PIN 2016 Retreat Program - Graduate Program In Life Sciences

Transcription

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19th Annual Program in Neuroscience Retreat
Cover Illustrations
Cover design by Stephan Vigues
Special Thanks
Pre-doctoral Neuroscience Training Program
(PI: Margaret McCarthy)
And, the continued support of the School of Medicine, the
Dental School and the Graduate School.
About the Program in Neuroscience
The Program in Neuroscience at the University of Maryland offers research
training in a wide range of brain sciences, including cellular, molecular and
integrative neuroscience. Program faculty consists of more than 100
neuroscientists with laboratories located in the Medical, Dental,
Nursing and Pharmacy Schools, and the Maryland Psychiatric Research
Center. The program is affiliated with the Graduate Program in Life
Sciences in the School of Medicine. Investigators utilize a wide variety of
state-of-the-art techniques to investigate topics whose scope ranges from the
single molecule to the human brain.
The University of Maryland campus is located in the heart of historic,
downtown Baltimore, offering all the amenities of city life while
maintaining easy access to the countryside and the irresistible appeal of the
largest estuary system in the world, the Chesapeake Bay.
To learn more about our program and to keep updated on upcoming
seminars, retreats and other exciting program events, please visit:
http://neuroscience.umaryland.edu
PIN Gratefully Acknowledges
• Jessica Mong for her outstanding service as Director of Graduate
Education
• Todd Gould for chairing the Retreat Committee.
• Mary Kay Lobo for coordinating Neuroscience Journal Club
• Norbert Myslinski for his work on the National and International Brain
Bee
• Tom Blanpied chairing the PIN Seminar Committee
• Sarah Metzbower for serving as student representative on the PIN
Training Committee and chairing the PIN Student Training Committee.
• Quinton Banks, Sarah Rudzinskas, and Amanda Labuza for serving on the
GSA and Jonathan Van Ryzin for serving as Vice President of the USGA
• Amanda Labuza f o r s e r v i n g a s P r e s i d e n t o f NOVA
(Neuroscience Outreach and Volunteer Association)
• Special thanks to all Faculty, Students and Staff whose time and effort
ensured the recruitment of an outstanding group of new PIN students
• Dudley Strickland and Tom McHugh for leadership and service to GPILS
• The continued support of The School of Medicine, The Dental School, and
The Graduate School
• PIN’s graduate students and postdocs - the point of it all!
And to the Program in Neuroscience
Standing Committees:

Retreat
Todd Gould, Chair
Sherrie Lessans
Brian Mathur
Brian Polster
Dennis Sparta
James Waltz
Sarah Metzbower
Training
Jessica Mong, Chair
Tom Abrams
Joe Cheer
Greg Elmer
Reha Erzurumlu
Bruce Krueger
Marta Lipinski
Mary Kay Lobo
Frank Margolis
Brian Polster
David Seminowicz
Paul Shepard
Michael Shipley
Brian Mathur
Matt Trudeau
Sarah Metzbower
Michael Donnenberg, ad-hoc
Student Training Committee
Sarah Metzbower–Chair
Kara Cover
Poorna Dharmasri
Chase Francis
Alex Klausing
Amanda Labuza
Janell Payano Sosa
Nisha Pulimood
Austin Ramsey
Sarah Rudzinskas
Sara Stockman
Jonathan Van Ryzin
Seminar
Tom Blanpied, Chair
Leo Tonelli,
Todd Gould
Iris Lindberg
David Seminowicz
Sandra Jurado
19th Annual Neuroscience Retreat
Tuesday, June 7th, 2016
Notre Dame of Maryland University
Baltimore, MD 21210
Schedule
8:30-9:15
Registration and Breakfast
9:20-9:30
Introductory Remarks
Jessica Mong, Ph.D.
Director of Graduate Education, Program in Neuroscience
Associate Professor, Department of Pharmacology
9:30-10:00
Special Presentation
Elyse Sullivan., Ph.D.
Science Policy Analyst
Ripple Effect Communications, Inc
Pathways from PhD to consulting
10:00-11:10
Neuroscience TED Talks
Amanda Labuza- Understanding regulation of intracellular calcium levels
Panos Zanos, Ph.D.- Why does ketamine make sad mice happy?
Alberto Castro, Ph.D.- Treating pain: Are we close to developing an efficient
treatment?
Sarah Rudzinskas- Sex, drugs, and rock n' roll: Meth and the neuroscience of
women's sexual motivation
11:10-11:25
Coffee Break
11:25-12:15
Faculty Panel
Brad Alger, Ph.D. Professor Emeritus, Department of Physiology
Robert Buchanan, M.D., Director, Maryland Psychiatric Research Center and
Professor, Department of Psychiatry
Asaf Keller, Ph.D, Professor, Department of Anatomy and Neurobiology
Peg McCarthy, Ph.D., Professor and Chair, Department of Pharmacology
Scott Thompson, Ph.D., Professor and Chair, Department of Physiology
12:15-1:00
Lunch Break
1:00-2:15
Poster Session and Voting
2:15-2:45
Special Presentation
Scott Thompson, Ph.D.
Professor and Chair, Department of Physiology
Spread the good news: communicating your science with non-scientists
2:45-3:00
Afternoon Break
3:00-3:40
Lab Olympics
3:40-4:00
Awards
4:00-5:00
Keynote Address
Yavin Shaham, Ph.D.
Branch Chief, Intramural Research Program, NIDA-NIH
Incubation of drug craving after prolonged voluntary abstinence: behavior and
circuit mechanisms
Biographical details of the
Special Presenters
2015 Glaser Prize in Imaging
winning image mapping the
movement of molecules in a dendritic
spine done by Peter Li, an MD/PhD
PIN student.
Elyse Sullivan, Ph.D.
Project Manager and Science Policy Analyst
Ripple Effect Communications, Inc.
Dr. Sullivan received her Ph.D. in Neuroscience from the University of Maryland,
Baltimore where she created a novel neurophysiological technique for recording neural
activity from behaving animals. In her current position as Project Director and Science
Policy Analyst with Ripple Effect Communications, she uses her background in biomedical
research to support a wide variety of projects for the National Institutes of Health, the
Department of Health and Human Services Office of Human Research Protections, and the
Center for Medicare & Medicaid Services. In 2016, Dr. Sullivan co-led the NIH
Application Guide Working Group in a high-visibility initiative to simplify and reinvent the
NIH Application Guides. Dr. Sullivan also acted as lead analyst for two large-scale studies
investigating the racial and ethnic makeup, as well as the educational backgrounds, of the
U.S. biomedical research workforce in order to inform future NIH policy.
Scott Thompson, PhD
Professor and Chair
Department of Physiology
Program in Neuroscience
University of Maryland School of Medicine
Dr. Thompson received his Ph.D. at Stanford University doing his research in the
laboratory of Dr. David Prince. He was a Postdoctoral Fellow in Zurich, Switzerland, and
then at Columbia University. He returned to the University of Zurich, Switzerland, as an
Assistant Professor at the Brain Research Institute before being promoted to Associate
Professor. In 1998, he joined the Department of Physiology of the University of Maryland,
Baltimore, School of Medicine. He received tenure in 2002, and became Chair of the
Physiology Department in 2011.
Dr. Thompson’s lab is interested in how and when synapses change their strength during
learning and memory formation. Additionally, they have looked synapse formation that
can lead to epilepsy after head injury and central pain syndrome after spinal cord injury.
Recent research in their laboratory has extended synapse formation and strengthening
work and has shown the benefits of a new class of potential antidepressant medications, a
class of compounds that work similarly to ketamine but through more selective targeting
of a precise signaling molecule. The fast-acting nature of these compounds and the lack of
side effects make them attractive alternatives to more traditional anti-depressants.
Dr. Thompson is a member of the John Brick Mental Health Science Network Advisory
Panel. He also serves as Chair of the Society for Neuroscience Public Education and
Communication Committee as well as participating in several SfN Hill Days. He is also a
proud member of the band Pinkie and the Brain.
Yavin Shaham, Ph.D.
Branch Chief, Intramural Research Program
NIDA-NIH
Yavin Shaham received his BS and MA from the Hebrew U, Jerusalem, and his PhD from
the Uniformed Services University of the Health Sciences, Bethesda, MD. His postdoctoral
training was at Concordia U, Montreal, in the laboratory of Dr. Jane Stewart. Prior to
joining the NIDA Intramural Research Program as a tenure-track investigator, he was an
investigator at the Addiction Research Center in Toronto. He is currently a tenured Branch
Chief and a Senior Investigator. In 2001 he received the NIDA Director’s Award of Merit,
in 2006 he received the Society of Neuroscience Jacob Waletzky award for innovative
research in drug and alcohol addiction, and in 2016 he received the NARSAD
Distinguished Investigator Grant Award. He has published over 170 empirical papers,
reviews, and commentaries, and his papers were cited over 19,800 times (h-factor: 77;
Google Scholar). He currently serves as a Senior Editor for The Journal of Neuroscience
and as a Handling (Reviewing) Editor of Neuropsychopharmacology. He is also an editorial
board member of Biological Psychiatry, Psychopharmacology, and Addiction Biology. His
group currently investigates mechanisms of relapse to heroin, oxycodone, cocaine, and
methamphetamine, as assessed in rat models developed in his lab. Most recently, his lab
has developed an operant aggression reward model to study mechanisms of relapse to
aggressive behavior in mice.
TED Talks
Understanding regulation of intracellular calcium levels
Amanda Labuza1,2, Patrick F. Desmond1,3, Joaquin Muriel1, Michele L. Markwardt1, Mark A.
Rizzo1, Robert J. Bloch1
1 Department of Physiology, 2 Program in Neuroscience, 3 Program in Biochemistry and Molecular
Biology, University of Maryland, Baltimore, MD 21230 USA
Regulation of intracellular calcium is vital for healthy cellular function. Cell growth,
metabolism, and apoptosis are just some of the functions regulated by calcium (Berridge et
al, 2000). The endoplasmic reticulum (ER) is the primary internal storage site for calcium.
Alterations in normal ER calcium regulation can result in a variety of diseases ranging from
muscular dystrophy to diabetes to Alzheimer’s disease (Mekahli et al., 2011). Therefore,
understanding the mechanisms that regulate calcium levels in the ER and cytoplasm is an
important goal.
Sarco(endo)plasmic reticulum Ca2+-ATPase 1 (SERCA1) is an enzyme that actively pumps
calcium from the cytosol back into the SR and ER of skeletal muscle. SERCA1 activity is
inhibited by sarcolipin (SLN), a small transmembrane protein. We recently showed that
SERCA1 activity can also be regulated by another transmembrane protein, small ankyrin 1
(sAnk1) (Desmond, 2015). Here we show that sAnk1 binds to SLN. Coimmunoprecipitation, anisotropy based FRET (AFRET) and bimolecular fluorescence
complementation (BiFC) were used to show close association of sAnk1 and SLN. We are
currently using BiFC of sAnk1 and SLN and a cerulean form of SERCA in AFRET assays
to test the possibility that sAnk1, SLN, and SERCA1 form a 3-way complex.
In preliminary studies, we also show that sAnk1, SLN, and SERCA are all expressed in the
brain, suggesting that SERCA activity may be regulated similarly in neurons as in muscle.
