Neuroscience 2014 - Presentations from Winnipeg scientists

Abstracts are below the schedule.

  Session Start Time
Presentation Time
Location
Pres #
& Type
Poster Board # Authors & Institutions Abstract Title
Session # & Title
Sat 11/15 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
90.09
Poster
QQ24 W. M. SNOW1,4, C. CADONIC2,4, S. K. ROY CHOWDHURY4, E. THOMSON4, S. ALASHMALI3, E. PLATT4, M. SUH3, P. FERNYHOUGH1,4, *B. C. ALBENSI1,2,4; 1Pharmacol. & Therapeut., 2Biomed. Engin., 3Human Nutritional Sci., Univ. of Manitoba, Winnipeg, MB, Canada; 4Div. of Neurodegenerative Disorders, St. Boniface Hosp. Res., Winnipeg, MB, Canada Spatial learning capabilities and mitochondrial function in creatine-supplemented mice

090.Memory Consolidation and Reconsolidation
Sun 11/16 8:00 AM
8:00 AM - 12:00 PM

Halls A-C
156.06
Poster
DD21 *T. A. LAZAR1, S. SHOMSTEIN2, J. J. MAROTTA1; 1Perception and Action Lab, Dept. of Psychology, Univ. of Manitoba, Winnipeg, MB, Canada; 2George Washington Univ., Washington, DC Parts-based processing in autism

156.Visual Attention
Sun 11/16 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
211.02
Poster
B8 *Y. WANG1, H. TAN1, Z. ZHOU1, Y. SUN1, S. ZHU1, X.-M. LI2, J.-F. WANG1; 1Neurosci. Res. Program, Univ. of Manitoba, Winnipeg, MB, Canada; 2Dept. of Psychiatry, Univ. of Alberta, Edmonton, AB, Canada Nitrosylation of vesicular neurotransmitter transporters

211.Neurotransmitter Signaling in Health and Disease
Sun 11/16 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
243.01
Poster
EE5 *T. A. MCIVER1, J. KORNELSEN2, S. D. SMITH3, R. L. BOSMA1, H. S. KHAN1, A. I. COTOI1, R. H. Y. LEUNG1, P. W. STROMAN1; 1Ctr. for Neurosci. Studies, Queen's Univ., Kingston, ON, Canada; 2Dept. of Radiology, Univ. of Manitoba, Winnipeg, MB, Canada; 3Psychology, Univ. of Winnipeg, Winnipeg, MB, Canada Functional MRI reveals emotional modulation of pain processing in the human brainstem and cervical spinal cord

243.Pain Imaging and Perception
Mon 11/17 7:00 AM
7:00 AM - 8:00 AM

209A
SAT80
Satellite/Ancillary Events
  Kunjumon Vadakkan, Winnipeg, MB, Canada Satellite Organizer


SAT80.Exploring the Mind Using the Semblance Hypothesis
Mon 11/17 8:00 AM
8:15 AM - 8:30 AM

152A
286.02
Nanosymposium
  A. ALIZADEH, S. DYCK, D. NGUYEN, S. KALLIVALAPPIL, E. PROULX, E. EFTEKHARPOUR, *S. KARIMI-ABDOLREZAEE; Physiol., Univ. of Manitoba, Winnipeg, MB, Canada Neuregulin-1 therapy moderates reactive astrogliosis and scar formation following spinal cord injury

286.Spinal Cord Injury: Therapeutic Strategies
Mon 11/17 8:00 AM
9:30 AM - 9:45 AM

152A
286.07
Nanosymposium
  *E. EFTEKHARPOUR1, M. IQBAL2, N. PANDIAN3; 1Physiol., Regenerative Med. Group, and Spinal Cord Res. Cntr , Univ. of Mani, Winnipeg, MB, Canada; 2Physiol., 3Univ. of Manitoba, Winnipeg, MB, Canada Intracellular delivery of thioredoxin enhances neuroprotection in in vitro model of oxidative stress and in vivo model of spinal cord injury

286.Spinal Cord Injury: Therapeutic Strategies
Mon 11/17 8:00 AM
10:00 AM - 10:15 AM

152A
286.09
Nanosymposium
  *S. M. DYCK, A. ALIZADEH, E. PROULX, S. KARIMI-ABDOLREZAEE; Physiol., Univ. of Manitoba, Winnipeg, MB, Canada Chondroitin sulfate proteoglycans negatively modulate the properties of adult spinal cord neural precursor cells by signaling through LAR and PTPσ receptors and activation of the Rho/ROCK pathway

286.Spinal Cord Injury: Therapeutic Strategies
Mon 11/17 8:00 AM
8:00 AM - 12:00 PM

Halls A-C
299.13
Poster
B37 *J. E. RASH1, K. G. VANDERPOOL1, T. YASUMURA1, J. I. NAGY2; 1Biomed. Sci., Colorado State Univ., Fort Collins, CO; 2Physiol., Univ. of Manitoba, Winnipeg, MB, Canada KV1 coupling to connexin-29 to form "xenotypic" junctions between axons and myelin: Mechanism for removing K+ may contribute to increased conduction velocity

299.Ion Channels and Disease States II
Mon 11/17 8:00 AM
8:00 AM - 12:00 PM

Halls A-C
305.29
Poster
D29 *L. LU1,2, A. HOGAN-CANN1,2, C. ANDERSON1,2; 1Univ. of Manitoba, Winnipeg, MB, Canada; 2Neurosci. Res. Program, Kleysen Inst. for Advanced Med., Winnipeg, MB, Canada Astrocyte-induced cortical vasodilation is dependent on endothelial NMDA receptors

305.Astroglial Homeostasis and Function
Mon 11/17 8:00 AM
8:00 AM - 12:00 PM

Halls A-C
308.20
Poster
F5 *S. ZHU1, R. SHI1, J. WANG3, V. LI4, R. ZHANG5, A. TEMPIER3, J. KONG2, J.-F. WANG1, X.-M. LI3; 1Dept. of Pharmacol. and Therapeut., 2Dept. of Human Anat. & Cell Sci., Univ. of Manitoba, Winnipeg, MB, Canada; 3Dept. of Psychiatry, Univ. of Alberta, Edmonton, AB, Canada; 4Univ. of British Columbia, Vancouver, BC, Canada; 5Dept. of Psychiatry, The Fourth Military Med. Univ., Xi'an, China Quetiapine attenuates glial activation and proinflammatory cytokines in APP/PS1 transgenic mice

308.Microglia and Inflammatory Mediators in Neurodegeneration
Mon 11/17 8:00 AM
8:00 AM - 12:00 PM

Halls A-C
316.11
Poster
U30 *P. LU1,2, A. KAMBOJ2, C. M. ANDERSON1,2; 1Neurosci. Res. Program, Kleysen Inst. For Advanced Med., Winnipeg, MB, Canada; 2Univ. of Manitoba, Winnipeg, MB, Canada Neurotoxic NAD+ depletion leads to PARP-1-dependent PGC-1α hyperacetylation and mitochondrial dysfunction

316.Neurodegeneration II
Mon 11/17 8:00 AM
8:00 AM - 12:00 PM

Halls A-C
352.12
Poster
OO26 *A. D. HOGAN-CANN1,2, L. LU1,2, C. M. ANDERSON1,2; 1Univ. of Manitoba, Winnipeg, MB, Canada; 2Neurosci. Res. Program, Kleysen Inst. for Advanced Medicine, Hlth. Sci. Ctr., Winnipeg, MB, Canada Brain endothelial cells express NMDA receptors functionally linked to nitric oxide generation

