SHUG) made primarily of s i l icone is being developed. Previous studies demonstrated its ability to block the flow of spermatozoa when placed in the monkey vas deferens . The s i l i cone material must be tested for chronic target organ Two hundred toxicity prior to clinical t r i a l s . one group rats were divided into two groups : had sham operations and the other group had s i l icone noodles implanted in the vas deferens and secured by a suture. At s i x months and one year after implantation animals were sacrificed and the vas deferens removed, fixed, and histologically evaluated. Changes in the vas deferens attributed to the presence of the s i l icone were minimal-to-mild reactive hyper plasia in the proxima l , surgical and distal segments of a few animal s . An increased incidence of spermatoceles, aspermatic granulomas and chronic inflammatory changes within the inner walls of the vas deferens were also observed. These changes in the vas deferens are considered of minor toxicological Supported by the Program for importance, Applied Research on Fertility Regulation (Agency for International Development, PARFR 339) App l i ed Research on F e r t l l i t y Re su l a t l o n , N o r t h""e s t er n U n I v e r s I ty <AIDIDPE-0546-A-00-1003>. 1' P-45 I Journal of Andrology , · January/February 1987 Vol. B I I! I 117 ANDROGEN HDVEME!ff INTO TilE RAT SEHINIFEROUS AND EPI DIDYMAL TUBULES PERIFUSED IN VIVO. T. T. Turner, Univ, of Virginia School of Medicine;- Charlottesville, VA 22908 Proluninal, net transport of Jn-testosterone ( 3H-T) froin peritubular to intratubular fluids of the adult rat testis and epididymis was studied by in vivo perifusion and sub sequent nicropuncture of se111iniferous tubules (SllT ' s ) and caput and cauda epididymidal tubules (CPT ' s and CDT ' s , re l Tubules were perifused with KRP containing spectively . 14 c-polyethyleneglycol ( 14c 13 uC!/ral H-T (50 C i/m.�) . PEG) was included as a dilution marker ( 0 , 7 uCi/nl) and a In some control for conta�ination of intraluminal fluid, experiEents lx10-3H DNP or unlabelled T at lOx or lOOx the concentration of 3H-T was included in the perifusion fluid. Radioactivity of 3tt-T and 14 c-PEG in these fluids was de t e rmined after 1 and 2 hrs and the percentage of peritubu lar 311-T appearing in intralurninal fluid was determined. Net entry of isotope into the SNT's was 10",.. (X=l5%) , but in the epididymis there was a large proluminal �overnent of 1he 1 and 2 hr isotope against a concentration gradient, values for the CPT 1 s were 303,9f69.2% and 2 2 2 , 0 + 6 1 . 0 % , re spectively, and for CDT ' s were 180. 7+37 . 8 and 1 8 3 . 4+30,8, The uphill JnOl!lement of isotope into CPT ' s was significantly and progressively inhibited by lOx and lOOx increases of 119 llATURAT10?1-RELATED GLYCOPROTEHlS Ill THE HOUSE EPIDIDYHIS llwan-Yun Liu* and Charles H . Huller, Oepartnent of Biological Structure, SM-20, University of HashinRton, Seattle, UA 98195 During epididymal trans i t , sper��tozoa undergo both s t ructural and functional naturation which is essential for spern to becone fertile. Anong those ��difications, are of particular interest . Ho-.· the surface alterations ever, the roles that tt€ttbrane molecules play are not well defined. In this study, we try to identify surface glyco proteins which are related to naturation by using Wheat Germ Agglutinin (WGA) as a probe. WGA is a lectin which specifically binds 11-acetyl glucosamine and sialic acid. Live spero recovered fron different portions of C57Bl/B6 mouse epididymides are probed by RGA-FITC. The binding is localized on plasma �embrane overlying anterior acrosome and entire tail, and increases as spera beco�e more ma ture. To identify menbrane proteins which bind HGA, NP-�0 extracted proteins are separated by SDS PAGE and then electrotransferred onto nitrocellulose paper. Recognized by RGA-Avidin-Biotin-Peroxidase �ethod, a 49±1 Kd glyco protein (GP-49) is identified on corpus spern. Besides GP-49, a 83+3 Kd glycoprotein (GP-83) is found on cauda spero. By the same approach, both GP-49 and GP-83 are re cognized in corpus fluid, cauda fluid and NP-40 homo unlabelled T in the perifus ion fluid, but not by DNP, Also, !,!!. vitro perifusion of epididyoal tubules with HEH removed native ltw:en fluid and significantly reduced 311-T uptake. Steady-state binding o f 3H-T to electrophoresed epididymal buffer and i s likewise not easily renoved by centri fu lumi Q al protein with competitive binding characteristics for -11-androgen, i . e . ABP, is important for epidid}'tQal up washed away fro� the spern surface by several changes of buffer. I n conclusion, h'GA-binding glycoproteins, GP-49 and GP- 8 3 , appear on spern surface during spero matura proteins demonstrated a single androgen binding peak in It is concluded that an intra CPT and CDT lumen fluid, take of peritubular androgens. Supported by HD 09490, 118 EPIDIDYMAL B Y . RAT SECRETION PROTEIN EPITHELIAL CELLS GRO\VING IN BICAMERAL CULTURE CHAMBERS. S.W. Byers. Department of Anatomy and Cell School, 11.fedical-Dental University Georgetown Biology, Washington D.C. Epididymal epithelial cells growing on extracellular matrix coated filters in bicameral culture chambers form confluent polarized monjlayers (Byers et al, J.Androl. 7, 59 1986). In the present study H-Jeucine incorporation into cellular and secreted proteins was investigated. Epithelial cells isolated from 50 day old rats were grown for 3 days in serum free defined medium (SFDf\f). After an overnig t incubation in leucine·free medium the cells were exposed to H-leucine (50uCi/ml) in SFDf\f for 6 hours. Total TCA precipitable material from the apical medium, the basal medium, and the cell layer was collected. Cells isolated from both caput·corpus and cauda epididymis secreted 50% of total synthesized protein with an apical:basal ratio of 2:1 and 3:1 respectively. Sub-confluent monolayers or monolayers disrupted by scraping did not secrete proteins in a polarized fashion. SDS· PAGE analysis of radiolabeled proteins in apical and basal medium revealed that the difference in apical-basal secretion was qualitative as well as quantitative. Proteins of 25kD and 68kD were secreted apically and basally from monolayers of both caput·corpus and cauda cells. In contrast, a number of proteins in the 30·55kD range were secreted only into the apical chamber. In particular, a prominent protein of 32kD was secreted apically from both caput-corpus and cauda cells. There were also differences in protein secretion between cells from the two regions of the epididymis with a protein of 55kD being secreted only by cells from the caput-corpus and a protein of 40kD secreted predominantly by cells from the cauda. (supported by NIH HD 20028) � 46-P genates of corpus tissue. Further investigat ions reveal that GP-49 i s not extractable by high salt (lM NaCl) gation and washing of sperm. In contrast , GP-83 i s easily tion in the mouse epididynis. These molecules nay be se creted by epitheliun into luminal fluid and bound to sperm. I 120 OJRRELATIONS BE'ThlEEN CHANGES I N RAT SPERM MEMBRANE LIPID 00!-IPOSITIO?l AND THE TEMPERATURE-DEPEtlDENCE OF THE MEMBRANE PHYSICAL STATE DURING EPIDIDYMAL MATURATION . * and Laura M . Edgerly Department o f Joseph C . Hall � Biological Sciences, Kean College of New Jersey, Union, New Jersey 07083. Chemical analyses of rat caput, corpus , and cauda sperm plasma rrembrane-enriched fraction revealed a s ignificant (P :$ 0 . 