Welcome to the Thirty-Fourth Annual Winter Conference on Brain Research

The Winter Conference on Brain Research (WCBR) was founded in 1968 to promote free exchange of information and ideas within . It was the intent of the founders that both formal and informal interactions would occur between clinical and laboratory based . During the past thirty years neuroscience has grown and expanded to include many new fields and methodologies. This diversity is also reflected by WCBR participants and in our program. A primary goal of the WCBR is to enable participants to learn about the current status of areas of neuroscience other than their own. Another objective is to provide a vehicle for scientists with common interests to discuss current issues in an informal setting. On the other hand, WCBR is not designed for presentations limited to communicating the latest data to a small group of specialists; this is best done at national society meetings.

The program includes panels (reviews for an audience not necessarily familiar with the area presented), workshops (informal discussions of current issues and data), and a number of posters. The annual confer- ence lecture will be presented at breakfast on Sunday, January 21. Our guest speaker will be Dr. Gerald Fischbach, Director of the National Institute for Neurological Disorders and Stroke at the National Institutes of Health. On Tuesday, January 23, a town meeting will be held for the Steamboat Springs community. Here Dr. Charles O Brien, and WCBR participants will discuss the neurobiological basis of mental illness. Also, participants in the WCBR outreach program will present sessions at local schools throughout the week to pique stu- dents interest in science. Finally, the banquet, including music and dancing, will be held on Friday evening. Generous donations from sponsors have permitted us to continue the WCBR Fellowship Award Program. These awards are given to young neuroscientists who are on the program and who are newcomers to WCBR. Congratulations and a warm welcome to this year s fellows: Bill Carlezon, David Chambers, Gersham Dent, Cecilia Flores,

1 Timothy Gomez, Ryder Gwinn, Andrew Jenkins, Gregory Kinney, Oliver Manzoni, Haley Melikian, Claudio Mello, David W. Miller, Derek Molliver, Cheryl L. Stucky, and Kevin Wickman. Please plan to attend the business meeting at 6:30 p.m. on Wednesday, January 24. We will elect a Facilities Chair-Elect and three members of the Board of Directors. A sign-up sheet will be available on the message board to indicate your interest in running for a Board of Directors position. Other important matters will be discussed including proposed changes in the by-laws and the selection of future conference sites.

Don’t forget to visit the exhibit area.

2 Conference Chair Allan Basbaum Michael S. Levine, Chair-Elect Program Committee Paul C. Letourneau, Chair Elizabeth Abercrombie, Christine Konradi Chair-Elect Michael S. Levine Allan Basbaum Joseph F. Lipinski, Jr. Jill B. Becker Wendy B. Macklin Marc D. Binder David J. Perkel Marie-Francoise Chesselet Bruce R. Ransom Thomas V. Dunwiddie Suzanne K. Roffler-Tarlov Neil L. Harrison Oswald Steward Karl Herrup James Surmeier Katherine Kalil Thomas H. Swanson Paul S. Katz Treasurer Elizabeth Abercrombie Facilities Committee Karen Greif, Chair Monte Westerfield, Chair-Elect Murray Blackmore Stanislav (Stas) Kholmanskikh Hsin Chu Simona Neumann Bill Cobb Karla Petersen-Zeitz Scott Gehler James Zackheim Board of Directors Marjorie Ariano Kristen Keefe Martha Bohn Joel E. Kleinman William E. Bunney Bruce Ransom Karen Gale James Surmeier Bart Hoebel Claude Westerlain Thomas M. Hyde Donald Woodward Fellowship Program James Surmeier, Chair Ed Hall Ray Bartus Phil Skolnik

3 Exhibits Michael Levine Wendy B. Macklin School Outreach Kim Topp, Coordinator Joseph LaManna Victor Denenberg Hugh McIntyre Paula Dore-Duffy Marsha Melnick Helene Emsellem Gene Palmer Janet Finlay Bruce Ransom Kyle Frantz Chris Ransom Karen Greif Suzanne Roffler-Tarlov A. Chistina Grobin Brad Stokes Samil Harik Tom Swanson Luc Jasmin Frank Welsh Town Meeting Janet Finlay Laura Peoples Conference Arrangements Scott C. Miller, Program Director Nancy Mulvany, Program Secretary Conferences and Institutes University of Illinois at Urbana-Champaign Suite 202 University Centre 302 East John Street Champaign, IL 61820 2001 Fellowship Awardees Bill Carlezon Oliver Manzoni David Chambers Haley Melikian Gersham Dent Claudio Mello Cecilia Flores David W. Miller Timothy Gomez Derek Molliver Ryder Gwinn Cheryl L. Stucky Andrew Jenkins Kevin Wickman Gregory Kinney

4 Winter Conference on Brain Research Fellowship Sponsors Benefactors: $1000+ Aventis Dystonia Foundation Eli Lilly and Co. Fine Science Tools Janssen Pharmaceuticals Pfizer, Inc. WCBR Board of Directors Patron: $500-$999 Alkermes, Inc. Dov Pharmaceuticals Dupont Parmaceuticals Pharmacia UpJohn Schering Plough Research Institute Wyeth-Ayerst Pharmaceuticals Exhibitors Academic Press 525 B Street, Suite 1900 San Diego, CA 92101-4495 Contact: Sue Boyer / Tari Paschall Tel (619) 699-6820 Email: [email protected] Association Book Exhibit 639 South Washington St. Alexandria, CA 22314 Contact: Mark Trocchi Tel (703) 519-3909 Fax (703) 519-7732 Email: [email protected] Carl Zeiss One Zeiss Drive Thornwood, NY 10594 Contact: Troy Tholen Tel (801) 484-8872 Voice mail (800) 543-1033 x7981 Email: [email protected]

5 Durect Corporation 10240 Bubb Road Cupertino, CA 95014 Contact: Stacey Matsuoka / Clarisa Peer Tel (408) 367-4038 Fax (408) 865-1406 Email: [email protected] Elsevier Science London 84 Theobalds Road London WC1X 8RR, ENGLAND Contact: Emily Miles Tel 44-020-761-4378 Fax 44-020-761-4401 Email: [email protected] Elsevier Science London The Online Community of Neuroscientists Middlesex House 34-42 Cleveland Street London W1P 6LE, ENGLAND Contact: Samar Syed Tel 44-207- 611-4362 Fax 44-207-611-4201 Email: [email protected]

ESA Inc. 22 Alpha Road Chelmsford, MA 01824-4171 Contact: Brent Morrison / Michael Baker / Damian Freeze Tel (978) 250-7081 Fax (978) 250-7087 Email: [email protected]

Fine Science Tools 373-G Vintage Park Drive Foster City, CA 94404 Contact: Daron Brown / Jeff Wiley Tel (800) 521-2109 Fax (800) 523-2109 Email: [email protected] / [email protected]

Instrutech Corporation 20 Vanderventer Ave., Ste 101E Port Washington, NY 11050-3752 Contact: Milan Kesler Tel (516) 883-1300 Fax (516) 883-1558 Email: [email protected]

6 S. Karger AG Allschwilerstrasse 10 Postfach 4009 Basel, SWITZERLAND Contact: Angela Gasser Tel 41-61-306-1264 Email: [email protected] Leica Microsystems Inc. Harbor Tech Center 724 Anthony Trail Northbrook, IL 60062 Contact: Wayne A. Buttermore / Andy Lee Email: [email protected] MED Associates Inc. PO Box 319 St. Albans, VT 05478 Contact: Karl Zurn Tel (802) 527-2343 Fax (802) 527-5095 Email: [email protected] Mini-Mitter Co., Inc. PO Box 3386 Sunriver, OR 97707 Contact: David or Judy Osgood Tel (541) 593-8639 Fax (541) 593-5604 Email: [email protected] The MIT Press NE25-4093 5 Cambridge Center Cambridge, MA 02142 Contact: John Costello Tel (617) 258-5764 Fax (617) 253-1709 Email: [email protected]

Molecular and Cellular Research Branch Division of Neuroscience and Basic Behavioral Science National Institute of Mental Health 6001 Executive Blvd., Room 7168 Bethesda, MD 20892-9641 Contact: Beth-Anne Sieber Tel (301) 443-5288 Fax (301) 402-4740 Email: [email protected]

7 Natural Research Council/National Academy of Science 2101 Constitution Ave NW (TJ2114) Washington, DC 20418 Contact: Jane Dell Amore Tel (202)334-2768 Fax (202) 334-2759 Email: [email protected] Nikon Inc. PO Box 2464 Evergreen, CO 80437 Contact: Gerald Benham Colorado Office Ph.&Fax (303) 674-1569 NY Tel: (516)547-8596 Fax (516) 547-4033 Email: [email protected] Olympus America Two Corporate Center Drive Melville, NY 11747 Contact: Angela Goodacre Tel (800) 645-8100 Fax (972) 625-6439 Email: [email protected] Contact: Kathleen Karmel Tel (800) 645-8100 Box 6462 Fax (972) 625-6439 Email: [email protected] Oxford University Press 198 Madison Avenue New York, NY 10016 Contact: Lanvi Tran / Fiona Stevens Tel (212) 726-6079 Fax (212) 726-6441 Email: [email protected] Pacer Scientific 5649 Valley Oak Drive Los Angeles, CA 90068 Contact: Richard and Kathy Bellamy Tel (323) 462-0636 Fax (323) 462-1430 Email: [email protected]

8 General Information

Headquarters is the Sheraton Steamboat Resort and Conference Center. All scientific activities will be held there.

WCBR Information Desk and Message Center are in Sunset A just to the right off the Main Lobby when entering from the front entrance. The desk hours are as follows: Morning Afternoon Evening Saturday 1/20 9:00—11:00 AM 3:30—5:30 PM 7:30—10:00 PM Sunday 1/21 7:00—8:00 AM 3:30—6:30 PM Monday 1/22— 7:00—8:00 AM 3:30—4:30 PM. Friday 1/26 The telephone number for messages is 970-879-2220, ext. 1112. The Sheraton Steamboat Resort and Conference Center fax number is 970- 870-9278. The person sending or receiving faxes is responsible for all charges.

Each registration packet contains: a conference badge; registration receipt; tickets for breakfasts, mid-week lunch, and banquet; program book. The packets should be picked up at the WCBR Information Desk. PLEASE NOTE that your housing reservation must be shown before these items can be issued. Conferees who did not accept WCBR-assigned accommodations are charged a facilities supplement of $100 as stated in the WCBR announcement. No exceptions can be granted. Attendance at this conference is strictly limited to PRE- REGISTERED participants. On-site registration is not available.

Posters will be available for viewing throughout the week in the Sunshine Peak. Poster presenters will be by their posters for discussion from 3:30—4:30 PM on the first day of each of the three poster ses- sions: Poster Session 1 Sunda y/Monday; Poster Session 2 T ues- day/Wednesday; and Poster Session 3 T hursday/Friday. Presenters may put up their posters after 9:30 AM on the day their session starts.

9 Presenters should take down their posters by 10 PM on the second day of their session. Please see Poster Sessions section in program for titles and names of presenters.

Exhibits and Lounge are in the Sunshine Peak. Coffee is available from 7:30—10:30AM Monday through Friday. Refreshments are provided by the exhibitors from 3:30—4:30 PM, Sunday through Friday.

Breakfast is served to all registrants on Sunday 7:30—8:20AM, in the Grand Ballroom, and on Monday through Friday, 6:30—7:30AM, in both Buddy s Run and Sevens Restaurant. (Social guests may breakfast until 10:00 AM in Sevens). The tickets in your registration packet are required for admission to all meals. On Saturday morning (January 26) before departure continental breakfast is available in the foyer of the Sheraton Steamboat Resort and Conference Center.

Ski Lift Tickets will be available from the WCBR Information Desk in Sunset A. Daily tickets can be purchased or prepaid tickets can be picked up during desk hours.

Don’t forget to visit the exhibit area.

10 Special Events Saturday, January 20: Welcome Wine and Cheese Party ¥ 7:30—10:30 PM, Grand Ballroom. Sunday, January 21: Conference Breakfast and Opening Address ¥ 7:30 AM, Grand Ballroom (Your required ticket is in your registration packet). Our plenary keynote speaker will be Gerald Fischbach, Director of the National Institute for Neurobiological Disorders and Stroke at NIH. The title of his plenary lecture is Neur oscience at the New Millenium. Meeting of Panel and Workshop Organizers ¥ 9:30—10:30AM, Grand Ballroom. The meeting will be brief but important. Organizers and WCBR staff please attend. Monday, January 22: First Meeting of Board of Directors ¥ 6:30—8:30AM, Ante Board Room. Tuesday, January 23: Town Meeting ¥ 8:30—10:30 PM Wednesday, January 24: Smitty Stevens Memorial (NASTAR) Ski Race ¥ 1:30—3:00 PM. NASTAR registration cards to be completed no later than Monday, January 22, 8:00 AM at WCBR Information Desk. Mountain Lunch ¥ 11:00—1:00, Bear River Bar and Grill at base of gondola. (Required lunch ticket is in your registration packet). Non-skiers, requiring transportation, should sign up at the WCBR Information Desk. Business Meeting ¥ 6:30 PM, Mt. Werner, Election of Facilities Chair-Elect and three Members of the Board of Directors. Friday, January 26: Second Meeting Board of Directors ¥ 6:30AM, Ante Board Room. Banquet and Dance ¥ 8:00 PM, Grand Ballroom. (Required ticket is in your registration packet). Cash bar opens at 7:00 PM in the foyer of the Sheraton Steamboat Resort and Conference Center.

11 Preamble to the Program

The 2001 WCBR Program consists of panels, workshops, and posters. Please consult the Pro- gram Booklet and posted announcements for details regarding the scientific presentations, as well as information regarding the school outreach program and the town meeting.

12 Sunday, January 21 7:30 AM PANEL ¥ The Biological Clock: Of Molecular Gears, Timing Belts BREAKFAST ADDRESS and Post-transcriptional Modifi- Guest Speaker: Gerald Fischbach cations Grand Ballroom R. Baler, D. Virshup, P. Lowrey, A. Keesler 3:30–4:30 PM Skyline POSTER SESSION 1 ¥ Presenters PANEL ¥ Evolution of Neurohypo- available for discussion physeal Peptide Systems: Why Sex Matters Sunshine T.R. Insel, M.S. Grober, F.L. Moore, L.J. Young, G.J. De Vries 4:30–6:30 PM Sunset PANEL ¥ Neurotrophic Factor- Induced Dopaminergic Plasticity in Animal Models of Parkinson s 8:30–10:00 PM Disease and Drug WORKSHOP ¥ Role of Noradrener- R.C. Pierce, K.B Seroogy, C.A. gic NTS in Drug Abuse: Altar, J.R. Taylor, C. Flores The Forgotten System Mt. Werner G. Aston-Jones, T. De Vries, Y. Shaham, G. Koob, C. O Brien PANEL ¥ Cytoskeletal Restructuring in the Formation of Dendrites Mt. Werner and PANEL ¥ Dopaminergic Projections R.H. Lenox, S. Halpain, D. and Neural Ensembles in Benson, A. Matus Prefrontal Cortical-Ventral Striatal Loops Storm Peak P. O’Donnell, A.A. Grace, L.L. PANEL ¥The Orbitofrontal Cortex Peoples, W. Schultz and the : Neuroimaging Studies Storm Peak B. Adinoff , L. Porrino, R. PANEL ¥ Glial Involvement in Anton, E. London Neurodegeneration: The Revenge of Unintended Consequences Twilight S. Griffin, S. Barger, L. Van PANEL ¥ Rhythms and Patterns in Eldik, J. Schwartz Neural Firing Twilight C.C. Chow, A. Bose, D. Terman, J. Rubin Rainbow

13 Sunday, January 21, continued

PANEL ¥ Positive Modulation of PANEL ¥ Success Comes from AMPA Receptors: Mechanisms Having the Right Connections: and Therapeutics Integrating the CNS and PNS E. Nisenbaum, D. Bleakman, K. D. Chambers, E. Bell, J. Begbie Partin, U. Staubli Sunset Rainbow PANEL ¥ New Teeth on an Old Saw: Evaluating Cholinergic Mecha- nisms in Brain Aging D. Ingram, N. Greig, R. Perez- Polo, J. Joseph Skyline

Monday, January 22 7:30–9:30 AM PANEL ¥The HPA Axis: A Para- digm for Both Short and Long PANEL ¥ Integrated Regulation of Term Plasticity in CNS Respon- ++ Synaptic Transmission by Ca siveness Channels, K+ Channels, and , J. Herman, S.L. Mitochondrial Ca++ Uptake G. Aguilera Lightman, J. A. Cidlowski W. A. Catterall, I. Forsythe, L. Rainbow Kaczmarek, T. Scheuer, D. Lipscombe PANEL ¥ Immune Regulation, CNS Pathogenesis and Therapeutic Mt. Werner Interventions in Multiple PANEL ¥ Group I Metabotropic Sclerosis Glutamate Receptors: Role in , G. Konat, P. Dore- Pain, Epilepsy, Movement R. Jones Duffy, H. Offner, A. Vandenbark Disorders, and Brain Injury. Skyline R. Schwarz, J. Conn, A. Faden, A. Kingston, C. Woolf PANEL ¥ The Raphe and Breathing: What Is the Connection? Storm Peak , E. Nattie, L. PANEL ¥ Exactly How Is G. Richerson Kubin, A. Berger, J. Ramirez Involved in Methamphetamine Neurotoxicity? Sunset T. Hastings, D. Sulzer, R. Gainetdinov, G. Ricaurte, Twilight

14 3:30–4:30 PM PANEL ¥The Bane of Cancer Pain: A Role for Peptides and Protons? POSTER SESSION 1 ¥ Posters , A. Beitz, G. Davar, P. available for viewing only G. Wilcox Reeh Sunshine Sunset 4:30–6:30 PM 8:30–10:00 PM PANEL ¥ New Vistas on Prefrontal WORKSHOP ¥ Gene Profiling in the Cortex: From Neuroanatomy to Using Nucleic Computational Modeling Acid Microarrays M. Banich, J. Giedd, A. Belger, , G. Dent, B. Meurers, R. O Reilly J. McGinty K. Merchant, G. Uhl Mt. Werner Mt. Werner PANEL ¥To rsin, Parkin, Alpha- WORKSHOP ¥ Glutamatergic Synuclein and Tau: Four Mechanisms in the Pathophysiol- Horsemen of the Basal Ganglia ogy of Drug Abuse D. Standaert, M. Schlossmacher, , F. Valenzuela, J. D. Miller, M. Hutton J. Weiner Kauer, O. Manzoni , A. Bonci Storm Peak Storm Peak PANEL ¥ Multimodal Multidimen- PANEL ¥ GeneticV ariation in sional Digital Developmental Human Catecholamine Systems: Atlas of the Mouse Brain Impact on Neurobiology and R.E. Jacobs, G. Burns, D. Susceptibility to Psychiatric Laidlaw, R. Moats, A. Toga Disorders Twilight D.R. Weinberger, M.F. Egan, D. PANEL ¥ Death and the Lyso- Goldman, J. Cubells some Ne w Insights into Twilight Neuronal Ceroid Lipofuscinoses WORKSHOP ¥ 5-HT2A Receptors H. Geller, S. Hofmann, P. Lobel, and Forebrain Function: From D. Pearce, B. Davidson the Cell to the Clinic Rainbow L. Parsons, G. Marek, B. Pehek, PANEL ¥ See No Evil, Hear No Evil, K. Cunningham, J.H. Krystal Smell No Evil The Origins of Rainbow Head Placodes PANEL ¥ Free Radicals in M. Westerfield, H. Chu, R. Alzheimer s Disease. Where Do Karlstrom, R. Grainger, They Come from? Skyline S. Richardson, A. Butterfield, M. Smith, M. Vitek Skyline

15 Monday, January 22, continued

WORKSHOP ¥A Decade of Hits R.J. Lukas, N.S. Millar, J.M. (and Misses): Heterologous Lindstrom, K.J. Kellar Expression of Nicotinic Acetyl- Sunset choline Receptors in Mammalian Cells

Tuesday, January 23 7:30–9:30 AM PANEL ¥ Epitope Spreading: A Novel Concept with Great PANEL ¥ Dimerization of G Protein- Relevance to Multiple Sclerosis coupled Receptors Functional , P. Lehmann, V. Implications M.C. Dal Canto Tuohy, S. Miller L. Devi, M. Bouvier, S. George, Skyline K. Mackie, Y. Patel PANEL ¥ Fragile X Mental Retarda- Mt. Werner tion Syndrome: Clinical Diagno- PANEL ¥Apoptotic Mechanisms: sis and Symptoms, Genetic and Are They also Used to Regulate Molecular Mechanisms and Synaptic Stability? Neural Correlates E.A.Jonas, J.M. Hardwick, M.P. W. Greenough, D. Nelson, R. Charlton, J. Zimmerberg Hagerman, E. Berry-Kravis Storm Peak Sunset PANEL ¥ Cell and GeneTherapy for Basal Ganglia Disorders 3:30–4:30 PM O. Isacson, J. Kordower, C. POSTER SESSION 2 ¥ Presenters Svendsen, K.S. Kim available for discussion Twilight Sunshine PANEL ¥The Price of Ecstasy : The Functional Consequences of MDMA Neurotoxicity 4:30–6:30 PM G. A. Gudelsky, B. Yamamoto, PANEL ¥ Origin and Fate of New G. Ricaurte, A. Fleckenstein, S. Neurons in the Adult Brain White H. Scharfman, A. Alvarez- Rainbow Buylla, T. Palmer, J. Parent Mt. Werner

16 PANEL ¥ New Horizons in the 8:30–10:00 PM Mechanisms Mediating the Effects of Estrogen in the Brain: WORKSHOP ¥ Mechanisms of The Evidence for Membrane Neuronal Migration and Receptors Stabilization in Neocortical Development J.B. Becker, C.S. Watson, D.M. Dorsa, M.J. Kelly S. Juliano, K. Nakajima, K. Herrup, A. Kriegstein Storm Peak Mt. Werner PANEL ¥ Dopamine and Serotonin Transporters: Siblings Do Not WORKSHOP ¥ Electrophysiology of Always Behave the Same Incentive Motivation: Can We Measur e the Affective States of , A. Galli, G. Gerhardt, A. Frazer Goal-directed Behavior? and M. Sonders K. Anstrom, T. Robinson, Twilight G. Schoenbaum, W. Schultz PANEL ¥ Proteases in the Storm Peak Neurodegeneration of Alzheimer Disease (AD): Chickens or Eggs? WORKSHOP ¥ Is Peer Review Peerless? R. Nixon, L-H. Tsai, K. Wang, C. Cotman C. Atwell, M. Martin, N. Steneck Rainbow Twilight PANEL ¥The Fruit Fly, Drosophila Melanogaster: A Model System PANEL ¥ Pediatric Epilepsy: From for the Study of Complex the Clinic to Basic Mechansims Behaviors. G.W. Mathern, M. Levine, S.N. J. Hirsh, R.L. Davis, J.C. Roper, J. Veliskova Hendricks, U. Heberlein Rainbow Skyline WORKSHOP ¥ Beyond Lozenges 2+ PANEL ¥ NGF and GDNF: The Yin and Dandruff: Zn Toxicity in and Yang of Unmyelinated the CNS Nociceptors I. Reynolds, E. Aizenman, D. Wright, D. Molliver, D. Choi, J. Weiss C. Stucky, K. Albers Skyline Sunset PANEL ¥V asopressin/Vasotocin: Key Regulators of Social Behavior Across Species? E. Albers, S. Boyd, D. Maney, J. Winslow Sunset

17 Wednesday, January 24 7:30–9:30 AM PANEL ¥ Stimulation, Lesioning and Transplantation: Application for PANEL ¥The Map Kinase Signaling the Treatment of Parkinson s Pathway and Synaptic Plasticity: Disease and Dystonia Activators, Regulators and , A. Lozano, J. Kordower, Effectors J. Vitek R. Bakay P. Chapman, D. Kuhl, J. Sunset Naranjo, J. Yin Mt. Werner PANEL ¥ Mechanisms of CNS 3:30–4:30 PM Regeneration and Repair POSTER SESSION 2 ¥ Posters available for viewing only W. J . Streit, W. Tetzlaff, T. Hagg, S.R. Whittemore Sunshine Storm Peak PANEL ¥ Mechanisms of Addiction: 4:30–6:30 PM The Brain Opioid System Ain t PANEL ¥ Cholinergic Modulation of Just for Opiates Anymore Cortical Function: Channels to T. S. Shippenberg, J.J. Frost, C.J. Circuits Evans, J.F. McGinty, C.T. Napier R.C. Foehring, R. Metherate, Twilight F. F. Ebner, R. Andrade PANEL ¥ Perinatal Cell Death in Mt. Werner Pathological Environments: PANEL ¥Axon Guidance: From Which Cells Are Most Suscep- Extracellular Cues to Growth tible and How/When Do They Cone Behaviors Respond? K. Kalil, T. O Connor, P. W. Macklin, M. Miller, S. Letourneau, T. Gomez Levinson, S. Vannucci Storm Peak Rainbow PANEL ¥ Recording from a Whole PANEL ¥ Peptide Nucleic Acid — Bunch of Neurons all at Once: New Life for Antisense (and Problems, Payoffs and Promise Antigene)? J.Walters, B.Waterhouse, P. Clarke, B. Tyler-McMahon, U. P. Janak, D. Kipke, D. Woodward Langel, L. Boffa Twilight Skyline

18 PANEL ¥ GABA Receptors as PANEL ¥ Role of Rhythmic Neural Anesthetic Targets: What We ve Activity in Cardiovascular Learned from Gene and Ion Behavior: From Molecular Channel Structure/Function Details to Black Diamonds Studies J. P. Horn, P. G. Guyenet, M.P. A. Jenkins, G. Houmanics, S.J. Gilbey, D. Weinreich Mihic, D.S. Weiss, C.M. Sunset Czajkowski Rainbow 6:30 PM PANEL ¥The Response to CO and 2 BUSINESS MEETING O2: From Ion Channels to Neural Networks. Mt. Werner J.M. Ramirez, M.M. Tamkun, N.R. Prabhakar, G.B. Richerson Skyline

Thursday, January 25 7:30–9:30 AM PANEL ¥ Neuroimmunophilins: Can Small Molecules Stimulate PANEL ¥ Kainate Receptors and Therapeutic Neurorestoration? Synaptic Plasticity E. Hall, P. Letourneau, E. G.L.Collingridge, R.A.Nicoll, Villafranca, B. Gold, D. Cole J.T.R. Isaac, D. Lodge Rainbow Mt. Werner PANEL ¥The Brain is a Cookie PANEL ¥ Microarrays in Neuropsy- Monster: Contribution of Food chiatric Research: A Delusion, Composition and Hedonic Illusion, or Reality? Circuitry to the Regulation of T.M. Hyde, C. Colantuoni, R. Food Intake. Coppola, W.J. Freed D. Lattemann, B. Levin, C. Storm Peak Billington, A.Kelley PANEL ¥ Regulation of Monoamine Skyline Transporter Characteristics and PANEL ¥ Mitotic Regulators Trafficking Prospero-us in Postmitotic A. Janowsky, M. Reith, H. Neurons from Become to Melikian, A. Eshleman, N. Succumb? Zahniser I. Vincent, H. Vaessin, R. Slack , Twilight P. Baas Sunset

19 Thursday, January 25, continued 3:30–4:30 PM PANEL ¥V estibular Adaptation , J. McElligott, M. POSTER SESSION 3 ¥ Presenters L. Young Shelhamer, H. Hecht available for discussion Sunset Sunshine 4:30–6:30 PM 8:30–10:00 PM WORKSHOP ¥ Is Enhanced PANEL ¥ Can Neuronal Glutamate Dopaminergic Neurotransmis- Release Be Measured In Vivo? sion the Primary Mediator of Microdialysis vs Biosensors Hedonic Reward? A Comparison N.Maidment, M. Boutelle, R. of Human and Animal Labora- Kennedy, A. Michael, B. Dunn tory Studies. Mt. Werner C. Bradberry, H. de Wit, G. PANEL ¥ SignalTr ansduction in Koob, T. Robinson Neuronal and Neuroendocrine Mt. Werner Cells: What the Mutant Mice WORKSHOP ¥W ithin-VTA Said to Me Differences in Drug Action: A M.R. Picciotto, I. Lindberg, P. House Divided? Allen, K. Wickman W. Carlezon, S. Ikemoto, R.A. Storm Peak Wise, D. van der Kooy PANEL ¥ Regulation of Voltage- Storm Peak Dependent Sodium Channels by WORKSHOP ¥ Excito-toxicity vs Protein Kinases and Phos- Excito-trophicity : Do Seizures phatases Switch the Set-point for Survival? T. Scheuer, A.R. Goldin, A.R. , R. Simon, R. Gwinn, D. Cantrell, P.C. Ruben K. Gale McIntyre Twilight Twilight PANEL ¥ Precur- WORKSHOP ¥What Really Is the sors in Development, Health and Food for Thought: Glucose or Disease Lactate ? J.W. Fawcett, W. D. Richardson, , G. Dienel, E. S. Goldman, J.M. Levine I.A. Simpson Novotny, M. McKenna, Rainbow L.Pellerin PANEL ¥ New Insights about Rainbow Abnormal Developmental Processes and Their Relevance to Epilepsy R.S. Sloviter, F.E. Jensen, K.S. Lee, D.H. Lowenstein Skyline

20 PANEL ¥ Neurotrophin-3: A Tool to WORKSHOP ¥ Multifunctional Query, and a Mechanism for Neurons and Networks for Sensory System Development Hearing and Vocal Control and Plasticity J.S. Kanwal, C.V. Mello, M.S. R. Lane, M. Jacquin, R. Fee, G.D. Pollak, Z.M. Fuzessery Rhoades, D. Wright Sunset Skyline

Friday, January 26 7:30–9:30 AM PANEL ¥ Epileptogenesis: To Fit or Not to Fit. PANEL ¥ Location, Location, , E. Dudek, K. Location—Scaffolding Proteins in C. Wasterlain Staley, T. Sutula the CNS Skyline D. Ron, C. Garner, H-J. Chung, R. Olsen PANEL ¥ Prospects for Remyelination Therapies Mt. Werner , J. Goldman, R. PANEL ¥The Glutamate-Glutamine T. Wood Armstrong, I. Duncan, R. Cycle: Potential Targets for Franklin Modulating Glutaminergic Function Sunset J.D. Erickson, A. Schousboe, M.P. Kavanaugh, R.H. Edwards 3:30–4:30 PM Storm Peak POSTER SESSION 3 ¥ Posters PANEL ¥The Role of Pregnane available for viewing only Neurosteroids in Brain Function Sunshine and Drug Action L. Morrow, R. Rupprecht, C. 4:30–6:30 PM Grobin, S. Mellon PANEL ¥ Ups and Downs in Twilight Synaptic Transmission: Factors PANEL ¥ Neurotrophins and that Affect Short-Term Synaptic Depression Efficacy P. Skolnick, C.A. Altar, L.A. R. E. Burke, L. Kaczmarek, B. Mamounas, A. Russo-Neustadt Walmsley, F. Weight Rainbow Mt. Werner

21 Friday, January 26, continued

PANEL ¥ New Mechanisms For An PANEL ¥ Orphan G Protein-coupled Old Question: How Does Receptors as Novel Drug Targets Prefrontal Cortex Affect Subcor- P. Iredale, O. Civelli, M. tical Dopamine Function? Yanagisawa, M. Fidock, S. B. Moghaddam, S. Sesack, J.A. Wilson Kauer, J. Krystal Skyline Storm Peak PANEL ¥The 5HT2b Receptor: A PANEL ¥ Microarray Profiling: Candidate for Serotonin Medi- Complex Diseases and Drug ated Morphogenesis Inside and Development Outside the Brain S. Potkin, W. Bunney, R. S.W. Schwarzacher, Thompson, N. Patil, M. Oliver J.M. Lauder, E. Fiorica-Howells, Twilight J.L. Maroteaux PANEL ¥GABA Transporter Sunset Localization, Development and Function 8:00 PM W. Spain, F. Conti, M. Quick, BANQUET AND DANCE G. Kinney, M. Jones Grand Ballroom Rainbow

Don’t forget to visit the exhibit area.

22 Poster Sessions Schedule and Abstracts Poster Session 1 • Sunday/Monday Sunshine Posters will be available for viewing from 3:30 PM Sunday to 4:30 PM Monday. Presenters will be with the posters on Sunday from 3:30 to 4:30 PM.

Trafficking of Zip Code Binding Protein and Binding to the ß-actin Zip Code Mediate the Regulation of ß-actin mRNA Localization to Neuronal Processes H.L.Zhang,T.Eom, Y.Oleynikov, S.M. Shenoy, R.H. Singer and G.J. Bassell*. Department of Neuroscience, Albert Einstein College of Medicine,Bronx, NY. The active transport of § -actin mRNA to neuronal processes may be a regulated mechanism to promote the enrichment of § -actin protein within growth cones and filopodia. Our previous results have demonstrated a microtubule-dependent § -actin mRNA localization mechanism which is regulated by the neurotrophin, NT-3. We have now identified cis-acting elements and trans-acting factors which are involved in this localization mechanism. Two zip code sequences, within the 3 UTR, which were previously identified for their role in fibroblasts, localized a heterologous reporter to neuronal processes. Antisense oligonucleotides to these zip codes, applied to the culture medium, were shown to inhibit the NT-3 stimu- lation of § -actin mRNA localization. Gel shift assays indicated that antisense oligonucleotides inhibited zip code function by preventing the formation of an RNA-protein complex. Zip Code Binding Protein, ZBP, was shown to bind the § -actin zip code in neuronal extracts using supershift, immunodepletion and UV- crosslinking experiments. RT-PCR, sequence analysis and Western Blot revealed the expression of ZBP in developing chick forebrain. Using high-resolution fluo- rescence in situ hybridization and immunofluorescence, ZBP was observed in granules which co-localized with § -actin mRNA in growth cones. ZBP granules were observed to colocalize along microtubules. Zip Code Binding Protein, fused to EGFP, was transfected into cultured neurons for analysis in live cells. ZBP- EGFP was observed in the form of particles which exhibited rapid anterograde and retrograde movements. These results suggest a novel mechanism for the regu- lated delivery of cytoskeletal precursors to growth cones, which involves the for- mation of an RNA-protein complex and its active transport to the growth cone, the site of local § -actin synthesis.

23 The Shaping of Presynaptic Calcium Transient by Mitochondria Brian Billups and Ian Forsyth, Department of Cell Physiology and , University of Leicester, Leicester LE1 9HN. UK Calcium entry into the presynaptic terminal triggers neurotransmitter release. This calcium must be swiftly removed to prevent neurotoxic accumulation and allow continuous high frequency synaptic transmission. Various mechanisms are thought to be responsible for terminating the calcium signal, including mobile and fixed calcium buffering proteins, pumps into intracellular organelles (ER and mitochondria) and pumps and ion exchangers on the plasma membrane. Residual calcium remaining in the terminal following stimulation is an impor- tant intracellular signal, contributing to long and short-term changes in synaptic efficacy. The location, affinity and kinetics of the various calcium clearance mechanisms are important in determining how the synaptic terminal will re- spond to rapid trains of action potentials. We investigated the role of mitochon- dria in calcium clearance at the rat Calyx of Held. This glutamatergic presynaptic terminal in the auditory brainstem is the output from the globular bushy cells in the anterio-ventral cochlear nucleus (AVCN) onto the contralateral medial nucleus of the trapezoid body (MNTB). It is specialized for accurate high-frequency trans- mission and plays a role in sound source localization. Its large size is suitable for whole-terminal patch-clamp recording and measurements of intracellular cal- cium concentration using fluorescent dyes. Conventional intracellular calcium dyes (Fura-FF and Fluo-3) were used to measure the calcium concentration in the cytoplasm of the presynaptic terminal. Rhod-2 was used to concurrently measure the free calcium concentration in the mitochondria. Short trains of 2ms step depolarizations applied to the terminal by the voltage-clamp pipette result in a calcium entry via a P-type calcium channel. A brief rise in intracellular cytoplasmic calcium concentration is observed, which decays with a time con- stant of less than 100ms at 37oC. A rise in mitochondrial free calcium is also observed that decays over several seconds. Inhibitors of mitochondrial function affect these calcium concentration changes. Rotenone inhibits mitochondrial elec- tron transport (at complex I), causing rundown of the mitochondrial membrane potential and thus inhibiting the membrane potential driven uptake of calcium into the mitochondria. The rise of mitochondrial free calcium concentration upon stimulation is almost completely blocked by the application of rotenone. At the same time, the cytoplasmic calcium transient is lengthened. The effect of roten- one on the cytoplasmic calcium transient is unlikely to be due to depletion of ATP since ATP is continually applied through the patch pipette and oligomycin which blocks mitochondrial ATP synthesis has no effect on the calcium tran- sient. TPP, a drug which blocks the mitochondrial calcium uniporter without inhibiting ATP synthesis, has similar effects to rotenone, lengthening the cyto- plasmic calcium transient and blocking the rise in mitochondrial calcium. These results suggest a role for mitochondrial calcium uptake in shaping the cytoplas- mic calcium transient in presynaptic terminals.

24 Regulation of the Effects of Chronic by CDK5 James A. Bibb1, Jingshan Chen2, Per Svenningsson1, Gretchen L. Snyder1, Zhen Yan3, Richard L. Huganir4, Angus C. Nairn1, Jane R. Taylor2, Eric J. Nestler5, Paul Greengard1 1Laboratory of Molecular and Cellular Neuroscience, The Rockefeller University, 1230 York Avenue, New York, NY, U.S.A. 10021, 2Department of Psychiatry, Yale University School of Medicine, New Haven, CT, U.S.A. 06520, 3Department of Physiology and Biophysics, School of Medicine and Biomedical Sciences, State University of New York at Buffalo, Buffalo, NY 14214, 4Department of Neuroscience, Howard Hughes Medical Institute, Johns Hopkins University School of Medicine, Baltimore, MD, USA 21205, 5Department of Psychiatry, University of Texas Southwestern Medical Center, Dallas, TX, U.S.A. 75235 Cocaine addiction is accompanied by long-term molecular and cellular changes in the brain. Chronic cocaine exposure causes accumulation of the _FosB in dopaminoceptive neurons of the striatum. DNA array analysis of inducible transgenic mice was used to identify Cyclin-dependent kinase 5 (CDK5) as a downstream target gene of _FosB. Moreover, _FosB over expres- sion or chronic cocaine administration raised CDK5 mRNA and protein levels. Intra-striatal injection of CDK5 inhibitors potentiated behavioral effects of re- peated cocaine administration. Chronic cocaine increased CDK5-dependent phos- phorylation of DARPP-32 (Dopamine and cyclic AMP-Regulated Phospho-Protein, Mr 32 kDa), a critical modulator of D1-dopamine/PKA sig- naling in striatal neurons. These results suggest that _FosB-mediated changes in CDK5 levels, and resulting alterations in D1/PKA/DARPP-32 signaling, con- tribute to adaptive changes in the brain related to cocaine abuse.

Heme Oxygenase-2 Prevents Neuronal Apoptosis in Brain Cultures and Following Transient Cerebral Ischemia Sylvain Dor , Johns Hopkins University/SOM, Dept Anesthesiology/Critical Care Medicine, Baltimore, MD Heme oxygenase (HO) cleaves the heme ring to form biliverdin, which is rapidly reduced to bilirubin, carbon monoxide, and iron. HO2 is constitutive and most highly concentrated in neural tissues. We demonstrated an association of amy- loid precursor proteins (APP) with HO which inhibits catalytic activity and aug- ments neurotoxicity in familiar Alzheimer s disease mutants. Bilirubin appears to be a physiologic neuroprotectant. Activation of HO2 by phorbol esters, aug- ments production of bilirubin which protects brain cultures from oxidative stress. Bilirubin itself in low concentrations is neuroprotective, while HO2 deletion (HO2-/-) leads to increased neurotoxicity in brain cultures and increased neural damage following in vivo transient cerebral ischemia (1h-middle cerebral artery occlusion followed by 23h-reperfusion). Mechanisms whereby HO2 provides neuroprotection have not been clarified including whether protection is prima- rily associated with apoptotic or necrotic cell death. Moreover, the generality of

25 neurotoxic stimuli influenced by HO2 has been unclear. We now demonstrate increased neuronal death in cerebellar granule cultures of HO2-/- mice with a selective augmentation of apoptotic death. We also demonstrate that cell lines overexpressing HO2 are resistant to induced apoptotosis. In intact mice, we show an increased incidence of apoptotic morphology in the penumbra area surround- ing the infarct core in HO2-/- mice undergoing transient focal ischemia. In the present study, we have provided new evidence supporting earlier indications for a protective role for neuronal HO2. (Supported by the American Heart and Stroke Association).