352.Brain Blood Flow
Mon 11/17 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
394.30
Poster
A30 *H. MARZBAN1, K. BAILEY1, M. RAHIMI BALAEI1, A. U.MANNAN3, S. GHAVAMI2; 1Dept. of Human Anat. and Cell Sci., 2Univ. of Manitoba, Winnipeg, MB, Canada; 3Inst. of Human Genetics, Univ. Med. Ctr. Goettingen, Goettingen, Germany Corticogenesis of the cerebellar cortex in lysosomal acid phosphatase (Acp2) mutant mice

394.Postnatal Neurogenesis: Molecular Mechanisms
Tue 11/18 8:00 AM
8:00 AM - 12:00 PM

Halls A-C
507.20
Poster
I6 *Y. DAI1, L. M. JORDAN2; 1East China Normal Univ., Shanghai, China; 2Dept. of Physioloy, Univ. of Manitoba, Winnipeg, MB, Canada Activation of muscarinic receptors underlies cholinergic modulation of serotonergic neurons in the brainstem of ePet-EYFP mice

507.Cellular Mechanisms of Modulation of Neuronal Firing Properties
Tue 11/18 8:00 AM
8:00 AM - 12:00 PM

Halls A-C
515.14
Poster
V18 *M. RASTEGAR; Biochem. and Med. Genet., Univ. of Manitoba, Winnipeg, MB, Canada MeCP2 isoforms and neurological disorders

515.Rett Syndrome
Tue 11/18 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
593.01
Poster
B29 *T. HABASH1,2, A. SALEH1, S. ROY CHOWDHURY1, P. FERNYHOUGH1,2; 1St-Boniface Hosp. Res. Ctr., Winnipge, MB, Canada; 2Pharmacol. and therapeutics, Univ. of Manitoba, Winnipeg, MB, Canada The proinflammatory cytokine, interleukin-17A, augments axonal plasticity and mitochondrial function of cultured adult sensory neurons

593.Axon Growth and Guidance: Intrinsic Mechanisms
Tue 11/18 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
593.03
Poster
B31 *E. D. SCHARTNER1,2, A. SALEH1, R. VIEIRA DA SILVA1, S. CHOWDHURY1, D. SMITH1, P. FERNYHOUGH1,2; 1St. Boniface Hosptial Res., Winnipeg, MB, Canada; 2Pharmacol. and Therapeut., Univ. of Manitoba, Winnipeg, MB, Canada Sirtuin 2 is a sensor of energy status and inducer of neurite outgrowth in adult sensory neurons

593.Axon Growth and Guidance: Intrinsic Mechanisms
Tue 11/18 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
595.01
Poster
C12 M. M. SIDDIQ1, S. S. HANNILA2, Y. ZORINA3, V. RABINOVICH1, E. NIKULINA4, *W. MELLADO5, R. IYENGAR1, M. T. FILBIN6; 1Mount Sinai Sch. of Med., New York, NY; 2Univ. of Manitoba, Winnipeg, MB, Canada; 3Acorda Therapeut., Ardsley, NY; 4The Feinstein Inst. for Med. Res., Manhasset, NY; 5Burke Med. Res. Inst., White Plains, NY; 6Hunter Col., New York, NY Extracellular histones, a putative inhibitor of cns axonal regeneration, are blocked by activated protein c (apc)

595.Extrinsic Mechanisms Controlling CNS Regeneration
Tue 11/18 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
609.08
Poster
T8 *S. S. HANNILA, N. JAHAN, S. GHAVAMI; Human Anat. and Cell Sci., Univ. of Manitoba, Winnipeg, MB, Canada Transforming growth factor beta-induced expression of chondroitin sulphate proteoglycans in reactive astrocytes: roles of non-Smad signaling pathways and autophagy

609.Spinal Cord Signaling in Trauma
Tue 11/18 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
621.10
Poster
BB23 *A. R. HIRSCH1, K. V. GAFTANYUK2; 1Smell & Taste Treatment and Res. Fndn., Chicago, IL; 2Intl. Univ. of the Hlth. Sci. Sch. of Med., Winnipeg, MB, Canada Aftertaste sans taste: ageusia with palinageusia

621.Taste
Tue 11/18 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
631.07
Poster
II16 *B. R. NOGA1, F. J. SANCHEZ1, C. O'TOOLE1, L. VILLAMIL1, S. STASIENKO2, S. KASICKI2, U. SLAWINSKA2, L. M. JORDAN3; 1Miami Project To Cure Paralysis, Univ. of Miami Sch. of Med., MIAMI, FL; 2Dept. of Neurophysiol., Nencki Inst. of Exptl. Biol., Warsaw, Poland; 3Dept. of Physiol., Univ. of Manitoba, Winnipeg, MB, Canada Theta frequency oscillations predominate in the mesencephalic locomotor region during voluntary treadmill locomotion

631.Afferent and Descending Control
Tue 11/18 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
654.06
Poster
UU10 H. WANG1, S. LI1, G. LIU1,2, *G. J. KIROUAC3; 1Oral Biol., Univ. of Manitoba, Winnipeg, MB, Canada; 2Intl. School, Guangdong Food and Drug Vocational Col., Guangzhou, China; 3Oral Biol. and Psychiatry, Fac. of Dent., Winnipeg, MB, Canada Changes in the orexin (hypocretin) system in rats following footshock exposure

654.Motivation and Emotions: Fear and Pain
Wed 11/19 8:00 AM
8:00 AM - 12:00 PM

Halls A-C
687.11
Poster
B14 L. ROBERTS, W. XIONG, N. LEVINE, M. F. JACKSON, *F. E. PARKINSON; Univ. Manitoba, Winnipeg, MB, Canada Localization of equilibrative nucleoside transporter 3 (ENT3) in mouse brain

687.Acetylcholine and Other Transporters
Wed 11/19 8:00 AM
8:00 AM - 12:00 PM

Halls A-C
756.06
Poster
VV5 *S. LI1, G. J. KIROUAC2; 1Oral Biol., 2Oral Biol. and Psychiatry, Univ. of Manitoba, Winnipeg, MB, Canada Sources of dopamine fibers in the paraventricular nucleus of the thalamus

756.Motivation and Emotions: Reward Circuitry
Wed 11/19 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
800.18
Poster
U24 *R. SHI1, S. ZHU2, V. LI4, S. B. GIBSON3, X. XU5, J. KONG1; 1Dept. of Human Anat. and Cell Sci., 2Dept. of Pharmacol. & Therapeut., 3Dept. of Biochem. and Med. Genet., Univ. of Manitoba, Winnipeg, MB, Canada; 4Fac. of Med., Univ. of British Columbia, Vancouver, BC, Canada; 5Inst. of Neurosci., Soochow Univ., Suzhou, Jiangsu Province, China BNIP3 interacting with LC3 triggers excessive mitophagy

800.Ischemia: Cellular Mechanisms and Neuroprotection V
Wed 11/19 1:00 PM
1:00 PM - 5:00 PM

Halls A-C
809.17
Poster
Z30 Z. ZHOU1, Y. WANG1, H. TAN1, Y. SUN1, Y. CHE2, *J.-F. WANG1; 1Dept. of Pharmacol. & Therapeutics, Fac. of Medicine, Univ. of Manitoba, Winnipeg, MB, Canada; 2Med. Col. of Soochow Univ., Suzhou, China Chronic treatment with mood stabilizer lithium inhibits amphetamine-increased risk-taking behaviour in rats