0 1 } decrease in total membrane phospholipid, cho l es terol and protein content during maturation. A more re fined analysis o f bulk rr.entbrane phospho lipid revealed that 56-68% of the phospholipid is represented by choline and ethanolamine phosphoglycerides, with the choline fraction decreasing relative to ethanolamine phosphoglycerides dur The bulk phospholipid-bound fatty ing epididyma l transit. ( C : 1 6 ) and stearoyl (C: l8) , and was highest in caput sperm and lowest in cauda sperm. Arrcng the various unsaturated fatty acids, (C l 8 : 1 ) , acids consisted primarily of panitoyl (C22 : 3 ) , (C2 2 : 4) and { C 2 2 : 5 } was corra.on in the phospholipid fraction of the three spern rr.errhrane populations, but de creased significantly as sperm traversed the epididynis . The lipid solubl e spin label, 5-doxyldecane (S?UO) was used to monitor the "physical state" of the three sperm membrane populations. While only one phase-transition temperature indicating the onset o f a change in the bu lk physical property of th·, plasma membrane was observed, the phase-transition temperature was di fferent . These data suggests that the bulk phase behavior (e . g . , membrane fluidity) of the plasma membrane changes as spern mature in the epididymis. J No. 1 TWELFTH ANNUAL MEETING 121 THE ROLE OF SUPBROXIOE OISMl.ITASE IN PREYENTI�G LIPID PEROXIDATIOli AND LOSS OF MOTILITY IN HUMAN EJACULATED * * J .C . Touchstone, Luis Juan G . Alvarez, SPERMATOZOA, Blasco, and Bayard T, Storey, Departments of Physiology and Obstetrics and Gynecology, University of 19104. Pennsylvania, Philadelphia, PA Superoxide disoutase (SOD) and glutathione peroxidase/reductase (GP/R ) are nost cells� Clajor enzy1ru1 tic defenses against lipid peroxidation. The function of these defenses in hul!!an spera was exaoined, Five replicate saoples from each of 8 norcal donors vere assayed for SOD and GP/R act ivity and for spontaneous lipid peroxidation, as oeasured by malonaldehyde (HA) production and loss of motility during aerobic incubation at 37oc, SOD activity in fresh spen:i ranged between 1 and 10 units/108 cells with a nean value of S . 5 + 2 , 2 (X The concentration of MA at the p(;"int of .± SD, n � 40 ) , complete loss of mot ility, defined as the lipoperoxidative lethal endpoint (LLE } , was 0 . 1 0 ± 0 . 03 Despite the nmo les KA/108 cells (X .±. SD , n "' 40 ) . variability in the time ( tL ) for comp lete loss o f motility (range: 1-10 h ) , the LLE reoained constant , When the rates of KA production for each individual sample, ss defined by (LLE ) / ( t L ) , were cocpared to the act ivity of SOD in the fresh samples, a linear correlation was obtained (r � 0 . 9 7 . GP/R activities were 21 ± 3 and 4 4 ± 3 nco l/min-10 cells respectively (X ..± SD, h "' 40 ) and showed no correlation with the rate of MA production in individual samples. These findings suggest that SOD rather than GP/R p lays the major role in protecting human spero from Oz toxicity due to lipid peroxidation. Supported by liIH grants HD-15842, HL-07027, § and llL-19737. 122 PATENCY AND PREGNANCY RATES FOLLOWING MICROSURGICAL EPIDIDYMOVASOSTO!>.IY FOR EPID IDYMO Medical Ross, S. Lawrence Cente r , 0. Charles VASAL OBSTRUCTION. Michael Chicago, Turk* , and and Reese Hos p i t a l 60616 IL Fourteen azoospermic ma,l e s with epididymal ob struction underwent microsurgical epididymovasos tomy . The s u r g i c a l mation of a single End proximal vas . technique used was tubule epididymis of approxi to the to s i d e anastomosis was per formed With s i n g l e layer 10-0 mono f i l ament Patency and pregnancy rates for the suture. entire group were additional 20% h ad 71% and 29% analysis w i t h n o pregnancy. cedure is described respe c t i v e l y . An a normal postoperative semen The surgical pro in detail and results are subgrouped by e t i o l o g y o f obstruction as e i t h e r primary conge n i t a l o r secondary t o previous in All patients having epididymovasostomy fection. for previous vasectomy were excluded . 123 RESUL'IS OF SPOCIFIC 'IUBULE MICRC6URGICAL W.SOEPIDIDil-U) 'RJ1Y, Sherman J. Silber, M,D. , St, Luke ' s Hospital & St. John's Mercy Medical Center, St. Louis, l4J 63017 A total of 541 patients underwent bilateral microsurgical vasoepididym::>stc:icrr/ with the specific tubule method origi nally described by the author. 