The Neurotoxicity of Nitroxyl Anion: Peroxynitrite-initiated Secondary Excitotoxicity Sandra J. Hewett* and Tracy F. Uliasz. Dept. of Neuroscience, The University of Connecticut Health Center, Farmington, CT 06030 Experimental evidence suggests that reactive nitrogen oxide species can contrib- ute significantly to cerebral ischemic injury. Recently, several in vitro studies have suggested that nitroxyl anion (NO-), the one electron reduction product of nitric oxide (NO), can be formed by NO Synthase either directly or indirectly. We investigated the neurotoxic potential of NO- using Angeli s salt (AS) as a sponta- neous NO- generating compound. When murine mixed cortical cell cultures were exposed to increasing concentrations of AS (1-5 mM; 1 hr), there was a decrease in survival that progressed markedly over the next 20-24 hr. Co-adminis- tration of AS (5 mM) with tempol (1 mM), a one electron oxidant that converts NO- to NO, completely prevented the toxic effects of AS. As NO- can react with O2 to form peroxynitrite (OONO-), we next set out to determine if OONO- was formed and responsible for the toxicity of AS. Dot Blot analysis of supernatants removed from cultures that had been exposed to AS (1-5 mM; 1 hr) revealed a concentration-dependent increase in 3-nitrotyrosine-immunoreactivity, a foot- print of OONO-. Further, AS-induced neuronal damage was prevented in a con- centration-dependent manner by the concomitant addition of FeTPPS (25-100 mM), an OONO- decomposition catalyst. Finally, addition of the glutamate re- ceptor antagonists, MK-801 (10 mM) and CNQX (30 mM), to the cultures pre- vented the delayed toxicity which occurred over the next 20-24 hr. Taken in toto, our data suggest that NO- neurotoxicity is initiated by OONO- and progresses via secondary excitotoxicity. Supported in part by NINDS grant NS 36812 (SJH).

Rac1A Mediates the Production of Reactive Oxygen Species in Motor Neurons in Response to Proinflammatory Cytokines M. V. Wright and T. B. Kuhn, Institute of Arctic Biology, University of Alaska Fairbanks Reactive oxygen species (ROS) have emerged as mediators in the pathogenesis of numerous neurological disorders and recently, also in normal cellular signal- ing. Increasing evidence implicates the neuronal plasma membrane as an impor- tant source of ROS production, yet the underlying biochemical mechanisms remain poorly characterized. Many NAD(P)H-dependent plasma membrane oxi-

26 doreductases (PMORs), some ligand-regulated, exist in various cell types including chick central nervous system (CNS) neurons. Subjecting plasma membranes de- rived from chick to non-denaturing gel electrophoresis followed by NBT reductase staining, we have obtained evidence for a rac1A-sensitive NADH-dependent PMOR, which may help maintain the antioxidant capacity of the plasma membrane. Second, chick motor neurons contain an NADPH- dependent PMOR, which exhibits strong similarity to the well-documented NADPH oxidase of neutrophils. Other reports suggest that this NADPH-depen- dent PMOR be regulated by rac1A analogous to the rac2-dependent regulation of the neutrophil NADPH-oxidase. Adenoviral-mediated expression of consti- tutively active (V12) rac1A in primary motor neurons significantly stimulated ROS production (1.67+0.06, p<0.01) compared to a normalized control (lacZ). Similarly, 100 ng/ml tumor necrosis factor _ (TNF__) or interleukin _ (Il-1__), key mediators of CNS inflammation associated with acute and chronic CNS trauma, increased ROS levels (1.25+0.05 and 1.3+0.06, respectively, p<0.01), which was negated upon expression of dominant negative (N17) rac1A. Anti- oxidants and the flavoprotein inhibitor diphenylene iodonium but not rotenone all abolished ROS production mediated by V12rac1A, TNF_, or Il-1_ suggesting the neuronal plasma membrane as the ROS source rather than mitochondria. Interestingly, actin filament reorganization accompanies both V12rac1A expres- sion and increases in intraneuronal ROS. Our data suggests that a rac1A-medi- ated generation of ROS in motor neurons constitutes a redox-signaling pathway possibly involved in redox homeostasis and cytoskeletal reorganization. Yet ab- errant function or overstimulation could result in oxidative damage to motor neurons as suggested by a significant increase in motor neuron cell death upon persistent exposure to TNF_, or Il-1_. [Supported in part by NINDS/NIMH/ NCRR and the Christopher Reeve Paralysis Foundation]

TAT-Facilitated Cell Entry of Peptide Nucleic Acid: A Novel Antisense Approach Joseph A. Ludwig, M.D., Dept of Internal Medicine, Mayo Clinic Jacksonville Peptide Nucleic Acids (PNAs) are DNA analogs that have potential to impede gene specific RNA polymerase II and ribosome elongation, thereby inhibiting transcription and translation, respectively. Inhibition of at this DNA/RNA level subsequently reduces protein expression of targeted genes while leaving expres- sion of other genes unaffected. This is important because effective antisense therapy may allow for suppression of pathogenic gene expression. In vivo effi- cacy of this antisense approach with unmodified PNAs has already been dem- onstrated in neurons. However, unmodified PNA treatment has several obstacles — PNAs are rapidly cleared in vivo and PNAs enter neurons poorly in vitro. It is proposed that these two obstacles can be overcome by covalently linking a car- rier peptide (TAT) to the utilized PNA. TAT is an eleven amino-acid carrier protein that exists in nature as a subcomponent of the larger transactivating re- sponse element produced by the human immunodeficiency virus (HIV). Several published reports demonstrate that TAT can facilitate transduction of large mol- ecules, previously impermeable, through the cell membrane of diverse cell types.

27 We demonstrate the first report that TAT-PNA hybrids have the ability to enter rat neurons in vitro and in vivo. Administration of the TAT-PNA hybrid to neu- ronal cells in vitro and in vivo occurred through supplementation to standard media and a one-time intraperitoneal injection, respectively. TAT- mediated PNA cell entry offers one approach to increase PNA cell entry. This technique for enhanced PNA uptake may improve antisense inhibition of patho- genic neuronal genes.

Dream Regulates Immediate Early Gene Expression in Neurons Britt Mellstr m, Dpto. Biolog a Molecular y Celular Centro Nacional de Biotecnolog a. CSIC Madrid Spain Induction of immediate early genes underlies short-term to long-term adapta- tion processes within the CNS in response to extracellular stimuli and is largely influenced by variations in the level of cAMP and Ca2+ ions. We have recently characterized the DREAM protein, the first known transcriptional repressor that is regulated by changes in the concentration of the second messengers cAMP and Ca2+ (Carri n et al. Mol. Cell. Biol. 18:6921-6929, 1998; Nature 398:80-84, 1999). Under basal conditions DREAM binds to the sequence-specific regula- tory element DRE and represses transcription of target genes including immedi- ate early genes c-fos, c-jun and ICERCREM. Upon neuronal stimulation, EF-hands present in the DREAM protein sense the rise in nuclear calcium in- ducing a conformational change of DREAM that results in unbinding from the DRE sequence and de-repression of the target gene. Moreover, de-repression after cAMP is cell-specific and depends on a direct protein-protein interaction between DREAM and aCREM (Ledo et al. Mol. Cell. Biol.Dec, 2000). To fur- ther understand the functional significance of DREAM as a transcriptional re- pressor of the immediate early response, we have prepared transgenic mice overexpressing dominant negative mutants of DREAM in neurons by the use of the neuron-specific CaMKIIa promoter. Work funded by CAM and DGICYT, Spain.

The Ultrastructural Basis for Synaptic Transmission at Large Calyceal Terminals in the Anteroventral Cochlear Nucleus Madeleine J. Nicol, The Synaptic Structure and Function Group, The John Curtin School of Medical Research, The Australian National University, Australia Auditory nerve fibers enter the mammalian brainstem and make large synaptic contacts with bushy cells in the anteroventral cochlear nucleus. Over the past several years, our laboratory has made an intensive electrophysiological investi- gation of the endbulb-bushy cell connection in rat brainstem slices. One of our recent studies (Oleskevich, S., Clements,J. and Walmsley, B., Journal of Physiol- ogy, 524(2), 513-523, 2000) has used variance-mean analysis to show that the mean quantal release probability is high (approx. p = 0.6) at endbulb release sites, and that there are many active release sites (in the order of several hun- dred). This study also showed that there are considerable differences between

28 connections in the mean release probability and release during paired stimuli. In the present study we have performed serial section electron microscopy of endulbs of Held to provide insight into our electrophysiological observations. Serial sec- tion reconstructions of endbulbs revealed a large number (several hundred) of separate synaptic specializations within a single endbulb. A wide range of shapes and sizes was observed in the presynaptic active zones, and in the number of morphologically docked vesicles at these active zones. Large active zones usu- ally exhibited multiple distinct clusters of vesicles, suggesting that a single pr- esynaptic active zone may contain more than one functional release site. Differences between active zones in the number of docked vesicles, and in the number of vesicles immediately adjacent (<300 nm) to these active zones, may provide a structural basis for the observed variability in release probability and the paired-pulse response at endbulb-bushy cell connections.

NMDA Receptor Phosphorylation Is Critical for Dopamine D1 Receptor-Mediated CREB Phosphorylation and Gene Expression A. Rajadhyaksha and C. Konradi. Laboratory of Neuroplasticity, McLean Hospital and , Belmont, MA 02178 Dopaminergic neurotransmission is a major mechanism of striatal plasticity. Dopamine D1 receptors phosphorylate the cAMP response element binding pro- tein (CREB) at Ser133. CREB is involved in many biological processes, includ- ing synaptic plasticity, learning, and memory formation. In primary striatal cultures, we have examined the intracellular pathway that leads from D1 recep- tor activation at the membrane to Ser133 CREB phosphorylation and CRE-me- diated gene expression in the nucleus. We find that D1 receptors recruit the NMDA second messenger pathway through an intracellular mechanism. Pro- tein kinase A (PKA) activated by D1 receptors phosphorylates the NMDAR1 (NR1) subunit of the NMDA receptor at Ser897 as seen with phosphorylation specific antibodies. We will present work that examines the importance of NR1 phosphorylation in the D1 pathway. Two approaches will be presented, trans- fecting phosphorylation site-specific mutants into primary striatal cultures using the calcium phosphate method and the use of the herpes simplex virus-1 (HSV- 1) as a vector for gene delivery. HSV-1 infection allows stable expression of re- combinant proteins and provides a powerful technique to examine signal transduction pathways in the brain.

Novel System A Transporters Mediating Na+:Glutamine/Alanine Cotransport in Glutamatergic Neurons Helene Varoqui, Bryan Mackenzie, Dongdong Yao, Matthias Hediger and Jeffrey D. Erickson, Neuroscience Center, LSUHSC, New Orleans, LA and the Membrane Biology Program and Renal Division, Brigham & Women s Hospital, Harvard Medical School, , MA cDNA clones encoding plasma membrane glutamine- and alanine-preferring transporters (SAT1 and SAT2) have been isolated from glutamatergic neurons in

29 culture and represent two novel members of the system A family of neutral amino acid transporters. SAT1 mRNA and protein expression are abundant in the brain, compared to peripheral organs, and display a widespread distribution with en- richment on neurons that use glutamate as their neurotransmitter. SAT2 displays a more widespread distribution and is expressed in most tissues, but in the brain it is also enriched on glutamatergic neurons. Both SAT1 and SAT2 are signifi- cantly upregulated during differentiation and depolarization of cerebellar gran- ule cells. Both isoforms are absent from astrocytes in primary culture. The functional properties of SAT1 and SAT2, examined using transfected fibroblasts and in cRNA-injected voltage-clamped Xenopus oocytes, show that small ali- phatic neutral amino acids are preferred substrates, transport is voltage and Na+- dependent (1:1 stoichiometry), pH-sensitive and inhibited by alpha- (methylamino)isobutyric acid (MeAIB), a specific inhibitor of system A. Kinetic analyses of alanine, glutamine and MeAIB uptake by SAT1 and SAT2 are satu- rable, with Michaelis constants (Km) of 200-500µM. In addition to their ubiqui- tous roles as substrates for oxidative metabolism and major vehicles of nitrogen transport, SAT1 and SAT2 may provide glutamine and alanine to glutamatergic neurons to function as precursors for neurotransmitter synthesis.

Agonist Induced Trafficking of EGFP-Tagged Dopamine Receptors Kim A.Neve, Tara A. Macey, and John N. Mason, Portland VA Medical Center Desensitization of the dopamine D1receptor is thought to be due to phosphory- lation of the receptor by cyclic AMP-dependent protein kinase (PKA) and G protein-coupled receptor kinases (GRKs). We constructed an enhanced green fluorescent protein (EGFP) tagged D1 receptor in which EGFP was fused to the C terminus of the D1 receptor, and we stably expressed the D1-EGFP fusion protein in NS20Y neuroblastoma cells, a cell line that normally expresses a low abundance of D1 receptors. The D1-EGFP receptor was indistinguishable from the wildtype D1 receptor in terms of affinity for several lagonists and antago- nists, stimulation of cyclic AMP accumulation, and agonist-induced desensiti- zation of cyclic AMP accumulation. D1-EGFP was phosphorylated when cells were treated with dopamine, a reaction catalyzed by PKA, since it was prevented by the PKA inhibitor H-89. Fluorescence was primarily localized to the cell membrane in NS20Y cells stably expressing the D1-EGFP receptor. Treatment with dopamine caused in increase in the ratio of intracellular-to-membrane fluo- rescence, due to the internalization of D1-EGFP in endocytotic vesicles, that was evident within 4 minutes and continued to increase for at least 20 minutes. Receptor internalization was prevented by inhibition of PKA. Mutant receptors were constructed in which two potential sites of phosphorylation by PKA, Thr- 268 and Ser-380, were converted to alanine. of Thr-268, but not Ser- 380, prevented dopamine-induced phosphorylation and internalization of the receptor. We conclude that phosphorylation of the D1 receptor on Thr-268 by PKA is a necessary step in dopamine-induced internalization of the D1 recep- tor. An EGFP-tagged D2 receptor has also been constructed, and the subcellular localization and agonist-induced internalization of this receptor will be described. (Supported by MH45372, MH12435, and VA Merit Review)

30 AMPA Receptor Trafficking in Nucleus Accumbens Neurons M.E. Wolf, S.Z. Chao, D.A. Peterson While dopamine (DA) receptors mediate acute effects of amphetamine and co- caine, neuroadaptations produced by chronic drug administration require glutamate transmission and are associated with changes in function. We have therefore hypothesized that addiction is a maladaptive form of neuroplasticity that may involve cellular mechanisms similar to those impli- cated in LTP and LTD. Very recently, it has been shown that AMPA receptor trafficking is a regulated process that plays an important role in LTP and LTD. We are interested in the possibility that chronic DA receptor over-stimulation (e.g., during repeated cocaine or amphetamine treatment) might influence AMPA receptor trafficking and thereby produce changes in synaptic strength in addic- tion-related neuronal circuits. Nucleus accumbens neurons play a critical role in addiction and receive convergent DA and glutamate inputs. We have therefore developed postnatal rat nucleus accumbens cultures as a model system to study interactions between DA and glutamate receptors. The present study examined the effect of D1 and glutamate receptor stimulation on surface expression of the AMPA receptor subunit GluR1 using fluorescence and confocal microscopy. Surface GluR1 was labeled by incubating live cultures with antibody recogniz- ing the extracellular portion of GluR1. Cultures were then fixed and incubated with fluorescent secondary antibody. Surface GluR1 labeling on neuronal pro- cesses was enhanced by D1 receptor stimulation and decreased by glutamate receptor stimulation. These results suggest that AMPA receptors on nucleus accumbens neurons may be subject to dynamic regulation in response to changes in the activity of DA and glutamate inputs. This may be important for triggering more enduring adaptations related to addiction. Support: DA09621, DA00453, DA13006, NARSAD (MEW); DA06024 (SZC); Alzheimer s Association (DAP).

Molecular Mechanisms and Modulation of Glial Activation: Ligand Modulation of Biological Responses Mediated by Gene-Regulating Protein Kinases D.M. Watterson, A.R. Sawkar, L. Guo, S. Mirzoeva, A. Velentza, T.J. Lukas, T. Koppal, M. Hibert, J.-J. Bourguignon, , J. Haiech and L.J. Van Eldik.Drug Discovery Program and Northwestern University Medical School, Chicago, IL, and IFR G.Laustriat, Facult de Pharmacie, ILLKIRCH, France Neuroinflammation in the CNS is a process that results primarily from an ab- normally high level or chronic activation of glial cells (astrocytes and microglia). Activation of glial cells leads to induction of pro-inflammatory cytokines like interleukin-1_ (IL-1_) and tumor necrosis factor-_ (TNF_), and oxidative stress- related enzymes like inducible nitric oxide synthase (iNOS) which generates ni- tric oxide (NO) and NO-derived neurotoxic species like peroxynitrite. While transient glial activation is beneficial for brain repairing processes, a chronic re- active state or an abnormally high proportion of activated may lead to neu- rotoxicity through propagation of inflammation and oxidative stress. Increasing basic science and clinical evidence suggests that neuroinflammation is a con-

31 tributor to pathology in neurodegenerative disorders, and that suppressing in- flammatory responses may be beneficial in terms of slowing progression of dis- ease. Therefore, elucidation of key signal transduction pathways and gene expression changes in activated glia is important to provide insight into mecha- nisms for down-regulation of these detrimental responses. In this regard, our previous investigations (Brain Res 1999, 844:126) found that certain anti-inflam- matory drugs and cell permeable kinase inhibitors could block iNOS induction in a microglial cell line, possibly through the inhibition of calmodulin protein kinase (CaMK) activity, and gene-regulating serine/threonine protein kinases have been demonstrated to be viable drug discovery targets in the regulation of some non-CNS inflammatory disorders (Drug Discovery Today 1999, 4:472). Therefore, we have initiated the discovery of new classes of cell-permeable ligands that are potential small molecule inhibitors of gene regulating CaMKs in order to explore the feasibility of this approach toward selective modulation of neuroinflammatory responses. We report here progress on exploring the role of CaMKs in mediating iNOS and IL-1_ induction in activated astrocytes and mi- croglia stimulated with cytokines, lipopolysaccharide, or beta-amyloid peptide, and the development of new classes of cell-permeable protein kinase inhibitors capable of downregulating iNOS and IL-1_ induction in response to glial acti- vating stimuli. Our results suggest roles for CaMKs in glial activation pathways that are independent of, and potentially complementary to, NF_B- mediated mechanisms. Current investigations using small molecule mimetics of peptide inhibitors (J.Med.Chem. 1999, 42:910; FASEB J. 2000, 14:306) and chemical scaffolds from smar t chemical libraries, designed with pharmacological and toxicological considerations, indicate that these new classes of compounds might be useful starting points for development of ligands capable of selectively modu- lating glial inflammatory responses and their neurotoxic consequences.

Neuronal/Glial Nitrogen Shuttling during Glutamate Synthesis in Retina Depends upon BCAT Enzymes Charles P Taylor, Kathryn F LaNoue, Neuroscience and Anatomy, Pennsylvania State Univ. College Med., Erich Lieth, SolOrganics, Susan M Hutson, Dept. Biochemistry, Wake Forest Univ. The glutamate/glutamine shuttle between neurons and glial cells has been stud- ied for about 30 years and is widely accepted as the major pathway for the recy- cling of glutamate in neuronal tissues. However, the de-novo iformation of glutamate in neuronal tissues is less clearly understood. The acutely isolated rat retina is a good system to study glutamate metabolism in vitro, because it is an anatomically normal neuronal/glial tissue with little acute injury. In isolated retinas, measurement of new synthesis of 14C-glutamate and 14C-glutamine from H14CO3- showed that amino acid synthesis increased 70% when medium pyruvate was increased from 0.2 to 5 mM. With addition of gabapentin (a weak but specific inhibitor of branched-chain aminotransferase-C [BCATc]), new syn-

32 thesis of 14C-glutamate and 14C-glutamine from H14CO3- was reduced by 31%. Furthermore, the oxidation of glutamate was increased by gabapentin treatment. These and additional results suggest that metabolism of pyruvate and subse- quent transamination of alpha-ketoglutarate to glutamate in glial cells (by BCATm) is rate limiting for de novo glutamine synthesis. They also suggest that selective inhibition of BCATc causes an increased rate of glutamate degrada- tion. Furthermore, branched-chain amino acids are an obligatory nitrogen source for de novo glutamate formation. A scheme relating the exchange of branched- chain amino acids for branched-chain ketoacids (mediated by BCAT isoenzymes, and existing in parallel to the glutamine/glutamate cycle) is described.

Modeling Huntington’s Disease in a Dish: Expression of Expanded Polyglutamine Huntington Rapidly Kills Neuronal (PC12) but Not Astrocytic Cells in Culture Erik S. Schweitzer, Brain Research Institute, UCLA We have generated stable PC12 cell lines that express exon 1 of huntingtin (Htt) containing either wild type (Q25) or expanded polyglutamine (Q103) tract, fused to enhanced green fluorescent protein (EGFP). We have placed these constructs in a plasmid containing the Bombyx mori ecdysone-responsive element, such that expression of the huntingtin protein is dependent on the presence of an ecdysone analog, tebufenozide. We have also generated parallel Htt-expressing cells from a conditionally-immortalized astrocytic cell line, BAS8.1. In the ab- sence of tebufenozide, the engineered PC12 cells grow normally and respond normally to NGF by exiting the cell cycle and differentiating morphologically. They express low-to-undetectable levels of the Htt transgene. When tebufenozide is added to the culture medium of cells engineered to ex- press HttQ25, there is a rapid accumulation of Htt that reaches steady-state within 24 hrs. Subsequent removal of the tebufenozide results in a return to baseline levels of Htt within 48 hrs. This removal of HttQ25 after removal of tebufenozide is completely blocked by proteasome inhibitors, indicating that proteasomes are solely responsible for the turnover of Htt. Direct fluorescence microscopy indi- cates that the HttQ25 is diffusely distributed throughout the cytoplasm, and is relatively excluded from the nucleus. Adding tebufenozide to cells carrying the HttQ103 construct leads initially to a diffuse cytoplasmic distribution of HttQ103, similar to HttQ25. However, this diffuse distribution rapidly is replaced by the presence of an intensely fluorescent perinuclear aggregate of HttQ103, reminis- cent of the intraneuronal aggregates seen in the brains of patients with Huntington s disease. Within a day or two, virtually all PC12 cells expressing HttQ103 die. We do not see this same toxic effect with either PC12 cells expressing HttQ25 or BAS8.1 cells expressing HttQ103. We have therefore replicated the selective neurotoxic- ity that is one of the hallmarks of Huntington s Disease in patients.

33 Volume-Actvated Cl– Currents Contribute to the Invasive Migration of Human Glioma Cells C.B. RANSOM1, J.T. O NEAL2*, AND H. SONTHEIMER1,2 1Dept. of Neurobiology, Univ. of Alabama School of Medicine; 2Transmolecular Inc., Birmingham, AL 35294 The vast majority of primary neoplasms in human CNS derive from glial cells. These tumors have a poor prognosis due to their invasive migration that renders surgical treatment untenable. Ion channels may contribute to this behavior by influencing salt and water movements between intracellular and extracellular compartments during shape and volume changes associated with migration through brain tissue. We used patch-clamp techniques and a transwell migration assay to examine the contribution of volume-activated Cl- channels to the inva- sive migration of human glioma cells from established lines (STTG-1, D54MG). Whole-cell currents evoked by cell swelling with hypotonic solutions were Cl- - selective and were sensitive to the Cl channel blocker NPPB. The IC50 for cur- rent inhibition by NPPB was »20 mM and near complete block (>90%) was ob- tained at 100 mM. In addition, the resting conductance of glioma cells was dominated by basal activation of these volume-activated Cl- currents, even in non-dialyzed cells (amphotericin perforated-patches). Migration of glioma cells in our transwell migration assay (8 mm pores) was extracellular matrix-depen- dent and was sensitive to NPPB. Inhibition of cell migration had a similar con- centration-dependence as NPPB inhibition of volume-activated Cl- currents. These results suggest that volume-activated Cl- currents are activated during, and contribute to, shape changes associated with glioma cell migration through nar- row spaces, such as would be encountered in the tortuous extracellular space of brain. This work was supported by RO1 NS36692.

Poster Session 2 • Tuesday/Wednesday Sunshine Posters will be available for viewing from 3:30 PM Tuesday to 4:30 PM Wednes- day. Presenters will be with the posters on Tuesday from 3:30 to 4:30 PM

Alpha 2C-Adrenoceptors Modulate Striatal Neurotransmitter Release and May Be Promiscuously Activated by Dopamine G.A. Ordway and W. Zhang, Dept of Psychiatry and Human Behavior, University of Mississippi The striatum has a high density of alpha 2C-adrenoceptors (alpha 2C-AR) rela- tive to other brain regions. However, the physiological significance of the striatal alpha 2C-AR is unknown. In our studies, a combination of molecular and phar- macological methods have been used to characterize the role of the alpha 2C- AR in striatal functions. To evaluate receptor function, mice with a targeted inactivation of the alpha 2C-AR gene (Adra2c-/-; KO mice) and genetically- related control mice expressing the wild-type 2C-AR (Adra2c+/+; WT mice)

34 were used. In addition, normal rat kidney (NRK) cells transfected with mouse alpha 2A-AR or ?2C-AR cDNA (NRK- alpha 2A or NRK- alpha 2C cells) were used to evaluate receptor affinities and functions. The alpha 2-AR antagonist, RX821002, enhanced forskolin-stimulated cyclic AMP accumulation in striatal slices from WT mice. In contrast, RX821002 did not produce this effect in stri- atal slices from KO mice or from monoamine-depleted WT mice. RX821002 increased electrically-stimulated [3H] release and reduced [3H]GABA release in striatal slices from WT mice, but these effects were absent in striatal slices from KO mice or from monoamine-depleted WT mice. RX821002 had no effect on [3H]dopamine release in striatal slices from WT mice. Given the paucity of noradrenergic innervation of the striatum, we considered that dopamine might act as an agonist at the striatal alpha 2C-AR. Dopamine inhib- ited forskolin-stimulated cyclic AMP accumulation in intact NRK- alpha 2A and NRK- alpha 2C cells, effects that were antagonized by RX821002. The affin- ity of dopamine for the mouse alpha 2C-AR in NRK- alpha 2C cells, determined in homogenate binding assays, was higher than the affinity of norepinephrine for the mouse alpha 2A-AR in NRK- alpha 2A cells. Together, these findings demonstrate that activation of the alpha 2C-AR regulates neurotransmitter re- lease in the mouse striatum, possibly by inhibiting adenylyl cyclase. Further- more, dopamine may be an endogenous activator of the striatal alpha 2C-adrenoceptor.

Dopamine Modulates Long-Term Depression in the Mesolimbic System Mark J. Thomas§, Robert C. Malenka§ and Antonello Bonci*, § Nancy Pritzker Laboratory, Department of Psychiatry and Behavioral Sciences, Stanford University School of Medicine, Palo Alto, California 94134, USA, * Ernest Gallo Clinic and Research Center, Department of , University, of California, San Francisco, California 94110, USA Long-lasting adaptations in the mesolimbic dopamine (DA) system in response to drugs of abuse appear to be crucial in mediating many of the behavioral changes that underlie drug addiction. Long-term changes in synaptic plasticity at excita- tory synapses in the two major components of this system, the nucleus accumbens (NAc) and ventral tegmental area (VTA), may be particularly important for the development of drug-related behaviors such as behavioral sensitization, as well as for normal response-reinforcement learning. Using whole-cell patch-clamp recording techniques from in vitro slice preparations, we have examined the ex- istence and basic mechanisms of long-term depression (LTD) at excitatory syn- apses on both GABAergic medium spiny neurons in the NAc and dopaminergic neurons in the VTA of C57 mice. Our results indicate that both sets of synapses express LTD, but that their basic triggering mechanisms differ. Furthermore, DA blocks the induction of LTD in the VTA via activation of D2-like receptors but has minimal effects on LTD in the NAc. The existence of LTD in mesolimbic structures and its modulation by DA represent mechanisms that may contribute to the modifications of neural circuitry that mediate reward-related learning as well as the development of addiction.

35 Postinhibitory Rebound Firing in Vestibular Nucleus Neurons Alters Firing Dynamics Chris Sekirnjak and Sascha du Lac, The Salk Institute for Biological Studies, 10010 N Torrey Pines Rd, La Jolla, CA 92037 Vestibular nucleus neurons encode information about head movements in modu- lations of firing rate. The transformation of incoming spike trains to output fir- ing is governed by both the nature of the input signals and intrinsic cellular properties. Here we attempt to link a cellular phenomenon, postinhibitory re- bound firing (PRF), to the in vivo firing behavior of medial vestibular nucleus (MVN) neurons and show how this intrinsic membrane property can account for aspects of signal processing dynamics that have previously been thought to arise purely from network connectivity. A subset of MVN neurons in mouse brainstem slices respond to brief periods of membrane hyperpolarization with a strong transient increase in firing rate (up to 70 spikes/s), lasting several seconds. PRF could be evoked either with inhibitory synaptic stimulation or with intrac- ellular current injection and depended strongly on hyperpolarization amplitude and duration. From studying MVN cells at physiological firing rates we con- clude that many cells routinely experience PRF under normal conditions in the living animal. Contributions from three currents underlying PRF were identi- fied: the cesium-sensitive H-current, a calcium current blocked by cadmium, and a TTX-sensitive Na-current. We compared the neuronal response to simulated head movements (sinusoidal intracellular current injection) in different MVN cell types and found that neurons with pronounced PRF displayed an average phase lead of about 10 degrees compared to low-PRF cells. A similar phase dif- ference between functionally distinct MVN neurons has been reported in vivo. This phase advance could aid in overcoming phase lags introduced by the oculo- motor plant and sharpen the time-varying motor output of the vestibulo-ocular system. The phase of the premotor signal would hence be regulated by both intrinsic and network factors.

A 5-HT6 Receptor Antagonist Facilitates an Amphetamine-Induced Increase in Frontal Cortical Dopamine K. J. Frantz, D. G. Stouffer, I. Polis, L. H. Parsons. Department of Neuropharmacology, The Scripps Research Institute, 10550 N. Torrey Pines Rd. La Jolla, CA 92037 Serotonin6 receptors are localized in high density in limbic and motor brain regions, and are occupied readily by several atypical antipsychotic agents. These characteristics suggest that 5-HT6 receptors may be involved in reward-related behaviors and motor activity. In previous experiments from our laboratory, a novel 5-HT6 receptor antagonist (SB 258510A) potentiated the locomotor ef- fects of amphetamine. It also altered lever-pressing maintained by amphetamine on both fixed- and progressive-ratio schedules of reinforcement, in a manner

36 suggestive of an enhancement of the reinforcing efficacy of amphetamine. In the present study, dual-probe microdialysis was employed to investigate possible mesocorticolimbic mechanisms that could mediate these effects in male Wistar rats. A 3 mm probe in the frontal cortex revealed that 5-HT6 receptor blockade facilitated an increase in dopamine induced by peripherally-administered am- phetamine. A 2 mm probe in the nucleus accumbens showed a much less robust facilitation of the amphetamine-induced increase in dopamine. This study points to the frontal cortex as an important locus where 5-HT6 receptor activation may serve to limit the stimulant and reinforcing effects of amphetamine. Recent lit- erature indicates that glutamate and acetylcholine neurotransmission may also be involved in this regulatory role of 5-HT6 receptors. Supported by NIDA: DA11004 (LHP) and DA05950-02 (KJF).

A Novel nNOS Knockout Mouse Lacking the Heme-Binding Domain Exhibits Preserved Brain Morphology, Infertility and Severe Pyloric Stenosis Robert Gyurko, Sonal Jhaveri* and Paul L. Huang, Cardiovascular Research Center, General Hospital, Boston, MA and *Department of Brain and Cognitive Sciences, MIT, Cambridge, MA The first nNOS knockout mice (KN1) lacking exon 2 show residual nNOS activ- ity in the brain due to the presence of beta and gamma splice variants. Here we report the deletion of the heme-binding domain (exon 6) of the nNOS gene us- ing Cre-Lox recombination system in embryonic stem cells (KN2). RT-PCR on brain RNA reveals residual nNOS transcripts in both KN1 and KN2 mice. How- ever, the absence of the heme-binding domain should result in complete inacti- vation of the nNOS protein even in the presence of alternatively spliced mRNAs. Heterozygote KN2 offsprings are viable, fertile, and undistinguishable from wild type littermates. Homozygote KN2 mutants are viable, but infertile. Mounting behavior is absent when either male or female KN2 s are paired with wild-type mice of the opposite sex. Histological examination reveals the absence of corpus luteum in the ovaries, and the presence of immature sperm cells in the epididy- mis. In the central nervous system, the gonadotrophs of the pituitary are intact, as evidenced by LH immunostaining. The macroscopic and microscopic struc- ture of the brain, spinal cord, pituitary and dorsal root ganglia appear to be pre- served. The barrel-field cortex is intact at 15 days of age, as evidenced by cytochrome c oxidase staining. In the enteric nervous system, KN2 appears to have a more profound defect in pyloric relaxation than KN1, resulting in enlarged stomachs in all KN2 s, as well as individual cases of acute complete functional pyloric blockade, re- sulting in rapid dehydration and death. In the absence of pyloric blockade, how- ever, KN2 are not starved, as evidenced by the absence of ketones in the serum, and the presence of abdominal fat. These data indicate that normal neural devel- opment is possible in the complete absence of nNOS, but the hypothalamic- pituitary-gonadal axis in both sexes requires nNOS for normal reproductive function.

37 Proteasome Involvement in Agonist-induced Down Regulation of Mu and Delta Opioid Receptors Richard D. Howells, Kirti Chaturvedi, Persis Bandari, Norihiro Chinen, Department of Biochemistry and Molecular Biology, University of Medicine and Dentistry of New Jersey-New Jersey Medical School, Newark, NJ 07103 This study investigated the mechanism of agonist-induced opioid receptor down regulation. Incubation of HEK 293 cells expressing FLAG-tagged delta and mu receptors with agonists caused a time-dependent decrease in opioid receptor lev- els assayed by immunoblotting. Pulse-chase experiments using [35S]methionine metabolic labeling indicated that the turnover rate of delta receptors was acceler- ated 5-fold by agonist treatment. Inactivation of functional Gi and Go proteins by pertussis toxin attenuated down regulation of the mu receptor, while down regulation of the delta receptor was unaffected. Pretreatment of cells with in- hibitors of lysosomal proteases, calpain, and caspases had little effect on mu and delta receptor down-regulation. In marked contrast, pretreatment with proteasome inhibitors attenuated agonist-induced mu and delta receptor down regulation. In addition, incubation of cells with proteasome inhibitors in the absence of ago- nists increased basal mu and delta receptor levels. Immunoprecipitation of mu and delta opioid receptors followed by immunoblotting with ubiquitin antibod- ies suggested that preincubation with proteasome inhibitors promoted accumu- lation of polyubiquitinated receptors. These data provide evidence that the ubiquitin/proteasome pathway plays a role in agonist-induced down regulation and basal turnover of opioid receptors. Supported by NIDA grant 09113.

Light Induced Potentiation of NMDA Receptors Enhanced by Protein Kinase Inhibition Daniel Leszkiewicz, BethAnn McLaughlin, Karl Kandler and Elias Aizenman, Dept of Neurobiology, University of Pittsburgh We have previously reported that certain ligand-gated ion channels in rat cortical and retinal neurons can be modulated by light. Both NMDA and GABA recep- tor-mediated currents are rapidly potentiated in response to brief focal light pulses, while non-NMDA receptors are unresponsive to light stimuli (Leszkiewicz et al. J. Physiol 524:365, 2000; Soc Neurosci Abstr 2000). In order to elucidate the molecular mechanism behind the light induced potentiation, we have utilized various phosphatase and protein kinase inhibitors. NMDA-mediated whole-cell responses in cortical neurons were measured both before and after a brief focal light pulse (0.5—1 s) directed at the cell body and proximal dendrites via an opti- cal fiber (50 (m). Light-induced potentiation of NMDA-induced currents was significantly greater in the presence of 40 (M K252c, a PKA/PKC inhibitor (111.5 % ( 19.3 enhancement, n=9), when compared to control (23.8 % ( 3.6, n=9). In contrast, vanadate (1 mM), a phosphatase inhibitor, had no effect on the light- induced potentiation of NMDA-mediated responses (19.5 % ( 4.0, n=12). These

38 results suggest that the light sensitive moiety may be closely related to protein phosphorylation sites on the NMDA receptor. Future research will characterize the modulation site using more specific protein kinase inhibitors, investigating the enhancement of GABA-mediated responses, and site-directed mutagenesis of recombinant receptors. Studying the modulation of NMDA receptors by light will further our understanding of the structure and function of the ion-channel and closely related structures.

Do Hebbian Pairing Protocols Alter the Release- dependence of Short-term Synaptic Depression? Murat Okatan and Michael Cohen, Department of Cognitive and Neural Systems and Center for Adaptive Systems, Boston University, 677 Beacon St., Boston, MA 02215 Short-term synaptic depression (STD) refers to a decrease in excitatory postsyn- aptic potential amplitude during repetitive stimulation. The exact biophysical mechanisms that underlie STD in cortical synapses are not known. Recently Matveev & Wang (J. Neurosci., 20, 4, p1575, 2000) suggested that release prob- ability in cortical synapses is too low to deplete the releasable pool to the extent that would explain the experimentally observed levels of STD (Markram & Tsodyks, 1996, Nature,382, p807). Using a stochastic synapse model, Matveev & Wang proposed that inactivation of release machinery (IRM) may be the ma- jor source of STD in synapses that couple cortical pyramidal neurons. In their implementation IRM was release-independent. Evidence suggests, however, that STD is release-dependent in these synapses (Thomson & Bannister, 1999, J. Phsyiol, 519.1, 57). To further investigate the mechanisms underlying STD, we developed an expanded stochastic synapse model that includes postsynaptic re- ceptor desensitization (Jonas et al., 1994, Neuron, 12,p1281), the number of synaptic contacts per axon, and implements IRM in release-dependent (a) and independent (b) scenarios. Using the model we fitted the pre- and post-Hebbian- pairing STD data of Markram & Tsodyks. The model predicted that IRM, and not depletion or desensitization, might be the main cause of STD. Moreover, it suggested that Hebbian pairing protocols alter the release-dependence of IRM: Pre-pairing STD data were fitted in scenarios (a) and (b) with comparable qual- ity, while the fit to post-pairing data was much better in scenario (b) than (a). We note that the experiments of Thomson and Bannister were conducted on neu- rons that were not subjected to a Hebbian pairing protocol. Thus, our results fill the gap between their results and those of Matveev and Wang by predicting that Hebbian pairing protocols may convert STD from being release-dependent in normal conditions to being release-independent after Hebbian pairing protocols, possibly by altering the sensitivity of the protein synaptotagmin that acts as the calcium sensor of the release machinery. These results raise the question of whether synaptic changes that are induced by artificial Hebbian pairing proto- cols are good approximations to naturally occurring synaptic plasticity. Supported by: Defense Advanced Research Projects Agency and the Office of Naval Re- search (ONR N00014-95-1-0409) and the Office of Naval Research (ONR N00014-95-1-0657).

39 Acute Sensitization to Cocaine after Single Injection of Orphanin FQ/Nociceptin into the Ventral Tegmental Area Shridhar Narayanan, Kabirullah Lutfy and Nigel Maidment. Department of Psychiatry and Biobehavioral Sciences, UCLA School of Medicine, Los Angeles, CA 90024. Orphanin FQ (OFQ), an endogenous neuropeptide with structural homology to dynorphin, has been shown to modulate dopaminergic mechanisms in the mesolimbic system. Administration of OFQ into the ventral tegmental area (VTA) produces hypolocomotion and also blocks the acute and sensitized motor effects of cocaine. However, repeated administration of orphanin FQ into the VTA results in a sensitized locomotor response to cocaine. The aim of the present study was to examine the potential for OFQ to produce a sensitized locomotor response to cocaine after a single intra-cerebral administration and to identify the locus of such an action. The bilateral administration of OFQ (30 mg/side) into the VTA on day 1 to male Sprague-Dawley rats resulted in an enhanced locomotor response to cocaine (10 mg/kg i.p) administered on day 2. This sensi- tization was short-lived and was not evident 7 days after the administration of OFQ. OFQ (3, 10 and 30 mg/side) administered on day 2, 5 mins prior to the administration of cocaine (10 mg/kg i.p) blocked the sensitized locomotor re- sponse, suggesting that there was no tolerance to the effect of OFQ. The admin- istration of OFQ into the substantia nigra or the nucleus accumbens did not produce a sensitized response to a cocaine challenge. These results demonstrate the ability of a single administration of OFQ to produce short-term changes in the mesolimbic dopaminergic system that is specific to the VTA.

Phosphorylation Regulates Cellulr Gain in Medial Vestubular Nucleus Neurons A. Nelson, Neurosciences Program, University of California-San Diego, La Jolla CA The vestibulo-ocular reflex (VOR) maintains visual acuity during head motion by transforming vestibular inputs into eye movement outputs. Sensory-motor learning produces adaptive changes in VOR gain (the ratio of eye and head move- ment) when image motion occurs persistently during head movement. As a site for integration of primary vestibular information and visual feedback, the me- dial vestibular nucleus (MVN) plays a role in plasticity of the VOR. MVN neu- rons demonstrate an extremely linear relationship between their inputs (synaptic or injected current) and output (firing rate). This cellular gain may be modulated during sensory-motor learning to generate behavioral gain changes. This study investigated protein phosphorylation as a candidate mechanisms for cellular gain control in the MVN. Whole-cell patch clamp recordings from MVN neurons in rat brainstem slices showed that cellular gain is increased in the presence of ATPgammaS, a non-hydrolyzable analog of ATP. ATPgammaS also reduced the range of firing rates attainable by MVN neurons. It has previously been shown that one of the primary determinants of MVN gain is the small-conductance calcium-dependent potassium (SK) channel, which regulates the interspike in-

40 terval in many cell types. Phosphorylation may be acting at the SK or other channels to modify firing properties. These findings suggest that phosphoryla- tion may play a role in adaptive modulation of the VOR.