These results have significant implications for the development of therapeutic approaches
to treat a variety of diseases linked to calcium misregulation such as muscular dystrophies
and neuropathies.
Why does ketamine make sad mice happy?
Panos Zanos, Ruin Moaddel, Patrick J. Morris, Polymnia Georgiou, Jonathan Fischell,
Greg I. Elmer, Manickavasagom Alkondon, Peixiong Yuan, Heather J. Pribut, Nagendra S.
Singh, Katina S.S. Dossou, Yuhong Fang, Xi-Ping Huang, Cheryl L. Mayo, Irving W.
Wainer, Edson X. Albuquerque, Scott M. Thompson, Craig J. Thomas, Carlos A. Zarate Jr.,
Todd D. Gould.
Department of Psychiatry, School of Medicine, University of Maryland, Baltimore, MD
Background: Major depressive disorder affects approximately 16 percent of the world
population at some point in their lives and is among the leading causes of death. Despite a
number of available monoaminergic-based antidepressants, these drugs require long-term
administration to be effective, and many patients never attain sustained remission of their
symptoms. Although the non-competitive glutamatergic N-methyl-D-aspartate receptor
(NMDAR) antagonist, (R,S)-ketamine (ketamine), exerts rapid and sustained antidepressant
effects, other NMDAR antagonists do not manifest identical actions, suggesting a distinct
ketamine mechanism of action. Ketamine is stereoselectively metabolised into a broad
array of metabolites, including norketamine, hydroxyketamines, dehydronorketamine and
the hydroxynorketamines (HNKs).
Methods: Tissue distribution and clearance measurements of ketamine and ketamine
metabolites were conducted with achiral liquid chromatography-tandem mass spectrometry.
Antidepressant efficacy in mice was assessed with the forced swim test, novelty suppressed
feeding test, learned helplessness test, social interaction following social defeat, and
reversal of anhedonia (sucrose preference and female urine sniffing preference) following
chronic corticosterone. Side-effect profiles were assessed with the open field test, pre-pulse
inhibition, drug discrimination, rotarod, and intravenous drug administration.
Electrophysiology assessing α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid
(AMPA) and NMDA receptor responses was conducted using field and whole-cell patchclamp recordings from rat hippocampal slices complemented by mouse in vivo EEG
recordings. Binding assays were used to assess NMDAR binding, and western blots to
assess activity of intracellular signaling pathways.
Results: (R)-ketamine, which has lower affinity for the NMDAR, exerted superior
antidepressant responses compared to (S)-ketamine. MK-801 (another NMDAR channel
blocker) did not induce the sustained effects observed following ketamine administration.
We showed that production of the (2S,6S;2R,6R)-HNK metabolite is essential for
ketamine’s antidepressant effects, and the (2R,6R)-HNK enantiomer exerts behavioral,
electroencephalographic, electrophysiological and cellular antidepressant actions in vivo.
These effects are NMDAR inhibition-independent but they involve early and sustained
AMPA receptor activation and increase in gamma EEG power. (2R,6R)-HNK did not exert
ketamine-associated self-administration, sensory dissociation or stimulant side effects.
Conclusions: Administration of (2R,6R)-HNK induces an acute increase in glutamatergic
signalling, followed by a long-term adaptation involving the upregulation of synaptic
AMPARs, as evidenced by an increase in GluA1 and GluA2 in hippocampal synapses. Our
results indicate a novel mechanism underlying ketamine’s unique antidepressant properties,
which involves the required activity of a distinct metabolite and which is independent of
NMDAR inhibition. These findings have relevance for the development of next generation,
rapid-acting antidepressants.
Treating pain, are we close to developing an efficient treatment?
Alberto Castro
Department of Anatomy and Neurobiology, School of Medicine, University of Maryland,
Baltimore, MD 21201
Chronic pain affects at least 116 million American adults with a national cost of $635
billion each year in medical treatment and lost productivity. Presently, chronic pain
treatments relive pain efficiently in only 50% of patients. Despite the huge effort in
research we have not been able to develop efficient treatments. What is failing and most
important, what can we do to improve? Join me to answer the question: Are we close to
developing an efficient treatment?
Sex, Drugs, and Rock n' Roll: Meth and the Neuroscience of Women's Sexual
Motivation
Rudzinskas, SA., Jurado, S., & Mong, JA.
Department of Pharmacology, University of Maryland, Baltimore, MD 21201 USA
Methamphetamine (MA) is a psychomotor stimulant of abuse which increases drive for
sexual activity. This increased sex drive results in ‘high risk’ sex behaviors, such as
unprotected intercourse and numerous sexual partners. In women, these behaviors pose
significant health concerns such as unplanned pregnancies and STD transmission, and
create an enormous societal economic burden. Previous work in our laboratory
investigating mechanisms underlying MA-facilitated female sexual motivation have given
us insights into the neural circuitry and components contributing to sexual motivation in the
normal female brain. Using cFos and lesion studies in a rodent model, we have
demonstrated that the medial amygdala (MeA) is the critical nucleus underlying MA’s
ability to increase sexual motivation in females. These studies resulted in the novel finding
that MA, in the absence of hormones, could increase progesterone receptor (PR) in the
MeA. Furthermore, a PR antagonist RU486 infused into the MeA after MA administration
blocks MA’s ability to increase sexual motivation. Taken together, this evidence led us to
hypothesize that PR upregulation in the MeA is responsible for increasing sexual
motivation. To test this hypothesis, we designed a lentivirus containing an ubiquitin-PRGFP-construct to overexpress PR, which was stereotaxically injected into the MeA two
weeks prior to sex behavior testing. Our preliminary results suggest that PR upregulation in
the MeA significantly increases overall number of sexually motivated behaviors in a
hormonally-primed female rat.
Poster Presentations
Author Index
Poster Number
1
2
3
4
5
6
7
8
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
31
32
33
34
35
36
37
38
Last Name
Guo
Chandra
Corsi
Engeln
Georgiou
Irving
Jarvela
Kilander
LeGates
Liu
Schenk
Shen
Song
Subramanian
Wenzel
Yang
Zlebnik
Bordt
Chen
Divakaruni
Francis
Hesselgrave
Joseph
Klimova
Konopko
Li
Ma
Metzbower
Patton
Peter
Pulimood
Shi
Stockman
VanRyzin
White
Gyawali
Ludman
Roberts
First Name
Wei
Ramesh
Nicole
Michel
Polymnia
James
Timothy
Michelle
Tara
Shuo
Lieven
Wenjuan
Chang
Krishna
Jennifer
Jiale
Natalie
Evan
Haiwen
Sai Sachin
Chase
Natalie
John
Nina
Melissa
Tuo
Elise
Sarah
Mary
Julia
Nisha
Da
Sara
Jonathan
Mike
Utsav
Taylor
Bradley
1. Further convergent observations on the antihyperalgesic effects of bone marrow
stromal cells in rodents
Wei Guo1, Yu-Xia Chu1,2, Satoshi Imai1,3, Jia-Le Yang1, Shiping Zou1, Zaid Mohammad1,
Feng Wei1, Ronald Dubner1, Ke Ren1
1
Department of Neural and Pain Sciences, School of Dentistry, & Program in Neuroscience,
University of Maryland, Baltimore, MD 21201, USA, 2Present Affiliation: Shanghai
Medical College, Fudan University, Shanghai, China, 3Present Affiliation: Department of
Clinical Pharmacology & Therapeutics, Kyoto University Hospital, Kyoto 606-8507, Japan
Bone marrow stromal cells (BMSC) have shown potential to treat chronic pain, although
much still needs to be learned about their efficacy and mechanisms of action under different
pain conditions. Here we provide further evidence on the effect of BMSC in rodent pain
models. While women exhibit higher prevalence of orofacial pain than men, it is unclear
whether BMSC produce pain relief in females as that in males. In a model of orofacial pain
involving injury of a tendon of the masseter muscle, we show in female rats that rat BMSC
(1.5 M cells, i.v.) attenuated behavioral hyperalgesia assessed by von Frey filaments and a
conditioned place avoidance test. The antihyperalgesia of BMSC in females lasted for 2856 d, which is shorter than that seen in males. To coincide preclinical findings with clinical
conditions, we extended our observation to human BMSC in rats after L5 spinal nerve
ligation (SNL). Human BMSC (1.5 M cells, i.v.) attenuated mechanical and thermal
hyperalgesia of the hind paw and suppressed SNL-induced aversive behavior, and the effect
persisted through the 8-week observation period. In an ex vivo trigeminal slice preparation,
BMSC treated-animals showed reduced amplitude and frequency of spontaneous miniature
excitatory postsynaptic currents. Electrical stimulation-evoked synaptic current was also
reduced in BMSC-treated animals. These results suggest inhibition of trigeminal neuronal
hyperexcitability and nociceptive transmission by BMSC. Finally, we observed that
GluN2A tyrosine phosphorylation and PKCγ immunoreactivity in the rostral ventromedial
medulla (RVM), a key site for descending pain modulation, was suppressed at 8-week after
BMSC in tendon-injured rats. As PKCγ activity related to NMDA receptor activation is
critical in opioid tolerance, these results help to understand long-term antihyperalgesia by
BMSC, which requires opioid receptors in RVM and apparently lacks the development of
tolerance. The present work adds convergent evidence to the growing preclinical literature
that support the use of BMSC in pain control.
2. Mitochondrial fission in nucleus accumbens projection neurons promotes cocaine
behavioral plasticity
Ramesh Chandra1, Michel Engeln1, Lace Riggs1, Chase Francis1, Shweta Das1, Kasey
Girven1, Ariunzaya Amgalan1, Leah Jensen1, Amy Gancarz2, Prasad Konkalmatt3, Sam A.
Golden4, Gustavo Turecki5, Scott J. Russo4, Sergio Iniguez6 David M. Dietz2, and Mary
Kay Lobo1
1
Department of Anatomy and Neurobiology, University of Maryland School of Medicine,
Baltimore, MD 21201, 2Department of Pharmacology and Toxicology and Institute on
Addictions, University at Buffalo, Buffalo, NY, USA.,
3
Division of Nephrology, University of Maryland School of Medicine, Baltimore, MD,
4
Fishberg Department of Neuroscience, Mount Sinai School of Medicine, New York, NY,
USA, 5Depressive Disorders Program, Douglas Mental Health University Institute and
McGill University, Montréal, Québec, Canada, 6Department of Psychology, California State
University, San Bernardino, California.