'l'Welve percent (77) requ ited an anastorrosis to the cap.it region because of exten sive disease distal to that site. Reliable followup for greater than three years was obtainable on 2/3 of those patients. "Patency" of the anastCUPSis was achieved in 81% of the patients undergoing anastom:>sis to the cap.lt. Forty-five percent of the patients with anastom::>sis to the cap.it ha:l a sperm count over 20,000, 000 per cc with directional ootility over 50%. Fifty-t\\O percent had gOOtl directional rrotility but socre of those had lw total ntn1bers of sperm. 'Ihe wives of 26% of men with anastorrosis to the cap.it epididymis achieved a normal full-term preg nancy (approximately 1/2 of those with normal semen) , Pa tients undergoing anastomosis to the corpus epididymis had a 91% "patency0 rate. Seventy-eight pere€nt had go<x1 rrotility, and 61% achieved pregnancy. Patients undergoing vasovasostany (epididymis presumably intact) had a 98% patency rate, and an 82% pregnancy rate. In rrost of the non-pregnancies with vasovasostany {or vasoepididynPstomy to the corpus) and normal serren pararreters, the wife was found to have either ovulatory dysfunction or tubal di sease. 'Itlis was not the case with cap..it epididymis anas tomosis, In conclusion, good success rates were achiev able with microsurgical vasoepididym::>stany. 'Ihe site of obstruction influenced the outcom::i . Meticulous microsur gical technique was iJ1lX)rtant for achieving a high success rate. 124 PREXNANCY AF'I'ffi. �. Sherrran J, Silber, M . D . , S t . Luk e ' s Hospital & S t . John 's V.ercy Medical Center, St, Louis, l1J 63017 This paper represents a 10 year study on our first 199 patients .,,no underwent vasova50$tany for reversal of vasectomy, Fifteen (8%) patients developed early marital difficulties, became separated or divorced , and were exclUded. One hundred fifty-three (83%) of the remaining 184 eventually inpregnated their wives . Patients with no sr:erm in vas are excluded , the adjusted pregnancy rate in a group of patients requiring si.m>ly vasovasostcmy ( i . e . epididymis intact) ...-oold be 89%, Twenty-five o f the 184 patients had a varicocoele (14%) . '!here was no signifi cant difference in pregnancy rate in patients who had varicocoele versus those •,.;ho did not, Sperm antibody titers were not associated with success or failure. serum irrm:i:>b lizing antibody titers were elevated in 25% of those who irrpregnated their wives , and 22% of the overall group, Serum agglutinating antibody titers were elevated in 40% of those who irrpregnated their wives and in 42% of the overall group. Detailed quantitative tes ticle biopsy analysis sho<t.'ed no testicular defect either in successful or unsuccessful patients. We concl\xle that microsurgical vasovasostomy has a high rate of anatomic success and also a high pre<Jnancy rate. Pre<JnancY is not influenced by sperm antibody titers, varicocoele, or pre vious failure at surgery, and is only rro::lerately affected by si:erm count, except at very lw levels. However epi didymal bl<r,,uuts with secondary epididyma.l obstruction is a major cause of failure if vasoepidid}'m'..)Stomy is not perfonred in appropriate cases. P-47 eAmerican Society of Andrology MEMBERSHIP APPLICATION FOR (Please type or print) Malllng Address: ------- Phone to List: Education (undergraduate, graduate, postdoctoral - in chronological order) Institution Year Degree Present Position: Speclallty or Area of Research Interest: ------- EACH APPLICATION MUST INCLUDE: * 0 * O Total Amount Enclosed: Completed form Check, payable in U.S. $ to U.S. $ Checks must be in U.S. $ written on a U.S. bank or a draft American Society of Andrology or money order payable in U.S. dollars. 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