Group Ii Metabotropic Glutamate Receptors Suppress Basal and 5-HT2a/2c-Induced C-fos Expression and Excitatory Responses in the Prefrontal Cortex Laura K. Nisenbaum, Eric S. Nisenbaum and Michael P. Johnson Eli Lilly and Company, Indianapolis, IN 46285 A common feature of psychotomimetic drugs, including PCP and DOI (a 5- HT2a/2c agonist), is their ability to increase glutamate release in the prefrontal cortex (PFC). Group II metabotropic glutamate receptors (mGluR 2/3) are known to reduce glutamate release. Previous electrophysiological studies have shown that mGluR 2/3 agonists suppress DOI-induced glutamate release. We exam- ined the ability of the mGluR 2/3 agonist, LY 379268, to reduce basal and DOI- induced EPSPs and c-fos mRNA expression in the PFC. Consistent with previous studies, application of DOI (3 mM) enhanced electrically-evoked EPSPs in PFC pyramidal neurons in a slice preparation. Subsequent application of LY379268 (0.01 — 1.0 mM) suppressed the enhanced response in a concentration dependent manner. This effect was blocked by the mGluR 2/3 antagonist LY341495 (1 mM). Furthermore, application of LY379268 reduced EPSPs in the absence of DOI. To examine the functional consequences of mGluR2/3 receptor activa- tion, we measured the effect of LY379268 on c-fos mRNA levels. Administration of DOI (12 mg/kg, i.p.) produced a significant increase in c-fos mRNA expres- sion in the PFC. Pretreatment with LY379268 (15 mg/kg, i.p.) blocked the DOI- induced enhancement. Furthermore, administration of LY379268 alone significantly decreased c-fos mRNA expression relative to control. Together, these data demonstrate that group II metabotropic glutamate receptors reduce both glutamate release as well as c-fos mRNA levels in the PFC. This study provides support for the potential therapeutic efficacy of mGluR 2/3 agonists as antipsychotics.

Calcium Transients in Higher Order Dendrites of Striatal Neurons during Spontaneous “Up” States J. N. D. Kerr*, and D. Plenz. Unit of Neural Network Physiol., LSN, NIMH, NIH, Bethesda, MD 20892 Most cortical regions project to the striatum, the major input structure of the basal ganglia. The corticostriatal axons predominantly contact spiny dendrites of striatal projection neurons. Finding the rules that govern the plasticity at these corticostriatal synapses will be necessary in order to understand how the stria- tum evaluates cortical activity. Upon synchronized cortical inputs, striatal pro- jection neurons display a fast transition from rest (-90 mV; Down state) to a stable, subthreshold membrane potential region (-60 mV, Up state) during which spiking is tightly controlled. As intracellular calcium levels are important for synaptic plasticity, we examined the relationship between Up states, dendritic

41 calcium transients and somatic spiking in striatal projection neurons. Spontane- ous striatal activity was analyzed using calcium imaging combined with whole cell patch recording in cortex-striatum-substantia nigra organotypic cultures grown for 5 6 w eeks. Single, spontaneous Up states were detected and changes in intra- cellular calcium levels were measured at time resolutions of 20 and 40 ms. Over- all, calcium transients followed the time course of the membrane potential during Down and Up states. Monitoring up to 60 striatal neurons simultaneously re- vealed that large calcium transients in single neurons indicate an overall activa- tion of the striatal network. Furthermore, in single neurons, sharp calcium transients at soma and higher order dendrites were correlated with spontaneous action potentials. Finally, action potentials evoked by current injection at the soma elicited sharp calcium transients throughout the neuron, whereas subthresh- old membrane depolarizations did not. We suggest that spatially heterogeneous calcium levels indicate local corticostriatal processing in striatal dendrites that is linked to firing success at the soma through back propagated calcium signals.

Correlated Multisecond Oscillations in Globus Pallidus Spiking and EEG Theta Rhythm: Prominent Effects of Apomorphine after Nigrostriatal Lesion David N Ruskin , Debra A Bergstrom, Lauren E Freeman, Patrick L Tierney, Judith R Walters, Experimental Theurapeutics Branch, NINDS, NIH Multisecond oscillations in firing rate with periods of 2 to 60 s are present in ~75% of globus pallidus neurons in awake rats, and are modulated by dopamine agonists [1]. To further explore the modulatory role of dopamine, and to assess the relationship of these subcortical oscillations to cerebral cortical activity, the effects of apomorphine on these globus pallidus spiking oscil- lations were ex- amined with extracellular single unit recording in the globus pallidus during si- multaneous EEG recording. Unilateral nigrostriatal dopamine neuron lesions were made by 6-hydroxydopamine infusion under chloral hydrate anesthesia (400 mg/kg, i.p.). Recording were performed several weeks after lesion. Because gen- eral anesthesia greatly attenuates dopamine responsiveness of basal ganglia neu- rons, recordings were performed in rats immobilized with gallamine and locally treated at surgical sites with injected (carbocaine s.c.) and topical (lidocaine) an- esthetics. Multisecond oscillations in globus pal- lidus firing rate often coincided with bursts of 4-7 Hz EEG rhythm: correla- tions between firing rate oscillations and theta rhythm bursts were found in ~40% of recordings in baseline in intact rats. In nigrostriatal-lesioned rats, a low dose of apomorphine (0.05 mg/kg, i.v.) significantly decreased the frequency (from 20 s to 10 s) and increased the strength (89%) of globus pal- lidus firing rate oscillations. Apomorphine also significantly increased the correlation of slow activity between globus pallidus spiking and EEG theta power (to 91%). These effects were reversed by haloperidol (0.2 mg/ kg, i.v.). Although the multisecond pattern of theta rhythm power was altered, apomorphine did not change the dominant frequency or the total power within the theta band. In intact animals, this low dose of apomorphine did not signifi- cantly alter any of these measures. However, as previously reported, a larger dose (0.32 mg/kg i.v.) increased the frequency and strength of multisecond oscil-

42 lations in the globus pallidus and other basal ganglia nuclei in intact animals [1]. The data show that the activity in basal ganglia-thalamocortical networks can be syn- chronized on multisecond time scales. Also, dopamine receptors modulate this synchronization of slow activity, in a manner demonstrating supersensitiv- ity after nigrostriatal lesion. 1. Ruskin et al, J. Neurophysiol. 81:2046; J. Pharmacol. Exp. Ther. 290:1493.

Metabotropic Receptors In Rat Cortical Astrocytes Exhibit Pharmacology Consistent With mGluR5 J.J. Schmidt, L.M. Rowse, K. Spiegel, R.D. Schwarz, P.A. Boxer; Pfizer Global Research & Development, Ann Arbor, MI. Group I metabotropic receptors (mGluRs), comprised of the mGluR1 and mGluR5 subtypes, appear to be expressed in astrocytes as well as neurons. We have chosen to study the Group I mediated calcium responses in primary rat astrocytes using FLIPR. Astrocytes from basal forebrain, cerebellum, cortex, and striatum were exposed to quisqualate and glutamate. Only astrocytes from cortex and striatum demonstrated responses that were consis- tent with Group I mGluR expression. The mGluR response in cortex was larger than that in striatum. Basal forebrain, cerebellum and hippocampus were gener- ally unresponsive to mGluR1 agents. The rank order potency of known mGluR Group I agonists on cortical astrocytes was: L-quisqualate> S- DHPG>glutamate>1S,3R-ACPD. Western blot analysis of cortical astrocyte membrane preparations showed cross-reactivity with antibodies to mGluR5 but not mGluR1. New selective mGluR5 antagonists allow pharmacological differ- entiation between mGluR1 and mGluR5, which is not readily made on the basis of agonist profile or antibody reactivity alone. These antagonists, SIB-1757, SIB- 1893 and MPEP, inhibited 300nM quisqualate responses with IC50s of 6.9µM, 9µM and 31nM, but were inactive in CHO cells expressing mGluR1 receptor. The potency, selectivity and complete inhibition exhibited by these compounds implicate the mGluR5 subtype as the predominant Group 1 mGluR receptor in rat cortical astrocytes. The presence of the mGluR5 receptor on rat primary cor- tical astrocytes suggests that mGluR5 receptor might be present on astrocytoma or glioma cell lines as well. We examined rat C6 glioma cells and 5 human astro- cytoma/glioblastoma cell lines. No evidence for functional Group I receptor was found. The absence of functional receptor in these cell lines and certain rat brain regions suggests that mGluR5 expression is not a general feature of cells of glial origin.

AMPA Receptor Phosphorylation during Cocaine Withdrawal GL Snyder, S Galdi, JA Bibb, RL Huganir*, AC Nairn, and P Greengard, The Rockefeller Univ., New York, NY 10021;*HHMI, The Johns Hopkins Univ., Baltimore MD 21205 Chronic cocaine exposure results in sensitization-enhanced neurochemical and behavioral responses to subsequent acute drug challenges. AMPA-type glutamate receptor antagonists block sensitization in rodents. The mechanism by which AMPA receptors facilitate sensitization is unclear. However, it is possible that

43 cocaine induces long-term changes in AMPA receptor activity mediated by re- ceptor phosphorylation via dopamine D1 receptors. Using phospho-specific an- tibodies, we show that cocaine treatment increases AMPA receptor phosphorylation in vivo. Acute cocaine (10 mg/kg) induced a rapid, reversible, and selective increase in phosphorylation of striatal GluR1 at Ser845, a site that controls AMPA channel open time. Cocaine also increased phosphorylation of DARPP-32 (dopamine and cAMP-regulated phosphoprotein, Mr32kD) at Thr34, a PKA site that converts DARPP-32 into a protein phosphatase 1 inhibitor and decreased phosphorylation at Thr75, a cdk5 site that converts DARPP-32 into a PKA inhibitor. Drug-induced phosphorylation of GluR1 was lost in DARPP-32 knockout mice, indicating that control of PP1 and/or PKA by DARPP-32 is required. Moreover, control of GluR1 phosphorylation by dopamine was altered after chronic cocaine. Mice were injected daily with cocaine for 7 days. Striatal slices prepared from these mice during cocaine withdrawal (1 or 7 d) were treated in vitro to assess D1/PKA-mediated regulation of DARPP-32 and GluR1 phos- phorylation. D1-stimulated phosphorylation of DARPP-32 at Thr34 and GluR1 at Ser845 was reduced, compared to controls, at 1 d, but not 7 d after withdrawal. In fact, the phosphorylation of GluR1 in response to D1 agonists or PKA activa- tors was 2-fold greater in slices 7 d after withdrawal in cocaine-treated compared with saline-treated mice. These data identify Ser845, a PKA-dependent site on the GluR1 AMPA receptor, as a striatal target regulated by psychostimulants in vivo. The results suggest that repeated cocaine use results in progressive and per- sistent changes in the coupling of AMPA receptors with PKA-signaling path- ways that may underlie certain aspects of sensitization. Supported by USPHS Grant DA-10044 (GLS, ACN, and PG).

Regulation of Cortex by Striatal Dopamine H. Steiner, Dept. of Cell. and Mol. Pharmacology, Finch University of Health Sciences/Chicago Medical School, North Chicago, IL 60064 Dopamine agonists produce increased immediate-early gene (IEG) expression throughout wide parts of the cerebral cortex, suggesting a widespread influence on cortical function. In our earlier studies, blocking dopamine action in the stria- tum by local drug infusion resulted in attenuated IEG expression in the cortex, indicating a role for striatal dopamine receptors in such cortical effects. Here, we further investigated the influence of striatal dopamine on cortical IEG expres- sion and examined the dopamine receptor subtypes involved. Thus, the effects of dopamine depletion by 6-OHDA, or systemic dopamine agonist treatment in combination with intrastriatal infusion of a D1 receptor antagonist on basal and sensory stimulation-evoked IEG expression in the cortex were assessed. Rats examined 2 days after unilateral 6-OHDA lesion showed a bilateral decrease in basal IEG mRNA levels across sensorimotor, insular and piriform, but not cin- gulate, cortex that was more pronounced on the lesioned side. Moreover, whis- ker stimulation-evoked IEG expression in the ipsilateral barrel cortex was largely abolished. Such rats displayed pronounced akinesia. Three weeks after the le- sion, both basal and sensory-evoked IEG expression in the cortex were back to control levels. This recovery of cortical IEG expression was accompanied by a

44 recovery in some, but not all, motor behaviors, and may be the result of neuroadaptive changes after the lesion. Injection of the D1/D2 dopamine recep- tor agonist apomorphine in normal rats produced increased IEG expression across sensorimotor, insular and piriform cortex, but not in the cingulate cortex, as well as behavioral activation. Whisker stimulation-evoked IEG expression in the barrel cortex was also en- hanced. Unilateral intrastriatal infusion of the D1 antagonist SCH-23390 blocked such cortical IEG expression bilaterally and inhibited most behavior, thus mim- icking acute lesion effects. These results suggest that striatal dopamine, by acting on D1 receptors, exerts a general bilateral, facilitatory effect on cortical function which seems to be necessary for normal behavior. (Supported by USPHS Grant DA11261).

CB2 Receptor Localization in the Retinas of the Rat and the Monkey Alex Straiker, U.C.S.D. San Diego,California Recent work (Straiker et al., 1999) has indicated the presence of a CB1 cannab- inoid receptor-based signaling system in the retinas of a wide variety of verte- brates. The other known cannabinoid receptor, CB2, is generally thought to be present in the immune system but not in the CNS, although a recent paper (Luet al., 2000) reported the presence of CB2 mRNA in retinas of the rat and the mouse. Using a subtype specific antibody to the CB2 cannabinoid receptor, I detected CB2-like cannabinoid receptor labeling in the retinas of the rat and the monkey. Labeling in the rat was highly consistent with previous in situ hybrid- ization studies demonstrating the presence of CB2 mRNA in all three nuclear layers of the rat retina. Double-labeling studies with calbindin and parvalbumin in rat retina demonstrated localization to specific cell types. This provides fur- ther evidence for CB2 receptor presence in the CNS and suggests that CB2 repre- sents part of a novel cannabinoid signaling system in the mammalian retina.

Characterization of Dopamine Neurotransmission in the Mesolimbic and Mesostriatal Pathways of Mutant Mice Lacking 5-HT2C Receptors O Dell L.E., *Tecott L.H., and Parsons L.H. The Scripps Research Institute, Department of Neuropharmacology, 10550 North Torrey Pines Road, La Jolla, CA 92037 *The University of California at San Francisco, Department of Psychiatry, 401 Parnassus Ave, San Francisco, CA 94143 Previous research suggests that serotonin2C (5-HT2C) receptors play an inhibi- tory role in mediating dopamine (DA) neurotransmission. Moreover, evidence suggests that 5-HT2C receptors exert a relatively greater influence on mesolimbic versus mesostriatal DA transmission. The present study compared phenotypic differences in DA neurotransmission in the terminal regions of these DA path- ways in wildtype and mutant mice lacking 5-HT2C receptors. Microdialysis probes were implanted into the nucleus accumbens (NAcc) and ipsilateral dorsal stria- tum. The next day, DA levels were monitored in both regions 1) under tonic

45 basal conditions using conventional dialysis sampling and the no-net-flux method 2) during placement into a novel open field and 3) following peripheral adminis- tration of cocaine (15 mg/kg). There were no phenotypic differences in DA lev- els during basal or no-net-flux conditions. Placement into the open field did not alter DA levels in the NAcc of either genotype. However, this manipulation pro- duced an elevation of DA levels in the dorsal striatum of wildtype mice that was attenuated in knockout mice. Cocaine produced a more robust elevation of DA levels in the NAcc of knockout mice relative to wildtype controls. However, there were no phenotypic differences in the dorsal striatum following cocaine admin- istration. The latter finding supports the hypothesis that 5-HT2C receptors play an inhibitory role in mediating DA neurotransmission, and that these sites exert a greater influence in the mesolimbic versus mesostriatal pathway.

Binding of an AMPA Receptor Potentiator ([3H]LY395153) to Native and Recombinant AMPA Receptors H. Yu1, A-M. Lind n1,2,3, H. Zarrinmayeh1, W. J. Wheeler1 and P. Skolnick1,2 1Lilly Research Laboratories, Lilly Corporate Center, DC 0510, Indianapolis, IN 46285; 2Department of Pharmacology & Toxicology, Indiana University School of Medicine, Indianapolis, IN 46202; 3A.I. Virtanen Institute, University of Kuopio, 70210 Kuopio, Finland Compounds that modulate AMPA receptor-mediated currents may be benefi- cial in a variety of neuropsychatric disorders. LY395153 is a member of a newly described class of arylpropylsulfonamide AMPA receptor potentiators. In elec- trophysiological assays, LY395153 is ≥ two orders of magnitude more potent than the prototypic AMPA receptor potentiator cyclothiazide. [3H]LY395153 binding was characterized in both native AMPA receptors from rat cerebral cor- tex and recombinant human GluR4flip receptors expressed in HEK293 cells. Glutamate and AMPA increased [3H]LY395153 binding to both native and re- combinant AMPA receptors in a concentration dependent and stereoselective manner. This effect of AMPA receptor agonists reflects an apparent increase in ligand affinity; in the presence of saturating concentrations of glutamate, [3H]LY395153 binding is saturable, with a significantly higher affinity at human GluR4flip (Kd = 55.6 ± 5.3 nM) than rat cortical receptors (Kd = 110 ± 15.1 nM). NBQX competitively inhibited glutamate-induced increases in [3H]LY395153 binding in both native and recombinant receptors, whilst LY303070 (the active isomer of GYKI53655) noncompetitively inhibited this effect in native, but not recombinant receptors. The prototypic AMPA receptor potentiator cyclothiazide competitively inhibited [3H]LY395153 binding with a potency (Ki ~7 mM) comparable to EC50 values reported in electrophysiologi- cal studies. In contrast, the structurally unrelated AMPA receptor potentiator CX 516 did not inhibit [3H]LY395153 binding at concentrations of up to 600 3 mM. Further, thiocyanate competitively inhibited [ H]LY395153 binding (Kb ~ 1.9 mM) at concentrations reported to facilitate AMPA receptor desensitization, but significantly lower than the concentrations (50-100 mM) routinely included

46 in [3H]AMPA binding assays to enhance the apparent affinity of this ligand. [3H]LY395153 binding was unaffected by a variety of structurally (and mecha- nistically) diverse compounds at 10 mM, including atenolol, baclofen, buspirone, caffeine, chlordiazepoxide, imipramine neomycin, phenylpropanolamine and rolipram. These data indicate [3H]LY395153 is a useful probe for labeling a unique modulatory site on both native and recombinant AMPA receptors.

Overactivation of AMPA and NMDA but Not Kainate Receptors Inhibits Phosphatidylcholine Synthesis before Excitotoxic Neuronal Death Ramon Trullas, Neurobiology Unit, IIBB/CSIC, Barcelona Spain Glutamate receptor overactivation induces excitotoxic neuronal death, but the contribution of glutamate receptor subtypes to this excitotoxicity is unclear. We have previously shown that excitotoxicity by NMDA receptor overactivation is associated with choline release and inhibition of phosphatidylcholine synthesis. We have now investigated whether the ability of non-NMDA ionotropic glutamate receptor subtypes to induce excitotoxicity is related with the ability to inhibit phosphatidylcholine synthesis. AMPA induced a concentration-dependent in- crease in extracellular choline and inhibited phosphatidylcholine synthesis when receptor desensitization was prevented. Kainate released choline and inhibited phosphatidylcholine synthesis by an action at AMPA receptors, because these effects of kainate were blocked by the AMPA receptor antagonist LY300164. Selective activation of kainate receptors failed to release choline, even when kainate receptor desensitization was prevented. The inhibition of phosphatidyl- choline synthesis evoked by activation of non-desensitizing AMPA receptors was followed by neuronal death. In contrast, specific kainate receptor activation, which did not inhibit phosphatidylcholine synthesis, did not produce neuronal death. Choline release and inhibition of phosphatidylcholine synthesis were in- duced by AMPA at non-densensitizing AMPA receptors well before excitotoxicity. These results show that AMPA, but not kainate, receptor overactivation induces excitotoxic cell death, and that this effect is associated with inhibition phosphati- dylcholine synthesis. Moreover, these results indicate that inhibition of phos- phatidylcholine synthesis is an early event of the excitotoxic process, downstream of glutamate receptor mediated Ca2+ overload, that may represent a specific target for neuroprotection.

47 Poster Session 3 • Thursday/Friday Sunshine Posters will be available for viewing from 3:30 PM Thursday to 4:30 PM Friday. Presenters will be with the posters on Thursday from 3:30 to 4:30 PM

Exercise That Alters Hippocampal-related Behavior and Metabolic Capacity in Motor Structures Does Not Alter Metabolic Capacity in the Hippocampus Brenda J. Anderson, Daniel P. McCloskey, David S. Adamo, and Pamela S. Coburn- Litvak, Dept of Psychology, SUNY-Stony Brook Exercise has numerous effects on the hippocampus. Hippocampal pyramidal cell discharge rates and theta rhythm correlate with speed of locomotion, and move- ment increases hippocampal glucose utilization. Exercise has been reported to increase hippocampal growth factor mRNAs and cell proliferation. In the first experiment, we tested whether or not exercise improves hippocampal-dependent behavior in a radial maze. Control rats and rats that voluntarily exercised for 3 months took the same average time to traverse an arm of the maze, but the exer- cising rats required significantly fewer trials to reach criterion-level performance. If exercise causes an increase in the level of sustained neural activity in the hip- pocampus, either directly or through a form of plasticity, then the hippocampus should exhibit an increase in the capacity to meet the additional metabolic de- mand. Alterations in behavior and sustained levels of neural activity are known to alter cytochrome oxidase (COX) activity, which is coupled to the production of ATP. Therefore, we tested whether or not exercise increases cytochrome oxi- dase (COX) reactivity after rats voluntarily exercised for 6 months with daily rates equivalent to those seen in the first experiment. Exercising rats relative to control rats had greater COX reactivity in limb representations of the motor cortex, and in limb, but not face, representations in the triatum. In contrast to our hypothesis, exercise did not affect COX reactivity in any region of the hip- pocampus. Voluntary exercise affected the rate of 8-arm radial maze acquisi- tion, and COX reactivity in motor structures, but did not increase the hippocampal capacity for oxidative metabolism.

Genetics and Age Alter Binge Drinking Induced Neurodegeneration F. T. Crews, Center for Alcohol Studies, University of North Carolina Human alcoholics are known to suffer from a loss of brain mass and function. Multiple day binge drinking is common among alcoholics as well as college stu- dents. Two factors that contribute risk to becoming alcoholic are genetics and age of onset of drinking. To determine if genetics and age contributed to binge drinking induced neurodegeneration we used a 4-day i.g. ethanol binge model in rats and identified neuronal damage using silver stain and other methods. Two inbred strains of rats, alcohol-preferring (P) and nonalcohol-preferring (NP), were used to study the relationship of a genetic predisposition for alcohol preference and neurodegeneration. Both P and NP rats had significant neurodegeneration in the olfactory bulbs, posterior perirhinal cortex, and posterior entorhinal cor-

48 tex following a 4-day ethanol treatment. P rats had significantly more neurodegeneration in the posterior perirhinal and posterior entorhinal cortices being 239% + 50% (p<0.02) and 219% + 46% (p<0.01) of NP, respectively. Im- munohistochemical studies showed COX2 and phospho-MAPK immunoreac- tivity consistent with glutamate signaling contributing to the differential damage. A component of the genetic factors that lead to risk of addiction could involve neurodegenerative responses. Heavy binge drinking in adolescents is becoming increasingly common. To determine if binge-induced neurodegeneration differs between adolescent (PN35) and adult (PN80-90) rats, the effects of a 4-day binge treatment were investigated. The olfactory bulbs were equally damaged in both groups. The anterior portions of the piriform and perirhinal cortices were dam- aged to a greater extent in adolescent rats. For example, adolescent rats showed 586 + 162 mm2 of argyrophilic area vs. 145 + 39 mm2 in the anterior piriform. The increased sensitivity of adolescents to ethanol induced neurodegeneration could contribute to the increased risk of adolescent drinkers becoming alcohol dependent. These findings indicate that both genetic and age differences contrib- ute to ethanol-induced neurodegeneration. (Supported by NIAAA).

Hibernation, a Model of Neuroprotection K. Drew, F. Zhou, X. Zhu, M.A Smith, R. Castellani Hibernation, as a natural model of tolerance to cer ebral ischemia , represents a state of pronounced fluctuation in cerebral blood flow where no brain damage occurs. Numerous neuroprotective aspects may contribute in concert to such tolerance. The purpose of this study was to determine if hibernating brain tissue is tolerant to penetrating brain injury modeled by insertion of microdialysis probes. Guide cannulae were surgically implanted in striatum of arctic ground squirrels before any of the animals began to hibernate. Microdialysis probes were then inserted in some animals after they entered hibernation and in others while they remained euthermic. Analysis of dialysate glutamate concentrations, collected at quantitative and non-quantitative flow rates, showed no difference between hibernating and euthermic animals. The brain tissue from hibernating and euth- ermic animals was examined 3 days after implantation of microdialysis probes. Tissue response, indicated by examination of H&E stained tissue sections and immunocytochemical identification of activated microglia, astrocytes and HO-1 immunoreactivity, was dramatically attenuated around probe tracks in hibernat- ing animals compared to euthermic controls. No difference in tissue response around guide cannulae was observed between groups. Further study of the mecha- nisms underlying neuroprotective aspects of hibernation may lead to novel thera- peutic strategies for stroke and traumatic brain injury.

Time Dependent Cue-induced Reinstatement during the First Month of Withdrawal from Cocaine J. W. Grimm, B. Hope, R. Wise, Y. Shaham Clinical research indicates that the probability of relapse to cocaine taking may become increasingly under the influence of conditioned stimuli over the course of drug withdrawal. Here we report a characterization of such a phenomenon in

49 a rat model wherein drug-paired cues provoke relapse to cocaine seeking during the first month of drug withdrawal. Rats were trained to press a lever for IV cocaine for 10 days (0.5 mg/kg/infusion, two 3-hr sessions/day). Each response on the drug-associated lever resulted in a cocaine infusion, and a presentation of a tone+light cue. The rats were then tested for reinstatement of cocaine seeking after 1, 2, 4, 7, 15, or 29 days of withdrawal (n=8-11 per group). Rats were given 60-min sessions (separated 5 min apart) during which lever presses were not re- inforced and the cue was absent (extinction). After a minimum of 6 sessions and after reaching a criterion of less than 15 responses/60 min on the cocaine associ- ated lever (rats reached criterion in 6-8 sessions), the effect of the cue on rein- statement was assessed in a session wherein lever presses resulted in cue delivery. Cue presentation resulted in a large increase in responding on the cocaine-asso- ciated lever after 15 or 29 days (p<0.05), but only a weak increase 1, 2, 4, or 7 days into withdrawal. Furthermore, at one day of withdrawal, the rate of non- reinforced lever pressing prior to the cue session was markedly lower than that observed after 7, 15, or 29 days of withdrawal from cocaine (p<0.05). This effect was replicated in a separate group using a within-subject design (n=6) where again, both extinction and cue responding at the 15 and 29 day time points were significantly greater than at the 1 day time point (p<0.05). The present findings provide an animal model in which the neural adaptations associated with time dependent increases in cocaine craving in abstinent cocaine addicts can be stud- ied.

The Role of Amino Acid Neurotransmission in the Acquisition and Expression of Conditioned Defeat K. L. Huhman and A. M. Jasnow, Dept of Pyschology, Georgia State University Conditioned defeat is a long-lasting and profound behavioral response of male Syrian hamsters following a brief defeat by a larger, more aggressive opponent. Following this initial defeat, hamsters fail to show normal territorial aggression, but instead display only submissive and defensive behaviors even though a smaller, non-aggressive opponent (NAO) is used for subsequent testing. It has been hy- pothesized that changes in the balance between glutamatergic and GABAergic neurotransmission in the amygdala are critical for regulating behavioral responses to anxiety provoking and aversive stimuli. In Experiment 1, we tested the hy- pothesis that increases in GABAergic neurotransmission in the amygdala would block the acquisition and expression of conditioned defeat. Adult male, Syrian hamsters were implanted with bilateral cannulae aimed at the central nucleus of the amygdala. After a one week recovery, hamsters were bilaterally injected with muscimol (0.0, 4.4 or 8.8 nM in 300 nl saline) either before training (the initial defeat by a more aggressive hamster) or before testing the following day with a smaller NAO. Muscimol dose-dependently and selectively reduced submissive and defensive behavior exhibited during testing when the drug was given either before training or before testing. This effect was not due to a general sedation caused by muscimol as other behaviors were not effected by the injections, and muscimol given before training did not alter the behavior of the aggressive trainer. The data indicate that an increase in GABAergic transmission in the amygdala

50 can block the acquisition and expression of conditioned defeat. In Experiment2, we tested the hypothesis that decreased glutamatergic neuro-transmission in the amygdala would block the acquisition of conditioned defeat. Hamsters were implanted with cannulae as in Experiment 1. Following recovery, the animals were injected with the glutamate (NMDA) antagonist AP5 (0.0, 2.5, 5.0 and 10.0 µg in 300 nl saline) before training. All doses of AP5 selectively reduced submissive/defensive behaviors exhibited during testing. The highest dose also decreased the amount of aggressive behavior shown by the trainer males toward the experimental males. Together, these data indicate that changes in amino acid neurotransmission in the amygdala can alter response to anxiety provoking and aversive stimuli in hamsters.

Lesions of the Amygdala Impair Sexual Behavior and Attenuate Levels of Extracellular Dopamine in the MPOA J. Dominguez, J.V. Riolo, Z. Xu, and E.M. Hull, Department of Psychology, SUNY Buffalo, Buffalo, NY, USA The medial preoptic area (MPOA) plays an important role in the regulation of male rats sexual behavior. Dopamine (DA) agonists or antagonists microinjected into the MPOA have been shown to facilitate or impair sexual behavior, respec- tively. Observations using microdialysis have shown an increase of extracellular DA in the MPOA of copulating animals. A major source of input to the MPOA is the medial amygdala (MeA). Lesions of the amygdala impair sexual behavior of male rats; microinjection of a dopamine agonist into the MPOA reverses this effect. The present experiment tested whether MeA lesions inhibit extracellular dopamine release in the MPOA of male rats, in addition to impairing copula- tion. Rats received either MeA lesions or sham lesions, and received guide can- nulae for microdialysis. Two weeks later microdialysis samples were collected and behavioral measures were obtained. Samples were later assayed using HPLC- EC. MeA lesions attenuated the increase of extracellular DA in the MPOA dur- ing exposure to an estrous female and during subsequent copulation. In addition, MeA lesions impaired measures of copulation, compared to shams. These data suggest that one means by which the MeA facilitates copulation in male rats is by increasing dopamine release in the MPOA in response to an estrous female and during copulation. Furthermore, these data confirm the close relationship between extracellular dopamine in the MPOA and copulatory ability in male rats. Supported by: NIMH grant#MH40826 to EMH

Life Without the Adrenaline Rush? Pain and Other Behaviors in Dopamine Beta-hydroxylase Knock-out Mice Luc Jasmin, Department of Neurological Surgery, University of California, San Francisco 505 Parnassus Avenue, Box 0112 San Francisco, CA94143-0112 Based on the available literature, it is unclear if a chronic lack of noradrenaline (NA) could be responsible for a state of hyperalgesia and decreased response to opiates.  To address these questions we have used mice in which the gene for dopamine beta-hydroxylase (DBH) has been deleted leading to complete lack

51 of NA. When compared with DBH +/+ mice (n = 6), DBH -/- mice (n = 6) showed clear hyperalgesia to heat and cold.  None of the animals in either group displayed signs of allodynia to innocuous heat or cold (42 and 20oC re- spectively) after 20 min.  Administering DOPS-Carbidopa (1 mg/g and 0.125 mg/g respectively) to restore noradrenaline in the CNS reestablished nor- mal withdrawal latencies in DBH -/- mice (p<0.01) while having no effect in the DBH +/+ mice. The antinociceptive effect of increasing doses of morphine (0.5, 2.5, 5, 10, 15, 20 and 25 mg/kg s.c.) was tested on the hot plate.  Mor- phine had a decreased potency in the DBH -/- mice since 3 times the dose of morphine was needed to reach the ED50 (25 mg in the DBH -/- mice vs. 7 mg in the DBH +/+ mice).  The antinociceptive effect of morphine was reversed by the opiate antagonist natrexone 10 mg/kg s.c. in both groups of mice. The finding of hyperalgesia in the DBH -/- mice is consistent with a previous sugges- tion that noradrenaline exerts a tonic inhibition of spinal nociceptive transmis- sion, and that decreasing noradrenaline or administration of noradrenergic antagonists results in behavioral hyperalgesia.  Increasing NA in normal animals, however, does not alter the baseline nociceptive threshold.  Also, these findings are consistent with previous reports that NA potentiates the antinociceptive effect of morphine.

Aging Induced Changes in Orexin a Neural Signaling C. Kotz, M. Mullett, A. Levine, C. Billington, Veterans Affairs Medical Center, Minneapolis MN Appetite and sleep disorders are common among the elderly, and may contribute to the high prevalence of anorexia in this population. Orexin A is a recently characterized neuropeptide that influences feeding behavior and plays a role in sleep disorders. Orexin A is predominant in the lateral hypothalamus (LH) and LH-injected orexin A dose-dependently stimulates feeding and activates neu- rons in several feeding-regulatory brain areas. In the current study, we tested the effect of LH-injected orexin A on feeding behavior and cfos-immunoreactivity (cFos-ir) in several brain regions in young and old male Fisher F/344 rats. We hypothesized that advancing age diminishes the feeding and cFos-ir response to orexin A. LH-cannulated rats at ages 3, 12 and 24 mo. were given graded doses of orexin A (0, .5, 1 and 2 nmol in 0.5 µl) in a repeated measures, counterbal- anced design. Food intake was measured at 1, 2 and 4 h following injection. For the cFos experiment, LH-cannulated rats at age 3 mo. and 24 mo. were injected with either artificial cerebrospinal fluid (aCSF) or orexin A (1 nmol/0.5 µl). One hour following injection, rats were perfused and brains processed for cFos im- munohistochemistry. The feeding response to orexin A was significantly dimin- ished and delayed in the 24-mo. rats, and the 24-mo. rats exhibited significantly attenuated or absent cFos-ir response to orexin in all brain regions examined. These data indicate that delayed and diminished feeding responses observed in the older animals may be the result of ineffective neural signaling. In summary, these results provide the first evidence of aging-induced decreases in functioning of orexin A neural signaling pathways, and may have implications for sleep and appetite disorders observed in the elderly.

52 Nucleus Accumbens Unit Activity in Rats Performing Radially Symmetric Tasks for Food Reward C.H. McCandless*; W.E. Skaggs, Depts Neuroscience and CNBC, Univ Pittsburgh, Pittsburgh, PA, USA The shell and core of the nucleus accumbens (NA) receive significant input from the hippocampal region (CA1 and subiculum). This input strongly affects NA activity and responses to prefrontal inputs (O Donnell & Grace, J.Neurosci. 15:5, 1995). Hippocampal cells exhibit spatially tuned activity, which suggests oppor- tunities for NA neurons to integrate positional with reward information. To in- vestigate this possibility, independently movable tetrodes were placed in NA core and shell in rats running for food reward on a circular platform. Rats were re- quired to contact any point at the edge of the platform before returning to the center for reward. This apparatus presents certain advantages over the radial maze. A variety of activity patterns were recorded. Many cells appeared to respond to the reward location, a pericentral radial zone, or in association with the edge of the platform. Most firing fields were radially symmetric. Apparently spatial ac- tivity was most often differential with respect to inbound vs. outbound trajecto- ries. A small subset of cells showed strong theta-frequency modulation, consistent with significant input from hippocampus. To account for the observed patterns, we hypothesize that NA neurons integrate information about context and behav- ioral state to encode the behavioral proximity of reward. The data support pre- vious suggestions that the NA participates in navigation by transducing the hippocampal place code into goal-relevant context. Data from a second experi- ment will also be presented which allows comparisons a) between internally- and externally-guided trajectories, b) with results obtained on a radial maze, c) between spatial and temporal determinants of firing, as well as correlations be- tween NA firing patterns and hippocampal EEG during relevant behavioral epi- sodes. Supported by: NINDS NS07433-02 and NSF 9720350

Prenatal Stress of Rats Causes Changes in Laterality of Adult Behaviors, but Only When Tested in Anxiogenic Situations S. E. Welcome, A. Garner, and W. O. McClure, Department of Biological Sciences and Keck School of Medicine, University of Southern California, Los Angeles 90089-2520 Prenatal stress causes both behavioral and neuroanatomical changes in the re- sulting adults. We have previously shown that a specific amount of stress applied to rats at a specific time of gestation causes neuroanatomical changes that mimic those seen in human schizophrenics. We now report that animals treated with prenatal stress also exhibit several behavioral changes, and that these behaviors are increased to statistical significance when the animals are tested under condi- tions of mild anxiety. Pregnant dams were treated with mild immobilization stress (90 sec) on days e11 — e14. Control animals were born to mothers that were not stressed. Litters were culled to 4 males and 4 females at post-partum day 3 (p3). Animals were tested from p13 until sacrifice at p105. A previous experiment suggested that both locomotion and rearing in the open field were

53 affected by the presence of other rats in the testing room. To confirm this obser- vation, we carried out 5 trials with all the other rats (40 animals) in the testing room, and 8 trials in which only the cage of 2 animals being tested was present. The presence or absence of conspecifics changed the results of testing. In the absence of other animals, rearing of the prenatally treated animals became later- alized: these animals preferentially used one paw or the other to support them- selves at the wall. Locomotion was also significantly increased by the absence of conspecifics. Neither effect was seen with control animals. To clarify the stimu- lus involved, animals were tested three times with only one cage at a time in the testing area while a tape of the noise of conspecifics was played. In all three trials noise of other animals completely inhibited both lateralization and increased locomotion of the treated animals. To test the hypothesis that anxiety was in- volved in these effects animals were tested for laterality in the presence of con- specifics using a step-down paradigm. Treated animals were not lateralized in control situations, but were lateralized after exposure to either shaking or re- peated intense auditory pulses. Control animals were never lateralized. These results suggest that anxiety is involved in generating the observed laterality. The increased laterality seen in prenatally stressed animals is highly reminiscent of the increased sinistrality reported in schizophrenics. It is possible that these pre- natally stressed animals will provide an useful model system for schizophrenia.

Behavioral Resetting of Circadian Rhythms Is Modulated by Caffeine: Role of Adenosine A1 Receptors R. E. Mistlberger, M. C. Antle, N. M Steen,Department of Psychology, Simon Fraser University, Burnaby BC V5A 1S6,Canada. Circadian rhythms in Syrian hamsters can be phase shifted by 2-4 h by proce- dures that stimulate arousal during the subjective day (the usual rest period of their daily rest-activity cycle). Phase resetting is thought to be mediated by neu- ropeptide Y (NPY) and possibly serotonin (5HT) inputs to the suprachiasmatic nucleus (SCN), the site of the circadian clock. Circadian rhythms can also be shifted by light exposure during the hamster s subjective night (active period), a response that is mediated by retinohypothalamic glutamate release. During the subjective day, light or glutamate can attenuate phase shifts induced by behav- ioral arousal, NPY or 5HT. Light-induced shifts early in the night are dependent on mobilization of intracellular Ca++ via ryanodine receptors within SCN cells. Caffeine is an agonist at this receptor, and mimics the phase shifting effects of glutamate in vitro. Caffeine can also potentiate glutamate release by blocking presynaptic adenosine A1 receptors, leading to the prediction that caffeine, a stimulant, might attenuate phase shifts induced by behavioral arousal. We found that caffeine (7.5, 37.5, 75 mg/kg, i.p.) did dose-dependently inhibit the phase shifting effects of either sleep-deprivation or wheel exercise in the mid-subjective day. However, the selective A1 receptor antagonist CPT (1.25, 6.25 mg/kg, i.p.) did not, suggesting that ryanodine receptors may mediate this action of caffeine.

54 The A1 agonist nCHA (1.25, 2.0, 2.5 mg/kg, i.p.), in the mid-subjective day induced dose-dependent phase shifts similar to arousal, but this may be a sec- ondary effect of the prolonged (up to 48 h) hypoactivity evident atthese doses. Caffeine, a widely used psychostimulant, has chronobiotic actions, but these ap- pear to be restricted to high dose effects at receptor sites to be determined.