Altered brain energy homeostasis is a key adaptation occurring in the cocaine-addicted
brain. However, the underlying mechanisms that govern these homeostatic adaptations are
not clearly defined. One such mechanism, which is a fundamental component of energy
homeostasis and has not been addressed in cocaine abuse are mitochondrial dynamics;
which can include mitochondrial biogenesis, fission, and fusion. To provide insight into this
we assess mitochondrial dynamics in the two nucleus accumbens (NAc) projection medium
spiny neuron (MSN) subtypes, those enriched in dopamine D1 vs. D2 receptors in mice that
self-administer cocaine. Using a Cre inducible adeno-associated virus (AAV)-double
inverted floxed open reading frame (DIO)-mito-dsRed combined with D1-Cre and D2-Cre
mouse lines we label mitochondria in MSN subtypes. Cocaine self-administration caused an
increase in the frequency of smaller mitochondria in D1-MSN dendrites, implicating
enhanced mitochondrial fission in D1-MSNs. Consistent with our data we observe an
increase in mRNA of Dynamin-related protein 1 (Drp1), a mediator of outer mitochondrial
membrane fission, in NAc of rodents that self-administer cocaine and in postmortem NAc
of cocaine dependent individuals. Using the RiboTag approach, we observe an upregulation
of ribosome-associated mRNA of Drp1in D1-MSNs and a decrease in D2-MSNs after
repeated cocaine. Additionally, the activated form of Drp1 protein is increased in NAc of
mice after repeated cocaine injections. We next used a small molecule inhibitor of
mitochondrial fission, Mdivi-1, which blocks the activated form of Drp1. Mdivi-1 treatment
blocked seeking after cocaine conditioned place preference and the expression of cocaine
locomotor sensitization. Finally, we generated Cre-inducible Drp1-constitutively active
(CA) and Drp1-wild-type (WT) AAVs. Overexpression of Drp1-CA in D1-MSNs increased
cocaine taking during acquisition to self-administration and seeking behavior after
abstinence. Our findings demonstrate a novel role for altered mitochondrial fission in NAc
in cocaine abuse and implicate that blockade of mitochondrial fission has potential
therapeutic treatment for cocaine addiction.
3. Beliving or not in treament’s side effects: Behavioral outcome and personality traits
associated to a nocebo effect in motor performance.
Nicole Corsi1,2, Michele Tinazzi2, Andani Mehran Emadi2,3, Mirta Fiorio2.
1
Pain and Translational Symptom Science Department, University of Maryland Baltimore
(USA), 2 Department of Neurological and Movement Sciences, University of Verona
(Italia), 3 Department of Biomedical Engineering, University of Isfahan (Iran)
Despite subject’s belief about treatment’s efficacy being crucial for the nocebo effect, no
study until now has tackled this issue. With this study we investigated whether the
persistence of belief and personality traits could account for individual differences in the
magnitude of the nocebo response. 27 volunteers were asked to perform a motor task. After
a training, participants underwent a nocebo procedure, in which an inert treatment was
applied together with verbal instructions about its negative effects on force. In a
conditioning session, subjects were exposed to the (fake) effects of the treatment. Finally, in
a test session, subjects received the same treatment and performed the motor task without
reduction of the feedback. Crucially, two different patterns have been observed in terms of
beliefs: some subjects gave higher scores in the test compared to the conditioning session
(high-responders, N=15), whereas other subjects did the opposite (low-responders, N=12).
Results showed that high-responders had a stronger nocebo effect, as evidenced by lower
levels of force (p < 0.001), higher feeling of weakness (p < 0.001) and higher sense of effort
(p = 0.036), compared to low-responders. Personality questionnaires revealed that highresponders had lower level of optimism (p = 0.008) and self-directedness (p = 0.048), but
higher anxiety trait (p = 0.008) and harm avoidance (p = 0.008) than low-responders. These
findings show that the magnitude of the nocebo response can be modulated by different
factors, such as the persistence of subject’s belief about the efficacy of the treatment and
personality traits.
4. Juvenile onset of stereotypy with loss of BDNF signaling in D1R expressing striatal
neurons
M. Engeln1, R. Chandra1, A. La2, T.C. Francis1, M.K. Lobo1
1
Department of Anatomy and Neurobiology, School of Medicine, University of Maryland,
Baltimore, MD 21201, 2 University of Maryland, College Park, MD 20742
Imbalance between D1- vs. D2-receptor containing medium spiny neuron (MSN) basal
ganglia output-pathways is implicated in stereotyped disorders including Tourette
Syndrome (TS). Surprisingly, there is little information on the molecular role of MSN
subtypes in TS or other stereotypy disorders. We have a mouse model with a deletion of
TrkB (the BDNF receptor) in D1-MSNs (D1-Cre-flTrkB mice), in which a subset of mice
displays involuntary stereotypic behaviors beginning around 3 weeks of age. Consistent
with an impaired GABAergic system in TS, these mice display a decrease in striatal
GABA-A subunits accompanied by reduced inhibition in striatal D1-MSNs. We first
characterized repetitive behaviors in D1-Cre-flTrkB mice with stereotypy (S), or with no
stereotypy (NS), and D1-Cre control mice. Complete turns, head tics, rearing, and grooming
are assessed weekly from ages 3 to 8 weeks. We found that D1-Cre-flTrkB-S mice display
more complete turns at all ages compared to D1-Cre-flTrkB-NS and control mice. D1-CreflTrkB-S mice exclusively display head tics, which decline from juvenile to adult ages.
Since the transcription factor, early growth response 3 (Egr3) is regulated by BDNF and
Egr3 transcriptionally regulates a subset of GABA-A subunits, we are examining striatal
Egr3 mRNA in all groups. Our preliminary data demonstrates decreased Egr3 mRNA in the
striatum of D1-Cre-flTrkB mice. To confirm that Egr3 is decreased in D1-MSNs, we are
using D1-Cre-flTrkB-RiboTag mice to isolate ribosome-associated mRNA from these mice,
at both juvenile (4 weeks) and adult (8 weeks) ages. In addition, D1-Cre-flTrkB-RiboTag
mice allow us to examine GABA-A receptor subunit expression in D1-MSNs to examine
specific GABA-A subunit alterations that may underlie stereotypy. We are also examining
Egr3 transcriptional regulation of GABA-A subunits, using chromatin immunoprecipitation
(ChIP) at these two age time points. Finally, we overexpress Egr3 in D1-MSNs with AAVDIO-Egr3-eYFP at 2 and 6w of age to rescue these stereotyped behaviors. Our findings
demonstrate that D1-MSNs through dysfunctional BDNF signaling play a role in juvenile
onset of stereotypy behaviors. The enhanced stereotypy behaviors potentially occur through
reduced inhibition in D1-MSNs via altered Egr3 regulation of GABA-A subunits. Our
ongoing studies can provide novel insight into the cell subtypes and molecular mechanisms
underlying stereotypy disorders.
5. Increased susceptibility of hypogonadal male mice to social stress is mediated by
estradiol
Polymnia Georgiou1, Panos Zanos1, Margaret M. McCarthy3,5, Mary Kay Lobo1,4, Istvan
Merchenthaler6, Lazlo Prokai2, Todd D. Gould1,3,4
1
Department of Psychiatry, School of Medicine, University of Maryland, Baltimore,
Department of Pharmacology &Neuroscience, University of North Texas, 3Department of
Pharmacology, School of Medicine, University of Maryland, Baltimore, 4Department of
Anatomy & Neurobiology, School of Medicine, University of Maryland, Baltimore,
5
Department of Physiology, School of Medicine, University of Maryland, Baltimore,
6
Department of Epidemiology and Public Health, School of Medicine, University of
Maryland, Baltimore
2
Hypogonadism has been associated with increased propensity of developing depression in
men. Although testosterone exerts antidepressant effects on its own, it is not clear whether
these effects are mediated by direct actions of testosterone on the androgen receptor or by
its aromatase-dependent conversion to 17β-estradiol (E2), and activation of estrogen
receptors (ER).
Male sham or orchiectomized mice received vehicle, testosterone, 17β-estradiol, or
10β,17β-dihydroxyestra-1,4-dien-3-one (DHED; brain-selective E2) implants 10-days prior
to subthreshold social defeats to assess their effects on hypogonadism-induced susceptibility
to develop social avoidance and anhedonia.
While orchiectomy did not induce behavioral changes on its own, combined hypogonadism
and social stress induced social interaction deficits and anhedonia. Although testosterone
replacement reversed these deficits, blockade of the androgen receptor did not induce social
interaction deficits or anhedonia following social defeat, suggesting an androgenindependent mechanism to mediate the observed susceptible phenotype in male mice. In
contrast, aromatase inhibition induced a susceptible phenotype indicating that these
maladaptive symptoms of social stress under hypogonadal conditions are primarily driven
by the conversion of testosterone to E2. In support of this, both E2 and DHED treatment
reversed the affective behaviors induced by hypogonadism/social stress. We also show that
this effect is mediated by the ERβ since mice lacking this gene manifested a behavioral
phenotype identical to the one caused by hypogonadism/social stress.
Our findings demonstrate that hypogonadism increases the risk to develop maladaptive
responses following a stressful event and highlight a novel role for E2 in males to modulate
depressive-like behaviors via an ERβ-dependent mechanism.
6. Optogenetic and pharmacogenetic interrogation of central amygdala corticotropin
neurons on ninge ethanol drinking
J. M. Irving, C. J. Maehler, D.R. Sparta
University of Maryland School of Medicine, Baltimore MD 21201
Binge ethanol drinking is an increasing problematic component of alcoholism and has been
linked to the progressive dysfunction of various organs. While the initial appetitive effects
of ethanol consumption may attract someone to binge drink at first, over time this appetitive
feeling diminishes and a new aversive feeling to the absence of ethanol emerges. This
negative emotional state drives the alcoholic to consume ethanol for relief. Understanding
the specific neural circuit mechanisms that contribute to binge drinking is critical because
dysregulation of these neural processes is hypothesized to underlie many of the maladaptive
behaviors in neuropsychiatric illness, including alcoholism. Corticotropin releasing factor
(CRF) neurons within the central amygdala (CeA) participate in both the negative affect and
binge drinking, but the precise circuit mechanisms sufficient for driving these behaviors
remains poorly understood. Here we used optogenetics coupled with in vivo
electrophysiology and pharmacogenetics to determine how binge drinking alters CeA CRF
neurons. We hypothesized that increased activity of CeA CRF neurons drives excessive
ethanol consumption. First, we recorded the activity of CeA CRF neurons in a binge
drinking paradigm. Preliminary analyses revealed that CeA CRF neurons exhibited
increased firing activity with repeated binge drinking sessions. Ongoing experiments are
examining whether pharmacogenetic inhibition of CeA CRF neurons can reduce excessive
ethanol intake. These data suggest that repeated binge drinking leads to progressive and
persistent increases in the activity of CeA CRF.
7. The neural chaperone proSAAS blocks α-synuclein fibrillation and neurotoxicity
Timothy S. Jarvela1, Hoa A. Lam2, Michael Helwig1, Nikolai Lorenzen3, Daniel E. Otzen3,
Pamela J. McLean4, Nigel T. Maidment2 and Iris Lindberg1*
1
University of Maryland-Baltimore; 3University of Aarhus, Denmark; 4Mayo Clinic,
Florida; and 2University of California, Los Angeles
Emerging evidence strongly suggests that chaperone proteins are cytoprotective in
neurodegenerative proteinopathies involving protein aggregation; for example, the
accumulation of aggregated α-synuclein into the Lewy bodies present in Parkinson’s
disease. Of the various chaperones known to be associated with neurodegenerative disease,
the small secretory chaperone known as proSAAS has many attractive properties. We show
here that proSAAS, widely expressed in neurons throughout the brain, is associated with
aggregated synuclein deposits in the substantia nigra of Parkinson’s disease patients.
Recombinant proSAAS potently inhibits the fibrillation of α-synuclein in an in vitro assay;
residues 160-180, containing a highly conserved element, are critical to this bioactivity.