809.Amphetamine Reinforcement II


ABSTRACTS

90.09/QQ24. Spatial learning capabilities and mitochondrial function in creatine-supplemented mice
W. M. SNOW1,4, C. CADONIC2,4, S. K. ROY CHOWDHURY4, E. THOMSON4, S. ALASHMALI3, E. PLATT4, M. SUH3, P. FERNYHOUGH1,4, *B. C. ALBENSI1,2,4;
1Pharmacol. & Therapeut., 2Biomed. Engin., 3Human Nutritional Sci., Univ. of Manitoba, Winnipeg, MB, Canada; 4Div. of Neurodegenerative Disorders, St. Boniface Hosp. Res., Winnipeg, MB, Canada
Objective: Dietary supplementation with creatine, a neuroprotective organic acid, has been shown to improve memory in humans and animals. Whether creatine enhances the acquisition and retention of spatial memory in the Morris water maze (MWM), a well-characterized rodent memory assay, is unknown. Further, the physiological mechanisms for the memory enhancements reported thus far are unclear. The transcription factor nuclear factor kappa B (NF-κB) has been implicated in creatine-induced enhancements in cultured neurons. CREB signaling is implicated in creatine-enhanced spatial memory assayed using the Barnes maze in mice. We have shown a role for NF-κB in maintaining synaptic plasticity, memory, and neuronal energy homeostasis. Thus, this study sought to investigate: 1) the effects of creatine supplementation on hippocampal-dependent spatial memory in the MWM; 2) the effects of creatine on neuronal mitochondrial function in vitro; 3) CREB and NF-κB levels in the hippocampi of creatine-fed mice; and 4) the NF-κB complex as a possible target of creatine-induced memory enhancements and mitochondrial function. Methods: C57BL/6 male mice (age 7 mos) were fed either a creatine-supplemented (3% w.w.) or control diet for 8-9 wks. During wk 8, mice underwent MWM training to assess spatial learning and memory. In a subset of animals, mitochondrial function was assessed (i.e., oxygen consumption rates (OCR); XF24 Analyzer, Seahorse Biosciences)) following MWM training. Cortical neurons from embryonic mice were cultured. Mitochondrial bioenergetics were measured in creatine-treated (1 mM) vs. untreated neurons alone and in the presence of sulfasalazine, an NF-κB blocker. Western blot experiments were used to determine hippocampal protein levels of CREB and NF-κB with and without creatine. Results: In vivo, creatine dietary supplementation significantly enhanced spatial learning in the MWM (p<0.05) relative to control-fed mice. In vitro, creatine increased OCR in cultured cortical neurons vs. untreated neurons (p<0.05). Blockade of NF-κB with sulfasalazine, however, resulted in diminished mitochondrial OCR (p<0.05). Western blot experiments to determine NF-κB and CREB levels are ongoing. Conclusions: These data establish creatine-induced enhancements in learning and neuronal cellular bioenergetics in mice and suggest the involvement of NF-κB as a key regulator of energy homeostasis. Such research has important implications for the treatment of disorders affecting memory, including Alzheimer´s disease. Funding from Natural Sciences and Engineering Research Council (NSERC), the St. Boniface Hospital Foundation, and the Everett Endowment Fund



156.06/DD21. Parts-based processing in autism
*T. A. LAZAR1, S. SHOMSTEIN2, J. J. MAROTTA1;
1Perception and Action Lab, Dept. of Psychology, Univ. of Manitoba, Winnipeg, MB, Canada; 2George Washington Univ., Washington, DC
Research suggests that individuals with autism spectrum disorders (ASD) process their environment in a local, parts-based manner (Happé & Frith, 2006; Plaisted et al., 1999). This local bias could possibly be driven by difficulties in grouping stimuli based on Gestalt principles, an early pre-attentional perceptual system required for perception of global structures (Han & Humphreys, 1999). We tested this hypothesis by adopting a Moore & Egeth, 1997 paradigm, in which the Ponzo illusion was induced by a set of background dots that surrounded two solid lines. The illusion could only be induced by the virtue of perceptually grouping the background dots into lines based on color similarity. We hypothesized that adults with high functioning ASD are less susceptible to the effects of the illusion than typically developing controls, due to a tendency to process the items in a parts-based manner (i.e., failing to group the background). A line discrimination task was administered in which two horizontal lines were superimposed on a background of black and white dots. On occasion, black and white background dots were presented so that if automatically grouped and integrated, they should influence the perceived length of the horizontal lines. Results showed that on average, individuals with ASD were significantly less likely than controls to report an illusion-based response. In fact, the mean percent correct for responding consistently with the illusion did not differ significantly from 50% in the ASD group, indicating at chance performance. These results suggest that individuals with ASD experience deficits in perceptual grouping abilities at early, pre-attentive levels of visual processing. Difficulties in grouping stimuli according to Gestalt principles may explain why individuals with ASD are less inclined to use global context very early in perception and tend to rely on local visual processing strategies. These findings may have broader implications in forming behavioral therapies and intervention strategies that help address the unique way that individuals with ASD process the world, leading to improved quality of life in the classroom, workforce, and society in general.



211.02/B8. Nitrosylation of vesicular neurotransmitter transporters
*Y. WANG1, H. TAN1, Z. ZHOU1, Y. SUN1, S. ZHU1, X.-M. LI2, J.-F. WANG1;
1Neurosci. Res. Program, Univ. of Manitoba, Winnipeg, MB, Canada; 2Dept. of Psychiatry, Univ. of Alberta, Edmonton, AB, Canada
Brain is the main site for oxygen consumption (80%), making the brain cells vulnerable to production of reactive oxygen/nitrogen species (ROS/RNS). Vesicular transporters such as vesicular monoamine transporter 2 (VMAT2) vesicular glutamate transporter 1 (VGluT1) and vesicular glutamate transporter 2 (VGluT2) are key proteins for packaging and uploading neurotransmitters into vesicles. These transporters have high density of cysteine residues that are susceptible to ROS/RNS attack. Therefore we determined whether nitric oxide radical donor S-nitrosoglutathione (GSNO) induces nitrosylation of VMAT2, VGluT1 and VGluT2 and whether GSNO regulates vesicular uptake activity of glutamate in mouse brain. Cysteine nitrosylation of the transporters was measured by biotin-switch method followed by immunoblotting analysis, and tritium labeled glutamate was used to measure the uptake of glutamate by vesicles. We found that GSNO at 80 uM GSNO treatment significantly increased nitrosylation of VMAT2, VGluT1 and VGluT2 and inhibited the uptake of glutamate by vesicles. Our finding suggested that nitrosylation of vesicular transporters may play a role in the regulation of monoamine and glutamate neurotransmission.