Patients with Complex Regional Pain Syndrome (Reflex Sympathetic Dystrophy/Causalgia) after Unilateral Injury Can Have Bilateral Loss of Nociceptive Neurites in Skin Anne Louise Oaklander, Harvard Medical School Our understanding of chronic neuropathic pain is poor because of the paucity of objective findings. Nerve conduction studies reflect activity of large myeli- nated fibers, and electromyographic abnormalities are present only with motor damage. Patients with injury to unmyelinated and thinly myelinated fibers can have pain as their sole symptom and no abnormalities on exam or laboratory tests. Patients may be viewed with suspicion and have difficulty obtaining treat- ment and/or compensation. Patients with Complex Regional Pain Syndrome (CRPS, formerly known as Reflex Sympathetic Dystrophy or Causalgia) are es- pecially perplexing since their pain is excess for their injury, and they can report pain distant to (or even contralateral from) the injury site. Small punch skin biop- sies immunolabeled against PGP9.5 allow LM visualization of cutaneous sen- sory nerves and quantitation of epidermal nerve endings. These represent exclusively nociceptors (pain-sensing neurons). In an ongoing IRB-approved study, we compared the density of epidermal neurites in punch skin biopsies taken from the site of CRPS pain (P) on the arms, legs, or torso, an adjacent ipsilateral non-painful control site (I), the contralateral mirror-image homologue of the painful site (M) and the control site (C). To date, 11 consenting adults (3 with bilateral pain) and no risk for other peripheral nerve diseases have participated in this study with these results: 1. There was no statistical difference in density of epidermal neurites when comparing the ipsilateral (I) and contralateral control punches (C). 2. The average density of epidermal neurites at the painful site (P) was almost 40% less than the density at the ipsilateral control site (I). 3. The average density of epidermal innervation at the contralateral site mirror-image to the painful site (M) was almost equally reduced when compared to densities at the contralateral control site (C). These data suggest the following preliminary conclusions: 1. Punch skin biopsies can provide objective evidence of focal neu- ral damage in CRPS. 2. CRPS may be associated with damage to cutaneous nociceptive neurites. 3. Humans with unilateral injuries producing CRPS may have bilateral nerve damage. 4. Loss of cutaneous neurites contralateral to a CRPS-inducing injury is restricted to the area mirror-image to the painful area. 5. The contralateral site mirror-image to an injury should not be used as a con- trol site. Supported by the Beatrice and Roy Backus Foundation, the Paul Beeson Physician Faculty Scholarship in Aging Research, and the Ruth and Maurice Freeman Fund for Pain Research.

55 Topographic Map for Visual Space In Association Cortex of Behaving Monkey R.M. Siegel, R.E. Phinney, J.A. Turner, G. Jando Center for Molecular and , Rutgers University Classical studies of the primate inferior parietal lobule began with shrapnel inju- ries during World War I. Modern refinements using electrophysiological mea- surements revealed the cortical fields and the neuronal properties are crucial in constructing representations of surrounding visual space. Unlike striate and other extrastriate cortical regions, there has been almost no evidence from any mea- surements of single cells for a mapping of visual space across the inferior pari- etal lobule s subfields in the behaving monkey or human. Although attempts have been made to determine if there are organized topographic map of retinal or eye position across the inferior parietal lobule, the day-to-day variability in- herent in electrode positioning in a chronic monkey study over a typical two year course has no revealed a map. Such maps would be expected to serve as a basis for subsequent regions to plan and guide motor behaviors has been demonstrated. Intrinsic optical imaging, which uses changes in neuronal metabolism at a sub- millimeter scale, was used to assess neuronal tuning at thousands of locations across the cortex. The visually evoked optical signal time course in behaving monkey is comparable in size and amplitude to those in anesthetized monkeys, however it is superimposed on a second waveform arising from the execution of the task itself. We have found two maps in the association cortex. There is a functional architecture that has a reproducible gain field representing eye posi- tion. This map is confirmed with single electrode recording. The second repre- sentation of retinal position was variable over the two months of imaging. The mutability of the retinal topography across days may indicate modulation as a result of past experience, or present attentional and intentional states. Supported by the Whitehall Found. Foundation, NIH/NEI EY09223, NIH/NCRR 1S10RR12873 (RMS), NIH/NEI EY06738 (REP), NIH/NEI EY06738 (JAT), Hungarian Scientific Research Foundation OTKA/T023657 (GJ).

Investigation of Novel Neurotensin Analogs in Brain L. Warrington, Birdsall Research, Mayo Clinic Jacksonville FL Schizophrenia is a disease that affects about 1% of the world s population, and has devastating effects on patients and their families. Many theories exist regard- ing the etiology of schizophrenia. The most likely candidate neuropeptide with evident alteration in schizophrenia is neurotensin (NT), found in CNS and GI tissue primarily. It shows pharmacologic interactions with dopamine and in some brain regions colocalization and corelease. NT shares certain properties with antipsychotic drugs (APDs), especially the newer atypical APDs. Both NT and APDs cause increased dopamine turnover rate and blockade of apomorphine induced climbing behavior as well as amphetamine and cocaine induced loco- motor activity and rearing in rats. NT displays some differences from APDs. Certain APDs antagonize the stereotypic behavioral effects of dopamine ago- nists such as sniffing and licking while NT does not. Typical APDs cause cata-

56 lepsy in rats, NT does not. Overall, NT resembles atypical APD s. Numerous APDs exist and fall into two categories, older typical APDs and newer atypical APDs. Typical APDs suffer from debilitating side effects such as dystonia and tardive dyskinesia. Atypical APDs have fewer side effects but the most successful atypical, clozapine, can cause potentially fatal agranulocytosis. NT has been tried as an APD but was found to be quickly degraded by proteases in the periphery and must be given ICV. To combat this problem various analogs of NT, such as NT69 and NT77, have been developed in our laboratory with a high degree of stability in the periphery, and which can cross theblood brain barrier intact. Through the use of microdialysis in freely moving rats, we have found that NT69 shows a similar neurochemical profile to clozapine in striatum and prefrontal cortex, while NT77 shows a more similar profile to haloperidol in those same brain regions. It is the ultimate hope that we can develop a NT analog which maintains the beneficial effects of atypical APDs with fewer side effects.

Does Mild Cognitive Impairment Lead to Synapse Loss in the Hippocampal Dentate Gyrus? Stephen Scheff, Sanders-Brown Center on Aging, U. Kentucky, Lexington KY It is well documented that significant synapse loss occurs in many regions of the neocortex and hippocampus in Alzheimer s disease (AD), however it is unknown whether similar changes occur in patients with mild cognitive impairment with- out AD. The outer molecular layer of the hippocampal dentate gyrus receives a direct input from the ipsilateral entorhinal cortex, an area known to play an im- portant role in the progression of dementia. The present study was designed to assess the total number of synaptic contacts in this region of the hippocampal formation. Tissue was examined from individuals in the ongoing Religious Or- ders Study who underwent detailed clinical evaluation within 12 months of death and categorized as no cognitive impairment (NCI), mild cognitive impairment (MCI), or AD. Systematic random sections throughout the entire extent of the hippocampal formation were obtained at autopsy and processed for standard transmission electron microscopy. Unbiased stereological techniques employing the physical disector method were used to estimate the total number of synapses in the outer molecular layer of the hippocampal dentate gyrus. Results demon- strate a substantial loss of synapses in individuals with AD but no difference between individuals with MCI and normal aging controls. The cognitive decline observed in the MCI group may be the result of a more global loss in connectiv- ity and not localized to a single structure such as the hippocampus.

Emotional and Cognitive Responses to Mild and Strong Aversive Stimuli in Mice Lacking Dopamine D4 Receptors Marcelo Rubinstein, Instituto de Investigaciones en Ingenieria Genetica y Biologia Molecular, CONICET and University of Buenos Aires, ARGENTINA The frontal cortex receives a major dopaminergic input from the ventral tegmen- tal area that plays an important role in the integration of diverse neuronal signals that ultimately determine behavioral responses to novel environmental stimuli. Emotional and cognitive dimensions participating in these responses are depen-

57 dent on mesofrontal dopamine transmission through the stimulation of multiple receptor subtypes. The dopamine D4 receptor (D4R) is expressed at highest lev- els in the frontal cortex and is the predominant D2-like receptor localized in this brain area. To investigate the functional significance of the D4Rs in DA-medi- ated responses, and in the absence of pharmacological compounds with proven selectivity and efficacy in vivo, we have analyzed a strain of mutant mice that lack D4Rs (Drd4-/-) on a C57Bl/6J x 129sv/Ola F2 genetic background. We first challenged wild type, heterozygote and Drd4-/- in two different approach/ avoidance conflict paradigms. In the elevated plus maze the Drd4-/- mice dis- played increased levels of anxiety as demonstrated by a significantly reduced number of entries and time spent in the unprotected open arms of the maze. This behavior was completely reversed by ethanol and the benzodiazepine midazolam, two well-known anxiolytic agents that do not increase novelty seek- ing behavior. In addition, when compared to their wild-type littermates, Drd4-/ - mice exhibited significantly longer latencies with respect to their exploration of the bright, aversive compartment in a light/dark preference exploration test. Be- cause heightened anxiety may lead to an increase in ethanol consumption and the rewarding effects of ethanol depend on D2-like receptor stimulation we chal- lenged wild type and Drd4-/- mice in a two bottle choice preference test. No differences were observed in total EtOH consumption or preference between the two genotypes. To investigate whether another emotional and cognitive responses to stressful stimuli were affected in Drd4-/ mice we performed a fear condition- ing test using contextual and auditory cues paired with an aversive uncondi- tioned foot shock and a passive avoidance test with another cohort of mice. In both behavioral paradigms all parameters evaluated showed no differences be- tween wild type and Drd4-/- mice. Taken together these results suggest that the absence of D4Rs lead to hypervigilance under mild stressors but normal fearful responses to more aversive stimuli.

Divergent GABAA Receptor Mediated Synaptic Transmission in Genetically Seizure-prone and -resistant Rats M.O.Poulter1, B. Hutcheon2, K. Schwabe1 and D.C. McIntyre1 Neuroscience Research Inst., Carleton University, Ottawa Canada Recent evidence suggests that the abnormal expression of genes coding for GABAA receptors underlies the development of epilepsy. By analysing individual miniature inhibitory postsynaptic currents (mIPSCs) in the neurones of normal, seizure-prone and resistant rats we have found that mIPSCs decayed at different rats and had different amplitudes. These mIPSCS kinetics were correlated, as well, to cell morphology. In the seizure-prone rats, the mIPSC amplitudes were smaller (» 30 pA) than in normal (» 45 pA and seizure prone (»60 pA). These differences were independent of morphology. However, in non-pyramidal neurones of the seizure prone rat mIPSCs were significantly slower (» 400 %) that in normal or seizure resistant rats. In seizure-resistant rats, on the other hand, in comparison to normal rats the kinetics of mIPSC deactivation was faster in both pyramidal and non-pyramidal neurones alike. Despite substantial differ- ences in peak amplitude, the total charge transfer of mIPSCs did not differ be-

58 tween the strains. Thus the timing of the inhibition and not the total amount was different between the strains. These data suggest, therefore, that epilepsy is asso- ciated with a dysregulation of inhibitory timing, primarily on non-pyramidal neurones in the brain.

Synaptic Inhibition Generates Long Lasting Increases in Firing Rate of MVN Neurons Claudia M. Krispel, Alexandra B. Nelson,Chris Sekirnjak, Sascha du Lac, The Salk Institute, La Jolla Ca Medial vestibular nucleus (MVN) neurons fire spontaneously and stably in the absence of synaptic input. We investigated how the history of synaptic input affects spontaneous firing rate in MVN neurons recorded intra- and extracellu- larly in mouse brainstem slices. Following five minutes of high frequency stimu- lation of inhibitory synaptic inputs, the majority of recorded neurons demonstrate a substantial increase in spontaneous firing rate. This potentiation of firing rate appears stable and lasts in excess of thirty minutes. Substitution of intracellular current injections for synaptic inhibition also elicits a stable increase in firing rate. MVN neurons encode vestibular information with changes in firing rate and receive high frequency inhibitory synaptic inputs from several sources in vivo. Firing rate potentiation may have important physiological implications for compensation following lesions to the vestibular system or homeostatic regula- tion of spontaneous firing rates. While reported here for the first time in the vestibular system, we expect similar forms of firing rate plasticity to be present in other spontaneously active cell types of the CNS.

Carbon Monoxide Is an Endogenous Mediator of Neonatal Cerebral Circulation C.W. Leffler, H. Parfenova, J.S. Robinson, Z. Fan, M. Pourcyrous, R.A. Neff III, A. Nasjletti, R.A. Johnson, and A.L. Fedinec, Dept of Physiology, University of CO is a potentially important paracrine relaxing factor in the neonatal cerebral circulation. Heme oxygenase-2 (HO-2) is highly expressed in the cerebral mi- crovasculature of piglets and CO production by cerebral microvessels is high compared to that of other vessels. CO and heme-l-lysinate, applied topically, cause dilation of neonatal pig pial arterioles. The dilation to heme-l-lysinate, but not to CO, is blocked by chromium mesoporphyrin. Metal porphyrin inhibitors of HO attenuate vasodilations to topically applied glutamate, to bicucullin-in- duced seizures, and to hypoxia. The putatively CO dependent dilations to glutamate and glutaminergic seizures could involve endothelium because glutamate increases CO production by cerebromicrovascular endothelial cells, in vitro. CO induced dilation appears to involve KCa channels. In vivo, KCa channel inhibitors block cerebral vasodilations to CO and heme-l-lysinate. CO or heme-l-lysinate hyperpolarises cerebral microvascular smooth muscle cells in primary culture. The effect of heme-l-lysinate membrane potential, but not of CO, is blocked by chromium mesoporphyrin. Cerebral vasodilatory responses to CO involve interactions with both the prostanoid and NO systems. Either in-

59 domethacin or nitro-L-arginine (LNA) can block cerebral vasodilatory responses to heme-l-lysinate or to CO. Following indomethacin, topically applied iloprost restores dilation to CO and to heme-l-lysinate, but sodium nitroprusside does not. Following LNA, sodium nitroprusside restores vasodilation to CO, but iloprost does not. Both prostacyclin and NO act upstream of KCa channels be- cause neither iloprost nor NO can restore dilation to CO following KCa channel inhibition. CO appears to be important in regulation of neonatal cerebral micro- circulation and may contribute to coupling of cerebral blood flow to neuronal activity. Furthermore, prostacyclin and NO appear to play permissive enabling roles in CO-induced cerebral vasodilation in piglets.

Pharmacological Manipulation of Locus Coeruleus Tonic Output Alters Information Transfer Through Forebrain Sensory Circuits D. M. Devilbiss and B. D. Waterhouse Microdialysis studies have shown that extracellular levels of norepinephrine (NE) in the forebrain increase in direct proportion to increased tonic output from the nucleus locus coeruleus (LC). Tonic activity of NE-containing LC neurons is increased by systemic administration of idazoxan (alpha-2a antagonist; IDZ) and decreased by clonidine (alpha-2a agonist; CLON). Reports from our labora- tory as well as others have revealed a variety of actions of synaptically or iontophoretically released NE on individual forebrain sensory neurons. The goal of the present study was to pharmacologically manipulate the tonic output of the LC by systemic administration of IDZ or CLON and measure changes in the response properties of simultaneously recorded sensory neurons in terminal field regions of the LC efferent system. Spike train activity was recorded from mul- tiple, single neurons in whisker-related areas of the thalamus and cerebral cortex of awake, quietly resting male rats. A bipolar electrode implanted beneath the whisker pad was used to activate the sensory afferent path to thalamic and corti- cal neurons. Individual neuronal responses to somatosensory pathway activa- tion were monitored before, during, and after i.p. administration of saline, CLON (2 mg/kg), or IDZ (2 mg/kg). Thirty min. after CLON administration, stimulus- evoked activity of thalamic neurons was suppressed from control conditions, whereas responses of cortical neurons were either suppressed or enhanced. CLON administration also increased trial-to-trial variability of response latency for in- dividual neurons and increased spontaneous activity. By contrast, IDZ (30 min. inter-drug interval) administration either facilitated or suppressed stimulus-evoked discharge of individual neurons in both the thalamus and cortex. Additionally, IDZ reduced spontaneous discharge and reduced trial-to-trial variability in re- sponse latency. Our preliminary conclusion from these data is that pharmaco- logical manipulation of LC output produces corresponding changes in the responsiveness of individual thalamic and cortical neurons to peripheral sensory stimulation. Such changes in neuronal response properties are not uniform across LC-innervated cell populations and, thus, most likely reflect differential actions of synaptically released NE on a variety of postsynaptic receptor targets.

60 Micro-Infusion of NMDA into the Amygdala Induces Cocaine- Seeking Behavior Stanislav R. Vorel*, Eliot L. Gardner**, *Dept of Neuroscience, Albert Einstein College of Medicine, Bronx, NY 10461 **Intramural Research Program, NIDA, NIH, Baltimore, MD The reinstatement paradigm of cocaine-seeking behavior is an animal model of relapse to cocaine abuse. We addressed the question which neuroanatomical site(s) underlie cocaine-seeking by applying electrical brain stimulation. We found that electrical stimulation of the ventral subiculum (VSUB) of the hippocampal formation and the basolateral nucleus of the amygdala (AMY) induces cocaine- seeking behavior in anatomically selective and frequency-dependent fashion. We hypothesized that this effect of VSUB stimulation may depend upon the VSUB connection with the AMY. We therefore implanted an electrode in the VSUB, bilateral cannulae in the AMY and an intravenous catheter in rats(n=5). After recovery from surgery the rats were trained to self-administer cocaine (0.5mg/ kg/inf) in operant chambers equipped with two levers. Active lever presses re- sulted in cocaine delivery, inactive lever presses had no programmed conse- quence. After the acquisition of stable cocaine self-administration for 10 days the rats underwent a period of extinction during which saline was substituted for cocaine. When the extinction criterion was met (three consecutive sessions with no more than ten lever presses), the rats received a priming electrical burst stimu- lation of the VSUB (400uA, 200 pulses, 20 trains of 10 pulses at 20Hz, 200ms between trains) 15 minutes after micro-infusion of 0.5uL vehicle (VEH) or kynurenic acid (KYN)(10ug) bilaterally into the AMY. There was no statistically reliable difference in the number of reinstated lever presses after VEH or KYN treatment. However, when the rats met the extinction criterion again, micro- infusion of 0.5uL NMDA (0.25ug) bilaterally into the AMY reliably induced robust and selective cocaine-seeking behavior. We conclude that (1) glutamatergic neurotransmission in the AMY is not necessary for the reinstatement observed after VSUB electrical burst stimulation; (2)glutamatergic neurotransmission in the AMY is sufficient to induce cocaine-seeking behavior. This conclusion ex- tends our previous finding that electrical stimulation of the AMY induces co- caine-seeking behavior.

Don’t forget to visit the exhibit area.

61 Session Abstracts

Panel • Sunday • 4:30-6:30 PM • Mt. Werner Neurotrophic Factor-induced Dopaminergic Plasticity in Animal Models of Parkinson’s Disease and Drug Addiction Organizer: R.C. Pierce; Participants: K.B Seroogy, C.A. Altar, J.R. Taylor, C. Flores Neurotrophic factors are proteins that promote the development, differentiation, maintenance and survival of neurons. Whereas an overwhelming body of evi- dence indicates that neurotrophic factors play a critical role in neuronal survival during development, only recently were neurotrophic factors implicated in plas- ticity in the adult mammalian brain. This panel will focus on the influence of neurotrophic factors on mesotelencephalic dopaminergic systems. Kim Seroogy will outline the short- and long-term plasticity of brain-derived neurotrophic fac- tor (BDNF), neurotrophin-3, transforming growth factor-alpha, and their receptors in the dopaminergic nigrostriatal system following a neurotoxic insult. Tony Altar will review the numerous physiological effects of BDNF on dopaminergic neurons, including electrical, biochemical, behavioral and struc- tural changes. Jane Taylor will focus on the ability of BDNF and glial-derived neurotrophic factor to enhance the locomotor activating and conditioned reward- ing effects of cocaine. Cecilia Flores will discuss the role of basic fibroblast growth factor in amphetamine-induced behavioral sensitization. Collectively, these pre- sentations will outline the profound effects of several classes of neurotrophic factors on dopaminergic systems in the brain, which may play a role in the pro- gression of Parkinson s disease as well as the development of drug addiction.

Panel • Sunday • 4:30-6:30 PM • Storm Peak Cytoskeletal Restructuring in the Formation of Dendrites and Synapses Organizer: R.H. Lenox; Participants: S. Halpain, D. Benson, A. Matus Alterations in neural function in both the mature and developing nervous system often involve activity-dependent changes in neuronal morphology. Cytoskeletal proteins are integral to changes in actin-membrane plasticity and may constitute intriguing future targets for therapeutic interventions in neuropsychiatric diseases. While much as been learned in the past about axonal morphogenesis, the gen- eration of dendritic architecture is just beginning to be characterized. This ses- sion will focus on molecular aspects of the regulation of plasticity at the synapse. Robert Lenox will discuss the role of the actin and membrane-associated MARCKS protein in neuroplastic events in the developing and adult brain and as a target for psychotropic drugs. Shelley Halpain will present evidence for an expanded role of MAP2 in signal transduction at both the microtubule and actin

62 cytoskeletons. Deanna Benson will discuss the different roles of actin in emerg- ing versus established synapses. Andrew Matus will present evidence that den- dritic spines exhibit rapid actin-based motility that is altered by glutamate receptors and volatile anesthetics. Together these investigators will initiate debate and dis- cussion among audience members on the key molecular events affecting den- drite growth and synapse formation.

Panel • Sunday • 4:30-6:30 PM • Twilight The Orbitofrontal Cortex and the Addictions: Neuroimaging Studies Organizer: B. Adinoff; Participants: L. Porrino, R. Anton, E. London Approaches to the treatment of substance abuse have focused upon pharmaco- logical antagonism of drug-induced reward and craving, and ameliorating with- drawal and dysphoric states. More recently, interest in cognitive deficits as well as neuroimaging studies of ¥ substance abusers have highlighted deficits in the prefrontal lobe par ticularly the orbitofrontal cortex. This alternate model of addiction suggests that, in an addicted individual, there is an imbalance between the urge to consume drugs and the resistance to that behavior similar to aspects of obsessive compulsive disorders (OCD). This panel will discuss both primate and human neuroimaging studies exploring the orbitofrontal dysfunction in sub- stance use disorders and the relationship of these findings to impulse disorders and OCD. Linda Porrino will discuss the unique position of the orbitofrontal cortex as a site of long-term neuroadaptation following chronic stimulant self- administration. In the non-human primate, imaging data will be presented de- scribing the topography of the functional response in the orbital and medial prefrontal cortex during the initial, later, and withdrawal phases of cocaine self- administration. Ray Anton will present the Obsessive Compulsive Drinking Scale (OCDS) as a measure of craving and addiction severity in humans. Using fMRI, Anton has assessed brain activity during alcohol cue exposure, observing areas of activation similar to those implicated in OCD. Eydie London will discuss selective deficits on the Gambling Task, which is affected by the integrity of the orbitofrontal cortex, in cocaine addicted subjects. These subjects also show im- paired activation of orbitofrontal cortex using PET. Bryon Adinoff will discuss studies in cocaine addicted subjects. Following intravenous administration of the limbic-stimulant procaine, these subjects demonstrate increased rCBF in the orbitofrontal cortex compared to controls, suggesting hyperresponsivity of the orbitofrontal cortex following stimulation.

Panel • Sunday • 4:30-6:30 PM • Rainbow Rhythms and Patterns in Neural Firing Organizer: C.C. Chow; Participants: A. Bose, D. Terman, C.C. Chow, J. Rubin The brain is known to exhibit synchronous oscillations and spatiotemporal acti- vation patterns with correlates to behavior and cognition. These states are pre-

63 sumed to arise from the collective dynamics of interacting neurons. Several mod- eling approaches both theoretical and computational have been developed to unravel the neural mechanisms underlying these states. The approaches are gen- erally based on either networks of spiking neurons or averaged population rate models. Bose will begin by demonstrating how the output of small networks of neurons can be understood by identifying controlling elements. He will show how phase precession and the emergence of place fields of rat CA3 hippocampal pyramidal cells can be understood by determining how control of interneurons in CA3 changes during different behaviors. Terman will then explore the mecha- nisms behind the generation of synchronous oscillations in the basal ganglia in Parkinsonian patients. Insights from the model may help to resolve controversies surrounding the role of the indirect pathway of the basal ganglia in the genera- tion of resting tremor. Chow will follow with a model of localized self-sustained neuronal activity as seen in working memory. The state requires asynchronously firing neurons and synchrony may actually be used as a turn off mechanism. Rubin will round out the discussion with a model of the spontaneous formation of activity patterns in V1 that generate images in the visual field consistent with some of the common forms of geometric visual hallucinations. The model in- corporates biologically plausible architecture in which the connections between iso-orientation patches in V1 are locally isotropic but non-locally anisotropic. In the all the presentations, modeling yields new explanations about the functional relationships within the network responsible for the observed neural activity which were not evident from experiments alone. These efforts also suggest new sets of experiments to test proposed hypotheses.

Panel • Sunday • 4:30-6:30 PM • Skyline The Biological Clock: Of Molecular Gears, Timing Belts and Post-Transcriptional Modifications Organizer: R. Baler; Participants: D. Virshup, P. Lowrey, A. Keesler A complex transcriptional loop lies at the core of the molecular mechanisms that constitute a circadian master oscillator. Essentially, positive factors can turn on transcription of state variable genes whose products, once they reach a cer- tain threshold, can travel to the nucleus where they can modulate their own as well as their regulator s transcription. Variations on this theme have now been reported in a wide range of organisms, including cyanobacteria, arabidopsis, neurospora, drosophila and mice. Given the astonishing reproducibility of pe- riod length one should expect a carefully orchestrated series of regulatory events contributing to the production of precise, self sustained, cycles. Expectedly, re- search on the role of post-transcriptional regulatory events in molecular chronobiology is advancing rapidly. As a result, significant progress is being made on the effect ¥ of specific phosphorylation, guided translocation and degradation events in establishing the delay devices required for the emergence of a ~24-hr cycle cellular timekeeper.

64 Panel • Sunday • 4:30-6:30 PM • Sunset Evolution of Neurohypophyseal Peptide Systems: Why Sex Matters Organizer: T.R. Insel; Participants: M.S. Grober, F.L. Moore, L.J. Young, G.J. De Vries More than any other neurotransmitters/modulators, neuropeptides show strik- ing sex differences in their distribution. Prime examples are the mammalian neu- rohypophyseal peptides arginine vasopressin (AVP) and oxytocin (OT), which are among the most sexually dimorphic systems in the brain. Males of most mammalian species have more AVP cells in the bed nucleus of the stria terminalis and the medial nucleus of the amygdala and denser AVP fiber projections to the lateral septum than females. Consistent with these neuroanatomic sex differ- ences, AVP has been implicated in a series of male species-specific behaviors, such as territorial displays and paternal behavior. Recently, several studies have begun to explore the evolutionary origins of these gender differences. AVP and OT are the mammalian representatives of a family of nine amino acid peptides found throughout vertebrate evolution. Arginine vasotocin (AVT) is the ances- tral form of AVP; and isotocin (IT) in fish and mesotocin (MT) in tetrapods are the ancestral forms of OT. Matthew Grober has used a sex changing fish, the marine goby, to demonstrate the relationship between AVT cell size and func- tional gender differences (Neuroreport 7:2945-2949, 1996). Frank Moore pio- neered research on sex differences in AVT systems, using the rough skinned newt to explore the relationship between the number and distribution of AVT cells and the modulation of reproductive behavior (Gen and Comp Endo, 117: 281- 298, 2000). Larry Young has used a comparative approach in mammals to de- fine the molecular mechanisms for the diversity of OT and AVP effects, showing that natural variations in promoter structure may lead to different patterns of neuropeptide receptor gene expression (Nature, 400: 761, 1999). Geert De Vries was the first to describe sex differences in AVP pathways. His research on bipa- rental behavior in rodents (PNAS 91: 400, 1994) suggests that neural sex differ- ences may not only induce sex differences in behavior, they may compensate for physiological sex differences, and thereby avoid sex differences in behavior (Beh. Brain Res, 1998). Taken together these presentations will provide an evolution- ary perspective on the origin and function of sex differences in the brain. This panel, focused on neuroanatomical sex differences, was designed to complement a proposal from Elliott Albers on the evolution of AVT-AVP effects on social behavior.

65 Workshop • Sunday • 8:30-10:00 PM • Mt. Werner Role of Noradrenergic NTS Neurons in Drug Abuse: The Forgotten System Organizer: G. Aston-Jones; Participants: T. De Vries, Y. Shaham, G. Koob, C. O Brien The A2 norephinephrine (NE) cell group located in the nucleus tractus solitarius (NTS) provides a major noradrenergic innervation of several forebrain areas, including the amygdala, bed nucleus of the stria terminalis (BNST), nucleus accumbens and hypothalamus. Although NE is implicated in the abuse of sev- eral classes of drugs, and many of the targets of the A2 neurons are involved in drug abuse, this major NE system has received little attention until recently for its possible role in drug addiction. This workshop will discuss new results from several laboratories indicating a prominent role for A2 NE cells in opiate and stimulant abuse. Taco De Vries will present data showing that accumbens NE from the NTS is not regulated by autoreceptors, but is regulated by D1 and D2 receptors. Also, he will show that following repeated amphetamine administra- tion, accumbens, but not prefrontal cortex NE release (from the locus coeruleus, LC) becomes supersensitive to electrical stimulation. These results show impor- tant differences between the LC and A2 NE systems that may be relevant to drug abuse. Yavin Shaham will present recent data showing that low doses of alpha-2 adrenoceptor agonists attenuate stress-induced reinstatement of opiate or stimu- lant self-administration. These effects are not due to actions in the LC system, but instead are mimicked to some degree by lesions of the A2 fiber bundle in the ventral midbrain. George Koob will discuss corticotropin-releasing hormone (CRH)/NE interactions in the basal forebrain that contribute to negative emo- tional states associated with drug dependence. Prior observations and his lesion work indicate that this NE is from the A2 cell group. Finally, Chuck O Brien will discuss clinical studies in heroin and cocaine addicts where medications influ- encing the NE system have been used for relapse prevention. The beta-blocker propranolol has been used in controlled studies and the enhancer of NE trans- mission desipramine has been studied. The results of these trials will be related to the available data with laboratory animals.

Panel • Sunday • 8:30-10:00 PM • Storm Peak Dopaminergic Projections and Neural Ensembles in Prefrontal Cortical-Ventral Striatal Loops Organizer: P. O Donnell; Participants: A.A. Grace, L.L. Peoples, W. Schultz The prefrontal cortex and limbic striatal areas are involved a variety of func- tions, including attention, reward and cognition. These functions involve the activation of neural ensembles in those circuits, and require appropriate levels of dopamine receptor activation. How is information processing in these circuits modulated by dopamine? Are neural ensembles in the accumbens and prefrontal cortex driven or modulated by their dopamine inputs? Is there a correlation be-

66 tween dopamine cell firing and accumbens or prefrontal cortical neuron firing? How do such actions of dopamine translate in behavior? These and related ques- tions will be addressed in this panel. Patricio O Donnell will talk about the role of accumbens and PFC neuron membrane potential states in defining neuronal ensembles, as well as on synchronous activity between dopamine cells and accumbal or prefrontal neurons. Tony Grace will address cortico-subcortical in- teractions, specifically those involved in withdrawal from long-term DA agonists and recovery from DA depletion; these experiments reveal that the system is capable of undergoing modulation at a network level as a means of responding to long-term perturbations, which put the system into a new steady-state con- figuration. Laura Peoples will present data on accumbens neurons recorded from behaving animals, focusing on neuronal firing during instrumental behavior re- inforced by cocaine. Accumbens neurons exhibit changes in background firing and phasic changes correlated with specific events. Wolfram Schultz will talk about the relative roles of primate orbitofrontal and striatal neurons in the adap- tation of reward expectation during learning. Overall, the panel will bring to- gether important issues focusing on the role of the simultaneous activity of DA, accumbens and cortical neurons on specific behaviors. In short, DA neurons participate in reinforcement mechanisms by defining ensembles of active PFC and accumbens neurons. As a consequence, a particular behavioral context may be associated with relative enhancement of excitability of neural ensemble ap- propriate to that context.

Panel • Sunday • 8:30-10:00 PM • Twilight Glial Involvement in Neurodegeneration: The Revenge of Unintended Consequences Organizer: S. Griffin; Participants: S. Griffin, S. Barger, L. Van Eldik, J. Schwartz It is becoming increasingly clear that glial-neuronal interactions that evolved to promote functionality and survival of neurons in the short-term can ultimately contribute to neuronal dysfunction and death when they become chronic. In many cases, the primary insult impacts on the neurons, made vulnerable by the extremes of their metabolic and physical attributes. Glia respond in ways that aid immediate survival and regenerative attempts. However, repetitive activation or unchecked persistence of such responses may have consequences detrimental to neuronal function and viability. The studies presented by this panel will ad- dress the mechanisms responsible for such neurodegenerative consequences. Using the molecular components of Alzheimer s disease as a framework, Sue Griffin will introduce the theme with examples of stimuli that initiate the glial response. Joan Schwartz will follow with data from animal and in vitro models demon- strating the neuroprotective contributions of cytokines and glia-derived neu- rotrophic factors. Linda Van Eldik will discuss molecular events involved in the autoregulation of glial reactions, including autocrine feedback loops. Steve Barger will discuss neurodegenerative sequelae propagated of activated glial cells, in- cluding the ability of microglia to both release and respond to excitotoxins in

67 ways that interact with cellular antioxidant defenses. Finally, Dr. Griffin will draw from these data sets to illustrate the key testable hypotheses of the overall concept of glial contribution to neurodegenerative cascades.

Panel • Sunday • 8:30-10:00 PM • Rainbow Positive Modulation of AMPA Receptors: Mechanisms and Therapeutics Organizer: E. Nisenbaum; Participants: D. Bleakman, K. Partin, U. Staubli Glutamate ionotropic AMPA receptors are tetramers formed by the assembly of one or more distinct subunits (GluR1-4 or GluRA-D), yielding homomeric or heteromeric receptor-channels, respectively. Additional complexity among AMPA receptors is conferred by alternative splicing and RNA editing of each subunit. Although all AMPA receptors form cation selective channels, these differences give rise to a heterogeneous distribution of receptors in brain with diverse bio- physical and pharmacological properties. Recent data indicate that positive al- losteric modulators of AMPA receptors can dramatically enhance currents conducted through these channels. Moreover, the modulation of AMPA recep- tors may be restricted to specific subtypes using compounds that act preferen- tially on particular subunits or splice variants. In addition, electrophysiological experiments have shown that AMPA potentiators can augment glutamatergic synaptic responses and facilitate the induction of NMDA-dependent, long-term potentiation, suggesting that they may enhance memory formation. Consistent with this hypothesis, an accumulating body of clinical and experimental evi- dence indicates that positive modulators of AMPA receptors may be therapeuti- cally effective in the treatment of cognitive deficits in a variety of disorders. The goal of this panel is to provide an overview of positive modulators of AMPA receptors. K. Partin will review the AMPA receptor family and present her re- search investigating the molecular mechanisms of action of AMPA receptor potentiators in recombinant cell lines. E. Nisenbaum will discuss the activity of several classes of AMPA receptor potentiators at native receptors and their im- pact on synaptic transmission. U. Staubli will describe her studies demonstrating that AMPA potentiators can promote synaptic plasticity and enhance memory encoding. D. Bleakman will discuss the therapeutic implications of positive modu- lation of AMPA receptor function.

Panel • Sunday • 8:30-10:00 PM • Skyline New Teeth on an Old Saw: Evaluating Cholinergic Mechanisms in Brain Aging Organizer: D. Ingram; Participants: N. Greig, R. Perez-Polo, J. Joseph Since the late 1970 s the cholinergic hypothesis (ACh) has provided a useful av- enue for research on brain aging by elucidating mechanisms involved in the age- related decline in cognitive function as well as the neurodegeneration observed in Alzheimer s disease (AD). The three compounds used clinically for treating

68 AD (tacrine, donepezil and rivastigmine) are cholinesterase inhibitors produced directly from research supporting this hypothesis. Despite this therapeutic progress, many investigators and clinicians recognize the limited effectiveness of these treatments for AD and thus question the further utility of the ACh hypoth- esis. To address this issue, our panel will discuss new twists and turns for the hypothesis that argue for it s enduring viability within a multifactorial interactive system. To this end, D. Ingram will discuss the interaction of the ACh system with the glutamatergic (Glu) system and the possibilities for polypharmaceutical approaches for cognitive enhancement. Specifically, combined indirect stimula- tion of the ACh and Glu system synergistically enhances learning in aged rats compared to stimulation of either system separately. N. Grieg will describe a new class of acetyl-cholinesterase and butrylcholinesterase inhibitors that not only provide cognitive enhancement in rodent models but also reduce produc- tion of the amyloid precursor protein and its putatively toxic peptide, b-amyloid, implicated in the pathogenesis of AD. Interestingly, the mechanism for this ac- tion may be through non-ACh actions. R. Perez-Polo will expand the hypothesis to include the involvement of oxidative stress (OS) that increases the vulnerabil- ity of ACh neurons to neurodegeneration in AD compared to other neuronal types. A major factor is the NF-kB transcription factor, which is highly sensitive to OS, and can bind to sites on the promoters for both choline acetyltransferase and Bcl-x. Alterations in NFkB binding in the aging brain can promote commit- ment to apoptosis but also reduce inflammation. Finally, J. Joseph will also dis- cuss the involvement of OS in brain aging and its actions through the cholinergic system. Using COS7 cells as a model system, he will describe how there may be regionally selective vulnerability to OS that is the result of differences in OS sensitivity among muscarinic ACh receptor subtypes.

Panel • Sunday • 8:30-10:00 PM • Sunset Success Comes from Having the Right Connections: Integrating the CNS and PNS Organizer: D. Chambers; Participants: E. Bell and J. Begbie A primary goal of developmental neurobiologists is to elucidate the mechanisms underlying the initial specification of both the central and peripheral nervous systems (CNS/PNS), how these early events are further elaborated upon and the two systems subsequently integrated. Recent work has begun to shed light on both the developmental processes and molecular effectors responsible for many aspects of establishing and connecting neuronal circuitry. The CNS arises from the neural plate, which folds upon its anteroposterior (AP) axis to form the neu- ral tube. The imposition of AP identity on the neural tube is crucial for the later spatial ordering of differentiated neurons. David Chambers will present a brief review of the current understanding of how the nervous system is patterned along the AP axis. This will be followed by a presentation of the results of a cDNA screen designed to identify novel players involved in the specification of fore/mid and hindbrain territories. In particular, we will focus upon the identity

69 and functional role of some of the novel genes implicated in the specification of rhombomere 1 (r1) in the hindbrain, the developmental precursor of the adult cerebellum. Significant studies have demonstrated that genes implicated in re- gional patterning of the CNS not only establish pattern in the CNS by assigning neuronal identity, but also impart pattern to the periphery. Esther Bell will present evidence indicating that individual Hox gene action (Hoxb1) is sufficient for the specification of distinct motor neuron identity as well as establishing the appro- priate connectivity of motor axon projections between the hindbrain and its pe- ripheral targets. The molecular mechanisms involved in the formation of the PNS have until recently remained elusive. With the advent of new molecular markers and recombinant proteins significant advances have been made in the understanding of the induction of the cranial neurogenic placodes, source of cranial sensory neurons. Jo Begbie will firstly describe the molecular nature of the induction of the epibranchial placodes. This will be followed by recent data that has cast light upon the mechanisms involved in the delamination and guid- ance of neuroblasts to the ganglion, and their subsequent innervation of appro- priate hindbrain entry point. Together, this panel will draw together recent advances in our knowledge of the molecular events driving the specification and integration of the embryonic CNS and PNS.

Panel • Monday • 7:30-9:30 AM • Mt. Werner Integrated Regulation of Synaptic Transmission by Ca++ Channels, K+ Channels, And Mitochondrial Ca++ Uptake Organizer: W. A. Catterall; Participants: I. Forsythe, L. Kaczmarek, T. Scheuer, D. Lipscombe Neurotransmitter release is critically dependent on the Ca++ transient induced by propagation into the presynaptic nerve terminal, having third power dependence on Ca++ entry. The level of Ca++ achieved in presynaptic terminals is influenced by the duration of the action potential, the extent of acti- vation of presynaptic Ca++ channels, and the activity of intracellular organelles such as the endoplasmic reticulum and mitochondria that actively accumulate Ca++. This panel will consider the integrated regulation of Ca++ levels and syn- aptic transmission by each of these processes. Catterall will serve as chair and will give a brief introduction to the structure of Ca++ channels and their role in synaptic transmission. Forsythe will describe recent work on synaptic transmis- sion at the calyx of Held in the auditory system, a synapse in the medial nucleus of the trapezoid body containing a large presynaptic terminal that is amenable to voltage clamp analysis. He will present recordings of presynaptic Ca++ currents and correlate them with efficacy of synaptic transmission as a function of the pattern of stimulation and changes in intracellular Ca++ concentration. He will also describe the effects of mitochondrial Ca++ uptake on calcium dynamics and synaptic transmission in this specialized synapse. Kaczmarek will present studies of voltage-gated K+ channels in the calyx of Held. He will describe regu-

70 lation of the high threshold Kv3.1 potassium channel and how phosphorylation of these channels by casein kinase 2 and protein kinase C may be necessary for the precise timing of transmitter release at different stimulus frequencies, as re- quired for correct processing of sensory information in the auditory system. Scheuer will describe the regulation of cloned and expressed P/Q-type Ca++ channels, the Ca++ channel type present in the calyx of Held. He will show that these Ca++ channels are regulated by interactions with SNARE proteins, G pro- tein bg subunits, and Ca++. Ca++ entering through these channels interacts with calmodulin and binds to a site in the C-terminal domain, causing facilitation and then inactivation as a function of stimulus duration and frequency. This mecha- nism may contribute to paired-pulse and post-tetanic facilitation at synapses. Lipscombe will show that the function of presynaptic Ca++ channels is also regulated by alternative splicing of their mRNAs. Alternative splicing of N-type Ca++ channels at multiple sites alters the voltage dependence and kinetics of their gating, providing an additional mechanism of functional diversity that is essential for signaling in complex nervous systems of higher organisms.