ProSAAS also exhibits a neuroprotective function; proSAAS-encoding lentivirus blocks αsynuclein-induced cytotoxicity in primary cultures of nigral dopaminergic neurons, and
recombinant proSAAS blocks α-synuclein-induced cytotoxicity in SH-SY5Y cells. Four
independent proteomics studies have previously identified proSAAS as a potential
cerebrospinal fluid biomarker in various neurodegenerative diseases. Coupled with prior
work showing that proSAAS blocks beta amyloid aggregation into fibrils, this study
supports the idea that neuronal proSAAS plays an important role in proteostatic processes.
ProSAAS thus represents a possible therapeutic target in neurodegenerative disease.
8. Autism-associated mutations in CEP290 disrupt cell proliferation process
Michaela B. C. Kilander and Yu-Chih Lin
Laboratory of Neuronal Connectivity, Program in Neuroscience, Hussman Institute for
Autism, Baltimore, MD 21201
Autism is classified as a neurodevelopmental condition, but the molecular mechanisms
contributing to the altered neurophysiology observed in individuals with autism are still
largely unknown. In large-scale genetic analysis for autism risk genes, CEP290, a
centrosomal protein has been identified as one of the candidate genes. Mutations in the
CEP290 gene are common in ciliopathies; severe multi-organ disorders caused by primary
cilia dysfunction. The primary cilium is a microtubule rich cell protrusion crucial for normal
cell migration, polarity and division. Moreover, Sonic Hedgehog (Shh) signaling, a
biological pathway necessary for proper tissue development and maintenance, is
preferentially localized to the primary cilium and is essential for proliferation of granule cell
progenitors (GCPs) during cerebellar development. Interestingly, defects in cerebellar
development occur frequently in CEP290-associated ciliopathies. However, how mutations
of CEP290 may contribute to autism phenotypes is still unknown. Here, we show that
CEP290 regulates cell proliferation and autism-associated mutations disrupt this process.
9. Chronic stress alters the electrophysiological properties of hippocampal-accumbens
synapses.
Tara LeGates and Scott Thompson
Department of Physiology, University of Maryland School of Medicine, Baltimore, MD,
21201.
Depression is a leading cause of disability with a lifetime prevalence of 12% and 20% in
men and women, respectively. A common risk factor that increases the likelihood of
depressive episodes is stress. The nature of the stress-induced neuronal changes that
promote depression remains unknown. There is increasing evidence that chronic stress
weakens neuronal communication in multiple brain regions including the hippocampus and
nucleus accumbens (NAc), perhaps accounting for the wide range of behaviors affected by
stress. The ventral hippocampus provides excitatory input to the NAc shell, and this synapse
has received increased attention due to its potential role in mood regulation, specifically in
response to reward and motivation to seek rewards.
We used whole-cell
electrophysiological recordings to characterize the properties of the connections from the
hippocampus to the NAc in the brains of mice, including the contributions of glutamate
receptor subtypes. We found that Hip-NAc synapses become strengthened in response to
high frequency activity due to classical activity-dependent long-term plasticity mechanisms
but independent of dopamine receptor signaling. These results were similar in D1- and D2MSNs. Furthermore, the properties of this synapse change in a stress-induced model of
depression, and the observed changes differ in D1- and D2-MSNs. This work defines a
specified neuronal circuit that is responsible for mood regulation and further our
understanding of excitatory synaptic strength as a critical mediator of this process.
Understanding the neuronal changes that underlie depression and antidepressant response
will provide key insight into developing new, more effective treatments for this disorder.
10. Disrupted autophagy after spinal cord injury is associated with necroptosis.
Shuo Liu1, Chinmoy Sarkar2, Eugene Y. Koh1, Junfang Wu2 and Marta M. Lipinski2
1
Department of Orthopaedics, University of Maryland School of Medicine, Baltimore, MD.
Department of Anesthesiology and the Center for Shock, Trauma and Anesthesiology
Research (STAR), University of Maryland School of Medicine, Baltimore, MD.
2
Necroptosis is a caspase-independent programmed cell death, which is mediated by
receptor-interacting protein 1 (RIP1). Autophagy is a catabolic mechanism facilitating
degradation of cytoplasmic proteins and organelles in a lysosome-dependent manner.
Current literature has shown that autophagy can participate in necroptosis. RIP1 inhibitor
(necrostatin-1) has demonstrated its benefit on functional recovery of spinal cord injury
(SCI). However, the mechanism of this improvement in SCI is still unclear. Using a rat
model of contusive SCI, we observed accumulation of LC3-II positive autophagosomes
starting at post-trauma day 1. This was accompanied by a pronounced accumulation of
autophagy substrate protein p62, indicating that early elevation of autophagy markers
reflected disrupted autophagosome degradation. Levels of lysosomal protease cathepsin D
(CTSD) and numbers of CTSD-positive lysosomes were also decreased at this time,
suggesting that lysosomal damage may contribute to the observed defect in autophagy flux
and further neuronal cell death. Consistently in our rat model of SCI, motor neurons
showing disrupted autophagy co-expressed necrosome element RIP1, suggesting disrupted
autophagy after SCI may contribute to necroptosis. In addition, motor neurons with a low
number of CTSD-positive lysosome co-localized with a higher number of RIP1-positive
puncta, suggesting that lysosomal damage may contribute to the observed RIP1
accumulation in motor neuron. In vitro study with PC12 cells and neurons also supports this
finding by showing an increased expression of RIP1 following chloroquin-induced
lysosomal inhibition. Together these findings indicate that SCI causes lysosomal
dysfunction that contributes to autophagy disruption and is associated necroptosis.
11. Differential neural processing of treatment and stimulus-expectancy.
Lieven A. Schenk1, 2, Selim Onat2, Christian Sprenger2, Christian Büchel2
1
Department of Pain and Translational Symptom Science, University of Maryland,
Baltimore, Baltimore, MD 21201, USA, 2Department of Systems Neuroscience, University
Medical Center Hamburg-Eppendorf, 20246 Hamburg, Germany
Background: The influence of contextual information on pain perception is a crucial part of
placebo analgesia. In two context groups, we compared neural pain responses associated
with treatment expectation against expectation regarding pain stimulus intensity.
Methods: A total of 48 healthy volunteers received heat pain stimuli along with two distinct
cues anticipating either treatment and/or pain (group 1: treatment/no treatment cue versus
group 2: low/normal pain cue). fMRI measurements were used to explore neural responses
during conditioning and test sessions while volunteers had to rate expected and experienced
pain. We used standard univariate analysis and multivariate Representational Similarity
Analysis (RSA) to investigate the neural differences between the two groups.
Results: We observed lower expectancy and pain ratings with the treatment/low pain cue in
both groups. Importantly, this reduction was stronger in the treatment context compared to
the stimulus intensity context during the test phase. At the neural level, we observed
univariate BOLD response differences between the groups in the rostral ACC and the
hippocampus during the conditioning phase and in the lateral PFC and the amygdala during
the test phase. In the multivariate RSA, we observed group differences in the bilateral
ventral striatum during the cue presentation and in the postcentral gyrus and the parietal
operculum during the pain stimulation.
Conclusion: Taken together, our data shows that a treatment context leads to larger
expectancy effects on pain. Both imaging analysis methods show significant differences in
neural processing between both contexts.
12. Autism-associated mutation of syntaxin binding protein 5 disrupts dendrite
arborization.
Wenjuan Shen, Jeannine Frei, and Yu-Chih Lin
Laboratory of Neuronal Connectivity, Program in Neuroscience, Hussman Institute for
Autism, Baltimore, MD 21201
Autism is a neurological condition that features marked qualitative differences in
communication and social interaction. As many as 1/3 of individuals with autism spectrum
condition also have epilepsy. Consistent with the extremely heterogeneous presentation of
autism, genetic studies have implicated numerous genes that may contribute to the autism
phenotype. Deletion and mutations of syntaxin binding protein 5 (STXBP5, also known as
tomosyn) are identified in association with autism and epilepsy. STXBP5/tomosyn is a
syntaxin binding protein that contains a WD40 domain at the N-terminus and a SNARE
motif at the C- terminus. STXBP5/tomosyn has a presynaptic role that negatively regulates
neurotransmitter release by forming a syntaxin-SNAP25-tomosyn complex.
STXBP5/tomosyn has also been shown to regulate neurite outgrowth in immature neurons.
WD40 as scaffolding domains, have a variety of functions such as signal transduction and
vesicle trafficking. Interestingly, two autism-associated variants of STXBP5 exhibit
missense mutations at the WD40 domain. Here, we show that wildtype (WT)- and mutant
tomosyn all localize to axons, dendrites and dendritic spines. Overexpression of WTtomosyn increased dendritic arbor complexity by increasing total dendrite length and branch
tip number, compared to control neurons. The autism-associated tomosyn mutant failed to
induce the dendrite complexity when compared to the WT-tomosyn. In conclusion,
STXBP5/tomosyn plays a role in regulating dendrite arborization. Mutations of STXBP5
found in individuals with autism may alter dendrite arborization and potentially disrupt
normal developmental processes in the brain.
13. Expansion of brain T cells in homeostatic conditions in lymphopenic Rag2-/- mice.
Chang Song1, James D. Nicholson1, Sarah M. Clark1,2, Xin Li1, Achsah D. Keegan3,2,
Leonardo H. Tonelli1,2
1
Laboratory of Behavioral Neuroimmunology, Department of Psychiatry, University of
Maryland School of Medicine, Baltimore, MD, United States, 2Research and Development
Service, Department of Veterans Affairs, VA Maryland Health Care System, Baltimore,
MD, United States, 3Center for Vascular and Inflammatory Diseases, University of
Maryland School of Medicine, Baltimore, MD, United States
Background/Objectives: The concept of the brain as an immune privileged organ is rapidly
evolving in light of new findings outlining the sophisticated relationship between the central
nervous and the immune systems. The role of T-cells in brain development and function, as
well as modulation of behavior has been demonstrated by an increasing number of studies.
Moreover, recent studies have redefined the existence of a brain lymphatic system and the
presence of T-cells in specific brain structures, such as the meninges and choroid plexus.
Methods: In the present study we employed the Rag2-/- mouse model of lymphocyte
deficiency and reconstitution by adoptive transfer to study the temporal and anatomical
expansion of T-cells in the brain under homeostatic conditions. Lymphopenic Rag2-/- mice
were reconstituted with 10 million lymphoid cells and studied at one, two and four weeks
after transfer. Moreover, lymphoid cells and purified CD4+ and CD8+ T-cells from
transgenic GFP expressing mice were used to define the neuroanatomical localization of
transferred cells.
Results: T-cell numbers were very low in the brain of reconstituted mice up to one week
after transfer and significantly increased by 2 weeks, reaching wild type values at 4 weeks
after transfer. CD4+ T cells were the most abundant lymphocyte subtype found in the brain
followed by CD8+ T-cells and lastly B cells. Furthermore, proliferation studies showed that
CD4+ T-cells expand more rapidly than CD8+ T-cells. Lymphoid cells localize abundantly
in meningeal structures, choroid plexus, and circumventricular organs. Lymphocytes were
also found in vascular and perivascular spaces and in the brain parenchyma across several
regions of the brain, in particular in structures rich in white matter content.
14. The basal ganglia has altered inhibitory receptor expression in autism.
Krishna Subramanian, Cheryl Brandenburg and Gene J. Blatt
Hussman Institute for Autism
Background: The basal ganglia (BG) is a collection of sub-cortical nuclei that contain
mainly inhibitory GABAergic medium spiny neurons (MSN’s). The BG projects to the
thalamus and has reciprocal connections with multiple cortical regions and the cerebellum.