243.01/EE5. Functional MRI reveals emotional modulation of pain processing in the human brainstem and cervical spinal cord
*T. A. MCIVER1, J. KORNELSEN2, S. D. SMITH3, R. L. BOSMA1, H. S. KHAN1, A. I. COTOI1, R. H. Y. LEUNG1, P. W. STROMAN1;
1Ctr. for Neurosci. Studies, Queen's Univ., Kingston, ON, Canada; 2Dept. of Radiology, Univ. of Manitoba, Winnipeg, MB, Canada; 3Psychology, Univ. of Winnipeg, Winnipeg, MB, Canada
Emotions can modulate pain perception. In line with Motivational Priming Theory, painful stimuli are generally experienced as more painful when accompanied by, or associated with a negative emotional influence, while pain ratings decrease with positive emotional influence. Functional magnetic resonance imaging (fMRI) has been used to investigate pain and its underlying neural mechanisms throughout the central nervous system. Although fMRI studies have examined emotional modulation of pain processing in the brain, this provides an incomplete representation of the process. Pain processing areas are widely distributed across the brain, brainstem, and spinal cord. To gain a complete understanding of how emotion influences pain, we need to characterize the neural response in the brainstem and spinal cord. The present study used fMRI of the spinal cord and brainstem to examine the effect of emotional modulation on thermal pain processing and neural function at this level. Behavioral testing identified healthy, heterosexual, right-handed adult females who exhibited emotional modulation of pain perception. Painful heat stimulation (via Medoc® TSA-II Thermal Sensory Analyzer) was calibrated for each participant to produce a moderately painful sensation and applied on the thenar eminence of the right hand, corresponding to the C6 dermatome. Imaging sessions entailed 12 functional runs, with the painful stimulus applied in a block paradigm. For the duration of each run, participants viewed images selected from the International Affective Picture System of negative, neutral, or positive valence, with image valence held constant throughout each run. Participants reported the sensation´s intensity and unpleasantness using pain rating scales ranging from 0 -100. Data analysis for fMRI included use of a general linear model, with subsequent contrasts to compare BOLD responses between conditions. We observed significant differences in pain perception, reflecting the influence of emotional valence of the visual stimuli on pain intensity and unpleasantness. Similarly, contrast analysis revealed significantly greater BOLD percent signal change, in the ipsilateral dorsal horn (C6) corresponding to the painful stimulus during negative picture viewing, relative to both positive and neutral picture viewing conditions. Image valence and arousal appear to have varying effects on BOLD responses to pain in different regions of the brainstem, including the periaqueductal gray, rostral medulla, and the nucleus tractus solitarius. These results provide evidence for descending emotional modulation of brainstem and spinal mechanisms involved in pain processing.



SAT80. Satellite Organizer
Kunjumon Vadakkan, Winnipeg, MB, Canada
Identifying the set of neurons that fire during a task can become meaningful if we discover how they are related to the formation of the inner sensations during the execution of the task. First-person properties of the mind can only be accessed by the owner of the nervous system. The testable Semblance Hypothesis is trying to find a mechanism that can explain most brain functions; replicating neurons firing in an artificial system and obtaining readouts of the inner sensations to make it accessible to a second person. Identifying the right frame of reference that can connect major findings by various faculties of brain sciences at different levels - molecular, cellular, electro-physiological, systems, and behavioral - is essential to understand the mind. We will explain the steps towards the artificial replication of the hypothesized mechanism.



286.02. Neuregulin-1 therapy moderates reactive astrogliosis and scar formation following spinal cord injury
A. ALIZADEH, S. DYCK, D. NGUYEN, S. KALLIVALAPPIL, E. PROULX, E. EFTEKHARPOUR, *S. KARIMI-ABDOLREZAEE;
Physiol., Univ. of Manitoba, Winnipeg, MB, Canada
Introduction: Reactive astrogliosis is a key pathophysiological event after spinal cord injury (SCI). Activated astrocytes secrete a myriad of pro-inflammatory cytokines, nitric oxide (NO) and inhibitory extracellular matrix components including chondroitin sulphate proteoglycans (CSPGs) that cause neurotoxicity and impede tissue repair and regeneration. Our recent evidence suggests that drastic downregulation of Nrg-1 after SCI may underlie astrocytes reactivity following injury. Here, using complementary in vitro and in vivo approaches, we examined the impact of Nrg-1 on astrocyte activation. We demonstrate that availability of Nrg-1 mitigates multiple detrimental consequences of reactive astrogliosis in SCI. Methods: We utilized an in vivo rat model of incomplete compressive SCI and two in vitro models of reactive astrogliosis. In SCI rats, recombinant human Nrg-1β1 (rhNrg-1β1) was delivered intrathecally after SCI using mini-osmotic pumps and spinal cord tissue was analyzed by western blotting, immunohistochemistry and stereological techniques at different intervals post-SCI. For in vitro astrogliosis, primary rat astrocyte cultures were activated using lipopolysaccharide (LPS) or transforming growth factor-beta (TGF-β) and then Nrg-1 with or without its neutralizing antibody was added to the cultures. Astrocyte conditioned media (ACM) and cell lysate were collected and analyzed using immunocytochemistry, enzymatic assays, and Western and slot blotting to assess cellular and molecular characteristics of astrocyte reactivity including oxidative stress markers, CSPGs and proinflammatory cytokines. Results: We report that Nrg-1 treatment significantly attenuates several inhibitory and toxic aspects of reactive astrogliosis including CSPG production and the release of proinflammatory cytokines (TNF-α and IL-1β) in ACM of LPS-activated astrocytes. Additionally, Nrg-1 activation can mitigate oxidative stress in activated astrocytes by reducing their NO production. Moreover, Nrg-1 availability attenuated cell proliferation and nestin upregulation, two cellular characteristics of reactive astrogliosis. In vivo administration of Nrg-1 following SCI reduced CSPG production and chronic scar formation. The effects of Nrg-1 were specific as inactivated Nrg-1 using its neutralizing antibody failed to exhibit such effects when used in vitro. Conclusion: Our work provides novel insights into the mechanisms of astrogliosis following an injury and suggests a positive role for Nrg-1 in ameliorating the outcomes of SCI.



286.07. Intracellular delivery of thioredoxin enhances neuroprotection in in vitro model of oxidative stress and in vivo model of spinal cord injury
*E. EFTEKHARPOUR1, M. IQBAL2, N. PANDIAN3;
1Physiol., Regenerative Med. Group, and Spinal Cord Res. Cntr , Univ. of Mani, Winnipeg, MB, Canada; 2Physiol., 3Univ. of Manitoba, Winnipeg, MB, Canada
Increasing evidence indicate the importance of oxidation/reduction balance (Redox status) in cell biology. A sudden rise of oxidizing reactive oxygen species (ROS) levels after injury will quickly interrupt the redox status of the cell resulting in oxidation of vital proteins and initiation of cell death pathways. It is therefore hypothesized that enhancement of cell reducing capacity may have protective effects in oxidative stress related conditions. During the last two decades antioxidant therapies have been extensively used for treatment of neurotrauma buthave failed to generate optimal results in clinical applications. This indicates the need for novel treatment options. Thioredoxin (trx) is a 12kD protein with a central role in regulation of oxidative stress through maintaining the cellular proteins in reduced state. Intravenous administration of high doses of Trx has been shown to be protective in models of brain ischemia. Additionally, recent evidence indicates the ability of Trx for enhancing cell proliferation in adult brain-derived neural stem cells, suggesting its potential role for regeneration. The underlying protective mechanisms of intravenous trx therapy remain to be identified. Plasma Trx has a relative short half-life and does not cross the blood brain barrier (BBB) and therefore the extent of Trx tissue deposition after intravenous delivery is very minimal. In these studies we hypothesized that intracellular delivery of Trx will enhance the cell reducing capacity of neural cells and will improve neuroprotection. Methods: Routine molecular biology techniques were used to generate Trx constructs containing a cell-penetrating peptide. We used adult spinal cord cultures of neural stem cells to investigate the effect of intracellular delivery of Trx on cell proliferation. Neuroprotective effects were tested in cultures of SH-SY5Y and our clip compression model of spinal cord injury (SCI). Results: Our results indicate that our novel Trx intracellular delivery enhances Trx tissue deposition and distribution and enhances neuroprotection after SCI. Further details will be discussed. Conclusion: Our data indicate the efficacy of our novel Trx intracellular delivery as a potential therapeutic approach for treatment of neurotrauma.