Panel • Monday • 7:30-9:30 AM • Storm Peak Group I Metabotropic Glutamate Receptors: Role in Pain, Epilepsy, Movement Disorders, and Brain Injury. Organizer: R. Schwarz; Participants: J. Conn, A. Faden, A. Kingston, and. C. Woolf The eight G-protein-coupled metabotropic glutamate receptors (mGluRs) are grouped into three families based upon similarities in sequence homology and signal transduction pathways. Group I receptors(mGluRs 1 and 5) are positively coupled to phospholipase C and their activation results in the mobilization of intracellular Ca++ and the production of diacylglycerol. In contrast, Group II (mGluRs 2 and 3)and Group III (mGluRs 4,6,7,and 8) are negatively coupled to adenyl cyclase. Experimental results, which include the use of newly identified subtype selective compounds, have shown that Group I receptors play a critical role in a variety of physiological processes, such as pain processing, memory and learning, motor function, seizures, and neurodegeneration following brain injury. Following a brief introduction to mGluR nomenclature and receptor char- acterization by Roy Schwarz, Clifford Woolf will discuss the role of Group I receptors within spinal pathways involved in pain perception and their function in various types of pain. Jeffrey Conn will focus on the localization of mGluRs within basal ganglia pathways and present data on their activity in the control of motor function and potential therapeutic utility in movement disorders. Alan Faden will present results showing that Group I mGluRs are involved in excitotoxic cell injury/death with differential effects being observed on the pro- cesses of necrosis and apoptosis. Ann Kingston will describe the pharmacology of potent and selective Group I agonists/antagonists and demonstrate their ac- tivity in various in vitro and in vivo models.

71 Panel • Monday • 7:30-9:30 AM • Twilight Exactly How is Dopamine Involved in Methamphetamine Neurotoxicity? Organizers: T. Hastings, D. Sulzer; Participants: R. Gainetdinov, G. Ricaurte, T. Hastings, D. Sulzer Methamphetamine (METH), is both a widely abused psychostimulant and a neurotoxin that produces selective degeneration of dopamine neurites in culture and in vivo. While glutamate and temperature modulate METH neurotoxicity, the initial step appears to be due to oxidative stress following oxidation of dopam- ine. Roles have been proposed for 1) formation of extracellular oxyradicals by dopamine released by METH or 2) cytosolic oxyradical stress following dopam- ine redistribution from synaptic vesicles. Raul Gainetdinov will discuss the roles of the dopamine uptake transporter (DAT) and the central vesicular monoamine transporter (VMAT2) in METH toxicity as determined from results with knock- out mice. George Ricaurte will also address roles for DAT and extracellular oxyradicals in rodent and primate models. Teresa Hastings will address the spe- cific neurotransmitter pools involved and the likely toxic downstream products of dopamine oxidation such as quinones and cysteinyl-dopamine, as well as the potential protein modifications that may lead to neurite loss. David Sulzer will demonstrate how postnatal neuronal cultures can be used to demonstrate intrac- ellular oxidation of dopamine, the differentiation of synaptic vesicle, cytosolic, and extracellular pools, and for elucidation of the role of VMAT2 sequestration of dopamine in axonal degeneration.

Panel • Monday • 7:30-9:30 AM • Rainbow The HPA Axis: A Paradigm for Both Short and Long Term Plasticity in CNS Responsiveness Organizer: G. Aguilera; Participants: J. Herman, S.L. Lightman, G. Aguilera, J. A. Cidlowski Glucocorticoids, the end hormone of the hypothalamic pituitary adrenal (HPA) axis, influence behavior, metabolism, growth, reproductive and immune func- tions, and precise regulation of their secretion is critical for homeostasis spe- cially under stress conditions. Regulation of the system involves the hypothalamic peptides, corticotropin releasing hormone (CRH) and vasopressin (VP), and an entire range of genomic and secretory changes through complex interactions among receptors, signaling systems and transcription factors. This panel will explore recent advances in the mechanisms underlying the plasticity of the HPA axis responses during adaptation to different types of chronic stress. This in- cludes analysis of how utilization of different neural pathways and regulators, and the interaction of multiple cellular signaling systems can modulate the ex- pression of components of the HPA axis and alter the sensitivity to glucocorti- coid feedback. After a brief overview of neural pathways, neuropeptides and

72 neurotransmitters activated by different stressors, James Herman will present studies showing biphasic changes in HPA axis activity after middle age in rats, which correlate with changes in GABA biosynthesis in stress-regulatory path- ways. Stafford Lightman will describe how specific subgroups of serotonergic neurons are activated during stress and can be sensitized by repeated stress. These neurons are especially responsive to CRH and seem to represent an important pathway for the mediation of the HPA response to stress. Greti Aguilera will show that regulation of pituitary receptors for CRH and VP play a critical role determining ACTH responses, and will demonstrate mechanisms by which the interactive actions of CRH, VP and glucocorticoids regulate levels of these re- ceptors at transcriptional and posttranscriptional sites. Finally, John Cidlowski will discuss the molecular mechanisms by which the glucocorticoid receptor re- presses gene expression, and how signaling pathways induced by activation of plasma membrane receptors could modulate feedback inhibition by glucocorti- coids during stress. Discussion of these topics will provide new insight on the multiple mechanisms determining plasticity of the HPA axis during stress and aging, as well as opening novel perspectives for the development of diagnostic and therapeutical tools for stress related disorders.

Panel • Monday • 7:30-9:30 AM • Skyline Immune Regulation, CNS Pathogenesis and Therapeutic Interventions in Multiple Sclerosis Organizer: R. Jones; Participants: G. Konat, P. Dore-Duffy, H. Offner, A. Vandenbark T lymphocyte-mediated immune responses directed against myelin antigens cause paralytic inflammation and demyelination in the CNS. Animal models have been used to elucidate many of the molecular and cellular mechanisms of pathogen- esis and this understanding has in turn facilitated a rationale approach toward development and testing of experimental treatments. This panel will present ex- perimental results demonstrating mechanisms of tissue damage, cellular and immune processes regulating the progression of disease, and potentially effective treatment strategies based on our understanding of these mechanisms. Richard Jones will describe studies demonstrating a direct relationship between cell-me- diated immune responses to non-neural antigens and susceptibility to T cell-me- diated CNS inflammation. Gregory Konat will present his current findings regarding molecular and cellular mechanisms of oxidative damage to the oligo- dendrocyte-myelin unit during CNS inflammation, and their implications for loss of function and potential treatments. Paula Dore-Duffy will discuss the role of the microvascular pericyte as it pertains to blood brain barrier function, im- mune regulation and therapeutic responses. Arthur Vandenbark will describe TCR therapy in MS patients and mechanisms of immune regulation involving TCR-specific T cells. Halina Offner will present experimental results demon- strating that estrogen potentiates the clinical benefit of the TCR vaccine in EAE, and the implications of these results for treating MS.

73 Panel • Monday • 7:30-9:30 AM • Sunset The Raphe and Breathing: What Is the Connection? Organizer: G. Richerson; Participants: E. Nattie, L. Kubin, A. Berger, J. Ramirez The raphe nuclei are the primary source of serotonin in the brain, and project to all major respiratory nuclei. The neurotransmitters of the raphe (serotonin, TRH and substance P) can each stimulate breathing. The specific role of the raphe in modulating respiratory output is the subject of active research whose implica- tions extend beyond the issues specific for the respiratory system. This panel will discuss the roles that the raphe play in regulating respiratory output based on recent data at both the system and cellular levels. Eugene Nattie will describe the effect on ventilation of local changes in CO2 and pH within the raphe, and how this is modified by the sleep/wake cycle. Leszek Kubin will review the role of serotonin receptors in activation of upper airway motoneurons and the expres- sion profiles of different neurotransmitter receptor mRNAs in dissociated hypo- glossal motoneurons and other brainstem cells. Albert Berger will describe mechanisms of serotonergic modulation of hypoglossal motoneuron excitabil- ity, and will also focus on recently uncovered mechanisms by which glycinergic synaptic transmission is reduced by 5-HT and potentiated by ethanol. Nino Ramirez will discuss the role of raphe transmitters, including serotonin and sub- stance P, on generation of rhythmic bursting in the PreBotzinger complex in brain slices. He will describe recent data indicating that putative respiratory out- put generated in this region is dependent on ongoing release of serotonin from coexisting raphe neurons within the slice. Taken together, these data indicate that raphe neurons modulate ventilation at multiple sites and through a variety of mechanisms, and disruption of their activity may impair respiratory rhythmogenesis and weaken respiratory output. Thus, the raphe nucleus may be critical for production of normal respiratory output. The mechanisms by which the raphe modulate the respiratory system may also occur in other motor and sensory systems.

Panel • Monday • 4:30-6:30 PM • Mt. Werner New Vistas on Prefrontal Cortex: From Neuroanatomy to Computational Modeling Organizer: M. Banich; Participants: J. Giedd, A. Belger, M. Banich, R. O Reilly The goal of this panel is to provide an overview of new insights into the organi- zation and function of prefrontal cortex from its cellular organization to the nature of its computational processes. Traditionally prefrontal cortex has been considered to be critical for working memory, from which its role in executive function and psychopathology has been conceptualized to arise. In this panel we will provide evidence that the role of prefrontal cortex can be better thought of as playing a larger role in the control of behavior, providing a mechanism for integrating contextual information and selecting information that is critical for

74 task performance. To examine this issue, we will discuss converging evidence from different levels of investigation: neuroanatomy, neurophysiology, neuroimaging, and computational neuroscience. First, Jay Giedd will describe recent findings from neuroimaging that suggest continuing morphological devel- opment in prefrontal regions in humans during adolescence. He will discuss the implications of these findings for changes in behavior and cognition during this developmental time period. Next, Marie Banich will summarize the recent flood of functional neuroimaging studies focusing on working memory and executive function in humans that yield activation in prefrontal regions. She will provide a conceptualization of prefrontal function that integrates the role it plays in these processes. Third, Randy O Reilly will present an overview of computational models of prefrontal function. He will discuss the insights that can be provided by these models as to how the underlying biology of frontal cortex can give rise to the various functions to which it has been ascribed. Finally, Aysenil Belger will discuss evidence that modulation of NMDA receptor function in prefrontal cortex is critical for executive control processes. She will present evidence from both healthy individuals and individuals with schizophrenia.

Panel • Monday • 4:30-6:30 PM • Storm Peak Torsin, Parkin, Alpha-synuclein and Tau: Four Horsemen of the Basal Ganglia Organizer: D. Standaert; Participants: D. Standaert, M. Schlossmacher, D. Miller, M. Hutton in several proteins cause diseases of the basal ganglia, including dys- tonia, Parkinson s disease, and frontotemporal dementia with parkinsonism linked to chromosome 17 (FTDP-17). This Panel will review the current state of knowl- edge of the function and pathogenetic mechanisms of these proteins. David Standaert will discuss torsinA, a 332 amino acid protein encoded by the DYT1 gene. A 3 base pair deletion in DYT1 is responsible for autosomal dominant early onset torsion dystonia. The function of torsinA is ¥ unknown, but it has homology to the AAA family of ATPases, which serve as chaperones in pro- tein trafficking. The gene is highly expressed in dopaminergic neurons of the human substantia nigra pars compacta, suggesting that the mutation may pro- duce impairment of dopaminergic transmission. Michael Schlossmacher will discuss the protein parkin. Mutations affecting the 500 kb parkin gene have been identified as the cause of autosomal recessive parkinsonism in families of di- verse ethnic backgrounds. He will discuss recent efforts to elucidate the function of parkin, including studies of the expression pattern of parkin in rodent and human brains, and transfection experiments. Two different point mutations in the gene for the protein alpha-synuclein have been found to cause autosomal dominant Parkinson s disease. In addition, aggregates of alpha-synuclein are found in non-genetic Parkinson s disease, diffuse Lewy body disease, and multiple sys- tem atrophy. David Miller will review the biology of alpha-synuclein, and the expression of the gene in normal brain and neurodegeneration. Tau is a phos-

75 phoprotein that binds to microtubules. Michael Hutton will discuss the identifi- cation of missense and splicing mutations in tau associated with FTDP-17, which has lead to a re-evaluation of the role of tau in neurodegenerative disease. He will also discuss the current understanding of the mechanisms by which the mutations lead to disease and recent efforts to model neurofibrillary tangle for- mation in transgenic mice.

Panel • Monday • 4:30-6:30 PM • Twilight Multimodal Multidimensional Digital Developmental Atlas of the Mouse Brain Organizer: R.E. Jacobs; Participants: G. Burns, D. Laidlaw, R. Moats, A. Toga Gene expression patterns, receptor domains, arrays of innervation in the devel- oping nervous system, cell lineage patterns, and a host of other types of biologi- cal processes in embryonic and adult animals occur in three spatial and one temporal dimensions. The creation of a comprehensive framework to encom- pass diverse imaging and genetic information about the mouse holds tremen- dous promise for integrating genotype and phenotype throughout the animal s development. A digital atlas provides such a framework. Digital atlases provide a means to put specific data within the context of normal specimen anatomy, analyze the information in three (or more) dimensions, and examine relation- ships between different types of information. In this panel we discuss initial ef- forts at collecting data through several methodologies (blockface imaging, classical histology, CT, & MRI), amalgamating the various data structures into a coher- ent whole, annotating the 3D images (i.e. delineating anatomical structures), incorporating non-anatomical data from other formats (e.g. gene expression data), viewing the information, and disseminating the information. Arthur Toga will discuss blockface imaging and data integration issues. Gully Burns will review the organization and dissemination of complex neuroanatomical information via databases and the web. David Laidlaw will show ways of visualizing com- plex multidimensional information. Rex Moats will discuss uses and abuses of microCT in the context of the mouse atlas and Russell Jacobs will do the same for microMRI.

Panel • Monday • 4:30-6:30 PM • Rainbow “Death and the Lysosome”—New Insights into Neuronal Ceroid Lipofuscinoses Organizer: H. Geller; Participants: S. Hofmann, P. Lobel, D.Pearce, B. Davidson Neuronal ceroid lipofuscinoses (NCLs) are a group of neurodegenerative lyso- somal storage diseases that are invariably fatal during childhood, adolescence or early adulthood. Within the past several years, the molecular basis for several of these diseases have been identified using positional cloning methods as well as enzymological and proteomic approaches. Several genes encode lysosomal en- zymes while others encode membrane proteins of unknown function. The pre-

76 senters will provide an overview of the discovery of these genes, their functions, hypotheses about why loss of function leads to neurodegeneration and discuss potential treatment strategies. Sandy Hofmann will talk about the molecular patho- genesis of infantile NCL, which is caused by a deficiency in palmitoyl-protein thioesterase (PPT). Peter Lobel will discuss late infantile NCL and congenital ovine NCL, which are caused by deficiencies in the lysosomal proteases CLN2p/ tripeptidyl peptidase I and cathepsin D. Dave Pearce will discuss the molecular characterization of the of CLN3 gene deficient in juvenile NCL (Batten s Dis- ease). Finally, Bev Davidson will review novel gene and other therapeutic ap- proaches for restoring the function of these proteins in affected individuals.

Panel • Monday • 4:30-6:30 PM • Skyline See No Evil, Hear No Evil, Smell No Evil—The Origins of Head Placodes Organizer: M. Westerfield; Participants: H. Chu, R. Karlstrom, R. Grainger, M. Westerfield The majority of cells in the nervous system derive from the neural plate. In the head, however, specialized sets of cells, termed placodes, arise at the lateral edge of the anterior neural plate and form the ear, nose, lens and anterior pituitary. The origins of these placodes and the inductive mechanisms that lead their speci- fication and morphogenesis are largely unknown. This panel will compare and contrast the results of recent embryological and genetic experiments aimed at understanding how placodes arise. Rolf Karlstrom will present genetic analysis that implicates the Hedgehog signaling pathway in specification of the anterior pituitary and lens placodes. Monte Westerfield will describe lineage analysis that has revealed the morphogenetic movements that lead to formation of the olfac- tory placode. Rob Grainger will describe embryological experiments that have defined critical intermediate tissue interactions leading to lens determination and the onset of lens differentiation. We will also discuss several transcription factors that have been implicated in the early stages of nose (Westerfield), lens (Grainger) and ear (Hsin Chu) determination.

Panel • Monday • 4:30-6:30 AM • Sunset The Bane of Cancer Pain: A Role for Peptides and Protons? Organizer: G. Wilcox; Participants: G. Wilcox, A. Beitz, G. Davar, P. Reeh The difficulty in treating cancer pain, particularly in late stages of disease and after metastasis to bone and other tissues, contributes significantly to patient morbidity and may derive in large part from a paucity of knowledge about its basic mechanisms. Two tumor types in which pain is a significant problem, pros- tate and breast cancer, often metastasize to bone. In order to understand how these bony metastases might contribute to pain, an animal model for metastatic cancer pain has recently been developed. Osteolytic fibrosarcoma cells (NCTC clone 2472 cells derived from a spontaneous connective tissue tumor found in

77 C3H mice) implanted in the femur or around calcaneous or humerus bones of C3H mice promote osteoclastic reduction of bone mass and release cytokines including TNF-alpha, ET-1 and NGF. The reduction of bone mass and release of cytokines is accompanied by hyperalgesia and nociceptor sensitization. George Wilcox will introduce the behavioral and electrophysiological characteristics of the model. Al Beitz will discuss the neurochemical characterization of the model, including tissue dialysis for TNF, ET-1 and NGF and immunocytochemistry. Gudarz Davar will present evidence that exogenously applied ET-1, a potent vasoconstrictor and mitogen that is oversecreted by metastatic prostate and breast cancer cells, can selectively drive nociceptor activity in a receptor-dependent fash- ion. Finally, Peter Reeh will discuss the possibility that tumor growth along nerve sheaths and into nervous tissue may activate archaic transduction mechanisms present all along unmyelinated nerve fibers. Finally, the panel will discuss the possibility that reduced pH accompanying osteoclast activation in bone may in- voke acid-sensing mechanisms, mediated either by ASIC or VR-1, in C-fiber axons, releasing CGRP and causing neur ogenic inflammation of the nerve. We expect a lively debate over the nociceptive vs. neuropathic nature of cancer pain.

Workshop • Monday • 8:30-10:00 PM • Mt. Werner Gene Profiling in the Nervous System Using Nucleic Acid Microarrays Organizer: J. McGinty; Participants: G. Dent, K. Merchant, B. Meurers, G. Uhl High density nucleic acid microarrays provide the ability to monitor gene ex- pression profiles in a given tissue for thousands of genes simultaneously. In con- trast to other hybridization techniques that examine one selected gene at a time, microarrays offer the advantage of examining changes without any pre-existing bias or selection process based on the current state of knowledge. Thus, rather than a single or a handful of genes, expression profiles of thousands of gene en masse can be generated and compared between two experimental conditions. With several iterations of this process, a focus on a few critical genes involved in a specific physiological/ pathological process can be obtained. In this workshop we will discuss the potential, the advantages, and the disadvantages of different types of nucleic acid microarrays1,2 in their application to neuroscience research. Technical issues to be addressed will include the sensitivity and reliability of cDNA vs. oligo arrays, radioactivity vs. fluorescence, and bioinformatics. Dr. Kalpana Merchant will discuss her studies of genomic events underlying psychostimulant sensitization in the prefrontal cortex and neostriatum, which have uncovered a potential role of the ubiquitin-proteasome pathway. Dr. Gersham Dent will describe an anatomical dissection of the mouse amygdaloid complex into subregions followed by microarray analysis of subregion-specific gene ex- pression. Dr. George Uhl will discuss murine chip evaluations from single and multiple gene knockout mice and acute or chronic responses to psychostimulants and opiates. Dr. Bernd Meurers will discuss his research combining T7-based

78 RNA amplification with microarray technology applied to identified neuronal cell types isolated with laser capture microdissection, particularly gene expres- sion patterns in basal ganglia nuclei after dopamine depletion. Audience participation will be encouraged. 1. The Chipping Forecast. Nature Genetics suppl. 21:1-60, Jan 1999. 2. Serafini et al., Of neurons and gene chips, Current Opin. Neurobiol. 9:641- 644, 1999.

Workshop • Monday • 8:30-10:00 PM • Storm Peak Glutamatergic Mechanisms in the Pathophysiology of Drug Abuse Organizer: J. Weiner; Participants: F. Valenzuela, J. Kauer, O. Manzoni , A. Bonci Converging lines of evidence suggest that glutamatergic mechanisms may play a central role in mediating the reinforcing effects of a wide variety of addictive drugs. For example, within the nucleus accumbens (NAc), ethanol, cocaine, amphetamine, and morphine have all been shown to depress glutamatergic exci- tation of the primary GABAergic output neurons of this brain region. Recent evidence also suggests that chronic exposure to a number of drugs of abuse can result in long-lasting changes in glutamatergic synaptic transmission. The goals of this workshop are to bring together researchers that study a variety of drugs of abuse to stimulate a lively debate regarding a) the mechanisms through which this diverse array of addictive drugs impinge upon glutamatergic neurotransmis- sion and b) the relevance of glutamatergic mechanisms to the addictive proper- ties of these drugs. F. Valenzuela will start the workshop off by providing a brief review of the various receptor classes that make up the glutamate receptor fam- ily and will then discuss the long-term effects of ethanol exposure on glutamate receptor expression and function in cultured hippocampal neurons. J. Weiner will discuss the role of kainate receptors in the NAc and hippocampus and present evidence that ethanol can inhibit kainate receptor activity in these brain regions. J. Kauer will describe the effects of amphetamine on long-term depression of glutamatergic synapses in the ventral tegmental area. O. Manzoni will present anatomical evidence of the expression of CB-1 receptors on excitatory afferents to medium spiny neurons in the NAc and the functional consequences of acute and chronic activation of these receptors.

Panel • Monday • 8:30-10:00 PM • Twilight Genetic Variation in Human Catecholamine Systems: Impact on Neurobiology and Susceptibility to Psychiatric Disorders Organizer: D.R. Weinberger; Participants: M.F. Egan, D. Goldman, J. Cubells Although genetic variation accounts for a major portion of susceptibility to many neuropsychiatric disorders, finding specific genes has been problematic because of genetic and clinical heterogeneity. The recent development of high through-

79 put variant detection methods has led to the discovery of non-conservative and/ or functional variants in a number of candidate genes, such as those involved in catecholamine neurotransmission. A major clinical challenge is to determine how these variants affect human neurobiology and susceptibility to psychiatric disorders. David Goldman will begin by describing variant detection methods, recently discovered polymorphisms in genes related to catecholamine neurotrans- mission and results of association studies in several psychiatric disorders. Michael Egan will review methods for detecting genes associated with psychiatric disor- ders and how intermediate phenotypes can improve power. As an example, he will present data suggesting that a functional variant in the COMT gene affects prefrontal function and risk for schizophrenia. Joe Cubells will present evidence that functional variation at the dopamine beta hydroxylase (DBH) locus affects vulnerability to cocaine-induced paranoia. Finally, Daniel Weinberger will re- view recent neuroimaging data demonstrating the in vivo effects of the dopam- ine transporter and COMT on dopamine neurotransmission and prefrontal physiology, respectively. He will close by summarizing the pitfalls involved in going from genetic polymorphism to understanding variation in human neurobi- ology to clinical applications that could improve the lives of patients.

Workshop • Monday • 8:30-10:00 PM • Rainbow

5-HT2a Receptors and Forebrain Function: From the Cell to the Clinic Organizer: L. Parsons; Participants: G. Marek, B. Pehek, K. Cunningham, J.H. Krystal A wide variety of serotonin (5-HT) receptors are involved in cognitive process- ing. Among these particular attention has been focused on 5-HT2A receptors based on evidence that these receptors mediate the psychotomimetic effects of hallucinogenic drugs and are involved in both the pathogenesis and treatment of neuropsychiatric disorders such as schizophrenia. 5-HT2A receptors have been observed on GABAergic cells in many brain regions, cortical pyramidal neu- rons, midbrain mesolimbic and mesocortical dopamine neurons, and cholinergic projections of the pontine nuclei. Accordingly there are a wide variety of neural mechanisms through which 5-HT2A receptors may participate in psychiatric dysfunction. The goal of this workshop is to create an interactive discussion of the role played by 5-HT2A receptors in the regulation of forebrain function, with particular regard to thalamo-cortico-striatal circuits. A multidisciplinary frame- work will be presented to foster discussion. Gerard Marek will present and dis- cuss electrophysiological evidence that activation of 5-HT2A receptors located on thalamocortical terminals stimulates cortical glutamate release. Evidence that group II mGluR activation in turn suppresses transmitter release induced by 5- HT or other presynaptic releasing agents will also be discussed. Betsy Pehek will discuss in vivo neurochemical studies on the role of 5-HT2A receptors in the regulation of the mesocorticolimbic dopamine pathway along with correspond- ing immunohistochemical data on the localization of relevant 5-HT2A recep-

80 tors. Kathryn Cunningham will focus on the involvement of 5-HT2A receptors in the regulation of the discriminative stimulus and locomotor stimulant effects of cocaine but not the psychotropic amphetamine derivative MDMA. The pos- sible cellular mechanisms through which these effects are mediated will be dis- cussed with regard to immunohistochemical and immediate early gene data. John Krystal will provide a clinical perspective on the involvement of 5-HT2A recep- tors in the etiology and treatment of schizophrenia.

Panel • Monday • 8:30-10:00 PM • Skyline Free Radicals in Alzheimer’s Disease. Where Do They Come From? Organizer: S. Richardson; Participants: A. Butterfield, M. Smith, M. Vitek Free radicals, long suspected of causing the deterioration of the body s organs in the aging process, have been demonstrated to play active roles in various neurodegenerative conditions as well. Over the past 10 years or so, evidence of excessive free radical activity has been found not only in situations of acute brain damage such as cerebral ischemia and head trauma, but also in chronic progres- sive neurodegenerative conditions such as Alzheimer s disease. Among the many questions suggested by these observations are: What is the initial source of the excessive free radicals? What keeps the level of oxidative stress elevated for de- cades in the chronic neurodegenerative diseases? and Are free radicals the cause or the consequence (or both) of neurodegeneration? Previous panels and work- shops at WCBR have not focused on these specific aspects of oxidative stress in neurodegeneration. This panel, and the ensuing discussion, will contribute to answering these questions with respect to Alzheimer s disease. Allan Butterfield will discuss oxidative stress in Alzheimer s disease and the formation of free radicals by _-amyloid. Mark Smith will describe other sources of free radicals in Alzheimer s disease and the effects of free radicals on biological processes. Mike Vitek will present the effects of apolipoprotein-E isoforms in Alzheimer s dis- ease ¥ and their oler in controlling oxidative stress by regulating nitric oxide pro- duction. Steve Richardson will outline the neurotoxicity of, and the formation of free radicals by, a 5 amino acid sequence that is found in _-amyloid, and in proteins from three viruses associated with neurodegeneration: HIV-1, Japanese encephalitis virus and Newcastle disease virus. Each panelist will also address the issue of free radicals as the primary aggressor in, or merely the byproduct of, neurodegeneration in Alzheimer s disease.

81 Workshop • Monday • 8:30-10:00 PM • Sunset A Decade of Hits (and Misses): Heterologous Expression of Nicotinic Acetylcholine Receptors in Mammalian Cells. Organizer: R.J. Lukas; Participants: N.S. Millar, J.M. Lindstrom, R.J. Lukas, K.J. Kellar Nicotinic acetylcholine receptors (nAChR) play critical physiological roles throughout the brain and body. They also are targets for the tobacco alkaloid, nicotine, dependence on which promotes use of tobacco products. nAChR also are potential targets for a number of novel therapeutic agents for treatment of mood, cognitive, or sensory disorders. nAChR exist as a diverse family of mol- ecules assembled as different combinations of genetically-distinct subunits. Het- erologous expression of nAChR in Xenopus oocytes has been very useful toward fundamental characterization of diverse nAChR subtypes. This session aims to summarize successes and failures to date in stable, heterologous expression of nAChR in mammalian cells. It will also elaborate theoretical and technical con- siderations generally relevant to stable, heterologous expression of any kind of complex, transmembrane molecule and advantages or limitations of the tech- nique. Neil Millar will introduce the problem of selection of a cell host. Jon Lindstrom will discuss strategies that led to the first success in heterologous ex- pression of non-muscle nAChR in a mammalian cell. Ron Lukas will describe alternative strategies that led to success in stable, heterologous expression of nAChR as functional entities. Ken Kellar will outline approaches for character- ization of a variety of heterologously expressed nAChR subtypes. All partici- pants and the audience will discuss how vector and host cell choices might influence success or failure in expression. Potential roles played by chaperones, cell type-specific(?) patterns of membrane protein folding, sorting and traffick- ing, and metabolic regulation (e.g., calcium buffering) in successful expression will be elaborated. Findings from studies of wild-type and mutant nAChR that help to elucidate basic principles of ligand-gated ion channel structure and func- tion also will be presented.

Panel • Tuesday • 7:30-9:30 AM • Mt. Werner Dimerization of G Protein-coupled Receptors—Functional Implications Organizer: L. Devi; Participants: M. Bouvier, S. George, K. Mackie, Y. Patel The ubiquity and functional importance of G protein-coupled receptor (GPCR) dimerization (or oligomerization) has only recently become appreciated. Dimer- ization of GPCR s affect their signaling, impacting phenomena that are of inter- est to a broad range of neuroscientists. Dimerization with an identical receptor

82 (homodimerization) is a widespread and important in ligand binding and activa- tion of several GPCR s. Dimerization with non-identical GPCR s (heterodimerization) is also common. Heterodimerization gives rise to receptors with novel binding and signaling properties. Heterodimerization may explain some of the discrepancies between the behavior of native receptors in brain mem- branes and cloned receptors in expression systems. The aim of this panel is to discuss the mechanisms and implications of GPCR dimerization using mem- bers of several different GPCR families as examples. Lakshmi Devi will intro- duce the field and give a brief overview. Michel Bouvier will speak on assessing GPCR oligomerization in living cells. Ken Mackie will discuss the role of CB1 cannabinoid receptor dimerization in cell lines and brain. Susan George will present work relating to the dimerization of dopamine and serotonin receptors and its implication in structure, trafficking, and signaling. Yogesh Patel will de- scribe the consequences of heterodimerization between somatostatin and dopam- ine receptors.

Panel • Tuesday • 7:30-9:30 AM • Storm Peak Apoptotic Mechanisms: Are They also Used to Regulate Synaptic Stability? Organizer: E.A.Jonas; Participants: J.M. Hardwick, M.P. Charlton, J. Zimmerberg The molecular mechanisms that commit a cell to survive or to die in response to external signals include the activation of Bcl-2 family proteins. These proteins, acting either alone or in combination with endogenous mitochondrial channel proteins, are believed to alter the permeability of mitochondrial membranes. With some Bcl-2 family proteins, this permeability change leads to the release of cal- cium and cytochrome c from the mitochondria and subsequent cell death. In contrast, other members of the Bcl-2 family are able to prevent these events, and confer resistance to subsequent apoptotic signals. Neurons, however, differ from many other types of cells in that death may not be an all or none decision, but can be localized to specific synaptic connections. In particular, in the absence of trophic factors from the environment or from their postsynaptic targets, axon terminals may retract and effectively die, while other terminals from the same neurons can become stabilized. The very high density of mitochondria in synap- tic terminals suggests that these organelles may play a role in the decision of whether a terminal dies or survives. Marie Hardwick will introduce Bcl-2 family proteins and their molecular modes of action in neurons. Milton Charlton will discuss the factors that determine different patterns of synaptic calcium dynam- ics and their relevance to synaptic stability. Liz Jonas will present recent findings on the actions of Bcl-2 family proteins on mitochondrial membranes in intact presynaptic terminals. Finally, Josh Zimmerberg will discuss potential mecha- nisms for the formation of the ion channels and the a poptotic pore by Bcl-2 family proteins.

83 Panel • Tuesday • 7:30-9:30 AM • Twilight Cell and Gene Therapy for Basal Ganglia Disorders Organizer : O. Isacson; Participants: J. Kordower, C. Svendsen, K.S. Kim The selective degeneration of dopaminergic (DA) synapses and cell loss seen in the basal ganglia system can produce Parkinson s disease and other types of basal ganglia dysfunction, such as Huntington s disease. In attempts to restore neu- ronal function for these systems, gene therapy allows a targeted delivery of neuroprotective transmitter related genes to provide enhanced or normalized function. In the exceptional case, new synapses could be produced by regenera- tive simulation or by implantation of naive cells from embryonic or stem cell origin. The aim of this panel is to present recent data on gene therapy using neuroprotective agents such as GDNF or neuroimmunophilins and new oppor- tunities available through gene transcription control of neural transmitter genes in normalizing basal ganglia function. Recent findings on stem cell biology and the capacity of transplanted embryonic cells to develop into phenotypic mature neurons will be discussed. First, Ole Isacson will describe the various neuronal and ES cell types used for repairing the dopaminergic systems in models of Parkinson s disease. Second, Dr. Jeff Kordower will present his primate work on the DA system and GDNF lenti-virus gene therapy. Third, Clive Svendson speaks on embryonic cell development, including humans, and transplantation para- digms with relevance to basal ganglia function. Fourth, Kwang-Soo Kim will speak on transcriptional regulators for catecholaminergic function in the central nervous system, specifically the (TH) and DBH promoters and how these can be affected by Nurr 1, Phox 2A/2B, and PTX3. This panel will provide an overview of novel new cell and gene therapy efforts at the very basic science level with insight into the way basal ganglia plasticity can be modified.

Panel • Tuesday • 7:30-9:30 AM • Rainbow The Price of ‘Ecstasy’: The Functional Consequences of MDMA Neurotoxicity Organizer: G. A. Gudelsky; Participants: B. Yamamoto, G. Ricaurte, A. Fleckenstein, S. White The single or repeated administration of large doses of the amphetamine analog 3,4-methylenedioxymethamphetamine, MDMA or Ecstasy, produces a long-term depletion of the brain concentration of 5-HT, as well as the number of 5-HT uptake sites. The issue of whether these persistent neurochemical effects of MDMA reflect neurotoxicity remains controversial. Important to an assessment of the potential for MDMA-induced neurotoxicity is an examination of the long- term functional consequences of the repeated administration of this drug. The participants in this panel will discuss electrophysiological, biochemical, behav- ioral, and human cognitive and PET imaging studies that document abnormal

84 pharmacological and physiological responses following the repeated administra- tion of MDMA. The effects of MDMA on amine transporters will be discussed by Drs. Fleckenstein and Ricaurte. Dr. Ricaurte also will discuss the extent to which behavior and cognition are altered in human abusers of MDMA. Dr. Yamamoto will focus on the effects of the repeated administration of MDMA on the subsequent neurochemical responses to stress. Finally, the effects of MDMA administration on postsynaptic neurotransmitter function will be ad- dressed by Dr. White. Through a discussion of these data from studies involving diverse methodological approaches in animals and humans the participants will encourage a discussion of the potential risk (i.e., consequences) for human abus- ers of MDMA.

Panel • Tuesday • 7:30-9:30 AM • Skyline Epitope Spreading: A Novel Concept with Great Relevance to Multiple Sclerosis Organizer: M.C. Dal Canto; Participants: P. Lehmann, V. Tuohy, S. Miller Epitope spreading is characterized by widening of the immune response to epitopes distinct from, and non-cross-reactive with, the initiating epitope. This phenomenon has great implications for a variety of immune-mediated human diseases, including Multiple Sclerosis (MS). The goal of this panel is to present both human and animal studies on epitope spreading, to explore its mechanisms, and to suggest future therapeutic approaches. Paul Lehmann will explore whether activation of T cells against secondary epitopes takes place at the periphery or in the target organ in an autoimmune model of MS. In addition, he will discuss the role of microbial products, such as pertussis toxin, as important second signal links. Vincent Tuohy will report on human studies of MS and will illustrate the concomitant occurrence of two events, i.e., the spontaneous regression of the primary established autoimmune repertoire associated with disease onset, and the emergence of secondary T cell repertoires that recognize new self-antigens as disease progresses. He will point at the importance of this innate plasticity of self-recognition as an opportunity to develop customized immunotherapy in the future. Steve Miller will illustrate the phenomenon of epitope spreading in the best recognized viral model of MS, induced by Theiler s murine encephalomy- elitis virus (TMEV). He will show how a first phase of demyelination, produced by by-stander type mechanisms, driven by CD4+ lymphocytes specific for viral epitopes, is followed by a chronic phase, in which new T cell specificities against a number of myelin epitopes participate in the protracted destruction of myelin. Dal Canto will follow up on these studies by showing that these new T cell speci- ficities are functionally relevant as they are able to produce demyelination in an organotypic culture system. These studies are relevant to MS in the context of the widely held hypothesis of combined viral-immune-pathogenesis in this hu- man disease.

85 Panel • Tuesday • 7:30-9:30 AM • Sunset Fragile X Mental Retardation Syndrome: Clinical Diagnosis and Symptoms, Genetic and Molecular Mechanisms and Neural Correlates Organizer: W. Greenough; Participants: D. Nelson, R. Hagerman, E. Berry-Kravis (FXS) is the leading cause of inherited mental retardation, yet its details remain remarkably unknown among both clinical practitioners and basic scientists. In large part this reflects the recency of widespread knowledge of the karyotype and even more recent cloning of the gene and discovery of the nature of the genetic disorder. FXS involves an trinucleotide repeat in the regula- tory region of the FMR-1 gene which is expanded from a longer-than-normal repeat number in the carrier (premutation) female to a still longer repeat in the affected offspring. Similar triplet repeat expansions have recently been described in the etiology of a number of genetic disorders. The past few years have seen a dramatic explosion in our knowledge of the clinical characteristics, brain abnor- malities and molecular biology associated with FXS. There is reason to believe that FMRP, the protein encoded by FMR-1, which is synthesized near synapses, may play a fundamental role in synaptogenesis and patterning of connections in development and possibly also in adulthood. FMRP is an RNA-binding protein and it is thought that it may be involved in RNA transport and/or protein syn- thesis regulation. This panel brings together clinician-researchers and basic sci- entists familiar with human clinical symptoms, the molecular and cellular biology of FMRP expression and synaptic protein synthesis, the neuropathology of the syndrome, experimental gene therapy, and the characteristics of a model FMR- 1 knockout mouse. The session is intended to inform a general audience.

Panel • Tuesday • 4:30-6:30 PM • Mt. Werner Origin and Fate of New Neurons in the Adult Brain Organizer: H. Scharfman; Participants: A. Alvarez-Buylla, T. Palmer, J. Parent, H. Scharfman One of the most exciting areas of Neuroscience in recent years has been the rapidly growing field of adult . It is now fully recognized that new neurons are born in the adult nervous system. In addition, research continues to elucidate the various ways multipotent stem cells arise, migrate, and can be ma- nipulated experimentally. This panel will discuss the fundamental aspects of these processes and their potential applications to neurological disorders such as epi- lepsy and stroke. Each speaker will provide an overview of fundamental cell biological principles related to their work, followed by new data. Arturo Alvarez- Buylla will discuss the identification of stem cells in the adult subventricular zone and subgranular layer of the hippocampus. New data will be presented on the organization of these germinal layers and on the role of bone morphogenic proteins (BMPs), and the BMP inhibitor Noggin, in the formation of new neu-

86 rons. Theo Palmer will focus on the fate of stem cells that are taken from various parts of the nervous system, amplified in culture, and then returned to novel sites within the adult brain. These studies emphasize the plasticity of stem cells and their potential use to repair or modify diverse areas of the nervous system. Jack Parent will describe the effects of prolonged seizures or focal ischemia on neu- ronal precursor proliferation and migration in the adult rat forebrain subventricular zone-olfactory bulb system, and potential implications for brain repair strate- gies. Helen Scharfman will discuss physiological characteristics of granule cells that are born after pilocarpine-induced status epilepticus. Recordings of the newly- born cells in hippocampal slices show that they develop intrinsic properties that are indistinguishable from normal adult granule cells, but synchronize abnor- mally with area CA3 hippocampus pyramidal cells. The unexpected findings concerning the origin, migration, and differentiation of young neurons that have resulted from studies of neural stem cells and neurogenesis in the adult brain will provide an atmosphere of controversy intended to stimulated debate. We will also discuss the potential use of adult neural progenitors to treat various brain diseases.