Parts of the BG are implicated in OCD, habit formation, motor, speech and language
disorders. Thus, the BG is an ideal region of interest to examine the neurochemical basis of
repetitive, stereotyped behavior and social communication difficulties observed in autism.
Objective: Determine levels of expression of inhibitory GABAA receptors in functional
subdivisions of the BG in autism. Specifically, the dorsal striatum consisting of caudate and
putamen, and the ventral striatum including the core and shell territories of nucleus
accumbens (NAcc) were quantified.
Results: Tritiated flunitrazepam binding was found to be significantly increased in the
caudate region of the dorsal striatum in subjects with autism compared to controls
(p=0.015). Additionally, significantly increased benzodiazepine binding site density was
found in the region of caudate that receives putative projections from the ACC (p=0.03).
Conclusions: The results from this initial study suggest that there are significant increases in
the expression of inhibitory GABAA receptors in the striatum of subjects with autism.
Normally, the limbic ACC input that specifically targets the striatal interneurons plays an
important role in BG efferent projections. In autism, there is likely a disturbance of
inhibitory/excitatory balance within BG circuitry.
15. Cannabinoid exposure in adolescence, but not adulthood, modulates subsequent
cocaine reward.
Jennifer M. Wenzel PhD1 and Joseph F. Cheer PhD1,2
1
Department of Anatomy and Neurobiology, 2Department of Psychiatry, University of
Maryland School of Medicine, Baltimore, MD, USA
Marijuana is the most commonly abused illicit drug among adolescents, and regular use of
cannabinoids (CBs) in this vulnerable population is associated with the development of
psychiatric disease including drug abuse. Human research indicates that adolescent CB use
predicts future cocaine (COC) abuse, however, the underlying cause of this phenomenon is
unclear. Adolescent CB exposure disrupts the dopaminergic response to COC in adulthood
suggesting that CB use may disturb the subjective experience of COC. It is well
documented that immediate euphoria produced by COC administration is subsequently
replaced by feelings of dysphoria and anxiety. Thus, it is likely that an individual’s
experience of either of these opposing processes may motivate successive COC use. Here
we utilize a modified place conditioning test to assess how CB exposure in adolescence
affects COC’s dual positive and negative effects. In this test rats are conditioned to
associate a unique environment with the effects of COC present either immediately or 15
min after IV COC injection. Characteristically following place conditioning rats exhibit a
preference for the environment paired with COC’s immediate/positive effects (a CPP), and
an aversion for the environment paired with COC’s delayed/negative effects (a CPA).
Therefore, we assessed the impact of CB exposure in adolescence on adult COC reward and
aversion. Adolescent male rats were treated with one of three doses of the synthetic CB
WIN 55,212-2 (WIN; 0.5mg/kg, 2mg/kg, 5mg/kg) or its vehicle (VEH) once per day for
eight days (PND35-42). Following treatment rats were left in their home cages until they
reached adulthood at which point they underwent place conditioning for either the
immediate/positive or delayed/negative effects of COC (PND77-86). Interestingly, while
rats treated with VEH during adolescence developed the typical pattern of COC CPP and
CPA, exposure to WIN during adolescence dose-dependently resulted in the development of
CPA for the immediate effects of COC (an outcome opposite to the canonical COC CPP),
while not disrupting CPA for COC’s delayed/negative effects. These data suggest that CB
exposure in adolescence diminishes COC’s rewarding effects in adulthood and, in fact,
results in a predominantly negative experience within the first 5 min after administration
(i.e. the length of each conditioning trial). It remains unclear how these alterations in COC
reward translate to COC seeking in adulthood. In animal models, however, reduced COC
reward is associated with increased COC intake, suggesting that these observed decrements
in COC reward might contribute to increased COC administration.
16. Chronic stress-induced lower back pain in a rat trigeminal neuropathic pain model
and its central mechanisms.
Jiale Yang, Yaping Ji, Jamila Asgar, Shiping Zou, Wei Guo, Ke Ren, Ronald Dubner,
Richard Traub and Feng Wei
Department of Neural and Pain Science, Dental School, & Program in Neuroscience,
The Center to Advance Chronic Pain Research, University of Maryland, Baltimore, MD
21201
Chronic pain and stress impact on each other and are potentially modifiable risk factors for
poor health outcomes. Clinical data have identified an association between perceived stress
and various pain syndromes including orofacial and abdominal pain. However, the
mechanisms underlying how psychological stress exacerbates pain experiences are poorly
understood. In the present study, we investigated the effects of chronic stress on behavioral
hypersensitivity in the rat trigeminal neuropathic pain model induced by chronic
construction injury to the infraorbital nerve (CCI-ION). Ten days after CCI or sham
surgery, rats were subject to 3 day forced swim and subsequently tested daily for
mechanical sensitivity. While CCI-ION induced orofacial mechanical hyperalgesia and
allodynia occurred, visceral hyperactivity and lower back pain were observed after forced
swim without significant changes in mechanical nociception at both fore and hind paws.
This behavioral visceral hypersensitivity persisted at least 11 days in female rats but only 7
days in male rats, suggesting that chronic stress results in overlapping visceral pain
conditions in the presence of an existing orofacial neuropathic pain. The current model
further demonstrates a potential sex difference of stress-induced comorbid pain. In order to
explore involvement of central mechanisms, functional silence of hypothalamus subnuclei,
genetic depletion of 5-HT in the rostroventral medulla (RVM), or intrathecal injection of the
selective 5-HT3R inhibitor Y25130 could attenuate visceral hypersensitivity at 7d after
forced swim. This finding indicates that neuronal circuit between hypothalamus and the
RVM may mediate stress-induced comorbid pain conditions by enhancement of 5-HTdependnet descending pain facilitation.
17. Cannabinoid modulation of the neurophysiological correlates of motivation in the
Q175 mouse model of Huntington's disease.
Natalie E. Zlebnik1, Houman Qadir1, Manuel A. Anaya1, Ellen A. Cole1, Iness Gildish1,
Dan P. Covey1, Joseph F. Cheer1,2
1
Department of Anatomy and Neurobiology, University of Maryland School of Medicine,
Baltimore, Maryland 21201, USA, 2Department of Psychiatry, University of Maryland
School of Medicine, Baltimore, Maryland 21201, USA
Huntington’s disease (HD) is an inherited neurodegenerative disorder caused by a
polyglutamine expansion in the huntingtin gene and results in the progressive loss of striatal
and cortical neuropil. Striatal neurodegeneration occurs first and mainly affects the indirect
pathway of the basal ganglia, causing dyskinesia followed later by akinesia. However, the
marked motor impairments characteristic of HD may often be preceded by motivational and
cognitive dysfunction. Recent studies report early loss of striatal endocannabinoid (eCB)
receptors and degradation enzymes in HD patients and rodent models, but how this relates
to motivational deficits is unclear. In order to characterize striatal network dynamics and
eCB function in HD progression, extracellular electrophysiological recordings were
collected at multi-electrode arrays implanted in the nucleus accumbens of wild-type (WT)
and Q175 knock-in mice during a task of motivation (progressive ratio schedule) and
following treatment with the CB1 receptor inverse agonist AM-251 and the MAGL
inhibitor JZL-184. Preliminary results indicate lower motivation for reward in Q175
compared to WT mice. These deficits in motivation were accompanied by significantly
different session-wide power distribution across several frequency bandwidths and in
reward-evoked power specifically in the gamma bandwidth among the genotypes. Raising
tissue levels of 2-AG with JZL-184 attenuated accumbal gamma power and rescued reward
motivation in the Q175 mice in a CB1-receptor-dependent manner. Overall, these results
suggest that elevating eCB tone in the nucleus accumbens may ameliorate the motivational
impairments inherent to HD.
18. The Influence of Oxygen Tension on Mitochondrial Respiratory Inhibition in
Inflammatory Microglia
Evan A. Bordt and Brian M. Polster
University of Maryland School of Medicine, Baltimore, Maryland 21201
Excessive or prolonged microglial activation exacerbates brain damage through the release
of nitric oxide (NO) and other proinflammatory factors. In vitro studies of microglial
neurotoxicity are performed at atmospheric O2 (160 mm Hg) despite a far lower O2 tension
in the brain (~3% O2, 23 mm Hg). NO is thought to impair mitochondrial respiration by
competing with O2 at complex IV. Surprisingly, the influence of pO2 on NO-mediated
respiratory inhibition has been largely overlooked in in vitro studies despite this O2
competition model. This study tested the hypothesis that microglial activation inhibits
microglial respiration in a pO2-dependent manner. We predicted that the higher pO2 would
lead to reduced effects of microglial NO on the bioenergetics of the cells at 21% vs 3% O2.
Basal and maximal O2 consumption by microglial cells decreased by 6-8 hr, concomitant
with increase in NO release. Inhibition of iNOS prevented inhibition at 3% O2, but only
modestly at 21% O2. At physiological O2, inhibition was reversed by scavenging NO, but
was unable to rescue impairment at atmospheric O2. The exogenous electron donors TMPD
and idebenone fully rescued respiration at 21%, but not at 3% O2. Data suggest that
respiratory inhibition in microglia activated with LPS/IFN-γ is mediated by NO at
physiological O2, while another mechanism appears to be mediating inhibition at
atmospheric O2. Complex IV appears to be the target of NO-mediated respiratory inhibition
at 3% O2, but appears functional at 21% O2. Our findings illustrate the importance of
modeling neuroinflammatory disease at physiological O2.
19. Distinct organization of evoked and spontaneous vesicle fusion sites within single
CNS active zones
Haiwen Chen1,2,3, Ai-Hui Tang1,2, Sarah Ransom Metzbower1,2, Thomas A. Blanpied1,2
1
Department of Physiology, 2Program in Neuroscience, and 3Medical Scientist Training
Program, University of Maryland School of Medicine, Baltimore, MD 21201, USA
Because glutamate receptors are inhomogenously distributed in the postsynaptic density, the
organization of vesicle docking and fusion sites within single active zones (AZs) will
strongly influence synaptic strength. However, there is extremely limited information about
where within the AZ action potentials trigger vesicle fusion, and even less about the
location of spontaneous vesicle fusion. Interestingly, there is controversy over whether
spontaneous and evoked release utilize different vesicle pools, involve different trafficking
and fusion machinery, or activate different groups of receptors. All these factors suggest
that spontaneous and evoked fusion could be organized at spatially distinct regions of the
AZ, but this has not been tested.
To address this, we first studied the subsynaptic distributions of key presynaptic proteins
likely to define vesicle fusion sites, including RIM1/2, Munc13, and Bassoon. Using 3D
dSTORM on immunostained cultured hippocampal neurons, we found that all three protein
distributions were distinctly heterogeneous with high local-density peaks (~80-110 nm in
diameter), which we identified as nanodomains (NDs). Interestingly, though these proteins
largely overlapped in the AZ, they were distinct in not only cluster volumes, but number
and size of NDs. Compared to the postsynaptic scaffolding protein PSD-95, RIM1/2 had
similar size and number of NDs, while Munc13 had similar size but greater number, and
Bassoon had larger size but lesser number of NDs.