286.09. Chondroitin sulfate proteoglycans negatively modulate the properties of adult spinal cord neural precursor cells by signaling through LAR and PTPσ receptors and activation of the Rho/ROCK pathway
*S. M. DYCK, A. ALIZADEH, E. PROULX, S. KARIMI-ABDOLREZAEE;
Physiol., Univ. of Manitoba, Winnipeg, MB, Canada
Multipotent neural stem/progenitor cells (NPCs) reside in the spinal cord and are capable of replacing lost oligodendrocytes following spinal cord injury (SCI). Despite this intrinsic capacity, adult spinal cord NPCs mainly differentiate into astrocytes, with only a limited number becoming oligodendrocytes. This evidence emphasizes a key role for the post-SCI niche in modulating the regenerative response of spinal NPCs. We recently reported that injury-induced upregulation of chondroitin sulfate proteoglycans (CSPGs) potently restrict the survival, integration and differentiation of transplanted and endogenous NPCs in SCI. In vivo administration of chondroitinase (ChABC) promoted the long-term integration of transplanted NPCs and enhanced the activation and oligodendrocyte differentiation of resident NPCs after subacute and chronic SCI. Given the long-lasting upregulation of CSPGs in NPCs niche after SCI, it is important to unravel the potential mechanisms by which CSPGs influence the properties of NPCs. Using an in vitro model of the extracellular matrix of SCI, we investigated the direct role of CSPGs on NPCs. In primary cultures of adult spinal NPCs, using cell viability, western blotting and immunocytochemistry assays, we show that CSPGs significantly decrease NPC growth and attachment, survival, proliferation and oligodendrocytes differentiation of adult spinal cord NPCs. Genetic down-regulation of CSPG receptors protein tyrosine phosphate receptor sigma (PTPσ) and leukocyte common antigen-related phosphatase (LAR) in NPCs attenuated the inhibitory effects of CSPGs on NPCs. CSPGs inhibitory effects were mediated through activation of the Rho/ROCK pathway and inhibition of Akt and Erk phosphorylation. Our data suggest the impact of CSPGs and its signaling receptors in governing the response of NPCs in their post-SCI niche, and identify new therapeutic targets for enhancing NPC-based therapies following SCI.



299.13/B37. KV1 coupling to connexin-29 to form "xenotypic" junctions between axons and myelin: Mechanism for removing K+ may contribute to increased conduction velocity
*J. E. RASH1, K. G. VANDERPOOL1, T. YASUMURA1, J. I. NAGY2;
1Biomed. Sci., Colorado State Univ., Fort Collins, CO; 2Physiol., Univ. of Manitoba, Winnipeg, MB, Canada
Saltatory conduction in mammalian myelinated axons results from the abrupt influx of Na+ via voltage-gated Na+ channels (NaV1-family) that are densely localized to nodes of Ranvier, followed within 0.5 mSec by efflux of K+ via voltage-gated K+ channels (KV1-family) that until recently were also thought to be localized to nodes of Ranvier. Historically, axonal action potentials were thought to require "temporal separation" of Na+ vs. K+ conductances, but also to require spatial co-localization. However, K+ efflux could not be detected at nodes of Ranvier in mammals, leading influential investigators to suggest that voltage-gated K+ efflux was not necessary for saltatory conduction to occur in mammals. Surprisingly, K+ efflux became strongly detectable after physically or enzymatically separating the paranodal loops of myelin from the axon, revealing that K+ channels were normally masked under myelin. A decade later, KV1 channels were first mapped immunocytochemically - not to nodes of Ranvier as originally presumed, nor beneath paranodal loops of myelin, where myelin detachment procedures had indicated - but instead, KV1 channels were localized solely to the juxtaparanodal axon expansions, as well as to a thin band along the entire inner mesaxon. Moreover, the KV1 channels were precisely co-localized with connexin[[unable to display character: &#8209;]]29 (Cx29), which in both CNS and PNS is expressed solely in myelin-forming cells, and is unique among connexins because Cx29 does not couple with itself or any other connexin to form gap junctions. We now show by confocal immunofluorescence microscopy and by simultaneous dual freeze-fracture replica immunogold labeling (FRIL) that KV1 channels are present in the axon plasma membrane as distinctive hexagonal rosettes of 10-nm E-face particles, and that these KV1 particles are precisely aligned (±2 nm) with similar hexagonal rosettes of 10-nm P[[unable to display character: &#8209;]]face particles in the innermost layer of myelin, which we also identify by FRIL as Cx29 "hemichannels". These dissimilar channels apparently link across the extracellular space to create a newly-discovered form of "xenotypic" intercellular junction. By ionically and osmotically coupling axons to innermost cytoplasmic myelin, these xenotypic junctions appear to represent the recently-described "potassium gateway" into the panglial syncytium. We are currently investigating the biophysical properties of these newly-discovered ion and water transport pathways using cell expression systems and using mice having knockouts of Cx29 and Cx32. We are also examining the effects of damage to this K+ entry portal into the panglial syncytium as a possible source for human neurological disease.



305.29/D29. Astrocyte-induced cortical vasodilation is dependent on endothelial NMDA receptors
*L. LU1,2, A. HOGAN-CANN1,2, C. ANDERSON1,2;
1Univ. of Manitoba, Winnipeg, MB, Canada; 2Neurosci. Res. Program, Kleysen Inst. for Advanced Med., Winnipeg, MB, Canada
Astrocytes play a critical role in neurovascular coupling by providing a physical linkage from synapses to arterioles and releasing gliotransmitters with vasomotor activity. We previously identified that astrocyte activation lead to local vasodilation in a manner mediated by D-serine, activation of N-methyl-D-aspartate (NMDA) receptors production of nitric oxide derived from endothelial nitric oxide synthase (eNOS). The objective of this study was to determine whether endothelial NMDA receptors (eNMDARs) are involved in astrocyte-mediated cortical vasodilation. To generate endothelial NMDA receptor null mice, we crossed driver mice expressing cre recombinase under the control of the endothelial promoter, Tie-2, with mice expression a deletion construct for the pan-NMDA receptor subunit, NR1. The deletion construct was flanked by Lox-p excision sites sensitive to cre recombinase such that offspring were either endothelial NR1-null (Cre+) or wildtype littermates (Cre-). Wildtype mice displayed widespread neuronal and endothelial NR1 expression. In contrast, endothelial (e) NR1-null progeny kept extensive neuronal NR1 immunoreactivity and had undetectable brain endothelial immunofluorescence signal. Peri-vascular astrocytes in acute cortical tissue slices were activated using bath application of metabotropic glutamate receptor agonist, trans-1, 3-dicarboxylic acid (t-ACPD), two-photon uncaging of astrocyte NP-EGTA or localized picoliter pressure ejection of a glutamate and D-serine mixture directly to arteriolar surfaces in 20% ambient O2. Astrocyte Ca2+ dynamics and changes in arteriolar diameter were subsequently measured using two-photon imaging. t-ACPD, astrocyte Ca2+ uncaging and local glutamate/D-serine application all led to astrocyte Ca2+ elevations that reached maximal levels within 5-10 seconds. In all cases, Ca2+ responses were followed by initiation of local vasodilation in wild type mice. Maximal vasodilatory responses were dramatically reduced in eNR1 deletion mice from 5.1 ± 1.5% to 1.8 ± 0.9% baseline in Ca2+ uncaging experiments, and from 4.9 ± 0.7% to 1.5 ± 0.9% baseline in experiments using pressure-ejected glutamate/D-serine. Cumulative dilatory responses over 5 min were assessed by measuring area under the curve (AUC) for relaxation time course plots. AUC was reduced to 30% and 61% control in eNR1-null slices in Ca2+ uncaging and glutamate/D-serine application experiments, respectively. Our results suggest that astrocyte-mediated cortical vasodilation is significantly dependent on NMDA receptors expressed by cortical arteriolar endothelial cells.