Panel • Tuesday • 4:30-6:30 PM • Storm Peak New Horizons in the Mechanisms Mediating the Effects of Estrogen in the Brain: The Evidence for Membrane Receptors Organizer: J.B. Becker; Participants: C.S. Watson, D.M. Dorsa, M.J. Kelly, J.B. Becker The steroid hormone estrogen has been shown to have rapid effects on neuronal activity that cannot be explained by its actions to induce transcription by binding intracellularly to ERa or Er_. This panel will provide an overview of the evi- dence that estrogen can act on neurons and pituitary cells at the level of the plasma membrane to induce rapid, transient, changes in cellular activity. Cheryl Watson will provide evidence that the membrane receptor for estradiol is actu- ally ERa that is associated with the extracellular membrane. One effect of estro- gen that is thought to be mediated by this mechanism is the rapid release of prolactin from pituicytes. Dan Dorsa will discuss the mechanisms through which estrogen modulates activity in growth factor and cAMP signaling pathways. He will also present evidence that the known n uclear estrogen receptors, alpha and beta, can act at the cell membrane to initiate these effects. These effects of estrogen are thought to play a role in the neuroprotective effects of estrogen. Martin Kelly will discuss the effect of estrogen to modulate the activity of G- protein coupled receptors on hypothalamic neurons, and the potential role this plays in regulation of the reproductive cycle. Jill Becker will conclude with a discussion of the mechanisms through which estrogen affects neural activity in the striatum, and the implications of these findings, in general, for our under- standing of how steroid hormones act in the brain. These rapid, non-traditional effects of estrogen may play a role in reproductive function and behavior. They may also be involved in the effects of estrogen on cognitive processes, and for gender differences in responses to drugs of abuse.

87 Panel • Tuesday • 4:30-6:30 PM • Twilight Dopamine and Serotonin Transporters: Siblings Do Not Always Behave the Same Organizer: A. Frazer; Participants: A. Frazer, A. Galli, G. Gerhardt, M. Sonders The serotonin transporter (SERT) and dopamine transporter (DAT) are key cel- lular targets for several classes of psychoactive drugs, including antidepressants or psychostimulants. These transporters are members of the same gene family. Nevertheless, important differences exist in both their regulation and the confor- mational changes elicited by drug binding. This panel will illustrate such differ- ences. Aurelio Galli will present data, using multiple techniques, showing that exposure of HEK-293 cells stably transfected with human DAT to a gonists such as amphetamine causes internalization of this transporter with a conse- quent decrease in transporter capacity. The effects of amphetamine are not mim- icked by a DAT antagonist (cocaine); rather, cocaine blocks the effects of amphetamine. By contrast, Alan Frazer will show that chronic exposure of rats to SERT antagonists (sertraline, paroxetine) cause a time-dependent downregulation of SERT binding sites that is accompanied by decreased SERT function in vivo. Greg Gerhardt will show, using both chronoamperometry and microdialysis, that acute administration of either antagonists of the SERT or terminal autoreceptors for 5-HT have a much more modest effect on the clear- ance of serotonin (5-HT) than antagonists of either the DAT or terminal autoreceptors for dopamine (DA) do on the clearance of DA. Dr. Sonders has been studying structure-activity relationships of substrates for the cloned DAT and SERT proteins, employing both electrophysiological and biochemical tech- niques. He will report that even though these related transporters carry many of the same substrates (amphetamines, phenethylamines, tryptamines, MPP+), DAT and SERT have evolved different recognition criteria for substituent features of the solute molecules. Additionally, each transporter appears to have multiple functional modes for translocating various substrates.. All the speakers will em- phasize how such differences between DAT and SERT could have marked ef- fects on the functional consequences of psychoactive drug administration.

Panel • Tuesday • 4:30-6:30 PM • Rainbow Proteases in the Neurodegeneration of Alzheimer Disease (AD): Chickens or Eggs? Organizer: R. Nixon; Participants: R. Nixon, L-H. Tsai, K. Wang, C. Cotman The demise of neurons in (AD) is a protracted process that takes months or even years after the first cytoskeletal pathology appears. While few disagree that pro- teases mediate this degenerative process, the question of which hydrolases, if any, are the most attractive therapeutic targets is unanswered. Are neurons so

88 dysfunctional by the time they clog with tau aggregates that blocking execution of the agonal event(s) is counterproductive? Are there pathogenic proteolytic events, other than those generating A_ peptide, that gradually cripple neuronal function before setting off the proteolytic chain reaction that kills the cell? Like the chicken and egg riddle, the issue of which protease is activated first in trigger- ing neurodegeneration is complicated. In different forms of cell death including apoptosis, calpains, cathepsins or other proteases may become activated either upstream or downstream of caspase activation. Further complicating the analy- sis in AD is evidence that end stage proteolytic events may occur in one part of the neuron, such as the synapse, well before the rest of the cell is irreversibly compromised. Calpains, cathepsins and caspases are all activated during the neu- ropathological progression of AD. The aim of this panel is to review data impli- cating these proteases in neurodegeneration and in the early development of pathologic lesions in AD. Cause and effect in their activation will be considered as a springboard for discussing the proteases as therapeutic targets. Ralph Nixon will present the concept of AD as a disorder of activated proteases and focus on the contribution of early and progressive activation of the lysosomal system to AD pathogenesis in humans and animal models. Li-Huei Tsai will present data linking calpains to the neurotoxicity of A_ and other AD-related stresses via cdk5-induced alterations of the cytoskeleton. Kevin Wang will discuss the inter- play of calpains and caspases in different patterns of neurodegeneration and its relevance to AD pathogenesis. Carl Cotman will present evidence supporting the involvement of apoptotic mechanisms in AD and the particular contribution of caspases to the degenerative process.

Panel • Tuesday • 4:30-6:30 PM • Skyline The Fruit Fly, Drosophila Melanogaster: A Model System for the Study of Complex Behaviors Organizer: J. Hirsh; Participants: R.L. Davis, J.C. Hendricks, U. Heberlein, J. Hirsh This panel will present recent advances using the fruit fly, Drosophila melanogaster, as a model for studying complex behaviors. A common theme linking each of these systems is that responses are modulated by cAMP signal- ing, and in each case there is ample evidence showing that the phenomena under study show similarity with behaviors seen in higher vertebrates. Ron Davis will discuss advances in the analysis of genes, signaling pathways and brain struc- tures involved in learning and memory. Joan Hendricks will talk about the mo- lecular genetics of rest, a sleep-like state in flies, and how it is regulated by cAMP signaling via CREB proteins. Ulrike Heberlein will discuss the molecular genet- ics underlying responses to drugs of abuse, focussing primarily on alcohol. Jay Hirsh will discuss progress in dissecting the pathways regulating cocaine respon- siveness and sensitization to repeated cocaine exposures.

89 Panel • Tuesday • 4:30-6:30 PM • Sunset NGF and GDNF: The Yin and Yang of Unmyelinated Nociceptors Organizer: D. Wright; Participants: D. Molliver, C. Stucky, K. Albers Small unmyelinated primary sensory neurons that convey nociceptive informa- tion are uniquely tied to nerve growth factor (NGF) for their embryonic survival. Recent studies have established that nociceptive neurons undergo dramatic changes during early postnatal development. The goal of this panel is to provide new information about nociceptive subpopulations using anatomical, electro- physiological and molecular approaches. Derek Molliver will illustrate how no- ciceptive neurons differentiate postnatally into subpopulations that are distinct in their neurochemistry, anatomy, and neurotrophin-responsiveness. Dr. Molliver will demonstrate that a peptidergic nociceptive population maintains its reliance on nerve growth factor, while a separate non-peptidergic population undergoes a dynamic shift in their neurotrophic responsiveness from NGF to glial cell-line derived neurotrophic factor (GDNF). Recently, it has been suggested that both peptidergic and nonpeptidergic nociceptors play an important role in the estab- lishment of chronic pain states, but that peptidergic nociceptors are particularly important in inflammatory conditions and the nonpeptidergic nociceptors may be more important in neuropathic states. To address this issue, Dr. Cheryl Stucky will provide new information about the functional role(s) of peptidergic and nonpeptidergic nociceptors using both in vitro and in vivo electrophysiological approaches. Next, Drs. Kathryn Albers and Doug Wright will discuss the bio- logical actions of GDNF on non-peptidergic nociceptors. Using new transgenic mice that overexpress GDNF in the skin, Dr. Albers will discuss the effects of elevated levels of target-derived GDNF on nociceptive subpopulations. Finally, using a murine model of diabetic neuropathy, Dr. Wright will present work on the fate of nonpeptidergic nociceptors in diabetes and provide evidence that GDNF has therapeutic potential in treating neuropathic nonpeptidergic nociceptors.

Workshop • Tuesday • 8:30-10:00 PM • Mt. Werner Mechanisms of Neuronal Migration and Synapse Stabilization in Neocortical Development Organizer: S. Juliano; Participants: K. Nakajima, K. Herrup, A. Kriegstein Factors that underlie neuronal migration, the formation of neocortical layers, and subsequent intrinsic circuitry are important features of CNS development. Interruption of these processes leads to human disorders ranging from seizures to lissencephaly. Clearly, a range of molecules provide cues that direct neurons along their path, prompt them to g o or stop , and guide the normal laminar formation of neocortex. This workshop focuses on a limited number of such

90 instructive molecules, including those identified as a result of the reeler and Cdk5 deficient mouse, and their roles in neocortical layer formation and stabilization of laminar properties. The speakers will provide an overview of several mecha- nisms that contribute to successful or disrupted migration and stabilization of neocortical layers. Kazunori Nakajima has identified several alternate splicing isoforms of reelin that may have unique roles in brain development. These isoforms are truncated to exclude a repeat region that contains the binding sites for receptors. These and several other forms of reelin will be discussed in relation to their function in cortical development, including participation in migration and laminar formation. Sharon Juliano will present a model of disrupted brain development in which Cajal Retzius (CR) cells containing reelin are disordered and misplaced from layer 1 following interruption of mitosis due to toxicity early during corticogenesis. She will discuss whether the reelin positive CR cells can result in abnormal cortical migration patterns due aberrant signals from unusu- ally placed reelin containing cells in the cortical plate and intermediate zone. Karl Herrup will discuss the significance of a different protein, the Cdk5 protein, in neuronal migration into the cerebral cortex. He reports that layer 1 and layer 6 neurons do not require Cdk5, nor do the GABAergic interneurons that migrate from the ganglionic eminence. A different set of neurons that reside in layers 2-5, however, are incapable of populating cortical plate in the absence of this kinase, suggesting its importance in guiding a subset of neurons into the neocortex. Arnold Kriegstein has identified a widespread, transient, GABAergic network in layer 1 during cortical development. Neurites of intrinsic GABAergic cortical neurons, including a subset of CR cells and neurogliaform cells potentially con- tribute to this plexus, but the primary source is a projection from the zona incerta of the ventral thalamus. Studies of the developing neocortex indicate a role for membrane depolarization as a necessary step in associative processes including the conversion of silent synapses to functional synapses. A widespread excita- tory GABAergic projection within layer 1 could provide the membrane depolar- ization necessary to induce long-term changes in synaptic efficacy and consolidation of the formation of neocortical layers. This workshop provides an overview of a specific constellation of molecules and neuronal elements, and will permit discussion of their differential roles in layer formation and stabiliza- tion of circuits in the neocortex.

Workshop • Tuesday • 8:30-10:00 PM • Storm Peak Electrophysiology of Incentive Motivation: Can We “Measure” the Affective States of Goal-directed Behavior? Organizer: K. Anstrom; Participants: T. Robinson, G. Schoenbaum, K. Anstrom, W. Schultz Historically, the coordinated activity of the corticomesolimbic system, most no- tably the VTA dopamine neurons and their projections to the nucleus accumbens, prefrontal cortex and amygdala, is thought of as the neural substrate driving goal-directed behavior. Classic reinforcement theories have cited the release of

91 dopamine in these projection areas as a common endpoint of both drug and natural reinforcement. However, this same dopamine system is activated by the cues predicting reward rather than the reinforcer itself, leading to the proposal that the corticomesolimbic system should be thought of as a lear ning system rather than a r eward system. Incentive motivation is defined as the process by which stimuli that predict future rewards, such as food, sex or drugs, shape be- havioral strategies during goal-directed behavior. But can we measure the value of a stimulus in discrete areas of the corticomesolimbic system? Electrophysiol- ogy in awake, behaving animals provides insight to the specific functions of in- terconnected areas of the corticomesolimbic system as stimuli acquire and are used to obtain reward in context-specific paradigms. Terry Robinson will elabo- rate upon the concept of incentive-motivation driving goal-directed behaviors. Geoff Schoenbaum will discuss the roles of interconnected regions of prefrontal cortex and amygdala in linking incentive value and action. Kristin Anstrom will demonstrate context-specific, orbitofrontal coding of taste cues in an alcohol self-administration paradigm. Wolfram Schultz will close the session with a dis- cussion integrating the roles of striatal and prefrontal components of the corticomesolimbic system in goal-directed behavior.

Workshop • Tuesday • 8:30-10:00 PM • Twilight Is Peer Review Peerless? Organizer: C. Atwell; Participants: M. Martin, N. Steneck Was your grant funded or your manuscript accepted for publication? If so, you probably think that peer review in science rewards the truly deserving and has competent, unbiased reviewers. If not,..... This workshop will address both facts and perceptions related to the current state of peer review in neuroscience. Staff from NIH and NSF will lead a discussion of the relative merits of panel vs. mail review and separation of review from program decisions. Initial results from the newly formed working groups to advise NIH on the effectiveness of the neuro- science study sections will be presented and discussed. Nick Steneck, an histo- rian of science and ethicist, will lead a discussion of research on the integrity and fairness of peer review, especially in publications. Discussion will center on how peer review can be improved to facilitate the advance of neuroscience.

Panel • Tuesday • 8:30-10:00 PM • Rainbow Pediatric Epilepsy: From the Clinic to Basic Mechansims Organizer: G.W. Mathern; Participants: M. Levine, S.N. Roper, J. Veliskova Pediatric epilepsy is a common neurological problem, and when refractory to medical management and from a symptomatic cause may need to be treated with surgery. The clinical and surgical treatment of children with refractory epi- lepsy from discrete etiologies has evolved considerable in the last 15 years, as has our understanding of age-specific basic mechanisms of seizure generation. This panel will focus on clinical and basic science mechanisms of epileptogenesis in

92 the developing brain. Mathern will describe the different pediatric epilepsy syn- dromes and substrates that often require surgical treatment. Emphasis will be placed on cortical dysplasia (CD) which is the most com- mon etiology in young children. Levine will describe re- cent findings using in vitro visualization and electrophysiology techniques in brain slices from human pediatric neocortical tissue removed at surgery that show hypo- and hyperexcitable cellular and synaptic features. Roper will discuss ani- mal models of CD, their mechanisms of epileptogenesis, and how the animal models relate to humans. Veliskova will highlight the important of deep brain structures, such as the substantia nigra pars reticulata in the control and synchro- nization of seizures, and how this region shows sex- and age-specific differences in rats.

Workshop • Tuesday • 8:30-10:00 PM • Skyline Beyond Lozenges and Dandruff: Zn2+ Toxicity in the CNS Organizer: I. Reynolds; Participants: E. Aizenman, D. Choi, J. Weiss Zn2+-mediated cell death has been suggested to contribute to neuronal damage associated with ischemia and epilepsy. Zn2+ is present in synaptic vesicles, can be released from neurons upon depolarization, and translocates from presynap- tic sites into postsynaptic neurons to induce cell death. Additionally, a large frac- tion of neuronal Zn2+ that may contribute to neurodegeneration is reversibly complexed to proteins such as metallothionein and Zn2+ finger-containing tran- scription factors. However, the mechanisms by which neurons manage Zn2+, and the cellular targets of Zn2+ mediated cell death pathways are poorly under- stood. This workshop will discuss various aspects of Zn2+-mediated damage in the CNS with a particular focus on the controversies surrounding the mecha- nisms of zinc homeostasis and the processes by which Zn2+ kills neurons. Ian Reynolds will discuss the problems associated with measuring free Zn2+ in neu- rons and introduce the concept of neuronal Zn2+ homeostasis. John Weiss will lead a discussion on the relationship between Zn2+ influx into mitochondria and generation of free radicals. Dennis Choi will give us his latest thoughts on the effects of Zn2+ on cellular energy status. Elias Aizenman will challenge the au- dience with a model that links the release of intracellular zinc with an apoptotic cascade. Zinc lozenges will be distributed to all participants that pose questions to the panelists, except for those who are using dandruff control shampoo, for whom such lozenges may be dangerous.

Panel • Tuesday • 8:30-10:00 PM • Sunset Vasopressin/Vasotocin: Key Regulators of Social Behavior Across Species? Organizer: E. Albers; Participants: S. Boyd, D. Maney, J. Winslow Vasopressin and vasotocin are members of a large family of structurally and functionally related neuropeptides. These two nonapeptides were initially identi-

93 fied as neurohypophysial hormones that regulate hydromineral balance. How- ever, more recently, these neuropeptides have been found to be important regula- tors of a variety of different types of social behavior in species ranging from fish to primates. In fact, agonists and antagonists of vasopressin and its non-mam- malian analog vasotocin have been found to turn certain social behaviors on or off by their actions in specific brain sites. This panel will review the rapid progress being made in identifying the role of these peptides in the control of reproductive, communicative and agonistic be- haviors across vertebrate taxa. The session will begin with Sunny Boyd review- ing the interaction of androgens and vasotocin in the control of reproductive behaviors in amphibians. Donna Maney will then discuss the anatomical and behavioral evidence implicating vasotocin in avian communication. Elliott Albers will review the role of vasopressin containing circuits in the control of several forms of agonistic and communicative behavior in hamsters and how hormones and social experience alter these behaviors by modulating vasopressin receptors. Finally, Jim Winslow will review efforts to describe the interaction of social ex- perience with brain nonapeptide systems for the normal development of social behavior in non-human primates.

Panel • Wednesday • 7:30-9:30 AM • Mt. Werner The Map Kinase Signaling Pathway and Synaptic Plasticity: Activators, Regulators and Effectors Organizer: P. Chapman; Participants: D. Kuhl, J. Naranjo, J. Yin We have know for quite some time that synaptic plasticity in the CNS of adult mammals can be induced by transient changes to the intracellular calcium con- centration of neurons. It is also clear that the establishment of plasticity depends upon the activation of protein kinases and phosphatases, many of which are calcium sensitive. More recently, evidence has suggested that long-lasting synap- tic plasticity requires modification of gene expression, and that the link between transient changes in calcium and long-term plasticity will involve the recruit- ment and/or modulation of inducible transcription factors. MAP kinase is a potentially important link in this pathway, as it can be recruited and modulated by both calcium-dependent and neurotrophin-dependent second messengers, and can in turn activate important transcription factors including the CREB family. Jerry Yin will describe evidence linking protein kinase C to activation of CREB and subsequent memory formation in both flies and mice. Paul Chapman will discuss the consequences of deleting upstream activators of MAP kinase on syn- aptic function and plasticity in several brain regions. JosT Naranjo will present data demonstrating the multiplicity of ways in which gene expression can be regulated by DREAM, a calcium-sensitive repressor of gene transcription which itself is subject to regulation by protein kinase A through CREM. Dietmar Kuhl will discuss an important class of immediate early genes that are induced in hippocampal neurons by stimuli that activate plasticity related signaling path- ways and translated locally at activated synapses. As a group, we will discuss

94 strategies for making sense of the ever-increasing number of molecules that have been implicated in synaptic plasticity, and how these might one day be related to learning and memory.

Panel • Wednesday • 7:30-9:30 AM • Storm Peak Mechanisms of CNS Regeneration and Repair Organizer: W.J. Streit; Participants: W. Tetzlaff, T. Hagg, S.R. Whittemore The limited ability of central neurons to regenerate after injury is due to a num- ber of factors including a lack of trophic support and an inability of injured neurons to express appropriate pro-regenerative genes. The goal of this panel is to summarize neuronal and glial cell responses that occur after CNS injury and discuss potential strategies for repair. Wolf Tetzlaff will describe how rubrospi- nal and corticospinal neurons respond to axotomy in terms of gene expression patterns and neuronal atrophy, and how these processes are affected by exog- enous application of BDNF. Jake Streit will contrast glial cell responses in regen- erating and non-regenerating systems, and suggest mechanisms by which glial cells may provide trophic support to injured neurons. He will emphasize the importance of neuronal-glial signaling and discuss potential signaling molecules. Theo Hagg will present strategies for the use of neurotrophic factors in attracting severed fiber tracts across injury and scar sites, guide lengthy outgrowth, induce appropriate innervation of the original target and induce (or prevent when nec- essary) collateral sprouting of uninjured fibers. Scott Whittemore will discuss the use of pluripotent stem cells for CNS repair. Specifically, he will describe how stem cells genetically modified to initiate lineage-specific differentiation into neurons, , or astrocytes are used to facilitate neuron replace- ment, remyelination of demyelinated axons, and engender axonal regeneration, respectively.

Panel • Wednesday • 7:30-9:30 AM • Twilight Mechanisms of Addiction: The Brain Opioid System Ain’t Just for Opiates Anymore Organizer: T.S. Shippenberg; Participants: J.J. Frost, C.J. Evans, J.F. McGinty, C.T. Napier Opiate drugs exert their effects by binding to opioid receptors, but it is increas- ingly recognized that non-opiate drugs of abuse modulate the activity of endog- enous opioid systems (EOS). Furthermore, their activation can profoundly alter the effects of psychostimulants and alcohol. Stimulants cause changes in opioid peptide concentration, opioid peptide mRNA, and opioid receptor regulation. The administration of _-opioid antagonists or knock-out of this receptor reduces the behavioral effects of stimulants. In contrast, k-receptor activation attenuates, whereas k-receptor deletion exacerbates the behavioral and neurochemical ac- tions of these agents. The effects of other abused drugs, such as THC and etha-

95 nol, also depend on intact EOS. This panel will address the role of _-, d- and k- opioid systems in modulating the neurochemical and behavioral effects of non- opiate drugs of abuse and provide an anatomic and mechanistic basis for their interaction with EOS. McGinty will provide evidence that k-agonists and d-an- tagonists suppress the behavioral effects of amphetamine and decrease amphet- amine-evoked extracellular levels of dopamine and glutamate in the dorsal and ventral striatum. Whereas k-receptors appear to directly affect glutamate release as revealed by their localization in axon terminals and inhibition of glutamate efflux in synaptosomes, the effects of d-receptor manipulation appear indirect. Napier will discuss how regulation of neurotransmission by the _- and k- EOS occurs at the level of limbic outputs and that EOS influence neuronal activity in basal ganglia, cortex and, other regions of the brain s motive circuit. The impli- cations of this regulation for understanding interactions of EOS with abused drugs will be discussed. Evans will address mechanisms by which these drugs could ultimately influence opioid system function, including the role of opioid receptor internalization and intracellular trafficking. Frost will discuss advances in PET imaging of opioid receptors in substance abuse, including studies of co- caine and alcohol dependence. Data demonstrating _-receptor up-regulation in brain regions involved in reward and a correlation between up-regulation, crav- ing and relapse will be presented. Changes in _-receptor binding after acute co- caine administration, likely due to cocaine-induced opioid peptide release, will also be discussed. Integrating cellular and neurophysiological data with PET studies of human substance abuse will be a key theme of this panel.

Panel • Wednesday • 7:30-9:30 AM • Rainbow Perinatal Cell Death in Pathological Environments: Which Cells Are Most Susceptible and How/When Do They Respond? Organizer: W. Macklin; Participants: M. Miller, S. Levinson, S. Vannucci Hypoxic/ischemic (H/I) insults to the fetal or neonatal brain have long-term consequences, in terms of cell death, altered cell migration and/or differentia- tion. The timing and duration of the insult clearly impacts the specific cellular responses. The susceptibility and responses of different cell types are important for the overall survival and function of the tissue. In this panel, responses of neurons, astrocytes, oligodendrocytes, oligodendrocyte progenitor cells and stem cells to hypoxic/ischemic events will be discussed. Dr. Miller will discuss cell responses in a perinatal model of repeated hypoxia, which produces animals with hypoxic ventilatory depression. Studies on cell death in this model using both TUNEL analysis and caspase immunoreactivity will be discussed, focusing on cells in the ventilatory centers of the brainstem, the nucleus tractus solitarius and the parapyramidal area. Dr. Vannucci will discuss the astrocytic responses

96 to a single perinatal H/I insult. There is a rapid loss of GFAP mRNA and likely astrocyte cell death in severe perinatal insults, which is followed later by signifi- cant gliosis. This contrasts with the transient induction of GFAP mRNA follow- ing milder insults. The mechanisms for these differential responses of astrocytes will be discussed. Dr Levison will discuss how perinatal H/I affects the stem cells in the subependymal zone (SZ). Cellularity counts in tissue sections reveal that 25% of the cells within the SZ are depleted within 48 h of recovery from a moderate H/I insult. Histochemistry and electron microscopy reveal that these cells undergo necrotic, apoptotic or hybrid cell deaths, with the majority of cell deaths being hybrid deaths. The susceptibility of these cells to death signals and the mechanisms responsible for their demise will be discussed. Dr. Macklin will discuss oligodendrocyte and oligodendrocyte progenitor responses to a fetal hy- poxic insult, along with altered cell migration. In this model, animals are ex- posed to 45 min hypoxia by occlusion of the uterine arteries at E18. Less dramatic cell death results from this insult, but significant changes in cell organization occur along with changes in the development of oligodendrocytes. The data dis- cussed in this panel will provide insight into the timing and mechanisms of cellu- lar responses to H/I, with a particular focus on mechanisms of cell death.

Panel • Wednesday • 7:30-9:30 AM • Skyline Peptide Nucleic Acid – New Life for Antisense (and Antigene) ? Organizer: P. Clarke; Participants: B. Tyler-McMahon, U. Langel, L. Boffa, P. Clarke For neuroscientists, the antisense approach has thus far failed to live up to expec- tations. There are currently two main problems associated with this approach. The first is low efficacy — an effective antisense sequence is hard to find, and protein expression is usually only partially inhibited. The second problem is tox- icity—administration of oligonucleotides can result in brain damage; almost cer- tainly, toxic antisense effects are inadequately investigated and under-reported. Peptide nucleic acids (PNAs) are synthetic oligodeoxynucleotide analogues that possess several important advantages over tr aditional antisense molecules (i.e. phosphodiester, phosphorothioate). Several recent publications suggest that PNA antisense work well in rat brain. The goal of the panel is to discuss the merits of PNA vs. other antisense treatments. Paul Clarke will describe the in vivo target- ing of delta opioid receptors by PNA, and recall past experience (mostly painful) with phosphodiester or phosphorothioate antisense. Beth Tyler-McMahon will report depletion of G-protein coupled receptors following central or systemic administration of PNAs. She will also show that PNA can exert an antigene effect in vivo. Ulo Langel will show that PNA antisense can be rendered more effective by conjugation to peptides that facilitate cellular uptake. He will also describe in vivo antisense effects in the spinal cord. Lidia Boffa will report how antigene effects of PNA can be enhanced by conjugation with a nuclear localiza- tion signal. She will describe several cellular model systems where appropriate PNA-conjugates can exert specific antigene effects.

97 Panel • Wednesday • 7:30-9:30 AM • Sunset Stimulation, Lesioning and Transplantation: Application for the Treatment of Parkinson’s Disease and Dystonia Organizer: J. Vitek; Participants: J. Vitek, A. Lozano, J. Kordower, R. Bakay Surgical therapy for the treatment of Parkinson s disease and dystonia has un- dergone a marked resurgence in the 1990 s and continues into the new millen- nium. Improved understanding of the pathophysiological basis underlying the development of the motor signs of PD has led to the refinement of old and implementation of new surgical approaches for the treatment of this disorder. With the advent of deep brain stimulation we now have a mechanism whereby patients may undergo bilateral procedures without the same risk associated with bilateral ablative procedures. Restorative methods continue to evolve with the refinement of fetal tissue transplantation techniques and the development of new approaches employing alternative tissues. The application of the same lesioning and DBS techniques used for patients with PD have now been employed for the treatment of patients with primary and secondary dystonia leading in some cases to an almost complete restoration of function. The purpose of this panel is to provide the audience with the most recent developments in the application of these techniques for the treatment of these disorders. Andres Lozano will dis- cuss the use of DBS and pallidotomy for the treatment of PD, the relative merits of each technique as well as the potential pitfalls and problems that one may encounter when performing these procedures. Jeff Kordower will discuss recent developments in fetal tissue transplantation for PD; problems that currently per- sist and methods that have been developed to address them. Roy Bakay will present recent data on the use of alternative tissues such as porcine cells in neurotransplatation, the rationale behind their use and the problems associated with this technique. Jerrold Vitek will discuss the reapplication of pallidotomy and the development of DBS for the treatment of dystonia. Results from a series of patients undergoing unilateral and bilateral procedures will be presented.

Panel • Wednesday • 4:30-6:30 PM • Mt. Werner Cholinergic Modulation of Cortical Function: Channels to Circuits Organizer: R.C. Foehring; Participants: R. Metherate, F.F. Ebner, R. Andrade Cholinergic afferents from the basal forebrain provide one of the major extrathalamic inputs to the neocortex. Cholinergic fibers ramify in all six corti- cal layers and both muscarinic and nicotinic receptors are expressed in pyrami- dal and nonpyramidal cells in cortex. Based upon fiber projections and receptor distribution, this cholinergic projection has been implicated in regulation of cor- tical activity, sleep-wake states, cognition, learning and memory, and plasticity. Consistent with these proposed functions, loss of the basal forebrain cholinergic projection to cortex occurs early in the progression of Alzheimer s disease. This panel will discuss recent advances from in vitro and in vivo preparations which

98 address cellular and circuitry mechanisms involved in acteylcholine s actions. R. Andrade will present evidence that nonselective cation currents are important contributors to cholinergic responses mediated via muscarinic receptors in pre- frontal cortex. He will also outline evidence indicating that one of these nonse- lective cation currents is also regulated by intracellular calcium, and hence can be impacted by concurrent synaptic activity and cell firing. R. Foehring will dis- cuss muscarinic modulation of voltage-gated calcium channels and calcium-de- pendent potassium channels in neocortical pyramidal cells. Special emphasis will be placed on the consequences of these modulations for synaptic transmis- sion and repetitive firing. R. Metherate will present evidence for differential cho- linergic regulation of thalamocortical vs. intracortical pathways in an auditory thalamo-cortical slice preparation. He will also present data suggesting a critical role for nicotinic cholinergic regulation of glutamatergic transmission in cortex during a specific postnatal period. F. Ebner will discuss cholinergic modification of cortical responses and plasticity in vivo. Both single cell response modulation and results of behavioral studies will be reviewed. The goal of the panel is to provide a forum for synthesis of ionic, cellular, and network data on modulatory actions of acetylcholine in neocortex and to present the audience with a current view of important cholinergic effects on cortical function.

Panel • Wednesday • 4:30-6:30 PM • Storm Peak Axon Guidance: From Extracellular Cues to Growth Cone Behaviors Organizer: K. Kalil; Participants: T. O Connor, P. Letourneau, T. Gomez In recent years molecular cues have been discovered that guide axons as they navigate toward targets. Axonal growth cones respond to cues such as semaphorins, netrins and ephrins by attraction or repulsion. These extracellular guidance cues function through surface receptors to control intracellular signal- ing events which are ultimately transduced through remodeling of the cytoskel- eton into effects on growth cone motility. The panel will present an overview of axon guidance mechanisms and will discuss exciting new findings from a variety of neural systems to explore some of these linkages between extracellular guid- ance cues and intracellular events driving growth cone behaviors. O Connor will demonstrate how spatial gradients of a secreted semaphorin axon guidance molecule can provide directional information to growth cones of grasshopper pioneer axons during pathfinding. Letourneau will discuss how growth cones integrate simultaneous interactions with multiple guidance cues. He will describe recent work on DRG and retinal neurons to demonstrate that neurotrophins can reduce the collapsing effects of molecules such as nitric oxide, semaphorin 3A and ephrins that induce growth cone collapse and will present evidence that neurotrophins appear to protect actin filaments against depolymerization induced by collapsing signals. Kalil will demonstrate dynamic changes in the actin and microtubule cytoskeleton that underlie cortical growth cone behaviors leading to axon branching and will discuss how some of these changes are affected by ap-

99 plication of growth factors, such as FGF-2. Gomez will show in frog spinal neu- rons how highly localized transient elevations of intracellular calcium that propa- gate down individual growth cone filopodia can serve as both frequency-coded signals that are integrated within the growth cone and as autonomous regulators of filopodial motility.

Panel • Wednesday • 4:30-6:30 PM • Twilight Recording from a Whole Bunch of Neurons All at Once: Problems, Payoffs and Promise Organizer: J.Walters; Participants: B.Waterhouse P. Janak D. Kipke D. Woodward Multichannel recording techniques offer significant promise for obtaining insight into mechanisms of information processing within the CNS. The ability to record simultaneously from many neurons in multiple brain circuits over extended peri- ods of time has challenged investigators to develop experimental strategies and data analysis routines that maximize the information obtained from this approach. Through examples from their current research, panel participants will discuss different approaches, problems and payoffs associated with many neuron record- ing techniques. Strategies for conducting large scale population recordings in primate brain motor areas and the use of ensemble spike train data to drive ro- botic devices will be discussed by Daryl Kipke. Challenges and rewards of study- ing the activity of neuronal populations in the mesocorticolimbic system in relation to drug- and food- reinforced behavior will be described by Patricia Janak. Barry Waterhouse will report on early surprises and ex- pectations from studies of brainstem monoaminergic sys- tems that project globally and exert modulatory effects on ensembles of neurons at multiple levels of intact sensory pathways. Donald Woodward will conclude with his vision of the future of this technology (i.e. analysis algorithms, software and hardware developments) and how it will be used to address a new generation of questions about multineuron interactions and basic neural network operations in behaving animals. Discussion will be moder- ated by Judith Walters. Contributions from the attendees will be encouraged. Bring a slide of your current many- neuron/multiple site recording problem, payoff or prom- ise!

100 Panel • Wednesday • 4:30-6:30 PM • Rainbow GABA Receptors as Anesthetic Targets: What We’ve Learned from Gene and Ion Channel Structure/Function Studies Organizer: A. Jenkins; Participants: G. Houmanics, S.J. Mihic, D.S. Weiss, C.M. Czajkowski Over the past decade, the GABAa receptor (GABAaR) has emerged as a promis- ing target for general anesthetic agents including ethers, benzodiazepines (BZD), barbiturates and alcohols. More recently, molecular biology has permitted GABAaR manipulation, revealing a wealth of information on expression, acti- vation and modulation. This panel aims to present different approaches to an- swering the questions Ho w do these drugs modulate receptor function ? and Ho w does this impact on the CNS ? Homanics will discuss the la test experi- ments utilizing genetically engineered mice that harbor mutations in GABAaR genes. Included will be alcohol and anesthetic responses in mice, which lack specific receptor subunits in either a global or tissue specific pattern. Mihic will examine the modulation of wild-type and mutated GABAa and Glycine recep- tors by general anesthetics, inhaled drugs of abuse and ethanol. Two amino ac- ids have been identified which regulate receptor-ligand sensitivity and allosteric modulation. These data suggest that inhaled drugs of abuse affect ligand-gated ion channels and that the molecular sites of action of all these compounds may overlap. Weiss will discuss the kinetics of receptor activation, modulation and ion-channel gating. [3H] binding experiments with single oocytes and site-di- rected mutagenesis experiments at a conserved leucine residue have led to the formulation of a novel activation mechanism for the receptor. These data sug- gest new pathways via which allosteric modulators may alter ion-channel func- tion. Czajkowski will discuss the mapping of the BZD binding and allosteric coupling domains of the GABAaR. Using the substituted cysteine accessibility method and by constructing chimeric subunits, two domains have been identi- fied which are necessary to impart full BZD sensitivity to chimeric receptors. These novel residues provide insight into the mechanisms underlying the allos- teric coupling of all ligand-gated ion channels. Jenkins will discuss evidence for a general anesthetic binding cavity. By comparing the effects of a number of anesthetics with different molecular volumes on a series of mutant receptors, it is proposed that these drugs bind to a ~250 +3 cavity located within the transmem- brane domain of the receptor.

101 Panel • Wednesday • 4:30-6:30 PM • Skyline

The Response to CO2 and O2: From Ion Channels to Neural Networks Organizer: J.M. Ramirez; Participants: M.M. Tamkun, N.R. Prabhakar, J.M. Ramirez, G.B. Richerson

The nervous system has to adapt to changes in CO2 and O2 to prevent irrevers- ible damage to the brain. Exposed to high-altitude, participants at the Winter Brain Conference have to face a hypoxic challenge while maintaining mental and physical power. In this panel WCBR participants have the chance to learn about the new developments in understanding how an organism adapts to alter- ations in CO2 and O2. The aim of this panel is to relate changes at the molecular level of ion channels with the adaptive response at the organismic level. Michael Tamkun will demonstrate how oxygen alters electrophysiological properties of voltage-gated K (KV) channels, which could lead to a hypoxic pulmonary vaso- constriction at the organismic level. The experimental use of cloned Kv channels enables the Tamkun laboratory to gain insights into the subtype-specific effect of hypoxia: Numerous ion channels localize to lipid rafts in nerve and muscle and these membrane domains act as signal transduction centers. How channel tar- geting to lipid rafts may be involved in CO2 and O2 sensing will be discussed. The role of calcium-dependent pathways in mediating the hypoxic response will be high-lighted in the presentation by Nanduri Prabhakar. Using RT-PCR tech- niques and reporter gene assays the Prabhakar laboratory demonstrates that the hypoxic modulation of L-type calcium channels leads to a cascade of cellular events involving CaMK activation and CREB phosphorylation. These changes result in an adaptive response of the carotid body, the major mammalian oxygen sensory organ. To address the role of the central nervous system in response to hypoxia, Jan-Marino Ramirez investigates the effect of O2 changes on the iso- lated respiratory network using brainstem slices. With the patch-clamp technique the Ramirez lab demonstrates that the hypoxic modulation of the N-type cal- cium channel leads to alterations in Kca and Ih conductances which result in an increased frequency of respiratory pacemaker activity and a reconfiguration of the respiratory network. George Richerson will discuss the effect of CO2 on serotonergic medullary Raphe neurons. Intracellular acidosis inhibits a novel calcium-activated cation current in these neurons with a reversal potential of -61 mV. This leads to an increased firing rate, which could be expected to stimulate breathing through release of serotonin, TRH and Substance P. The four different mechanisms described in this panel for detecting changes in blood gases would act together to maintain homeostasis, particularly during a hypoxic challenge at high-altitude.

102 Panel • Wednesday • 4:30-6:30 PM • Sunset Role of Rhythmic Neural Activity in Cardiovascular Behavior: From Molecular Details to Black Diamonds Organizer: J. P. Horn; Participants: P.G. Guyenet, M.P. Gilbey, J.P. Horn. D. Weinreich The autonomic nervous system enables one to ski downhill at high altitude. Al- though the best-known cardiovascular behavior, the baroreceptor reflex, can be viewed as simple, cardio-pulmonary adaptation during exercise is much more complex. How then does the nervous system orchestrate the appropriate motor elements during warm-up, the windy ride up the lift, and the rush down big bumps? Recent evidence suggests that rhythmic activity may play a central role not only for timing jumps through bumps, but also for coordinating and regulat- ing the control of peripheral autonomic targets. The panel will outline recent progress at different levels, from central circuitry to peripheral synaptic mecha- nisms. Patrice Guyenet will give a brief overview of the brainstem circuitry that integrates and drives the sympathetic and parasympathetic outflow to the car- diovascular system. He will describe how a new extracellular recording method can be used to mark physiologically identified neurons, and discuss how these neurons may participate in the pattern-generating network. Michael Gilbey will address how central pattern generators may coordinate activity in postgangli- onic sympathetic axons that control peripheral blood flow for thermoregulation. Activity recorded under different states of cardio-respiratory drive will be dis- cussed in terms of synchronization and motor binding. John Horn will discuss how synaptic integration in sympathetic ganglia contributes to cardiovascular function. He will outline a computational analysis, which indicates ganglia func- tion as use-dependent synaptic amplifiers whose gain can be adjusted by oscilla- tory activity. Danny Weinreich will discuss how changes in long-term potentiation in sympathetic ganglia correlate with blood pressure in normal and hypertensive animals.

Panel • Thursday • 7:30-9:30 AM • Mt. Werner Kainate Receptors and Synaptic Plasticity Organizer: G.L.Collingridge; Participants: R.A.Nicoll, G.L.Collingridge, J.T.R. Isaac, D. Lodge Fast excitatory transmission in the CNS is mediated by the ionotropic glutamate receptors that can be divided into three subtypes: AMPA, NMDA and kainate. Of these subtypes, the functions of kainate receptors have until recently remained largely unexplored. However due to the development of new ligands selective for kainate and AMPA receptors, a rapid advance in our knowledge of kainate re- ceptors has taken place. For example, kainate receptors have been shown to be activated at synapses and to mediate an excitatory postsynaptic current in a num- ber of regions of the mammalian CNS. One exciting emerging field is the in-

103 volvement of kainate receptors in the induction and expression of long-term synaptic plasticity, a process thought to underlie development, learning and memory, and some neurodegenerative diseases. The aim of this panel is to present recent data on the involvement of kainate receptors in synaptic plasticity. The potential role of these receptors in development and learning and memory, and the potential of the kainate receptor as a drug target for the prevention and treat- ment of certain neurodegenerative diseases will be discussed. Roger Nicoll will present his recent work on the role of kainate receptors in mediating and regulat- ing excitatory and inhibitory synaptic transmission in the hippocampus (Frerking et al., 1999 Proc. Natl. Acad. Sci. USA 96, 12917-12922.), and discuss the poten- tial for long-term modifications of this during synaptic plasticity. Graham Collingridge will present recent work on the involvement of kainate receptors in the induction of long-term potentiation (LTP) at the mossy fibre-CA3 synapse in the hippocampus (Bortolotto et al., 1999 Nature 402, 297-301). John Isaac will describe the involvement of kainate receptors in the expression of LTP at devel- oping synapses in primary sensory cortex (Kidd and Isaac, 1999 Nature 400, 569-573), and the implications of this for the development of receptive fields. David Lodge will describe advances in kainate receptor pharmacology (Clarke et al., 1997 Nature 389, 599-603) and present recent data on the suppression of epileptiform activity by kainate receptor antagonists. We anticipate that this highly topical panel will generate lots of interest and provide a useful discussion con- cerning the roles of kainate receptors at CNS synapses.

