Program Book - American Society of Andrology

Transcription

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
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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
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1979-1980
..............................Nancy).Alexander
1980-1981
. .............................Philip Troen
1981-1982
. .............................Richard M. Harrison
1982-1983
. .............................Richard). Sherins
1983-1984
Andrzej Bartke
. . . .. .
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1984-1985
Rudi Ansbacher
1985-1986
Anna Steinberger
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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
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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.
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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,
4:50
Insemination Techniques, Results of AID, and Other Facets to Consider-Kirtly Jones,
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
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
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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
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
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
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
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
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
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
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.
·
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
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
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
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
·
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
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
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
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
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
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
·
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
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
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
·
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
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
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
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
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
·
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
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
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
·
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
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
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
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
,
·
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
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
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Program Book - American Society of Andrology

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