Previous work has suggested that these proteins play different roles in evoked and
spontaneous release. For instance, while RIM1α knockout impairs only evoked but not
spontaneous release, Munc13 knockout impairs both. Given the distinct distributions of
these proteins, we hypothesized that the evoked and spontaneous vesicle fusion sites will
also differ spatially. To test this, we developed a novel technique to localize single-vesicle
fusion with high spatial resolution in live synapses by adapting single-molecule localization
for single-vesicle fusion signals obtained with vGlut1-pHluorin. We called this approach
“pHluorin uncovering sites of exocytosis” or pHuse. Using pHuse, we compared the pattern
of evoked and spontaneous vesicle fusion at individual presynaptic terminals. This revealed
that evoked fusion occurred over a significantly smaller area of the terminal than
spontaneous fusion. We conclude that differential protein organization in the AZ dictates
nanometer-scale distribution of vesicle fusion mode.
20. The regulation of Drp1-dependent mitochondrial fission during synaptic
potentiation
Sai Sachin Divakaruni1,2,3, Minerva Contreras3, Henry N. Higgs4, Thomas A. Blanpied2,3
1
Medical Scientist Training Program, 2Program in Neuroscience, 3Department of
Physiology, University of Maryland School of Medicine, Baltimore, MD, 4Department of
Biochemistry, Geisel School of Medicine at Dartmouth College, Hanover, NH
Healthy synapse function relies on the adequate presence of mitochondria. The heightened
energy utilization of activity-dependent processes supporting long-term potentiation (LTP)
places even greater a demand on mitochondria and, indeed, mitochondrial dysfunction is
implicated in many neuropsychiatric disorders including autism and schizophrenia.
Although the mechanisms that support dendritic mitochondrial reorganization are unclear, a
key process impacting mitochondrial distribution is mitochondrial fission. Thus, two central
questions that remain are whether LTP induction impacts mitochondrial fission, and
whether mitochondrial fission is required for LTP. Mitochondrial fission is accomplished
via a complex multi-step mechanism, and is regulated by a GTPase, dynamin-related
protein 1 (Drp1), which itself is tightly regulated by phosphorylation. Intriguingly,
CaMK1α and calcineurin, both required for LTP induction, also increase Drp1 recruitment
to mitochondria and mitochondrial fission by regulating Drp1 phosphorylation. Yet, despite
these indications, the link between LTP and mitochondrial fission remains unknown. Driven
by this curiosity, I used a protocol to chemically induce LTP (chemLTP) to test whether
LTP induction increases mitochondrial fission. My preliminary data show a rapid and
transient burst of mitochondrial fission after chemLTP stimulation, in addition to robust and
sustained canonical changes to dendritic spine morphology. Together, these observations
lead me to hypothesize that LTP requires an increase in dendritic mitochondrial fission via
direct regulation of Drp1 function. I will use a diverse arsenal of confocal and superresolution microscopy, molecular perturbations, and biochemistry techniques to determine
the relationship between LTP, Drp1 function, and mitochondrial fission.
21. Egr3 Expression in Ventral Striatal D1-MSNs Controls Behavioral Outcomes to
Social Defeat Stress
T. Chase Francis1, Ramesh Chandra1, Prasad Konkalmatt2, Michel Engeln1, Mary Kay
Lobo1
1
University of Maryland, Baltimore, Dept. Anat. and Neurobio, Baltimore, MD, 2George
Washington University, Washington, DC
The ventral striatum (vSt) is a principle integrator of reward related information within the
brain and is highly implicated in depression. The vSt consists primarily of two projection
neuron subtypes, medium spiny neurons (MSNs), which are differentiated by dopamine
receptor expression, either dopamine 1 receptors (D1) or dopamine 2 receptors. Social
defeat stress (SDS), a well-validated stress paradigm to induce depression-like symptoms,
promotes dichotomous behavioral, electrophysiological, and molecular outcomes in these
MSN subtypes. SDS produces two distinct behavioral phenotypes: mice susceptible to SDS
(i.e., displaying depression-like symptoms) or mice resilient to SDS. Using a cre-dependent
RiboTag method to isolate cell-type specific ribosome-associated mRNA, we discovered
increased Egr3 expression in D1-MSNs from susceptible, but not resilient mice. Injecting a
conditional double inverted open reading frame (DIO) adeno-associated virus (AAV)
expressing Egr3 in the NAc of D1-cre mice promoted susceptibility to subthreshold
(S)SDS. Oppositely, injecting a DIO-AAV expressing an Egr3-miRNA in the vSt of D1-Cre
mice prior to chronic (C)SDS enhanced resilience. Further, CSDS mice injected with a
scrambled miRNA displayed reduced frequency of miniature excitatory post-synaptic
currents (mEPSCs) and enhanced intrinsic excitability in D1-MSNs, an outcome prevented
by Egr3 knockdown. These defeat-induced changes were followed by dramatic alterations
in dendritic morphology and were blocked by Egr3 knockdown. Our results suggest Egr3
manipulation in D1-MSNs is sufficient to control outcomes to SDS. Further, these changes
are correlated to alterations in neuronal morphology of D1-MSNs which may underlie the
behavioral outcomes to SDS.
22. Oscillatory changes in the Hippocampus and Nucleus Accumbens following partial
inverse agonism of α5-GABA receptor.
Natalie Hesselgrave1 and Scott Thompson1
1
Program in Neuroscience, Department of Physiology, and MSTP at University of
Maryland School of Medicine
Depression is a debilitating disease that affects more than 20% of the population and is
associated with poor health and increased risk of suicide. Current first line antidepressants,
selective serotonin reuptake inhibitors (SSRIs), have a 3-8 week latency for clinical
improvement and remission of symptoms is reported in less than 50% of patients. Ketamine
has rapid antidepressant effects but is addictive and induces psychotomimetic
hallucinations. As an NMDA antagonist, ketamine’s effect is likely exerted by relieving
inhibition and promoting neural excitability. We have previously shown that subtype
selective negative allosteric modulators of GABA receptors (L-655,708 and MRK-016) also
strengthen synapses and have antidepressant effects, with minimal anxiolytic side effects, in
rodent models of depression. I hypothesized that L-655,708 and MRK-016 enhance neural
excitability by decreasing inhibitory activity of GABARs, thereby increasing synchronous
activity between the hippocampus and nucleus accumbens, two brain regions implicated in
stress-induced animal models of depression. To elucidate the in vivo electrophysiological
effects of L-655,708 or MRK-016, electrodes were chronically implanted in these two brain
regions in rats. Signals were recorded in awake animals following intraperitoneal
administration vehicle and L-655,708 or MRK-016. Signal analysis suggested increased
synchronous activity in the ventral hippocampus and nucleus accumbens in response to L655,708 but not MRK-016. The increased synchronicity may underlie the anti-depressant
effects of L-655,708 and support the role of GABARs in pathology and treatment of
depression.
23. Modality-specific mechanisms of PKC-induced hypersensitivity of TRPV1
John Joseph
Department of Neural and Pain Science, Dental School, & Program in Neuroscience,
The Center to Advance Chronic Pain Research, University of Maryland, Baltimore, MD
21201
TRPV1 is a non-selective cationic channel that can be activated by polymodal stimuli such
as capsaicin, proton and noxious heat. Under pathophysiological conditions, multiple
inflammatory mediators invoke the activation of multiple kinases that phosphorylate
TRPV1 to enhance functionality of TRPV1. Therefore, phosphorylation-induced
upregulation of TRPV1 functions could constitute specific signals underpinning pathological
nociception, and the residues of TRPV1 involved in the hypersensitivity may be targeted for
antihyperalgesic therapy. The residues involved in phosphorylation-induced hypersensitivity
of TRPV1 have been extensively studied by using the prototypical agonist capsaicin. But it is
rarely understood what residues contribute to hypersensitivity to other natural or
endogenous-stimuli, such as heat, acid or endocannabinoids. Since it has been suggested that
capsaicin, heat, and acid activate TRPV1 through distinct structural bases, it is possible that
differential residues are involved in sensitization to different modalities of agonistic stimuli.
In this study, we investigated common and distinct structural basis of Protein kinase C
(PKC)-induced hypersensitivity to capsaicin, proton and heat by mutagenic ablation of
PKC-associated phosphorylation sites followed by electrophysiological characterization.
TRPV1 S502 and S800 are known to be involved in PKC-induced hypersensitivity to
capsaicin. Unlike capsaicin, PKC-induced hypersensitivity to heat was attenuated by the
mutation of TRPV1 T704 and S800, but not S502. In contrast, PKC-induced
hypersensitivity to acid was attenuated only by the mutation of TRPV1 S800. These results
suggest that the contribution of the PKC-phosphorylation residues to hypersensitivity is
modality-specific and that TRPV1 S800 is a common multi-modality sensitizing site which
can be targeted for anti-hyperalgesic therapy.
24. Effect of nicotinamide mononucleotide on mitochondrial dynamics following
global cerebral ischemia.
Nina Klimova, Aaron Long, Tibor Kristian
Department of Anesthesiology, Center for Shock, Trauma and Anesthesiology Research,
School of Medicine, University of Maryland, Baltimore, Veteran Affairs Maryland Health
Care System
Mitochondria are organelles that undergo a cycle of fusion and fission (mitochondrial
dynamics) that is important for cell physiology. To examine the alterations of mitochondrial
morphology in mechanisms of ischemic brain injury a transgenic mouse model that
expresses a mitochondria targeted fluorescence protein in neurons was utilized. Z-stack
images from CA1 neurons were captured and mitochondrial organelles were reconstructed
in 3D using Volocity software. The data showed increase in mitochondrial fragmentation
(fission) at 2 HR and 24 HR recovery post ischemia. Administration of NMN, a
biochemical precursor of NAD+, in ischemic animals protected mitochondria from
excessive fragmentation and promoted normal mitochondrial morphology in both 2 HR and
24 HR recovery groups. This protective effect could be due to changes in levels and posttranslational modifications in proteins crucial for fusion and fission, DRP1/P-DRP-1,
OPA1, MFN-2, and FIS-1. Maintaining normal mitochondrial morphology is essential for
cell bioenergetics metabolism and survival and mechanisms of its regulation could represent
new therapeutic targets for neurobiological diseases.
25. Epigenetic modulation of BDNF transcription in the valproic acid model of autism.
Melissa Konopko, Allison Densmore, Clinton D. Roby, Bruce K. Krueger
Department of Physiology, Program in Neuroscience, University of Maryland Sch. of Med.,
Baltimore, MD, Department of Pharmacology, University of Maryland Sch. of Med.,
Baltimore, MD
Use of the drug, valproic acid (VPA) by pregnant women increases the risk of autism in
their children. Our lab has found that administration of a dose of VPA to pregnant mice
during the period of early neurogenesis causes a transient increase in Bdnf mRNA and
protein in the mouse fetal brain. Moreover, research has suggested that Bdnf dysregulation
may be involved in the development of autism. The goal of my research is to elucidate the
mechanistic link between VPA and the increased expression of this neurotrophin. VPA is a
histone deacetylase inhibitor. Changes in histone acetylation engage in molecular crosstalk
with other epigenetic marks and can result in alterations in histone and DNA methylation. I
hypothesize that the increased expression of BDNF is mediated by epigenetic mechanisms
including an increase in activating histone modifications and a decrease in inhibitory DNA
methylation. We analyzed several histone modifications using chromatin
immunoprecipitation. This revealed a two to five fold increase in acetylation of various
histone lysines associated with Bdnf promoters. Two histone trimethylation marks were
examined, H3K27me3 (transcriptionally inhibitory) and H3K4me (transcriptionally
stimulatory) were analyzed. H3K27me3 was unaffected by VPA, however, H3K4me3
levels were increased by the drug and showed sex differences in levels. I also analyzed
DNA cytosine methylation, a repressive modification. We found a significant, but modest
impact of VPA on DNA methylation. Together, these experiments help to elucidate the
epigenetic contributions to VPA’s ability to increase Bdnf expression in the brains of
fetuses exposed to the drug in utero.