Panel • Thursday • 7:30-9:30 AM • Storm Peak Microarrays in Neuropsychiatric Research: A Delusion, Illusion, or Reality? Organizer: T.M. Hyde; Participants: C. Colantuoni, R. Coppola, W.J. Freed Microarray technology holds great potential for deciphering the molecular basis of complex neuropsychiatric disorders that have defied conventional neuropatho- logical approaches. However, the utility of these arrays has only recently been validated in human post-mortem brain studies. This panel will discuss the appli- cation of a variety of microarray techniques to normal human brain tissue and such diverse disorders as schizophrenia and . Carlo Colantuoni will review the applications of competing microarray techniques to post-mortem brain tissue from patients with Rett Syndrome, a heritable disorder in females characterized by seizures, motor dysfunction, and cognitive impairment. He will also review recent applications of the same techniques to brain tissue from schizo- phrenic subjects. Richard Coppola will discuss an alternative approach to the use of microarrays in schizophrenia, especially the use of pooled samples from multiple cases. He will also review the bio-informatics issues that complicate the handling and interpretation of data derived from microarrays. Finally, William Freed will discuss the development of a novel cDNA neur oarray to probe for genes associated with CNS function, leading to the identification of a small num-

104 ber of genes involved in synaptic function, transcription, cell recognition, and signal transduction that are abnormally expressed in schizophrenia.

Panel • Thursday • 7:30-9:30 AM • Twilight Regulation of Monoamine Transporter Characteristics and Trafficking Organizer: A. Janowsky; Participants: M. Reith, H. Melikian, A. Eshleman, N. Zahniser Transporters are the major mechanism for inactivating biogenic amine neurotrans- mitters. Abused drugs such as cocaine and methamphetamine, and therapeutic agents such as antidepressants and drugs that are used to treat Parkinson s dis- ease bind to the transporters and block neurotransmitter uptake. Alteration of transporter function has been associated with euphoria, craving, symptoms of withdrawal, and exacerbation of symptoms that accompany schizophrenia and other neuropsychiatric disorders. This panel will describe new information on drug-transporter interactions, and molecular mechanisms involved in regulating transporter expression. Specifically, Maarten Reith will discuss conformational changes that occur during the interaction of substrates and non-substrate inhibi- tors with monoamine transporters, and how carrier oligomerization could regu- late transporter characteristics. He will also address the impact of posttranslational modification of monoamine transporters by glycosylation, utilizing experimen- tal evidence of dopamine transporter (DAT) mutants that lack glycosylation con- sensus sites. Haley Melikian will describe how the DAT undergoes a substantial level of constitutive trafficking under basal conditions, and how activation of protein kinase C (PKC) impacts particular elements of pre-existing constitutive trafficking, resulting in DAT downregulation. Amy Eshleman will explain differ- ences in the effects of PKC activation on internalization and on transporter char- acteristics among heterologously expressed DAT, serotonin and norepinephrine transporters. Nancy Zahniser will discuss the evidence that dopamine D2 autoreceptors regulate DAT function in brain. Shewill also present experimental evidence from expression systems that activation of D2 receptors increases DAT activity by altering their expression at the cell surface. Receptor signaling and transporter trafficking mechanisms involved in this regulation will also be dis- cussed.

Panel • Thursday • 7:30-9:30 AM • Rainbow Neuroimmunophilins—Can Small Molecules Stimulate Therapeutic Neurorestoration? Organizer: E. Hall; Participants: P. Letourneau, E. Villafranca, B. Gold, D. Cole Unlocking the mechanisms that regulate axonal plasticity and regeneration has long been a goal of neurobiology. This is based upon the hope of finding phar- macological means by which recovery of function (neurorestoration) could be stimulated after acute neurological injury and in neurodegenerative diseases. Sig- nificant advances have been made in our understanding of the ability of axons to 105 grow during development and after injury, and some of the ¥ molecular mecha- nisms involved. This has lead to the identification of small molecule pharmaco- logical targets by which axonal sprouting and regeneration might be enhanced. The aim of this panel is to review the neurobiology of axonal growth and one of the more promising of these pro-regenerative targets the neuroimmunophilins (neurophilins, NIMMs). The NIMMs are putative members of a family of en- zymes known as peptidyl-prolyl cis-trans isomerases (PPIases) believed to play a role in regulation of protein folding. The PPIases of interest are known as FK binding proteins (FKBPs), named for their binding of the prototype neurotrophic NIMM compound FK-506. In particular, the panel will review the neurorestorative mechanisms and efficacy of prototype NIMM-directed com- pounds including FK-506, GPI1046 and V-10,367, as well as other, novel NIMM compounds. Paul Letourneau will discuss the neurobiology of axonal sprouting and growth cone elongation and guidance as well as growth factor effects on axonal growth. Ernie Villafranca will introduce NIMM phar- macology focusing on the present mechanistic understand- ing of axonal growth-promoting effects and their dependence on neurotrophic factors. Bruce Gold will discuss the roles of different FKBPs in NIMM actions and interactions with steroid hormone chaperone/heat shock proteins. Addi- tionally, he will present data on the ability of NIMM compounds to stimulate neurite extension in cell-based systems and to promote neurorestoration in mod- els of peripheral nerve and spinal cord injury. Finally, Doug Cole will present work on the neuropharmacology of V- 10,367 and other, novel NIMM com- pounds.

Panel • Thursday • 7:30-9:30 AM • Skyline The Brain is a Cookie Monster: Contribution of Food Composition and “Hedonic Circuitry” to the Regulation of Food Intake Organizer: D. Lattemann; Participants: B. Levin, C. Billington, A.Kelley, D. Lattemann Studies over recent years have focused upon the role of the medial hypothalamus in the short- and long-term regulation of caloric intake. However, other studies are beginning to elucidate the CNS neuroanatomical and neurochemical bases for the under-appreciated observation that a calorie is not a calorie the compo- sition and the rewarding/hedonic qualities of food contribute significantly to the endpoint of total food intake, and its regulation. In this panel, we will summa- rize recent studies that evaluate the CNS regions and pathways that are impor- tant for mediating these attributes of food. Barry Levin will show how chronic intake of a palatable diet can alter the CNS neurochemical patterns of rats that are genetically prone or resistant to diet-induced obesity, a model of diet-induced neural plasticity. Charles Billington will discuss the involvement of brain opioid pathways in both ingestion of palatable foods and caloric-need driven ingestion. Ann Kelley will discuss the involvement of the ventral striatum in food intake, with particular focus on striatal enkephalin gene expression and intake of high fat/sweet food. Dianne Lattemann will discuss the interaction between periph-

106 eral adiposity signals and brain systems of reward/hedonics. A synthesis of how these systems may be integrated with the hypothalamic systems that regulate energy balance will be presented.

Panel • Thursday • 7:30-9:30 AM • Sunset Mitotic Regulators ‘Prospero-us’ in Postmitotic Neurons from Become to Succumb? Organizer: I. Vincent; Participants: H. Vaessin, R. Slack , P. Baas The downregulation of cell cycle regulators and the consequent inability of neu- rons to divide following their terminal differentiation have constituted a dogma of neuroscience that has fascinated neurobiologists for almost a century. Only recently is it becoming apparent that postmitotic neurons rely on mitotic regula- tors and mechanisms for some critical events in their life history, which has left some of us neurobiologists in awe as to how postmitotic these cells really are. In this panel, two participants will present new insights of the classical view of neuronal determination, and another two participants will highlight the role of mitotic mechanisms in postmitotic neurons. Drosophila prospero encodes a criti- cal regulator of the irreversible transition from mitotically active to differentiated neuron. Dr Harald Vaessin will expound the mechanism by which prospero or- chestrates the asymmetric mitosis leading to the birth of a daughter cell identical to the mother, and another cell committed to neuronal differentiation. Terminal mitosis and differentiation are tightly coupled and failure to exit the cell cycle results in impaired differentiation and neuronal cell death. Dr Ruth Slack will describe how pRb, a regulator of normal cell division, also acts as a developmen- tal switch to cause progenitor cells to exit the cell cycle and express a neuronal phenotype. Contrary to the dependence of neuronal differentiation on the purg- ing of mitotic machinery, Dr Peter Baas and colleagues have found that the mecha- nisms by which neurons generate axons and dendrites are modifications of the same mechanisms organizing mitotic spindles in dividing cells. Dr Baas will elabo- rate on this intriguing mechanism. Other evidence has suggested that reactiva- tion of mitotic regulators in mature neurons serves as a trigger for their degeneration in a variety of human neurological diseases, including Alzheimer s disease. A better characterized example is that of Ataxia-Telangiectasia caused by mutation of the Atm gene, whose protein product is a regulator of the cell cycle G2 and M checkpoints. Strangely, the nosologic feature of this disease is progressive degeneration of postmitotic cerebellar neurons. Dr Carrolee Barlow will describe recent findings from Atm deficient mice, an excellent model for understanding the role of ATM in normal neuronal function.

107 Panel • Thursday • 4:30-6:30 PM • Mt. Werner Can Neuronal Glutamate Release Be Measured in Vivo? Microdialysis vs Biosensors Organizer: N.Maidment; Participants: M. Boutelle, R. Kennedy, A. Michael, B. Dunn Controversy surrounds the use of microdialysis for measuring glutamate release in vivo. Unlike the case for monoamines, most reports indicate that the basal extracellular glutamate sampled with this technique is unperturbed by TTX or by removal of calcium from the perfusion medium. This has led many to con- clude that microdialysis has limited utility for sampling the neurotransmitter pool of glutamate. However, recent developments in rapid sampling techniques and in enzyme-based biosensor design suggest that neuronally released glutamate may be accessible in vivo. We will address several issues. 1. Are there conditions under which neuronally released glutamate can be measured with microdialysis? 2. If microdialysis is not sampling vesicularly released glutamate, from what pool is it sampling and is this information valuable? 3. Does the small size of glutamate biosensors enable them to sample closer to glutamate release sites thereby providing a better index of vesicularly released glutamate? Martyn Boutelle will discuss the origin of extracellular glutamate as sampled by microdialysis. He will also describe the combination of microdialysis with en- zyme-based sensors for on-line detection of microdialysis-derived glutamate—the enzyme bed electr ode and the dialtr ode . Robert Kennedy will describe the use of capillary electrophoresis for rapid measurement of extracellular glutamate, either in conjunction with microdialysis or as a direct sampling technique. Adrian Michael has developed a 10 um carbon fiber-based amperometric enzyme bio- sensor for direct measurement of glutamate in vivo. His design incorporates glutamate oxidase, horseradish peroxidase, an osmium-polymer complex together with a nafion coat resulting in a sensor that is uniquely selective for glutamate, and that provides TTX-sensitive signals in vivo. Bruce Dunn is developing a novel in vivo fiber optic sensor for glutamate employing sol-gel technology. Glutamate dehydrogenase is encapsulated in sol-gel at the tip of a fiber optic tube and the resultant production of NADH from NAD upon glutamate diffusion into the sol-gel is measured by a change in luminescence.

Panel • Thursday • 4:30-6:30 PM • Storm Peak Signal Transduction in Neuronal and Neuroendocrine Cells: What the Mutant Mice Said to Me Organizer: M.R. Picciotto; Participants: I. Lindberg, P. Allen, K. Wickman, M. Picciotto Signaling pathways are critical for transducing information from neurotransmit- ters outside the cell into changes in neuronal function. While molecular, bio- chemical and physiological techniques have been critical in identifying the players in intracellular signaling pathways, it has been difficult to develop highly spe-

108 cific, cell-permeable pharmacological agents to manipulate intracellular signal- ing molecules, and thus to elucidate their function at the cellular, systems and behavioral level. This is particularly true for molecules that are members of large gene families. Gene targeting in mice presents new opportunities to study the roles of proteins involved in signal transduction in the intact animal. The aim of this panel is to present experimental data that explore four different systems in which knockout mouse models have been used to elucidate the role of individual molecules in complex neurobiological processes. Marina Picciotto will introduce the topic of transgenic and knockout mouse models, and will use experiments on nicotinic receptor knockout mice as an example of how studies on signaling can start at the membrane. Kevin Wickman will present work on knockout mice lacking subunits of the G protein-gated family of potassium channels (GIRK) and will describe how these animals can be used to elucidate the roles of ion channel with no specific agonists or antagonists. Proceeding deeper into the cell, Iris Lindberg will talk about convertase pathways in neuroendocrine cells and how two convertase knockout mouse strains have shed light on the cell biology of peptide hormone production, suggesting novel roles for the binding proteins in neuroendocrine physiology. Patrick Allen will conclude the session with a description of how knockout mice lacking proteins that associate with protein phosphatase 1 contribute to regulation of the cytoskeleton, neurotransmission and synaptic plasticity.

Panel • Thursday • 4:30-6:30 PM • Twilight Regulation of Voltage-dependent Sodium Channels by Protein Kinases and Phosphatases Organizer: T. Scheuer; Participants: A.R. Goldin, A.R. Cantrell, P.C. Ruben Voltage-dependent sodium channels are the primary source of depolarizing in- ward current for neuronal action potentials. As such, their activity is critically important for the spread of excitation in neuronal cell bodies and dendrites, as well as for initiation and transmission of axonal action potentials. Dynamic regu- lation of sodium currents is expected to have important effects on these pro- cesses. Voltage-dependent sodium channels are targets for phosphorylation by a variety of protein kinases. However, the details of this phosphorylation and its effects, as well as entirely new modes of regulation are still being discovered. This panel will examine several aspects of regulation of sodium channels by phosphorylation. Sodium channels are known to be phosphorylated by protein kinase A (PKA) and protein kinase C (PKC) as well as by activa- tion. The best-characterized effect of PKA and PKC activation is to reduce the sodium current. However, increases in current by both PKA and PKC have been reported. Goldin will discuss mechanisms by which PKA can both increase and decrease sodium current through brain sodium channels. PKA, PKC and mem- brane potential interact in a cooperative manner to reduce sodium current. Cantrell will present evidence for phosphorylation at specific serines being responsible for reduction in sodium current by PKA and other, overlapping serines being

109 responsible for reduction in current by PKC when either kinase is activated alone. The cooperative effects of simultaneous activation of the kinases require addi- tional serines, some that are phosphorylated by PKA and others that are phos- phorylated by PKC. In addition to their rapid voltage-dependent activation and inactivation, sodium channels are subject to slow inactivation and the probabil- ity of slow inactivation depends on the particular sodium channel isoform. Those probabilities can be further modified in disease states and by association with sodium channel beta subunits. Ruben will discuss effects of phosphorylation on sodium channel slow inactivation that may underlie long-term changes in excit- ability. In addition to serine/threonine protein kinases, sodium channels are also subject to regulation by tyrosine phosphorylation. Scheuer will present evidence for reversal of tyrosine phosphorylation due to receptor tyrosine phosphatase beta that is bound specifically to sodium channel alpha and beta1 subunits.

Panel • Thursday • 4:30-6:30 PM • Rainbow Oligodendrocyte Precursors in Development, Health and Disease Organizer: J.W. Fawcett; Participants: W. D. Richardson, S. Goldman, J.M. Levine Oligodendrocyte precursors are responsible for myelination during development, but they do not disappear when this job is done. The adult CNS contains large numbers of cells with the antigenic profile of oligodendrocyte precursors. The function of these cells in the normal CNS is not fully understood, but they play a part in the CNS injury response, and probably in remyelination. The first speaker is Bill Richardson, who will talk about the oligodendrocyte precursor in develop- ment. Oligodendrocyte precursors have to spread throughout the nervous sys- tem to provide oligodendrocytes to all the myelinated pathways, so it has been a surprise to discover that their precursors have localized origins in the ventricular zones of the neural tube. In the embryonic spinal cord, oligodendrocyte progeni- tors are generated from a specialized region of the ventral VZ—the same egionr that at earlier times generates motor neurons. This suggests that the same neu- roepithelial precursors might first generate motor neurons then switch to pro- duction of oligodendrocyte progenitors. He reviews the evidence for a shared motor neuron-oligodendrocyte lineage and describes recent attempts to identify molecular components of the neuron-glial switch, together with more general questions—where do astrocytes fit into the picture, and what are the evolutionary implications of the close relationship between motor neurons and oligodendro- cytes? The second speaker is Joel Levine. He will talk about the adult oligoden- drocyte precursor. These cells are found in large numbers throughout the CNS. They contact nodes of Ranvier and also synapses, and may have roles in modu- lation of conduction and transmission at these sites. After injury large numbers of oligodendrocyte precursors are recruited to the lesion and activated, to be form major participant in the glial scar. Following demyelination these cells prob- ably provide the pool of cells for remyelination. Joel will review these various behaviours of the oligodendrocyte precursor and speculate about other possible functions. Steve Goldman will be speaking about the identification and isolation of a mitotically competent population of oligodendrocytic progenitor cells from

110 the adult human white matter. He will cover the promoter-specified, GFP-based extraction of these cells, their distribution and abundance, growth factor control and lineage potential. In addition, he will discuss the xenograft and integration of these cells into demyelinated adult brain, and will consider appropriate dis- ease models for their use. James Fawcett will talk about factors that might influ- ence the ability of the oligodendrocyte precursor and the Schwann cell to remyelinate after demyelination. The first step in remyelination is for the cells to migrate through the reactive astrocyte environment of the damaged CNS to- wards a demyelinated axon. However, the cells migrate poorly on astrocytes, inhibited by excessive adhesion mediated by N-cadherin. Myelination must then occur, and here again N-Cadherin plays a part, by mediating the first adhesive contacts between axon and myelin-forming cell.

Panel • Thursday • 4:30-6:30 PM • Skyline New Insights about Abnormal Developmental Processes and Their Relevance to Epilepsy Organizer: R.S. Sloviter; Participants: F.E. Jensen, K.S. Lee, D.H. Lowenstein, R.S. Sloviter The process of epileptogenesis has both intrinsic developmental and extrinsic acquired causes. Recent imaging data indicate that developmental anomalies and developmental processes play a much larger role than previously appreciated in subsequent epileptogenesis and other brain disorders, yet we do not understand how an abnormally developed network causes seizures. Dan Lowenstein will discuss new insights into the molecular basis of the development of the hippoc- ampus, and highlight potential developmental abnormalities that produce epi- leptogenic changes in the hippocampal network. Kevin Lee will address cases of human temporal lobe epilepsy that are associated with extratemporal develop- mental malformations of the neocortex. Although it is unknown exactly how these neocortical malformations impact the structure and/or function of the hip- pocampus, evidence from an animal model of neocortical dysplasia (the tish rat) suggests that developmental alterations in hippocampal circuitry precede the development of epilepsy, and may predispose the brain to seizures. Frances Jensen will address how the developmental regulation of neurotransmitter receptors modifies susceptibility to epileptogenesis in the immature brain, and how sei- zures during development can perturb receptor development to potentiate long term seizure susceptibility. Finally, Bob Sloviter will describe a previously unrec- ognized hippocampal malformation in human temporal lobe surgical specimens that may represent an unidentified developmental neurological defect long sus- pected of causing both febrile seizures in childhood and subsequent epilepsy.

111 Panel • Thursday • 4:30-6:30 PM • Sunset Vestibular Adaptation Organizer. L. Young; Participants: J. McElligott, M. Shelhamer, H. Hecht, L. Young The remarkable ability of the nervous system to interpret vestibular signals in unusual environments permits us to ski, to see look around with and without eyeglasses, and to adapt to the weightlessness of space flight. The adaptive char- acteristics of vestibulo-motor systems are examined with emphasis on recogni- tion of the context of the environment and the predictability of the stimulus. Young will introduce the problem of Coriolis stimulation in artificial gravity— when a centrifuge is used to overcome the debilitating effects of space flight. (Without vestibular adaptation the transitions between a rotating and non-rotat- ing spacecraft, popularized in the movie 2001 A Space Odyssey would be im- practical. Hecht will show how repeated head movements while rotating, although initially sickening, can lead to reductions of the vestibulo-ocular reflex (VOR), illusions of body tilt, and motion sickness itself. Shelhamer will deal with con- text-specific adaptation of the linear VOR and of saccadic eye movements. Con- text cues include head and eye orientation and gravity level. McElligott will introduce the predictability of the stimulus as an important factor in adaptation of the VOR.

Workshop • Thursday • 8:30-10:00 PM • Mt. Werner Is Enhanced Dopaminergic Neurotransmission the Primary Mediator of Hedonic Reward? A Comparison of Human and Animal Laboratory Studies Organizer: C. Bradberry; Participants: H. de Wit, G. Koob, T. Robinson, C. Bradberry This workshop will critically examine a concept which has wide acceptance within the neuroscience community, and is often referred to in the lay press, namely, that enhanced dopaminergic neurotransmission in the limbic striatum mediates the hedonic consequences of commerce with rewards. The majority of the evi- dence in support of dopaminergic mediation of reward derives from animal stud- ies. However, in human laboratory studies, which permit a verbalization of subjective effects of hedonic drugs, there are some results which appear inconsis- tent with this concept. This workshop will draw on both animal and human laboratory results and contrast them when possible. Harriet de Wit will present evidence from human laboratory studies examining whether dopamine antago- nists block the subjective effects of amphetamine, and will also briefly review other comparisons between human and animal laboratory results. George Koob will discuss evidence from animal studies and how it fits with a dopaminergic mediation of drug reward. Terry Robinson will present evidence that dopamin- ergic manipulations (including denervation) in rats do not alter hedonic responses to taste stimuli. Charles Bradberry will present data from primate microdialysis studies of cocaine and cocaethylene self-administration which, in comparison

112 with human laboratory studies, both support and contradict a dopaminergic mediation of cocaine reward.

Workshop • Thursday • 8:30-10:00 PM • Storm Peak Within-VTA Differences in Drug Action: A House Divided? Organizer: W. Carlezon; Participants: W. Carlezon, S. Ikemoto, R.A. Wise, D. van der Kooy The mesolimbic system, which originates in the ventral tegmental area (VTA) and projects to the nucleus accumbens (NAcc), is a neural substrate for the re- warding effects of many drugs of abuse. The NAcc has been divided into 2 main subregions, the nucleus accumbens core and the nucleus accumbens shell. These subdivisions have important anatomical and neurochemical differences, and ap- pear to be differentially involved in the rewarding effects of psychostimulants, opiates, and food. However, analogous functional subdivisions within the VTA have not been described. This workshop will address accumulating evidence that there are functional differences in drug action within the VTA. The panel will focus on studies involving drug reward mechanisms. Bill Carlezon (Harvard Medical School) will describe studies in which he used microinjections of viral vectors to elevate in the VTA expression of GluR1, an AMPA receptor protein that is regulated in this region by repeated drug exposure. Increased expression of GluR1 in the rostral portion of the VTA enhanced morphine reward, whereas increased expression of this protein in the caudal VTA enhanced morphine aver- sion. Satoshi Ikemoto (NIDA-Behavioral Neuroscience Branch) will describe intracranial self-administration studies in which he observes similar rostral-cau- dal differences in drug action. Rats self-administer GABA-A antagonists (but not agonists) into the rostral VTA, and GABA-A agonists (but not antagonists) into the caudal VTA. Roy Wise (NIDA-Intramural Research Program) will dis- cuss rostrocaudal differences in intracranial self-administration of endomorphin, a recently described endogenous opioid peptide. Derek van der Kooy (University of Toronto) will discuss two GABAergic reward phenomena involving the VTA, one that appears to be dopamine-dependent and one that appears to be dopam- ine-independent. Together, these observations suggest that there may be underappreciated within-VTA differences in drug action that can affect, and per- haps be affected by, motivational processes.

Workshop • Thursday • 8:30-10:00 PM • Twilight ‘Excito-toxicity’ vs ‘Excito-trophicity’: Do Seizures Switch the Set-Point for Survival? Organizer: K Gale; Participants: R. Simon, R. Gwinn, D. McIntyre Much attention has focused on neuronal cell death produced by prolonged con- tinuous seizures (status epilepticus). Less attention has been paid to the neu- rotrophic and neuroprotective action of repeated noninjurious brief seizures. This

113 workshop will explore the provocative concepts that seizures are a natural mecha- nism for inducing plasticity and neuroprotection, and that controlled seizures may have untapped therapeutic potential. We will discuss histopathological and molecular evidence that animals experiencing repeated seizures (evoked by elec- troconvulsive shock, kindling, or chemoconvulsants) are protected against apoptotic neuronal death induced by status epilepticus (in limbic structures), middle cerebral artery occlusion (in cortical regions), adrenalectomy (in dentate granule cells), and spinal cord contusion. Potential mechanisms for this neuroprotection will be considered, including induction of bFGF and upregulation of neuronal apoptosis inhibitory protein (NAIP). Several challeng- ing questions will be tackled with the help of the audience: Are trophic responses and cell death responses two sides of a coin or are they on a continuum? What determines the set-point for apoptotic reactions and how is this influenced by seizures? Is the seizure-evoked neuroprotection analogous to, or distinct from, ischemic tolerance ? What is the role of caspases and the bcl-2 genes? How can we find out if one key mediator or several tissue-specific factors is/are respon- sible for the enhanced survival after repeated seizures? Can subconvulsant stimu- lation work as well? Are there therapeutic implications? The participants will compare and contrast models and measurements in an effort to address these questions and generate others as part of a dynamic dialogue.

Workshop • Thursday • 8:30-10:00 PM • Rainbow What Really is the Food for Thought: Glucose or Lactate ? Organizer: I.A. Simpson; Participants: G. Dienel, E. Novotny, M. McKenna, L.Pellerin For many years glucose has been considered the obligate, if not sole, metabolic fuel for the brain. Studies with the non-metabolizable glucose analogue 2- deoxyglucose in experimental animals, and fluoro-deoxyglucose in humans (PET), provided a methodology for the measurement of regional glucose utilization associated with cerebral functions and the ability to diagnose pathological con- ditions such as Alzheimer s or Parkinson s Disease based on alterations in such utilization. Implicit in these observations was the assumption that the changes in accumulation of the labeled 2-DG occurred in neurons whose activity was un- der consideration. This assumption has been brought into question by the stud- ies of Magistretti and colleagues who have proposed that glucose is converted to lactate in glia, which it is subsequently exported to neurons for further oxidation. In the true spirit of a WCBR workshop, we propose an interactive forum includ- ing participants with diverse backgrounds and disparate view points. In the first hour the participants (3 slides, 5 minutes) will provide evidence to support or refute the neuronal use of glucose or lactate as primary energy substrate for the neuron. In the second hour, time will be provided for audience participation (1- 2 slides, 2-3 minutes) and more extensive discussion. Participants will also in- clude A. Gjedde and R.Greutter.

114 Panel • Thursday • 8:30-10:00 PM • Skyline Neurotrophin-3: A Tool to Query, and a Mechanism for, Sensory System Development and Plasticity Organizer: R. Lane; Participants: M. Jacquin, R. Rhoades, D. Wright Neurotrophins are involved in neuronal survival, proliferation, differentiation, growth and arborization of neuronal processes, synaptic refinement and plastic- ity. One of these factors, neurotrophin-3 (NT-3), has been identified at many sites during the development of the nervous system. Because of these features, the ability to manipulate the concentration of neurotrophins and specifically NT- 3 within the neuronal environment provides a valuable tool that can be utilized to study the development of the nervous system. The aim of this panel is to present data from recent experiments in which NT-3 concentrations are manipu- lated to explore specific aspects of somatosensory development. Mark Jacquin will outline the use of NT-3 to study trigeminal primary afferent growth and arbor formation in both in vivo and in vitro settings. Bob Rhoades will discuss the use of this neurotrophin to investigate the requirements for maintenance of the vibrissae-related pattern in the trigeminal brainstem complex of the rat. Doug Wright will discuss his use of transgenic mice that over express NT-3 in studying development of proprioceptive fibers and end organ morphogenesis associated with the mesencephalic nucleus of the trigeminal nerve. Rick Lane will describe NT-3 treatment as a method to investigate the relationship between the reorgani- zation observed in the brain stem with that seen in the primary somatosensory cortex of rats in response to a peripheral nerve injury.

Workshop • Thursday • 8:30-10:00 PM • Sunset Multifunctional Neurons and Networks for Hearing and Vocal Control Organizer: J.S. Kanwal; Participants: C.V. Mello, M.S. Fee, G.D. Pollak, Z.M. Fuzessery The functional organization of the nervous system has been classically exam- ined by neurophysiologists who study either a single sensory or a motor func- tion. Neuroanatomists also typically describe relatively stable components of neural organization, e.g., evolutionarily conserved lemniscal pathways, etc. These approaches have led to the view that parallel-hierarchical pathways in the central nervous system are dedicated to processing subsets of sensory information and that neurons become increasingly specialized to process specific parameters. This simplistic notion drives many neuroanatomical and physiological descriptions of the nervous system to date. A few studies, however, suggest that both, neurons and small networks of neurons can be involved in multifunctional roles. In fact, at various levels of sensory processing and especially within the cerebral cortex, different types of information may be multiplexed within the activity of single neurons and the configuration of local networks may change with the emotional

115 state of the animal, the sensory or motor signal and the recent history of neural activity. The aim of this panel is to present recent studies that illustrate multifunctional processing in auditory and vocal systems in vertebrates. Examples of this will be provided by experts working on songbirds and mustached bats: model species which have been extensively used for studies of the auditory system and in which recent research either directly or indirectly supports this idea. Thus, Claudio Mello working on budgerigars and songbirds has reported an overlap of auditory and vocal functions within audiovocal brain regions using gene expression analysis. Michale Fee has used intracellular recordings to show that single neurons can participate in different aspects of song production by functioning within mul- tiple time domains. In the laboratory of George Pollak, research at the level of the brainstem in bats shows that overlapping circuits are used for localizing single and multiple sounds. Zoltan Fuzessery and co-worker s recent research has shown that single neurons in the auditory cortex of pallid bats can play a role in both echolocation and passive sound localization. Finally, Jagmeet Kanwal s and oth- ers research has shown that single cortical neurons in mustached bats show a dual specialization for two distinct functions of the auditory system, namely, echolocation and sound communication. Whereas at subcortical levels multi- functional processing may be restricted to multiple dimensions of different as- pects of a single sensory modality, at the level of the association and sensory cortex this processing can become multisensory. More importantly, multifunc- tional processing within single neurons at higher cortical levels may be quite plastic. Thus, networks of neurons within the association areas may be frequently reconfigured so that single neurons process different types of stored and incom- ing neural signals.

Panel • Friday • 7:30-9:30 AM • Mt. Werner Location, Location, Location–Scaffolding Proteins in the CNS Organizer: D. Ron; Participants: C. Garner, H- J. Chung, R. Olsen, D. Ron Neurons function in a tightly orchestrated manner. In recent years it has become clear that scaffolding/anchoring proteins play a critical role in the molecular mechanisms that regulate synaptic function. Scaffolding/anchoring proteins are involved in clustering of ion channels, in coupling of receptors and ion channels to intracellular signaling pathways, in regulating the function of receptors and ion channels via phosphorylation and dephosphorylation, as well as connecting membrane proteins with the cytoskeleton. Scaffolding/anchoring proteins func- tion by compartmentalizing signaling proteins, receptors and ion channels or cytoskeletal proteins to distinct intracellular sites. Compartmentalization is usu- ally achieved via direct protein-protein interactions of well-defined protein bind- ing domains. Protein domains such as the PDZ domain enable scaffolding/ anchoring proteins to assemble signaling proteins such as kinases and phos- phatases from the same or from different cascades allowing tight regulation of cascades as well as crosstalk of signaling pathways. The speakers on the panel

116 will present examples that elucidate the significant role played by scaffolding/ anchoring proteins for both pre- and post synaptic events. Craig Garner (U. of Alabama at Birmingham) will present his work on the assembly of the presynap- tic active zone and on two presynaptic scaffolding proteins Piccolo and Bassoon that play an important role in synaptic vesicle trafficking at the active zone. Hee Jung Chung (Johns Hopkins) will present her work on the regulation of AMPA receptor internalization during synaptic plasticity by the anchoring proteins GRIP1 and PICK1. Richard Olsen (UCLA) will discuss the role of GABAA receptor interacting protein (GABARAP) in connecting the GABAA receptors to the cytoskeleton, and in the establishment of receptor clustering. Finally, Dorit Ron (Gallo Center, UCSF) will present work from her lab on RACK1 and its role as an inhibitory scaffolding protein for the NMDA receptor.

Panel • Friday • 7:30-9:30 AM • Storm Peak The Glutamate-Glutamine Cycle: Potential Targets for Modulating Glutaminergic Function Organizer: J.D. Erickson; Participants: A. Schousboe, M.P. Kavanaugh, R.H. Edwards, J.D. Erickson Glutamine is the preferred precursor for the neurotransmitter pool of glutamate. Upon synaptic release, glutamate is rapidly removed from the synaptic cleft by glutamate transporters that are located primarily on surrounding astroglial cells. There, glutamate is partially converted to glutamine by glutamine synthetase, an astrocyte-specific enzyme. To replenish the neurotransmitter pool of glutamate, glutamine is transported from astrocytes to glutaminergic neurons, converted to glutamate by phosphate-activated glutaminase, and then sequestered in synaptic vesicles by a vesicular transporter to make it available for regulated neurosecre- tion. This panel will revisit the glutamate-glutamine cycle and explore new tar- gets to modulate presynaptic glutaminergic function. First, Arne Schousboe will describe the evidence supporting the glutamate/glutamine cycle and the meta- bolic dependence of glutaminergic neurons upon glia to provide glutamine, as well as alternative precursors alpha-ketoglutarate and alanine for neurotransmit- ter synthesis. Michael Kavanaugh will consider astrocytic and neuronal glutamate transporters as potential targets for pharmacophore development and pharma- cologically review substrate and nonsubstrate inhibitors of the high-affinity, so- dium-dependent glutamate transporters. Robert Edwards will then present the identification of system N as a glutamate gated Na+/H+ exchanger linking glutamate efflux from astrocytes with neural activity through changes in intrac- ellular pH. Finally, Jeff Erickson will present novel system A glutamine/alanine transporters that are expressed on neurons that may be the molecular gateway for these substrates into glutaminergic neurons for neurotransmitter synthesis.

117 Panel • Friday • 7:30-9:30 AM • Twilight The Role of Pregnane Neurosteroids in Brain Function and Drug Action Organizer- L. Morrow; Participants: R. Rupprecht, C. Grobin, S. Mellon, L. Morrow Endogenous pregnane steroids, including 3a-hydroxy-5a-pregnan-20 one (3a,5a- THP, allopregnanolone) are present in brain in concentrations sufficient to modu- late ion channel receptor function during a variety of physiological events including stress, pregnancy, the estrus cycle and development. Exogenously ad- ministered 3a,5a-THP produces behavioral effects linked to potentiation of GABA-A receptor activity, including anxiolysis, sedation and anticonvulsant as well as antidepressant and neuroleptic-like actions. Mounting evidence suggests that neurosteroids may play a role in the psychopathology of depression and panic disorder. Furthermore, recent studies have shown that various psychoac- tive drugs, including fluoxetine, alcohol and olanzapine increase brain concen- trations of 3a,5a-THP. The goal of this panel is to present evidence that endogenous pregnane steroids modulate brain function and mediate the effects of various drugs. Rainer Rupprecht will present evidence that pregnane steroids interact with various ion channels and may play a significant role in depression, alcohol withdrawal and panic disorder. Chistina Grobin will present evidence that neurosteroids levels fluctuate across the ontogenetic development of the CNS and regulate the expression of GABA-A receptors and cortical neuron develop- ment in rats. Synthia Mellon will present evidence that specific serotonin reuptake inhibitors used for the treatment of depression and anxiety disorders directly enhance the biosynthesis of 3a,5a-THP. Leslie Morrow will present evidence that 3a,5a-THP contributes to ethanol action, tolerance and dependence and that novel neurosteroids may have therapeutic potential in the treatment of alco- hol-related problems.

Panel • Friday • 7:30-9:30 AM • Rainbow Neurotrophins and Depression Organizer: P. Skolnick; Participants: C.A. Altar, L.A. Mamounas, A. Russo-Neustadt Biogenic-amine based hypotheses of depression have dominated thinking in bio- logical psychiatry for more than four decades. However, during the past five years, converging lines of evidence have linked members of the NGF family of neu- rotrophic factors to both the pathophysiology of depression and the antidepres- sant actions of biogenic-amine based as well as electroconvulsive therapies. The objective of this panel is to overview neuroanatomical, neurochemical, and phar- macological studies that have linked alterations in neurotrophin expression to the pathophysiology of depression and chronic antidepressant treatment. Tony Altar will overview the plasticity of neuronal systems affected by neurotrophins and discuss the antidepressant-like actions of exogenously applied neurotrophins. L. Mamounas will describe her studies demonstrating the ability of neurotrophins

118 to promote the growth and function of intact serotonergic nerve terminals as well as the ability of these neurotrophins to facilitate the regrowth of serotoner- gic axons following chemical insult. A. Russo-Neustadt will review evidence that neurotrophin transcription is diminished by factors (e.g. stress) that can pre- cipitate depression and increased by chronic antidepressant. These findings will be discussed in the context of her work demonstrating that exercise can affect antidepressant-induced changes in BDNF transcription. P. Skolnick will describe strategies to develop novel, non-biogenic amine based antidepressants based on the data and principles described by the other participants.

Panel • Friday • 7:30-9:30 AM • Skyline Epileptogenesis: To Fit or Not to Fit Organizer: C. Wasterlain; Participants: E. Dudek, K. Staley, T. Sutula The nineteenth century dictum that seizur es beget seizures has recently re- ceived strong experimental support. Seizure-induced epileptogenesis is now rec- ognized as a common and widespread phenomenon, with several reliable experimental models in which status epilepticus induced chemically or electri- cally in healthy animals is followed after a silent period by the development of spontaneous seizures. However, the mechanisms and even the natural history of seizure-induced epileptogenesis are still largely unknown. Is the epilepsy that develops a molecular, cellular or network property? Is it a reflection of increased excitation, decreased inhibition or both? Is seizure-induced epilepsy a progres- sive disease in which each seizure adds to the likelihood of successive attacks, and does seizure control alter that natural history? What is the role of seizure- induced brain damage and of misdirected regeneration in that process? Is there a critical duration beyond which status epilepticus is epileptogenic, and are all agents that terminate status equally antiepileptogenic? The participants of this session are active investigators of epileptogenesis, and will review recent evi- dence that may suggest new answers to those questions. Wasterlain will describe the natural history of epilepsy following status epilepticus induced by perforant path stimulation in the rat, its temporal evolution and the factors that can modify it. Staley will examine the relationship of spike distribution to seizure develop- ment during the silent period . Sutula will discuss the role of NMDA receptor- dependent plasticity in the generation of granule cell spiking, and Dudek will present physiological evidence for the presence of a reverberating granule cell excitatory circuit in the epileptogenic hippocampus. The significance of these findings will be discussed in relation to the prevalent theories of epileptogenesis. Audience participation is strongly encouraged, and a limited number of slides ¥ can be shown from the audience (contact the organizer). BYOT (Bring your own theory)!

119 Panel • Friday • 7:30-9:30 AM • Sunset Prospects for Remyelination Therapies Organizer: T. Wood; Participants: J. Goldman, R. Armstrong, I. Duncan, R. Franklin Remyelination in the adult nervous system is critical to functional recovery from demyelinating diseases such as Multiple Sclerosis. Spontaneous remyelination of demyelinated axons occurs; however, this process is limited and ultimately fails after repeated demyelination events. Current research on promoting remyelinatio processes in the adult is focussed on investigations of endogenous mechanisms of spontaneous remyelination and on transplantation of myelin producing cells to promote remyelination. Studies on the endogenous mecha- nisms of remyelination include investigations of the adult oligodendrocyte pro- genitor cells and on the induction of growth and trophic factors in and around a demyelinating lesion. Recent evidence indicates that cycling progenitor cells in the adult mammalian CNS participate in endogenous remyelination. This en- dogenous remyelination is associated with the production of growth and trophic factors during the remyelination process. Jim Goldman will discuss the charac- teristics of the oligodendrocyte progenitor cells, including antigenic and mor- phologic heterogeneity, growth factor receptors, and responses to pathological circumstances. Regina Armstrong will talk about in vitro and in vivo studies of the PDGF and FGF pathways in the remyelination process in adult mice. Func- tional analysis of the role of PDGF and FGF2 in development and remyelination will be addressed by the use of pharmacologic and genetic manipulations of these pathways. Transplantation of myelin producing cells has emerged recently as a potential therapeutic approach to treat myelin disorders in humans. Which disorders could be treated and the cells that would be used, however, remains a matter of debate. Ian Duncan will discuss the major issues that need to be an- swered before therapeutic trials can be initiated. He will address key challenges that remain including issues of migration and myelination by transplanted cells and how focal lesions could potentially be repaired by either endogenous Schwann cells or primary oligodendrocytes. Robin Franklin will discuss an alternative for transplant-mediated remyelination, the use of olfactory ensheathing cells. Stud- ies will be presented on both rodent and human olfactory ensheathing cells, their phenotypes and remyelination potential in the context of a demyelinated envi- ronment.