308.20/F5. Quetiapine attenuates glial activation and proinflammatory cytokines in APP/PS1 transgenic mice
*S. ZHU1, R. SHI1, J. WANG3, V. LI4, R. ZHANG5, A. TEMPIER3, J. KONG2, J.-F. WANG1, X.-M. LI3;
1Dept. of Pharmacol. and Therapeut., 2Dept. of Human Anat. & Cell Sci., Univ. of Manitoba, Winnipeg, MB, Canada; 3Dept. of Psychiatry, Univ. of Alberta, Edmonton, AB, Canada; 4Univ. of British Columbia, Vancouver, BC, Canada; 5Dept. of Psychiatry, The Fourth Military Med. Univ., Xi'an, China
Introduction: In Alzheimer´s disease, growing evidence has shown that uncontrolled glial activation and neuroinflammation may contribute independently to neurodegeneration. Anti-inflammatory strategies may provide benefits for this devastating disease. Both in vivo and in vitro studies have shown that quetiapine, a novel atypical antipsychotic drug, inhibits microglial activation. The aims of this study are to address the issue of whether glial activation and proinflammatory cytokine increase can be modulated by quetiapine in vivo and in vitro and to explore the underlying mechanism. Methods: 4 month old APPK670N/M671L/PS1M146L transgenic and non-transgenic mice were treated with quetiapine (5 mg/kg/day) in drinking water for 8 months. Animal behaviours, total Aβ levels, and glial activation were evaluated by behavioural tests, ELISA, immunohistochemistry, and western blot accordingly. Inflammatory cytokines and NF-κB pathway were analyzed in vivo and in vitro. Results: Quetiapine improves behavioural performance, marginally affects total Aβ40 and Aβ42 levels, ameliorates glial activation, and reduces proinflammatory cytokines in APP/PS1 mice. Quetiapine suppresses Aβ1-42 induced activation of primary microglia by decreasing proinflammatory cytokines. Quetiapine also inhibits the activation of NF-κB p65 pathway in both transgenic mice and primary microglia stimulated by Aβ1-42. Conclusions: The anti-inflammatory effects of quetiapine in AD may be involved in NF-κB pathway. Quetiapine may be an efficacious and promising treatment for AD targeting of neuroinflammation.



316.11/U30. Neurotoxic NAD+ depletion leads to PARP-1-dependent PGC-1α hyperacetylation and mitochondrial dysfunction
*P. LU1,2, A. KAMBOJ2, C. M. ANDERSON1,2;
1Neurosci. Res. Program, Kleysen Inst. For Advanced Med., Winnipeg, MB, Canada; 2Univ. of Manitoba, Winnipeg, MB, Canada
Excessive excitotoxic activation of the nuclear enzyme poly(ADP-ribose) polymerase-1 (PARP-1) leads to mitochondrial dysfunction and neuron death, but the mechanisms of PARP-1-induced mitochondrial damage are poorly understood. Sirtuin deacetylases regulate activity of peroxisome proliferator-activated receptor gamma coactivator-1α (PGC-1α), which is a master regulator of mitochondrial biogenesis and respiration. PARP-1 consumes nicotinamide adenine dinucleotide (NAD+), which is required for sirtuin deacetylase activity. The central hypothesis of the present study was that PARP-1 leads to mitochondrial dysfunction in neurons by causing inhibition of sirtuins, hyperacetylation of PGC-1α and reduced mitochondrial respiratory capacity. Primary cortical neuron cultures were exposed to the DNA alkylating agent, N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) to initiate pathological PARP-1 activation. MNNG significantly reduced intracellular NAD+ levels in a manner sensitive to inhibition by the PARP-1 inhibitor, PJ34. PARP-1 activation augmented PGC-1α acetylation and reduced transcript levels of PGC-1α-responsive genes instrumental in regulating mitochondrial biogenesis and respiration, including nuclear respiratory factor 1 (NRF-1) and mitochondrial transcription factor A (TFAM). MNNG also significantly reduced mitochondrial mass in a fashion attenuated by PJ34 and exogenous NAD+ replenishment. In accordance with these findings, MNNG significantly reduced whole cell carbonylcyanide p-trifluoromethoxyphenylhydrazone (FCCP)-stimulated maximal OCR, spare respiratory capacity and respiratory control ratio. Taken together, these data demonstrate that PARP-1 activation induces NAD+ depletion severe enough to inhibit PGC-1α-directed mitochondrial mass and respiratory capacity in neuron cultures.



352.12/OO26. Brain endothelial cells express NMDA receptors functionally linked to nitric oxide generation
*A. D. HOGAN-CANN1,2, L. LU1,2, C. M. ANDERSON1,2;
1Univ. of Manitoba, Winnipeg, MB, Canada; 2Neurosci. Res. Program, Kleysen Inst. for Advanced Medicine, Hlth. Sci. Ctr., Winnipeg, MB, Canada
Brain is highly dependent on continuous oxygen and glucose delivery. A regulatory mechanism that ensures this consistency is coupling of neuronal activity to dynamic changes in local blood flow, or functional hyperemia. Glutamate neurotransmission causes functional hyperemia by leading to nitric oxide (NO)-dependent increases in penetrating arteriolar diameter in a manner dependent on activation of neuronal N-methyl-D-aspartate (NMDA) receptors coupled to neuronal NO synthase (nNOS). We observed that isolated middle cerebral artery segments free of neurons dilate in response to NMDA receptor agonists (LeMaistre et al. 2012, J Cereb Blood Flow Metab 32, p. 537). Vasodilation required intact endothelium and was eliminated in arteries from mice lacking expression of endothelial NOS (eNOS-/-). Thus, we hypothesized that eNOS-linked NMDA receptors expressed by cortical endothelial cells mediate glutamate-induced vasodilation. Expression of the pan-NMDA receptor subunit, NR1, was detected in fixed mouse cortical slice microvascular endothelial cells (immunofluorescence) and primary mouse brain endothelial cell cultures (western blot). Western analysis of anti-NR1 immunoprecipitates revealed that NR1 associates directly with the NR2C subunit. Functionally, glutamate exposure in the presence of co-agonist, D-serine but not on its own, significantly enhanced cumulative NO generation over a 5 hour time period in brain endothelial cell cultures, measured by confocal microscopy using a cell membrane-permeable fluorescent probe 4-amino-5-methylamino-2´7´-difluorofluorescein diacetate (DAF-FM diacetate). NO generation was significantly inhibited by NMDA receptor glutamate site competitive antagonist, D-2-amino-5-phosphonopentanoate (AP5), and D-serine/glycine site competitive antagonist, 5,7-dichlorokynurenic acid (DCKA). Glutamate/D-serine exposure to endothelial cultures at concentrations producing NO caused no detectable increases in intracellular Ca2+, measured by real-time fura-2 ratiometric fluorescence imaging at time points up to 15 minutes. Our findings establish that functional, NR2C-rich NMDA receptors linked to NO generation and arterial vasodilation are expressed by brain endothelial cells. Further work will determine the mechanistic linkage between NMDA receptors and NO generation, and whether endothelial NMDA receptors have a role in neurovascular coupling.