26. Control of transmembrane protein diffusion within the postsynaptic density
assessed by simultaneous single-molecule tracking and localization microscopy.
Tuo P. Li and Thomas A. Blanpied
Department of Physiology and Program in Neuroscience, University of Maryland School of
Medicine
Postsynaptic transmembrane proteins are critical elements of synapses, mediating transcellular contact, sensitivity to neurotransmitters and other signaling molecules, and flux of
Ca and other ions. Positioning and mobility of each member of this large class of proteins is
critical to their individual function at the synapse. One critical example is that the position
of glutamate receptors within the postsynaptic density (PSD) strongly modulates their
function by aligning or misaligning them with sites of presynaptic vesicle fusion. In
addition, the regulated ability of receptors to move in or out of the synapse is critical for
activity-dependent plasticity. However, factors that control receptor mobility within the
boundaries of the synapse are not well understood. Notably, PSD scaffold molecules
accumulate in domains much smaller than the synapse. Within these nanodomains, the
density of proteins is considerably higher than that of the synapse as a whole, so high that
steric hindrance is expected to reduce receptor mobility substantially. However, while
numerical modeling has demonstrated several features of how the varying protein density
across the face of a single PSD may modulate receptor motion, there is little experimental
information about the extent of this influence. To address this critical aspect of synaptic
organizational dynamics, we performed single-molecule tracking of transmembrane
proteins using uPAINT over PSDs whose internal structure was simultaneously resolved
using PALM. The results provide important experimental confirmation that PSD scaffold
density protein strongly influences the mobility of transmembrane proteins. Tracking a
protein with a cytosolic domain that does not bind PSD-95 still was slowed in regions of
high PSD-95 density, suggesting that crowding by scaffold molecules and perhaps other
proteins is sufficient to stabilize receptors even in the absence of binding. Because
numerous proteins thought to be involved in establishing PSD structure are linked to
disorders including autism and depression, this motivates further exploration of how PSD
nanostructure is created.
27. Altered brain-gut interactions in response to Citrobacter rodentium infection in
traumatic-brain-injured mice.
Ma, Elise L.1; Smith, Allen2; Desai, Neemesh1; Faden, Alan1; Shea- Donohue, Terez1
1
University of Maryland School of Medicine, Baltimore, MD, United States. 2United States
Department of Agriculture (USDA), Beltsville, MD, United States.
Introduction: Traumatic brain injury (TBI) induces persistent changes in cellular and
biochemical processes leading to chronic inflammation, white matter degeneration and
astrocyte activation in the brain. Secondary consequences of TBI extend to the systemic
circulation and peripheral organs, including the GI tract where enteric glial cells (EGC) play
a role in mucosal barrier function. Although human and animal studies have established a
link between TBI and intestinal dysfunction during acute stages after injury, long-term
consequences of TBI in the gut are unknown.
Aims: To investigate 1) TBI-induced changes in mucosal barrier function and glial cell
reactivity in the gut and brain, and 2) the effect of Citrobacter rodentium (Cr), an enteric
murine pathogen similar to human Escherichia coli, on these parameters.
Methods: Moderate TBI was induced by controlled cortical impact (CCI). Sham and injured
mice were infected with Cr 28 days post-CCI. Colonic paracellular permeability was
determined by labeled 3kD dextran flux. Brain astrocytic and colonic EGC reactivity was
measured by quantification of glial fibrillary acidic protein (GFAP) expression. Brain lesion
volume was quantified by stereological techniques.
Results & Conclusions: The association of chronic brain injury with impaired mucosal
barrier function and increased numbers of GFAP+ cells in both the colon and at the brain
lesion site indicates that CCI induces activation of glial cells to mediate tissue repair after
brain and gut injury. Further increases in colonic permeability in CCI-injured mice in
response to Cr infection was associated with significantly greater brain lesion volumes and
reduced numbers of GFAP+ cells suggesting that enteric pathogen-induced inflammation
impairs resolution of brain injury.
28. The role of synaptic nanostructure in regulating NMDA receptor activation.
Sarah Ransom Metzbower1, Sridhar Raghavachari2, and Thomas A. Blanpied1
1
Department of Physiology and Program in Neuroscience, University of Maryland School
of Medicine, Baltimore, Maryland 21201, USA,. 2Deptartment of Neurobiology, Duke
University Medical Center, Durham, NC, USA.
Within a single synapse, many factors influence NMDA receptor (NMDAR) activation,
including the number and subtype of receptors, their subsynaptic distribution, and their
relationship to sites of vesicle fusion. To examine regulation of NMDAR activation in
single synapses, we utilized spinning disk confocal imaging of cultured hippocampal
neurons expressing GCaMP6f. In the presence of TTX, 0 Mg2+, and 2 mM Ca2+, this
revealed miniature spontaneous NMDAR activation in individual dendritic spines.
It has been suggested that tens of NMDARs are present at a single synapse. However, a low
concentration of the high affinity antagonist CPP decreased response frequency with
minimal changes in amplitude. This suggests that very few NMDARs open following
spontaneous neurotransmitter release. The low maximal open probability of NMDARs
likely contributes to this phenomenon, but, in addition, subsynaptic position of NMDARs
away from glutamate release sites may minimize their activation. Previously, PALM has
revealed nanodomains within the PSD that contain a relatively high density of the NMDAR
subunit GluN2B. Simulating release over the model PSD revealed that the open probability
of GluN2B containing NMDARs sharply decreased as a function of distance from the site
of release (~50% in 120 nm). To study the relationship between PSD nanoscale
organization and NMDAR activation, we pioneered the use of PALM to map the
subsynaptic distribution of PSD-95 in spines imaged with GCaMP6f, thus enabling us to
measure both synaptic function and nanostructure at individual synapses. This revealed a
positive correlation between NMDAR activation at single spines and the fractional area of
the PSD that was incorporated into a nanodomain. Additionally, synapses with
nanodomains showed a wider range of average peak ΔF/F than those without. Together,
these data suggest that PSD nanostructure is a novel mechanism for regulation of NMDAR
activation.
]
29. Metaplastic control of non-canonical mesolimbic synaptic transmission
Mary H. Patton1, Bradley M. Roberts1, Titilola Akintola2, Brian N. Mathur1
1
Dept. of Pharmacology, University of Maryland School of Medicine, Baltimore, MD
21202, 2Dept. of Anatomy and Neurobiology, University of Maryland School of Medicine,
Baltimore, MD 21202
The nucleus accumbens (NAc) receives reward-encoding dopaminergic inputs from the
ventral tegmental area (VTA), which non-canonically co-release GABA onto NAc medium
spiny projection neurons (MSNs). The functional role of this inhibitory input onto MSNs
and whether it is modulated by drugs of abuse remains unexplored. Using whole-cell patchclamp electrophysiology and optogenetics in acute mouse brain slices, we test the
hypothesis that VTA-MSN inhibitory synapses undergo synaptic depression to disinhibit
NAc MSNs. We discovered a novel form of postsynaptically-expressed inhibitory longterm depression (iLTD) at these synapses that is dependent upon activation of TrkB, the
receptor for brain-derived growth factor (BDNF). We find that ethanol exerts metaplastic
control of NAc output by augmenting iLTD magnitude and that iLTD is occluded following
a single in vivo ethanol challenge. These findings are the first to demonstrate plasticity at
GABAergic VTA-MSN synapses and suggest a role for VTA-GABA release in mediating
reward and the hedonic response to alcohol.
30. The PARK10 gene USP24 is a negative regulator of autophagy.
Julia Peter1, Nivedita Hegdekar1, Ola Awad2, Chinmoy Sarkar1, Ricardo Feldman2 and
Marta M. Lipinski1
1
Department of Anesthesiology, University of Maryland Baltimore, 2Department of
Microbiology and Immunology, University of Maryland Baltimore
Autophagy plays an essential neuroprotective role during brain aging and its molecular
defects are linked to neurodegenerative diseases, including Parkinson’s disease (PD). Our
goal is to characterize autophagy dysregulation in all types of PD and identify targets for its
modulation. In a high-throughput screening for genes regulating mammalian autophagy,
Ubiquitin specific peptidase 24 (USP24) was identified as a negative regulator of
autophagy. USP24, located on chromosome 1 in the PARK10 locus, is associated with lateonset PD. Non-synonymous single nucleotide polymorphisms in the coding region of
USP24 affect predisposition to PD. We confirmed increased USP24 protein levels in the
substantia nigra of a subpopulation of non-familial idiopathic PD patients. In human cell
lines and human iPS cell derived neurons USP24 knock-down led to up-regulation in levels
of cellular autophagy, assessed by translocation of the GFP-LC3 autophagy reporter and by
increased autophagosome associated LC3-II. Knock-down of USP24 caused accumulation
of PtdIns3P (type III PI3 kinase product), shown by quantification of the FYVE-dsRed
reporter. Inducing autophagy by loss of USP24 function was attenuated in the presence of
type III PI3 kinase inhibitors. Lack of change in the phosphorylation of two mTORC1
targets after knock-down of USP24 indicate that USP24 acts independent of mTOR.
Induction of autophagy following USP24 knock-down did not cause loss of cell viability; it
lead to decreased accumulation of PD mutant α-synuclein A53T and enhanced long-term
survival of iPS cell derived dopaminergic neurons. Together, our data highlight the
importance of USP24 and its potential role in autophagy modulation in PD.
31. Role of CREB, SRF and MEF2 in ocular dominance plasticity.
Nisha S. Pulimood, Alexandre E. Medina
Program in Neuroscience, Department of Pediatrics, University of Maryland School of
Medicine
The transcription factors CREB (cAMP Response Element Binding factor), SRF (Serum
Response Factor) and MEF2 (Myocyte Enhancer Factor 2) play critical roles in neuronal
plasticity, and have been shown to differentially affect underlying plasticity mechanisms
like LTD and LTP. Additionally, a recent study showed that CREB, SRF and MEF2 control
the expression of the early gene Arc, which is required for the expression of ocular
dominance plasticity (ODP), by binding to the synaptic activity responsive element
(SARE). However the role of the activation of these transcription factors in the different
components of plasticity in vivo is not well known. We used Visually Evoked Potentials
(VEPs) in awake mice to investigate the role of CREB, SRF and MEF2 on the depression
and potentiation components of ODP (dc-ODP and pc-ODP). We infected animals with a
Herpes Simplex viral vector expressing dominant negative forms of CREB, SRF or MEF2,
then chronically implanted electrodes in the binocular zone of the mouse visual cortex. We
then recorded VEPs before and after a period of monocular deprivation (MD). Since the two
components of ODP express in a temporally distinct manner in the mouse visual cortex, we
used 3 days of MD to isolate dc-ODP and 7 days of MD to investigate pc-ODP. Our results
show that these three transcription factors have different effects on dc-ODP and pc-ODP.