Panel • Friday • 4:30-6:30 PM • Mt. Werner Ups and Downs in Synaptic Transmission: Factors that Affect Short-Term Synaptic Efficacy Organizer: R. E. Burke; Participants: L. Kaczmarek, B. Walmsley, F. Weight The efficacy of chemical synapses during repetitive activation is highly depen- dent of the frequency and patterning of input pulse trains. A great deal of work has been done on this topic over the last half-century not only because of its importance to CNS function but also because the behavior of different synaptic junctions during repetitive trains provides critical clues about the basic mecha- 120 nisms that control transmitter release. The development of in vitro models that provide access to pre- and postsynaptic elements for optical as well as electrical recordings. The resulting large body of data from a variety of synaptic systems have refined, and in some cases upset, classical views of short-term synaptic depression and facilitation. This panel will review some of this information, with particular emphasis on the multiple roles played by calcium in controlling pro- cesses of transmitter release and recovery during repetitive activation. Len Kaczmarek and Bruce Walmsley will discuss relevant findings made at the re- markably accessible calyx of Held in the brain stem auditory pathway in slice preparations. Forrest Weight will present studies on the short-term modulation of NMDA receptor-mediated responses by endogenous neuropeptides that are co-released with glutamate on repetitive presynaptic stimulation. Bob Burke will end the session with data on short-term depression of a synaptic system in the mouse spinal cord that, although not amenable to direct recording, also appear to involve similar mechanisms.

Panel • Friday • 4:30-6:30 PM • Storm Peak New Mechanisms for an Old Question: How does Prefrontal Cortex Affect Subcortical Dopamine Function Organizer: B. Moghaddam; Participants: S. Sesack, J.A. Kauer, J. Krystal Descending glutamatergic projections of the prefrontal cortex (PFC) are neces- sary for appropriate response of dopamine neurons in the ventral tegmental area (VTA) to stimuli within motivational and cognitive domains. Furthermore, clini- cal and basic investigations suggest that aberrant glutamatergic neurotransmis- sion in the PFC, leading to secondary disruption in the midbrain dopamine system, may contribute to the pathophysiology and etiology of schizophrenia. The aim of this panel is to present the most recent studies on the circuitry, plas- ticity, and neurochemical mechanisms underlying PFC- mediated changes in dopamine cell activity. These findings indicate that, contrary to previously hy- pothesized mechanisms, the PFC may exert a predominantly indirect and in- hibitory influence on subcortical dopamine function. Specifically, anatomical studies (Sesack) show that excitatory projections from the PFC synapse only onto dopamine cells that project back to the PFC and not those that project to the ventral striatum. In vivo neurochemical studies (Moghaddam) document dif- ferential tonic glutamate regulation of mesoprefrontal versus mesoaccumbens dopamine neurons, the latter being under tonic inhibitory AMPA control. Data from in vitro intracellular recording (Kauer) show that afferent stimulation of VTA dopamine neurons leads to long-term depression of glutamate synapses via influx of Ca2+ through voltage-gated channels. Finally, these basic findings will be integrated with recent clinical investigations (Krystal) employing imag- ing and pharmacological manipulations of glutamate and dopamine systems to help elucidate circuitry and physiological mechanisms that may have relevance to glutamate-dopamine interactions in healthy individuals versus those with schizophrenia.

121 Panel • Friday • 4:30-6:30 PM • Twilight Microarray Profiling: Complex Diseases and Drug Development Organizer: S. Potkin; Participants: W. Bunney, R. Thompson, N. Patil, M. Oliver This panel will review the recent developments in the area of expression array and gene chip technology, and their utilization to study complex diseases and drug development. This field has exploded especially in the last twelve months. In the past we have been able to compare the activity of single genes. With the new microarray technology we can analyze 15,000 genes simultaneously to con- trast gene expression in two different tissues or brain regions. Bunney will review the emerging application of microarray technology for the investigation of the differential expression of specific genes in complex diseases. Three approaches to microarray profiling will then be presented. Thompson will discuss the ex- pression array technology that starts with complimentary cDNA clones obtained from an expression sequence tagged (EST) database. New data utilizing this tech- nology will be presented comparing gene expression profiles obtained from de- fined brain regions in animals treated with different classes of antidepressants. These studies may contribute not only to our understanding of the mechanisms of action for existing antidepressants but also to the potential identification of novel antidepressant drug targets. Patil will contrast the advantages and disad- vantages of oligonucleotide arrays or gene chips. She will present an example of new data using Affymetrix GeneChip arrays to study neurogenesis, and to iden- tify a set of genes that are differentially expressed in cells isolated from the prolif- erating subventricular zone with the goal of identifying novel molecular markers differentially expressed in the neural precursors during various times of develop- ment. Oliver will present a new technology developed over the last year, the In- vader Assay. One of the critical issues in microarray profiling is the requirement of defining expression of different genes accurately, especially difficult with low levels of gene expression. The Invader Assay offers a new possibility to precisely quantify the expression levels of different genes even at low levels. At this mo- ment in science the extremely powerful combination of microarray technology combined with the recent progress in sequencing of the human genome provides hope for a new era in therapeutics, diagnostics and research.

Panel • Friday • 4:30-6:30 PM • Rainbow GABA Transporter Localization, Development and Function Organizer: W. Spain; Participants: F. Conti, M. Quick, G. Kinney, M. Jones Recent work on GABA transporters provides insight into the role of uptake mechanisms in regulating synaptic transmission. This panel will provide an over- view and recent developments in the study of these transporters. Bill Spain will provide a brief introduction of the subject. Fiorenzo Conti will discuss localiza- tion and development of GABA transporters in the CNS with a focus on GAT- 1, GAT-2 and GAT-3 in the cerebral cortex. Work on lower vertebrates will be related to localization of GAT-1 and GAT-3 in the human cerebral cortex. Also

122 data will be presented on altered expression of GAT-3 in some pathophysiologi- cal conditions. Mike Quick will discuss factors that regulate GAT-1 expression and function. He will review work on the redistribution of the transporter to/ from the plasma membrane as well as new data demonstrating that intracellular domains of the GABA transporter and protein-protein interactions govern sub- strate translocation, and identify a link between the machinery involved in trans- mitter release and re-uptake. Greg Kinney will discuss experiments using whole-cell recordings from astrocytes in rat neocortical brain slices to measure transporter currents during evoked synaptic release of GABA. The experiments provide insight into the time course and spread of GABA following release. The ultimate role of GABA transporters is to regulate the extracellular concentration of GABA. Matt Jones will present measurements of the concentration and time course of GABA released from single vesicles at synapses in hippocampal slices, the desensitization of synaptic GABA-A receptors resulting from transporter blockade, and Monte Carlo simulations illustrating the implications of these find- ings for GABAergic signaling.

Panel • Friday • 4:30-6:30 PM • Skyline Orphan G Protein-coupled Receptors as Novel Drug Targets Organizer: P. Iredale; Participants: O. Civelli, M. Yanagisawa, M. Fidock, S. Wilson G protein-coupled receptors (GPCRs) are a superfamily of plasma membrane proteins that are involved in a multitude of intracellular signaling pathways. His- torically, they have provided a rich source of targets for drug discovery which have subsequently resulted in numerous development candidates. To date ap- proximately 1000 GPCRs have been cloned of which about 1/5th are human sequences. Recently, following the advent of mass sequencing of expressed se- quence tags (ESTs) from cDNA libraries there has been a dramatic increase in the numbers of identified and cloned GPCRs. However, many of these remain as or phan GPCRs of unknown function with no identified ligand or apparent relationship to a disease indication. The aim of this panel is to review the current strategies for identifying cognate ligands for orphan GPCRs and to give examples of recent successes. After a brief introduction into the area by Phil Iredale, Olivier Civelli will continue by outlining the history of orphan ligand searching. He will begin by discussing the identification of orphanin FQ, focussing on the lessons learned, and subsequently how these have been applied to more recent examples. Next, Masashi Yanagisawa will present on the identification of the hypotha- lamic neuropeptides, orexin-A and -B, and their associated receptors. He will address findings suggesting a role for this novel peptide system in both the regu- lation of feeding and sleep. Mark Fidock will follow up with a review of the current methodologies that are being employed to search for ligands (including surrogates) at orphanDon’t GPCRs. forget Topicsto visit covered the willexhibit include area. the use of custom- ized G proteins, receptor internalization assays and constitutively active recep- tors. Lastly, Shelagh Wilson will discuss the role that orphan GPCRs play from a Pharmaceutical companies perspective, citing particular examples that have emerged as potential new drug targets. In summary, this panel will provide an overview of the current paradigms that have been developed to identify novel and interesting GPCRs and discuss some of the successes and lessons that have123 been learned along the way. 124 125 126 127 128 129 130 131 132 133 134 Index/Participant List Elizabeth Abercrombie Raymond Anton, 63 Rutgers University Medical University of South Carolina [email protected] [email protected]

James Adams Margie Ariano University Southern California Chicago Medical School [email protected] [email protected]

Bryon Adinoff, 63 Regina Armstrong, 119 University of Texas Southwestern Uniformed Services University [email protected] [email protected]

Greti Aguilera, 72 Andreas Arvanitogiannis National Institutes of Health Harvard Medical School [email protected] [email protected]

Elias Aizenman, 38, 93 Gary Aston-Jones, 66 University of Pittsburgh University of Pennsylvania [email protected] [email protected]

Elliot Albers Constance Atwell, 92 Georgia State University National Institute of Neurological [email protected] Disorders & Stroke [email protected] Kathryn Albers, 90, 93 University of Kentucky Peter Baas, 106 [email protected] University of Wisconsin [email protected] Patrick Allen, 108 The Rockefeller University Roy Bakay, 98 [email protected] Emory University [email protected] C. Anthony Altar, 62, 117 Otsuka America Pharmaceutical Inc Ruben Baler, 64 [email protected] National Institutes of Health [email protected] Arturo Alvarez-Buylla, 86 The Rockefeller University Marie Banich, 74 [email protected]. edu University of Illinois Urbana [email protected] Brenda Anderson, 48 SUNY Stony Brook Steve Barger, 67 [email protected] University of Arkansas bargerstevenw@exchange. uams.edu Rodrigo Andrade, 98 Wayne State University Allan I. Basbaum [email protected] University of California San Francisco [email protected] Kristin Anstrom, 91 Wake Forest University [email protected] 135 Anthony Basile Elizabeth Berry-Kravis, 86 National Institutes of Health RUSH Presbyterian St. Luke s Medical [email protected] Center [email protected] Gary Bassell, 23 Albert Einstein College James Bibb, 25 [email protected] The Rockefeller University [email protected] Steven Bealer University of Utah Charles Billington, 106 [email protected] Minneapolis VA Medical Center [email protected] Jill Becker, 87 University of Michigan Brian Billups, 24 [email protected] University of Leicester [email protected] Jo Begbie, 69 King s College London Murray Blackmore [email protected] University of Minnesota [email protected] Margery Beinfeld Tufts University David Bleakman, 68 [email protected] Eli Lilly and Company [email protected] Alvin Beitz, 77 University of Minnesota James R. Bloedel [email protected] Iowa State University [email protected] Aysenil Belger, 74 University of North Carolina Chapel-Hill Lidia C. Boffa, 97 [email protected] National Cancer Institute IST [email protected] Esther Bell, 69 The Rockefeller University Stephen Bondy [email protected] University of California Irvine [email protected] Gerald Benham Nikon Amitabha Bose, 63 [email protected] New Jersey Institute of Technology [email protected] Michael V.L. Bennett Albert Einstein College of Medicine Martyn Boutelle, 107 [email protected] King s College London [email protected] Deanna Benson, 62 Mount Sinai School of Medicine Michel Bouvier, 82 [email protected] University of Montreal [email protected] Albert Berger, 74 Sunny Boyd, 93 [email protected] University of Notre Dame [email protected] Craig Berridge University of Wisconsin [email protected]

136 Charles Bradberry, 111 Paul Chapman, 94 West Haven Veterans Administration Cardiff University [email protected] [email protected]

Daron Brown Dennis Choi, 93 Fine Science Tools Washington University [email protected] [email protected]

William Bunney, 121 Michael Chopp University of California Irvine Henry Ford Hospital [email protected] [email protected]

Robert Burke, 119 James Chou National Institute of Neurological Nathan Kline Institute Disorders & Strokes [email protected] [email protected] Carson Chow, 63 Gully Burns, 76 University of Pittsburgh University of Southern California [email protected] [email protected] Hsin Chu, 77 Allan Butterfield, 81 University of Oregon University of Kentucky [email protected] [email protected] Hee Jung Chung, 115 Angela Cantrell, 108 John Hopkins University University of Tennessee [email protected] [email protected] John A. Cidlowski, 72 Bill Carlezon, 112 National Institute Environmental Health Harvard Medical School Science [email protected] [email protected]

Susan Carlton Olivier Civelli, 122 University of Texas University of California Irvine [email protected] [email protected]

BJ Casey Paul Clarke, 97 Cornell University McGill University [email protected] [email protected]

William Catterall, 70 William Cobb University of Washington Rutgers University [email protected] [email protected]

David Chambers, 69 Vicki Coffin King s College London Shering Plough Research Institute [email protected] [email protected]

John Chapin Jonathan Cohen SUNY Health Science Center Brooklyn Princeton University [email protected] [email protected]

137 Carlo Colantuoni, 104 Kathryn A. Cunningham, 80 Johns Hopkins University of Texas [email protected] [email protected]

Douglas Cole, 105 Cynthia Czajkowski, 100 Vertex Pharmaceuticals University of Wisconsin [email protected] [email protected]

Graham L. Collingridge, 103 Mauro Dal Canto, 85 University of Bristol Northwestern University [email protected] [email protected]

Carol Colton Gudarz Davar, 77 Duke University Brigham and Women s Hospital [email protected] [email protected]

Paul Conn, 71 Beverly Davidson, 76 Emory University University of Iowa [email protected] [email protected]

Fiorenzo Conti, 121 Duff Davis Universita di Ancona Pfizer Inc [email protected] [email protected]

Richard Coppola, 104 Ron Davis, 89 National Institute of Mental Health Baylor College of Medicine [email protected] [email protected]

John Costello Geert De Vries, 65 The MIT Press Exhibits University of Massachusetts [email protected] [email protected]

Michael Costigan Taco De Vries, 66 Massachusetts General Hospital Harvard Research Institute Neurosciences Vrije [email protected] University [email protected] Carl W. Cotman, 88 University of California Irvine Harriet de Wit, 111 [email protected] University of Chicago [email protected] Fulton Crews, 48 University of North Carolina Chapel Hill Elizabeth Debski [email protected] University of Kentucky [email protected] Joseph Cubells, 79 Yale University Victor Denenberg [email protected] University of Washington [email protected]

138 Gersham Dent, 78 Robert Edwards, 116 University of Pennsylvania University of California San Francisco [email protected] [email protected] Michael Egan, 79 Lakshmi Devi, 82 National Institute of Mental Health New York University [email protected] [email protected] Mikhailova Elena Dennis Dickson Mental Health Research Center Mayo Clinic [email protected] [email protected] Helene Emsellem Gerald Dienel, 113 The Center for Sleep & Wake Disorders University of Arkansas [email protected] [email protected] Jeffrey Erickson, 116 Sylvain Dor , 25 Louisiana State University Johns Hopkins University [email protected] [email protected] Amy Eshleman, 104 Paula Dore-Duffy, 73 Oregon Health Sciences University Wayne State University [email protected] [email protected] Chris Evans, 95 Daniel M. Dorsa, 87 University of California Los Angeles University of Washington [email protected] [email protected] Alan I. Faden, 71 F. Edward Dudek, 118 Georgetown University Colorado State University [email protected] [email protected] Carolyn Fairbanks Ian Duncan, 119 University of Minnesota University of Wisconsin Madison [email protected] [email protected] James Fawcett, 109 Bruce Dunn, 107 Cambridge University University of California Los Angeles [email protected] [email protected] Michale Fee, 114 Tom Dunwiddie Bell Labs University of Colorado [email protected] [email protected] Mork Fidock, 122 Ford F. Ebner, 98 Pfizer Global Research and Development [email protected] [email protected] Howard Fields University of California [email protected] 139 David Fink William J. Freed, 104 University of Pittsburgh National Institute on Drug Abuse [email protected] [email protected] George Fink J. James Frost, 95 Pharmos Corp The Johns Hopkins University [email protected] [email protected] Denson Fujikawa Elena Fiorica-Howells, 123 University of California Los Angeles [email protected] [email protected] Zoltan Fuzessery, 114 Gerald Fischbach University of Wyoming National Institutes of Health [email protected] Raul Gainetdinov, 72 Cecilia Flores, 62 Duke University Harvard Medical School [email protected] [email protected] Karen Gale, 113 Francis Flynn Georgetown University University of Wyoming [email protected] [email protected] Aurelio Galli, 88 Bob Foehring, 98 University of Texas San Antonio University of Tennessee Memphis [email protected] [email protected] Craig Garner, 115 Ian Forsythe, 24, 70 University of Alabama Birmingham University of Leicester [email protected] [email protected] Don Gash Peter Fox University of Kentucky University of Texas [email protected] [email protected] Angela Gasser Robin Franklin, 119 S Karger AG University of Cambridge [email protected] [email protected] Scott Gehler Kyle Frantz, 36 University of Minnesota The Scripps Research Institute [email protected] [email protected] Herbert Geller, 76 Nigel Fraser Robert Wood Johnson Medical School University of Pennsylvania [email protected] [email protected] Alan Frazer, 88 Susan R. George, 82 University of Texas San Antonio University of Toronto [email protected] [email protected] Curt Freed Greg Gerhardt, 88 University of Colorado University of Kentucky [email protected] [email protected] 140 M. P. Gilbey, 102 Jeffrey Grimm, 49 Royal Free Campus National Institute on Drug Abuse [email protected] [email protected] Bruce Gold, 105 Matthew Grober, 65 Oregon Health Sciences University [email protected] [email protected] Alan Goldin, 108 Chistina Grobin, 117 University of California Irvine University of North Carolina Chapel Hill [email protected] [email protected] David Goldman, 79 Rolf Gruetter National Institute on Alcohol Abuse and University of Minnesota Alcoholism [email protected] [email protected] Gary Gudelsky, 84 James Goldman, 119 University of Cincinnati Columbia University [email protected] [email protected] Patrice G. Guyenet, 102 Steve Goldman, 109 University of Virginia Cornell University Medical College [email protected] [email protected] Ryder Gwinn, 113 Timothy Gomez, 99 Georgetown University University of Wisconsin [email protected] [email protected] Suzanne Haber Angela Goodacre University of Rochester Olympus America [email protected] [email protected] Randi Hagerman, 86 Anthony Grace, 66 University of Colorado Health Sciences University of Pittsburgh Center [email protected] [email protected] Robert Grainger, 77 Theo Hagg, 95 University of Virginia Dalhousie University [email protected] [email protected] Bill Greenough, 86 Edward D. Hall, 105 University of Illinois Urbana Pfizer Global Research and Development [email protected] [email protected] Karen Greif Shelley Halpain, 62 Bryn Mawr College The Scripps Research Institute [email protected] [email protected] Nigel Greig, 68 J. Marie Hardwick, 83 National Institute on Aging Johns Hopkins [email protected] [email protected] Sue Griffin, 67 University of Arkansas [email protected]. edu 141 Jay Hirsh, 89 Sami Harik University of Virginia University of Arkansas [email protected] [email protected] Bart Hoebel Neil Harrison Princeton University Cornell University [email protected] [email protected] Sandy Hofmann, 76 Teresa Hastings, 72 University Texas Southwestern University of Pittsburgh [email protected] [email protected] Gregg Homanics, 100 Edward Hawrot University of Pittsburgh Brown University [email protected] [email protected] John P. Horn, 102 Ulrike Heberlein, 89 University of Pittsburgh University of California San Francisco [email protected] [email protected] Richard Howells, 38 Heiko Hecht, 111 University of Medicine & Denistry of Massachusetts Institute of Technology New Jersey [email protected] [email protected] Johannes Hell Paul Huang, 37 University of Wisconsin Massachusetts General Hospital [email protected] [email protected] Hugh Hemmings Kim Huhman, 50 Weill Medical College of Cornell Georgia State University University [email protected] [email protected] Elaine Hull, 51 Joan Hendricks, 89 SUNY Buffalo University of Pennsylvania [email protected] [email protected] Michael Hutton, 75 Fritz Henn Mayo Clinic University of Heidelberg [email protected] [email protected] Thomas Hyde, 104 James Herman, 72 National Institute of Mental Health University of Cincinnati [email protected] [email protected] Satoshi Ikemoto, 112 Karl Herrup, 90 National Institute on Drug Abuse University Hospitals Case Western [email protected] Reserve [email protected] Donald Ingram, 68 National Institute on Aging Sandra Hewett, 26 [email protected] University of Connecticut Health Center [email protected]

142 Thomas Insel, 65 Elizabeth Jonas, 82 Emory University Yale University [email protected] [email protected]

Philip Iredale, 122 Mathew Jones, 121 Pfizer, Inc. University of Wisconsin Madison [email protected] [email protected]

John Isaac, 103 Reese Jones, 73 University of Bristol University of California San Francisco [email protected] [email protected]

Ole Isacson, 84 Richard Jones McLean Hospital Portland VA Medical Center [email protected] [email protected]

Michael Iuvone James Joseph, 68 Emory University Tufts University [email protected] [email protected]

Russell Jacobs, 76 Sharon Juliano, 90 California Institute of Technology Uniformed Services University HS [email protected] [email protected]

Mark Jacquin, 114 Kevin Ka-Wang Wang, 88 Washington University Pfizer Global R & D [email protected] [email protected]

Patricia Janak, 100 Leonard K. Kaczmarek, 70, 119 Univeresity of California San Francisco Yale University [email protected] [email protected]

Aaron Janowsky, 104 Katherine Kalil, 99 Portland VA Medical Center University of Wisconsin Madison [email protected] [email protected]

Luc Jasmin, 51 Jagmeet Kanwal, 114 University California San Francisco Georgetown University [email protected] [email protected]

Andrew Jenkins, 100 Rolf Karlstrom, 77 Weill Medical College University of Massachusetts [email protected] [email protected]

Frances Jensen, 110 Kathleen Karmel Children s Hospital Olympus America [email protected] [email protected]

Alan Kim Johnson Julie Kauer, 79, 120 University of Iowa Brown University [email protected] [email protected]

143 Michael Kavanaugh, 116 Gregory Kinney, 121 Oregon Health Sciences University University of Washington [email protected] [email protected]

Kristen Keefe Daryl R. Kipke, 100 University of Utah Arizona State University [email protected] [email protected]

George Keesler Joel Kleinman Aventis Pharmaceuticals National Institute of Mental Health [email protected] [email protected]

Ken Kellar, 82 Gregory W. Konat, 73 Georgetown University West Virginia University [email protected] [email protected]

Ann Kelley, 106 George F. Koob, 66, 111 University of Wisconsin Madison The Scripps Research Institute [email protected] [email protected]

Martin Kelly, 87 Jeffrey Kordower, 84, 98 Oregon Health Sciences University Rush Presbyterian Medical Center [email protected] [email protected]

John Kemp Conan Kornetsky Hoffmann-La Roche Ltd Boston University [email protected] [email protected]

Robert Kennedy, 107 Stephen Koslow University of Florida National Institute of Mental Health [email protected] [email protected]

Jason Kerr, 41 Catherine Kotz, 52 National Institute of Mental Health Veterans Administration Medical Center [email protected] [email protected]

Andrew Kertesz Arnold Kriegstein, 90 University of Western Ontario Columbia University [email protected] Claudia Krispel, 59 Stanislav Kholmanskikh University of California San Diego University of Minnesota [email protected] [email protected] John Krystal, 80, 120 Kwang-Soo Kim, 84 Yale University McLean Hospital Harvard Medical [email protected] School Leszek Kubin, 74 [email protected] University of Pennsylvania Ann Kingston, 71 [email protected] Eli Lilly Dietmar F. Kuhl, 94 [email protected] Hamburg University [email protected]

144 David Laidlaw, 76 Michael Levine, 92 Brown University University of California Los Angeles [email protected] [email protected]

Joseph LaManna Steve Levison, 96 Case Western Reserve University Penn State College of Medicine [email protected] [email protected]

Richard D. Lane, 114 Irwin Levitan Medical College of Ohio University of Pennsylvania [email protected] [email protected]

Ulo Langel, 97 Anita Lewin The Scripps Research Institute Research Triangle Institute [email protected] [email protected]

Dianne Lattemann, 106 Stafford Lightman, 72 University of Washington University of Bristol [email protected] [email protected]

Jean Lauder, 123 Iris Lindberg, 108 University of North Carolina Louisiana State University [email protected] [email protected]

Kevin Lee, 110 Jon Lindstrom, 82 University of Virginia University of Pennsylvania [email protected] [email protected]

Charles Leffler, 59 Diane Lipscombe, 70 University of Tennessee Brown University [email protected] [email protected]

Paul Lehmann, 85 Barbara Lipska Case Western Reserve University National Institute of Mental Health [email protected] [email protected]

Sarah Leibowitz Donald Lo The Rockefeller University Duke University Medical Center [email protected] [email protected]

Robert H. Lenox, 62 Peter Lobel, 76 University of Pennsylvania Robert Wood Johnson Medical School [email protected] [email protected]

Paul C. Letourneau, 99, 105 David Lodge, 103 University of Minnesota Lilly Research Centre [email protected] [email protected]

Barry Levin, 106 Edythe D. London, 63 VA Medical Center University of California Los Angeles [email protected] elondon@tracer

Joel Levine, 109 Daniel Lowenstein, 110 SUNY Stony Brook Harvard Medical School [email protected] [email protected]

145 Phillip Lowrey, 64 Oliver Manzoni, 79 Northwestern University CNRS [email protected] [email protected]

Joseph Ludwig, 27 Gerard Marek, 80 Mayo Clinic Jacksonville Yale School of Medicine [email protected] [email protected]

Ronald J. Lukas, 82 Neville Marks Barrow Neurological Institute Nathan Kline [email protected] [email protected]

William Lyman Luc Maroteaux, 123 Wayne State University Strasbourg University [email protected] [email protected]

Maria Maccecchini Michael Martin, 92 Annovis Inc National Institutes of Health [email protected] [email protected]

Bruce MacIver Susan Masino Stanford University University of Colorado [email protected] [email protected]

Ken Mackie, 82 Gary Mathern, 92 University of Washington University of California Los Angeles [email protected] [email protected]

Wendy Macklin, 96 Andrew Matus, 62 Cleveland Clinic Foundation Friedrich Miescher Institute [email protected] matus@fmi-ch

Daniel Madison Bob McCall Stanford University Pharmacia Corporation [email protected] [email protected]

Nigel Maidment, 107 Cyrus McCandless, 53 University of California Los Angeles University of Pittsburgh [email protected] [email protected]

Robert Malenka, 35 William McClure, 53 Stanford University University of Southern California [email protected] [email protected]

Laura Mamounas, 117 James McElligott, 111 Johns Hopkins Medical Institutions Temple University [email protected] [email protected]

Donna Maney, 93 Jacqueline F. McGinty, 78, 95 Johns Hopkins University Medical University South Carolina [email protected] [email protected]

Robert Mansbach Dan McIntyre, 58, 113 Pfizer, Inc. Carleton University [email protected] [email protected]

146 Hugh McIntyre David W. Miller, 75 University of California Los Angeles Massachusetts General Hospital [email protected] [email protected]

Mary C. McKenna, 113 Martha Miller, 96 University of Maryland Case Western Reserve University [email protected] [email protected]

F. Arthur McMorris Stephen Miller, 85 The Wistar Institute Northwestern University [email protected] [email protected]

Haley Melikian, 104 Rex Moats, 76 Harvard Medical School Childrens Hospital Los Angeles [email protected] moats @hsc.usc.edu

Claudio Mello, 114 Bita Moghaddam, 120 Oregon Health Sciences University Yale University [email protected] [email protected]

Synthia Mellon, 117 Derek Molliver, 90 University of California San Francisco Oregon Health Sciences University [email protected] [email protected]

Britt Mellstr m, 28 Eric Moody National Center Biotechnology Johns Hopkins [email protected] [email protected]

Marsha Melnick Frank Moore, 65 University California San Francisco Oregon State University [email protected] [email protected]

John Mendelson A. Leslie Morrow, 117 University of California San Francisco University of North Carolina Chapel Hill [email protected] [email protected]

Kalpana Merchant, 78 Yuko Munakata Pharmarcia Corporation University of Denver [email protected] [email protected]

Raju Metherate, 98 N. Eric Naftchi University of California Irvine New York University [email protected] [email protected]

Adrian C. Michael, 107 Angus Nairn University of Pittsburgh The Rockefeller University [email protected] [email protected]

John Mihic, 100 Kazunori Nakajima, 90 University of Texas Austin Jikei University [email protected] [email protected], [email protected] Neil Millar, 82 University College London [email protected]

147 T. Celeste Napier, 95 Edward Novotny, 113 Loyola University Yale University [email protected] [email protected]

Jose Naranjo, 94 Charles O Brien, 66 National Center Biotechnology University of Pennsylvania [email protected] [email protected]

Shridhar Narayanan, 40 Tim O Connor, 99 University of California Los Angeles University of British Columbia [email protected] [email protected]

Eugene Nattie, 74 Laura O Dell, 45 Dartmouth Medical School Scripps Research Institute [email protected] [email protected]

Alexandra Nelson, 40 Patricio O Donnell, 66 University of California San Diego Albany Medical College [email protected] [email protected]

David Nelson, 86 Randall O Reilly, 74 Baylor College of Medicine University Colorado Boulder [email protected] [email protected]

Kim Neve, 30 Anne Louise Oaklander, 55 Portland VA Medical Center Harvard Medical School [email protected] [email protected]

Madeleine Nicol, 28 Halina Offner, 73 Australian National University Oregon Health Sciences University [email protected] [email protected]

Roger Nicoll, 103 Murat Okatan, 39 University of California San Francisco Boston University [email protected] [email protected]

Eric Nisenbaum, 41, 68 Michael Olivier, 121 Eli Lilly & Company Stanford Human Genome Center [email protected] [email protected]

Laura Nisenbaum, 41 Richard W. Olsen, 115 Eli Lilly & Company University of California Los Angeles [email protected] [email protected]

Koichi Nishikawa Michelle Page Weill Medical College of Cornell University of Pennsylvania University [email protected] [email protected] Gene Palmer Ralph A. Nixon, 88 AstraZeneca R&D Boston Nathan Kline Institute [email protected] [email protected] Theo Palmer, 86 Stanford University Medical Center [email protected]

148 Marina Picciotto, 108 Jack M. Parent, 86 Yale University University of Michigan [email protected] [email protected] Chris Pierce Larry (Loren) Parsons, 80 Boston University The Scripps Research Institute [email protected] [email protected] Dietmar Plenz Kathy Partin, 68 National Institute of Mental Health Colorado State University [email protected] Yogesh C. Patel, 82 George D. Pollak, 114 McGill University University of Texas at Austin [email protected] [email protected] Nila Patil, 121 Linda Porrino, 63 Affymetrix Inc Wake Forest University [email protected] [email protected] Laura Pauples Steven Potkin, 121 University of Pennsylvania University of California Irvine [email protected] [email protected] David Pearce, 76 Nanduri Prabhakar, 101 University of Rochester Case Western Reserve University [email protected] [email protected] Robert Pearce Michael W. Quick, 121 University of Wisconsin University of Alabama Birmingham [email protected] [email protected] Betsy Pehek, 80 Remi Quirion Cleveland VA Medical Center McGill University [email protected] [email protected] Luc Pellerin, 113 Anjali Rajadhyaksha University of Lausanne McLean Hospital Harvard Medical [email protected] School Dale Pelligrino [email protected] University of Illinois Chicago Jan Marino Ramirez, 74, 101 [email protected] The University of Chicago Regino Perez-Polo, 68 [email protected] University of Texas Bruce Ranson [email protected] University of Washington Stephen Peroutka [email protected] iO Phamaceuticals Peter Reeh, 77 [email protected] University of Erlangen Pier Vincenzo Piazza [email protected] University of Bordeaux [email protected]

149 Peter H. Reinhart Utah State University Duke University Medical Center [email protected] [email protected] Jonathan Rubin, 63 Maarten E.A. Reith, 104 University of Pittsburgh University of Illinois Peoria [email protected] [email protected] Rainer Rupprecht, 117 Ian J. Reynolds, 93 Ludwig Maximilian University University of Pittsburgh [email protected] [email protected] muenchen.de

Robert W. Rhoades, 114 David Ruskin, 42 Medical College of Ohio National Institutes of Health [email protected] [email protected]

George Ricaurte, 72, 84 Amelia Russo-Neustadt, 117 Johns Hopkins University California State University Los Angeles [email protected] [email protected]

J. Steven Richardson, 81 Helen Scharfman, 86 University of Saskatchewan Helen Hayes Hospital [email protected] [email protected]

William Richardson, 109 Stephen Scheff, 57 University College London University of Kentucky [email protected] [email protected]

George Richerson, 74, 101 Todd Scheuer, 70, 108 Yale University University of Washington [email protected] [email protected]

Terry Robinson, 91, 111 Michael G. Schlossmacher, 75 University of Michigan Brigham and Women s Hospital [email protected] [email protected]

Dorit Ron, 115 Joann Schmidt, 43 Ernest Gallo Clinic and Research Center Pfizer [email protected] [email protected]

Oline K. Ronnekleiv Geoffrey Schoenbaum, 91 Oregon Health Sciences University Johns Hopkins University [email protected] [email protected]

Steven Roper, 92 Arne Schousboe, 116 University of Florida Royal Danish School of Pharmacy [email protected] [email protected]

Elliott Ross W. Schultz, 66, 91 University of Oklahoma University of Frihourg [email protected] [email protected]

Peter Ruben, 108 Joe Schwab

150 Pharmacia Ralph Siegel, 56 [email protected] Rutgers University [email protected] Joan P. Schwartz, 67 National Institute of Neurological Angela Siegling Disorders and Stroke Bayer AG [email protected] [email protected]

Roy Schwarz, 71 Eric J. Simon Pfizer Global Research & Development New York University [email protected] [email protected]

Stephan Schwarzacher, 123 Roger Simon, 113 University of Goettingen R.S.Dow Neurobiology Laboratories [email protected] [email protected]

Erik Schweitzer, 33 Ian Simpson, 113 University California Los Angeles Penn State University [email protected] [email protected]

Chris Sekirnjak, 36 Phil Skolnick, 117 Salk Institute for Biological Studies Eli Lilly and Company [email protected] [email protected]

David Self Ruth Slack, 106 University of Texas Southwestern University of Ottawa [email protected] rslack@uottawa

Kim B. Seroogy, 62 Robert Sloviter, 110 University of Kentucky University of Arizona [email protected] [email protected]

Susan Sesack, 120 Marion Smith University of Pittsburgh Stanford University [email protected] [email protected]

Yavin Shaham, 66 Mark A. Smith National Institute on Drug Abuse Case Western Reserve University [email protected] [email protected]

Mark Shelhamer, 111 Sherry Smith, 49, 81 Johns Hopkins University SUNY Health Science Center Brooklyn [email protected] [email protected]

Kent Shellenberger Gretchen Snyder, 43 Elan Pharmaceuticals The Rockefeller University [email protected] [email protected]

Toni Shippenberg, 95 Mark Sonders, 88 National Institutes of Health Oregon Health Sciences University [email protected] [email protected]

Beth-Anne Sieber Linda Sorkin National Institute of Mental Health University of California San Diego [email protected] [email protected]

151 William Spain, 121 C. N. Svendsen, 84 VA Puget Sound Health Care Systems University of Cambridge [email protected] [email protected]

Kevin Staley, 118 Thomas Swanson University of Colorado Midwest Neuroscience Inc [email protected] [email protected]

David G. Standaert, 75 Michael Tamkun, 101 Massachusetts General Hospital Colorado State University [email protected] [email protected]

Ursula Staubli, 68 Charlie Taylor, 32 Cortex Pharmaceuticals Inc Pfizer Global R & D [email protected] [email protected]

Donald Stein Jane Taylor, 62 Emory University Yale University [email protected] [email protected]

Heinz Steiner, 44 David Terman, 63 Finch University of Health Services Ohio State University [email protected] [email protected]

Nicholas Steneck, 92 Wolfram Tetzlaff, 95 University of Michigan University of British Columbia [email protected] [email protected]

Fiona Stevens Evelyn Thoman Oxford University Press University of Washington [email protected] [email protected]

Bradford Stokes Robert Thompson, 121 The Ohio State University University of Michigan [email protected] [email protected]

Alex Straiker, 45 Arthur W. Toga, 76 Salk Institute University of California Los Angeles [email protected] [email protected]

Wolfgang Streit, 95 Kimberly Topp University of Florida University of California San Francisco [email protected] [email protected]

Cheryl L. Stucky, 90 Mark A. Trocchi Medical College of Wisconsin Association Book Exhibits [email protected] [email protected]

David Sulzer, 72 Ramon Trullas, 47 Columbia University IIBB/CSIC [email protected] [email protected]

James Surmeier Li-Huei Tsai, 88 Northwestern University Harvard Medical School [email protected] [email protected]

152 Vincent K. Tuohy, 85 Jerrold Vitek, 98 Cleveland Clinic Foundation Emory University [email protected] [email protected]

Beth Tyler-McMahon, 97 Michael Vitek, 81 Mayo Clinic The Duke University [email protected] [email protected]

George Uhl, 78 Bruce Walmsley, 119 National Institutes of Health Australian National University [email protected] [email protected]

Harold Vaessin, 106 Judith Walters, 100 National Science Foundation National Institutes of Health [email protected] [email protected]

Carlos Valenzuela, 79 Claude Wasterlain, 118 University of New Mexico University of California Los Angeles [email protected] [email protected]

Derek van der Kooy, 112 Barry Waterhouse, 60, 100 University of Toronto Hahnemann University [email protected] b.waterhouse@drexel

Linda Van Eldik, 67 Cheryl S. Watson, 87 Northwestern University University of Texas [email protected] [email protected]

Arthur A. Vandenbark, 73 D. Martin Watterson, 31 Portland VA Medical Center Northwestern University [email protected] [email protected]

Susan Vannucci, 96 Daniel R. Weinberger, 79 Penn State Univeresity National Institute of Mental Health [email protected] [email protected]

Helene Varoqui, 29 Jeff Weiner, 79 Louisiana State Univeristy Wake Forest University [email protected] [email protected]

Jana Veliskova, 92 Daniel Weinreich, 102 Albert Einstein College of Medicine University of Maryland [email protected] [email protected]

J. Ernest Villafranca, 105 David Weiss, 100 Agouron Pfizer La Jolla University of Alabama Birmingham [email protected] [email protected]

Inez Vincent, 106 John Weiss, 93 University of Washington University of California Irvine [email protected] [email protected]

David Virshup, 64 David Wells University of Utah Yale University [email protected] [email protected]

153 Frank Welsh Donald Woodward, 100 University of Pennsylvania Wake Forest University [email protected] [email protected]

Monte Westerfield, 77 Clifford Woolf, 71 University of Oregon Massachusetts General Hospital [email protected] [email protected]

Susan White, 84 Doug Wright, 90, 114 Washington State University Kansas University [email protected] [email protected]

Scott Whittemore, 95 Bryan Yamamoto, 84 University of Louisville Case Western Reserve University [email protected] [email protected]

Kevin Wickman, 108 Massashi Yanagisawa, 122 University of Minnesota The University of Texas [email protected] [email protected]

George Wilcox, 77 F. Eugene Yates University of Minnesota University of California Los Angeles [email protected] [email protected]

John Williams Jerry Yin, 94 Oregon Health Sciences University Cold Spring Harbor Laboratory [email protected] [email protected]

Shelagh Wilson, 122 Larry Young, 65 SmithKline Beecham Emory University Shelagh [email protected] [email protected]

James Winslow, 93 Laurence Young, 111 Emory University Massachusetts Institute of Technology [email protected] [email protected]

Roy Wise, 112 Elmer Yu National Institute on Drug Abuse University of Pennsylvainia [email protected] [email protected]

Marina Wolf, 31 Hong Yu, 46 Chicago Medical School Eli Lilly & Company [email protected] [email protected]

Terri Wood, 119 James Zackheim Pennsylvania State University Rutgers University [email protected] [email protected]

Nancy Zahniser, 104 University of Colorado [email protected]

Michael Zigmond University of Pittsburgh [email protected]

154 Joshua Zimmerberg, 83 National Institutes of Health [email protected]