394.30/A30. Corticogenesis of the cerebellar cortex in lysosomal acid phosphatase (Acp2) mutant mice
*H. MARZBAN1, K. BAILEY1, M. RAHIMI BALAEI1, A. U.MANNAN3, S. GHAVAMI2;
1Dept. of Human Anat. and Cell Sci., 2Univ. of Manitoba, Winnipeg, MB, Canada; 3Inst. of Human Genetics, Univ. Med. Ctr. Goettingen, Goettingen, Germany
Introduction: The cerebellum is a highly organized centre of motor coordination and cognition. Structurally it consists of a three-layered cortex with distinct neuronal subtypes, such as granule cells (gcs) and Purkinje cells (Pcs). It is suggested that Pcs utilize reelin dependent pathways to form a monolayer in the cerebellar cortex. Reelin expressed by gcs, is required for Pcs distribution from the clusteric stage to establish a monolayer of Pcs between the molecular and granular layers of the cerebellar cortex. Proliferation of the gcs is influenced by Shh (Sonic hedgehog) expressed in Pcs. A mouse mutant called nax (naked-ataxia), resulting from a spontaneous mutation in lysosomal acid phosphatase (Acp2), presents multi-layered Pcs that ectopically invade the molecular layer. We hypothesize that the establishment of this mono layered Pcs is not dependent on the reelin pathway. Materials and Methods; Acp2 mutant mice were used for this study. Molecular expression and distribution were assessed by immunohistochemistry and Western blotting. Results; The cerebellar cortex of the Acp2 mutant mice which was characterized by the absence of the vermis, reveals the presence of Pcs in a randomized, dispersed manner spanning the entire molecular layer rather than a monolayer in the cerebellar cortex. The amount of gcs is severely reduced in the cerebellum which is more prominent rostrally as compared to their numbers in the caudal cerebellum. It is also observed that Pax6 expression follows the pattern of gcs proliferation and migration during postnatal development. The pattern of reelin expression is down-regulated in the Acp2 mutant cerebellum at around P12 and accompanies the down-regulation of Shh. Conclusion; The down-regulation of Shh that followed the declined reelin expression may be secondary to Pcs degeneration. However, the presence of reelin is comparable with wild type during early postnatal development, indicative of reelin effect during clustric stages however failed to form mono layered Pcs. Pcs differentiation is severely delayed inthe Acp2 mutant cerebellar cortex and it can be concluded that multilayer Pcs may be due to the failure of appropriate cross-talk between Shh and the reelin signalling pathway during postnatal cerebellar development.



507.20/I6. Activation of muscarinic receptors underlies cholinergic modulation of serotonergic neurons in the brainstem of ePet-EYFP mice
*Y. DAI1, L. M. JORDAN2;
1East China Normal Univ., Shanghai, China; 2Dept. of Physioloy, Univ. of Manitoba, Winnipeg, MB, Canada
5-HT neurons in the parapyramidal region (PPR) of the medulla play an essential role in generating locomotion. It has been shown that stimulation of the PPR produces locomotion in the isolated neonatal rat brainstem-spinal cord preparation. However, little is known about the properties of these locomotor 5-HT neurons. Using brainstem slices from ePet-EYFP mice in which EYFP was expressed in 5-HT neurons we were able to perform whole cell patch-clamp recordings on EYFP+ neurons of the brainstem (PND 1 - PND 15). We had previously shown that the 5HT neurons received excitatory cholinergic input and displayed location-related responses to acetylcholine (ACh). Bath application of 15-30 µM ACh produced depolarization of the membrane potential in 76% of the 5-HT neurons (n=50) in PPR while 67% of 5-HT neurons (n=33) in the midline raphe nuclei (MRN) of the brainstem did not respond to ACh. In this study, we further investigated the cholinergic modulation of 5-HT neurons in the brainstem of ePet-EYFP mice with bath administration of muscarine (10-20 µM) and nicotine (10-15 µM). Muscarine could mimic the effects of ACh on 5-HT neurons in the medulla while nicotine did not show any substantial effect on these neurons. Similar to ACh, muscarine induced depolarization of membrane potential in 70% of PPR 5-HT neurons (n=33) with decreases in rheobase (5.1±12 pA), input resistance (176.7±290 MΩ), action potential height (2.2±4 mV) and AHP depth (2.6±4 mV), whereas 72% of MRN 5-HT neurons (n=14) did not respond to muscarine. Similar to the observations with ACh, a small number of the PPR 5-HT neurons exhibited a hyperpolarization of membrane potential (6/33) or did not respond to muscarine (4/33), while a few MRN 5-HT neurons displayed depolarization (2/14) or hyperpolarization (2/14) in the presence of muscarine. Muscarine also induced or enhanced membrane oscillations in some of the 5-HT neurons. We further demonstrated that the effects of muscarine on PPR or MRN 5-HT neurons could be antagonized by 1-3 µM atropine (n=12), suggesting that the cholinergic modulation of 5-HT neurons in the brainstem of ePet-EYFP mice was mediated through activation of muscarinic receptors. Furthermore, we examined the subtype of muscarinic receptors with administration of antagonists of M1-M4 receptors (telenzepine 5-10 µM; methoctramine 5-10 µM; 4-DAMP 2-5 µM; PD102807 1-2 µM) in the presence of muscarine. Our data suggested that both M2 and M3 receptors were implicated in the muscarinic modulation of the 5-HT neurons. This study provides insight into the mechanisms underlying the cholinergic modulation of 5-HT neurons in the brainstem of ePet-EYFP mice.



515.14/V18. MeCP2 isoforms and neurological disorders
*M. RASTEGAR;
Biochem. and Med. Genet., Univ. of Manitoba, Winnipeg, MB, Canada
MeCP2 is an important protein in brain, and its altered expression causes neurodevelopmental disorders including Rett syndrome and autism. Alternative splicing of a single gene gives rise to two protein isoforms (MeCP2E1 and MeCP2E2). Deregulation of either isoform is implicated in neurological complications and impaired brain function. Currently, MeCP2-associated disorders have no cure and the mechanisms by which MeCP2 isoforms are controlled are not fully understood. We were the first group to develop isoform-specific MECP2 gene therapy vectors and to study the functional role of MeCP2E1 and MeCP2E2 in neuronal maturation. By developing isoform-specific anti-MeCP2E1 and anti-MeCP2E2 antibodies, we reported that MeCP2E1 and MeCP2E2 expression are cell type-, and brain region-specific. Further, we showed that DNA methylation impacts MeCP2 isoform-specific expression in differentiating brain-derived neural stem cells during brain development and in the adult murine brain in a brain-region specific manner. MeCP2 expression is important in different brain cell types, and disease-associated phenotypes are proven to be cell type-specific. Here, we will discuss our recent results on the MeCP2 isoform-specific regulation and functional role in neurons. We have exciting novel data on the epigenetic control of MeCP2 isoforms in brain-derived neural stem cells and different brain cells. Our results will have significant implications for future therapeutic strategies of MeCP2-associated neurological disorders.