These findings can elucidate the role of these key players in the two components of ODP,
thereby informing the search for therapeutic targets in diseases where ODP is defective.
32. Sex differences in the frontal lobe of the developing mouse brain
Da Shi1, Jiachen Zhuo1, Su Xu1, Jaylyn Waddell2, Rao P. Gullapalli1,3
1
Department of Diagnostic Radiology, 2Department of Pediatrics, 3Program in
Neuroscience, University of Maryland School of Medicine, Baltimore, Maryland
The frontal lobe controls many important brain functions and matures during postnatal life
in both humans and rodents, though the development of the frontal lobe is different between
males and females. The connections in the developing mouse frontal lobe was represented
with structural connectivity between regions of the frontal lobe to other cortical, gray and
white matter regions at postnatal day 20 (P20) and P30, measured using diffusion tensor
imaging with seed region in the frontal association cortex. Using probabilistic tractography,
our preliminary data revealed greater connectivity in the female cingulate to frontal
association cortex at P20 compared to the same regions in the male brain. Instead, greater
connectivity from the cingulate to the prelimbic cortex at P20 were found in the male brain
compared to the female brain. Sex differences were not observed at P30 between male and
female brain. The structural connectivity in white matter regions including the corups
callosum and anterior commissure did not vary between male and female at both ages. The
connectivity from the frontal lobe to posterior brain regions showed lower connectivities at
P20 and P30. The findings at P20 suggest that specific connections in the developing
structural brain network can vary depending on sex but these developmental sex differences
disappear with age.
33. Early predator odor exposure exerts sexually dimorphic effects on juvenile social
play behavior.
Sara L. Stockman2,3, Margaret M. McCarthy Ph.D.1
1
Deptartment of Pharmacology, 2Program in Neuroscience and 3Medical Scientist Training
Program, University of Maryland School of Medicine, Baltimore, MD
Juvenile social play behavior is one of the earliest sexually differentiated behaviors to
emerge through development. In rats, male animals engage in more rough-and-tumble play
compared to females. Participation in social play during the juvenile period is critical for
appropriate development. Exposure to early life adversity is a major driver of later health
and behaviors. Research suggests that adverse early life event exposure can have differing
effects on males and females, however, how such exposures alter social play behavior in the
juvenile period is poorly understood. To address this question, male and female neonatal
rats were exposed to predator odor (PO) for 5 min on PN1-PN3. When tested on an
additional fourth day, in the presence of PO, the rat pups were found to reduce both
ultrasonic vocalization (USV) call frequency and amplitude of emitted calls. Additionally,
video recordings over the three day exposure indicated that PO exposed pups display
elevated freezing behavior. This suggests neonatal rat pups modulate their behavior in
response to fearful stimuli. PO elevated corticosterone in male pups only, however, no
effects on dentate gyrus proliferation were found. Following exposure, animals were
normally raised to PN26 and social play and anxiety-like behaviors were tested. Social play
was evaluated for the frequency of pouncing, pinning, chasing and boxing behaviors, as
well as, the total time engaged in play and number of play events. Under the play paradigm
utilized, early predator odor produced sexually dimorphic alterations of play, whereby
males exposed to PO exhibited a decrease in all parameters measured and females exposed
to PO increased play behavior across all measures. PO exposure did not alter anxiety-like
behavior assessed in the open field test, the elevated plus maze or the light/dark box during
the juvenile period or at adulthood or social interaction of adulthood. Knowing that social
behaviors are disrupted in several neuropsychiatric disorders, this work provides some
insight into how sex may interact with adverse early life event exposure to contribute to the
etiology of these disease manifestations.
Supported by R01NS050525 to MMM
34. Role of Microglia in Sexual Differentiation of the Amygdala
Jonathan W. VanRyzin BS1, Kathryn J. Argue Ph.D.2, and Margaret M. McCarthy
Ph.D.1,2
Program in Neuroscience1 and Department of Pharmacology2, University of Maryland,
Baltimore, School of Medicine, Baltimore, MD, USA.
Microglia are the dominant resident immune cells of the brain and function in multiple ways
outside their traditional capacity of responding to insult. During development microglia
regulate tissue homeostasis, neuronal precursor populations, and synaptic circuitry. We
implicated microglia as an integral component of sexual differentiation of the preoptic area
and control of male copulatory behavior, suggesting these immune cells can also organize
sex-specific brain structure and function (Lenz et al. J Neurosci 33(7), 2013). The amygdala
is also a sexually dimorphic brain region that regulates social behaviors known to differ in
males and females. We reported a sex difference in the number of newly born cells in the
developing rat amygdala mediated by endocannabinoids, with females having higher
numbers of newly born cells than males and a lower endocannabinoid tone. The differences
in newly born cells correlated to behavioral changes, as treating newborn females with the
CB1/2 agonist WIN55,212-2 masculinized juvenile social play behavior and reduced cell
genesis in the amygdala (Krebs-Kraft et al. PNAS 107(47), 2010). Further investigation
suggests microglia may be central to establishing the observed sex difference in newly born
cells, as males have more Iba1+ microglia exhibiting phagocytic morphology compared to
females. We are now deciphering the relative contributions of microglia phagocytosis and
trophic signaling in the regulation of newly born cells. Ultimately, these studies will provide
valuable insight into new facets of immune regulation of brain sexual differentiation and the
development amygdala-dependent circuitry.
Funding: This work was supported by RO1 MH52716-018 to MMM.
35. The claustrum mediates cross-cortical communication for top-down attention.
Michael G. White, Matthew Panicker, Bradley M. Roberts, Brian N. Mathur
University of Maryland, School of Medicine, Department of Pharmacology, Baltimore, MD
Top-down (i.e. volitional) attention is critical for successfully interacting with our dynamic
and overwhelming sensory world. To exert top-down attention to visual stimuli,
executive/frontal cortices, such as the anterior cingulate cortex (ACC), modulate posterior
visual cortices to enhance processing of attended visual stimuli. The neural circuits that
mediate this frontal (top) to posterior (bottom) cortical modulation are poorly understood.
The claustrum possesses widespread cortical connectivity and is therefore positioned to
enable communication from frontal cortices to distal posterior cortical sites. However, the
role of the claustrum in top-down attention, or any cognitive process, is untested. We report
that the mouse claustrum receives a dense ACC  claustrum input that is 1) filtered by both
feedforward and feedback inhibitory microcircuits, and 2) propagated to posterior sensory
and association cortical areas involved in visual attention. Optogenetic manipulation of this
circuitry in vivo disrupts attentional performance on a five-choice serial reaction time task.
These results provide evidence for a novel circuit subserving volitional attention and have
important implications for understanding attentional impairments shared amongst several
neuropsychiatric disorders.
36. Thalamic intralaminar nuclei control of action initiation.
Utsav Gyawali, Mary H. Patton, Bradley M. Roberts, Michael G. White and Brian N.
Mathur
Department of Pharmacology, University of Maryland, Baltimore, MD 21201 USA
Successful reward acquisition in a dynamic environment requires attention to salient
sensory stimuli that instruct striatally-encoded actions. While striatally-projecting
intralaminar nuclei of the thalamus encode salient information, how the different thalamic
intralaminar nuclei govern striatal circuits to influence actions is poorly understood. Using
optogenetics, electrophysiology and fast-scan cyclic voltammetry in mouse brain slices we
find that individual intralaminar nuclei uniquely wire into striatal microcircuitry to axoaxonically control striatal dopamine release. Chemogenetic suppression of a thalamostriatal
pathway found to monosynaptically elicit dopamine release reveals a selective impairment
in action sequence initiation, but not performance once the action is initiated. Leveraging
this novel action initiation circuit, we find that optogenetic drive of this pathway restores
ambulatory behavior and action initiation ability in parkinsonian mice. These results shed
new light on the role of the thalamus in attentional command over action initiation and
suggest thalamostriatal circuits as a potential node of intervention in striatal dopaminerelated disorders such as Parkinson’s disease.
37. Prediction error-based model for estimating pain-related placebo effects and their
extinction rates.
Taylor Ludman1, Luana Colloca 1,2, Evan Livesey3, and Ben Colagiuri 3
1
University of Maryland School of Nursing, Baltimore, USA, 2University of Maryland
School of Medicine, Baltimore, USA, 3University of Sydney School of Psychology,
Sydney, Australia
The placebo effect (PE) has many beneficial applications, including placebo analgesia after
the induction of positive expectancies. Importantly not everyone experiences placebo
analgesia at the same magnitude; in fact, some individuals do not respond to placebos at all.
The extinction rate of the PE depends on many factors, such as how placebo analgesia was
induced (e.g. through conditioning, verbal suggestion, and social learning), expectancy, and
pain perception. To our knowledge a model has not been created that can predict how an
individual will respond to a placebo analgesic procedure and how long lasting these effects
will be. Therefore, the aims for this study were to: 1) predict pain-related placebo responses,
2) account for extinction rates, and 3) determine whether an individual will be a placebo
responder or non-responder. A prediction error-based model was developed and then
applied to the results of two separate human experiments. In both of these experiments, pain
expectancy and intensity was rated before and after, respectively, the delivery of noxious
electrical stimulations during a placebo manipulation procedure. The model has thus far
been proven effective for determining extinction rates and pain-related placebo responses
when positive expectancies are considered. Moreover, the model has successfully accounted
for the differences (for both PE and extinction rate) in the two experiments by estimating
the bias introduced by expectancy on pain perception. As for individual variations, findings
suggest that subject-by-subject differences in placebo responses can be successfully
established by assessing learning rates and pain expectancy biases.
38. Striatal fast-spiking interneurons synchronously encode action velocity dynamics.
Bradley M. Roberts1, Michael G. White1, Rong Chen2 and Brian N. Mathur1
1
Dept. of Pharmacology, 2Dept. of Diagnostic Radiology and Nuclear Medicine, University
of Maryland School of Medicine, Baltimore, Maryland 21202
The dorsal striatum is necessary for action learning and selection. Representing less than
5% of all striatal neurons, the GABAergic fast-spiking interneuron (FSI) population
powerfully regulates striatal output by perisomatically synapsing on striatal output neurons
and theoretically synchronizing with each other via gap junctions. Despite data implicating
FSIs in disorders such as Parkinson’s disease, Tourette syndrome and addiction, the
fundamental role of FSIs in encoding actions remains unsolved. Utilizing cell-type-specific
in vivo microendoscopic calcium imaging of FSIs in the dorsal striatum we provide the first
demonstration that the activity of parvalbumin-expressing FSIs is synchronized, supporting
theoretical network models. During free ambulatory behavior in an open field, we show that
synchronized FSI activity positively correlates with action velocity. Moreover, application
of a machine learning algorithm to FSI calcium imaging data significantly predicts action
velocity dynamics. Assessing behavior microstructure, we find that FSIs synchronously fire
immediately prior to and, to some extent, following action execution. Due to the their
powerful feedforward inhibitory influence in the dorsal striatum, these data suggest that
FSIs may be critical for suppressing the initiation of unwanted actions, while optimally
constraining those that are selected.

PIN 2016 Retreat Program - Graduate Program In Life Sciences

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