593.01/B29. The proinflammatory cytokine, interleukin-17A, augments axonal plasticity and mitochondrial function of cultured adult sensory neurons
*T. HABASH1,2, A. SALEH1, S. ROY CHOWDHURY1, P. FERNYHOUGH1,2;
1St-Boniface Hosp. Res. Ctr., Winnipge, MB, Canada; 2Pharmacol. and therapeutics, Univ. of Manitoba, Winnipeg, MB, Canada
Rationale and hypothesis: Diabetic neuropathy involves dying back of nerve endings that reflects impairment in axonal plasticity and regenerative nerve growth. Metabolic changes in diabetes can lead to a dysregulation of hormonal mediators, such as cytokines. Thus we studied the effect of interleukin-17A (IL-17A), a proinflammatory cytokine produced by T-cells, on the phenotype of sensory neurons derived from control or diabetic rats. We hypothesized that diabetes induces suboptimal cytokine levels in neurons that reduces axonal growth and this may underlie impaired axon regeneration and plasticity within the skin leading to sensory loss. Objectives: Determine the ability of IL-17A to enhance neurite outgrowth in cultured sensory neurons. Investigate the signalling pathways activated by IL-17A and mechanistically link to neurite outgrowth. Study the ability of IL-17A to improve mitochondrial function of sensory neurons (since axon outgrowth consumes high levels of ATP). Methodology: Cultured adult dorsal root ganglia (DRG) sensory neurons derived from age matched control or streptozotocin (STZ)-induced type 1 diabetic rats were fixed and stained for fluorescent imaging to determine total neurite outgrowth. Western blotting determined the levels of MAPK and PI-3K activation by IL-17A and for measuring levels of proteins of mitochondrial oxidative phosphorylation pathway. Mitochondrial bioenergetics function was tested in cultured DRG neurons using the Seahorse XF Analyzer. Results: We found that IL-17A (10 ng/ml; P<0.05) significantly increased total neurite outgrowth in cultures derived from both control and STZ-diabetic rat models. This enhancement was mediated by IL-17A-dependent activation of MAPK and PI-3K pathways with maximal effect at 15 minutes (P<0.05). Pharmacological blockade of one of these activated pathways led to total inhibition of neurite outgrowth. IL-17A improved mitochondrial bioenergetics function of sensory neurons. Bioenergetics function was associated with augmented expression of proteins of mitochondrial oxidative phosphorylation. Conclusion: IL-17A enhanced axonal plasticity through activation of MAPK and PI-3K pathways and was associated with augmented mitochondrial bioenergetics function in sensory neurons.



593.03/B31. Sirtuin 2 is a sensor of energy status and inducer of neurite outgrowth in adult sensory neurons
*E. D. SCHARTNER1,2, A. SALEH1, R. VIEIRA DA SILVA1, S. CHOWDHURY1, D. SMITH1, P. FERNYHOUGH1,2;
1St. Boniface Hosptial Res., Winnipeg, MB, Canada; 2Pharmacol. and Therapeut., Univ. of Manitoba, Winnipeg, MB, Canada
Introduction: Diabetic sensory neuropathy is a nerve disorder that results in distal dying back of nerve fibers. Neuronal mitochondrial function is impaired in diabetes and Sirtuin 2 (SIRT2) is a key sensor of redox state that regulates cellular bioenergetics. We hypothesized that defective SIRT2 signaling contributed to deficiencies in energy supply and nerve regeneration in diabetic neuropathy. Objectives: We studied the different mechanisms of SIRT2 signaling within adult sensory neurons that drive axon regeneration and how these pathways were impaired under diabetic conditions. Methods: Type 1 diabetes was induced in rodents by intraperitoneal injection of streptozotocin (STZ). Dorsal root ganglia (DRG) were isolated from age matched and 5 month STZ-diabetic mice and SIRT2 protein levels analyzed. DRG sensory neurons derived from control and STZ-diabetic rats were cultured in defined media with varying concentrations of neurotrophic factors and D-glucose. Quantitative Western blotting was performed to determine protein levels and immunocytochemistry utilized to quantify neurite outgrowth. Lentiviral transduction and plasmid transfection were instigated for shRNA knockdown of SIRT2 and overexpression of SIRT2 constructs. Results: SIRT2 isoforms 2.1 and 2.2 were reduced by 20-30% in DRG of diabetic mice (P<0.05). In cultured adult sensory neurons derived from age matched control or STZ-diabetic rats over-expression of wild-type SIRT2 significantly enhanced total neurite outgrowth. Overexpression of the dominant negative mutant (SIRT2-H150) or shRNA to SIRT2 blocked neurite outgrowth. Application of varying doses of AGK2, a selective inhibitor of SIRT2, significantly reduced total neurite outgrowth (P<0.05). Treatment with increasing doses of NAD+, an endogenous activator of SIRT2, significantly enhanced neurite outgrowth. After 72hrs in media with high D-glucose (25mM), cultured sensory neurons showed a significant 2-fold (P<0.05) decrease in the protein levels of SIRT2 isoforms. Conclusions: SIRT2 is a key component driving axon regeneration and this pathway was impaired in diabetic sensory neuropathy.



595.01/C12. Extracellular histones, a putative inhibitor of cns axonal regeneration, are blocked by activated protein c (apc)
M. M. SIDDIQ1, S. S. HANNILA2, Y. ZORINA3, V. RABINOVICH1, E. NIKULINA4, *W. MELLADO5, R. IYENGAR1, M. T. FILBIN6;
1Mount Sinai Sch. of Med., New York, NY; 2Univ. of Manitoba, Winnipeg, MB, Canada; 3Acorda Therapeut., Ardsley, NY; 4The Feinstein Inst. for Med. Res., Manhasset, NY; 5Burke Med. Res. Inst., White Plains, NY; 6Hunter Col., New York, NY
Axons in the injured adult CNS do not regenerate, in part due to the inflammatory response. Outside the nervous system, released histones are detected in response to inflammation. In the CNS, up-regulation of a cytoplasmic isoform of histone H1 was reported in neurons and astrocytes in a mouse model of prion disease and in humans with Alzheimer´s disease. Adult mouse ischaemic brain slices released histones H1 and H2B into the media. Examination of conditioned media from naďve astrocytic cultures revealed secretion of Histones H2A and H4. This suggests that histones are released extracellularly in the CNS under pathological conditions. Here, we show that primary rat cortical and hippocampal neurons extended long neurites when grown on permissive monolayers of CHO cells; however, when we simultaneously added exogenous histones (a mix of all isoforms) to the co-cultures, we observe significantly shorter neurites (up to 70% shorter). Using microfluidic chambers, a technique which isolates the cell bodies from the neurites, we plated cortical neurons on PLL and after one week we observed long neurites growing across the micro-grooves of the chamber. In contrast, when histones were applied to either the cell body- or neurite-containing parts of the chambers, we observed that neurites were unable to grow a significant distance past the micro-grooves. Furthermore, exogenous application of histones to primary cortical neurons activates Rho GTPase. Using western blot analysis, we determined that levels of extracellular histones are increased in vivo after dorsal column lesion and are released into the CSF within 24hrs. Together with our in vitro data, these findings suggest that extracellular histones may inhibit axonal regeneration in spinal cord injury as well. Extracellular histones can be cleaved by APC, an enzyme that is part of coagulation system. In microfluidic chambers with cortical neurons, the inhibitory effect of histones on neurite outgrowth can be blocked with the addition of APC. In addition, delivery of APC after dorsal column injury or optic nerve crush promotes axonal regeneration in vivo. Our data suggests that extracellular histones released after injury to the CNS are inhibitory to axonal regeneration and administration of APC can block this inhibitory effect and promote regeneration.



609.08/T8. Transforming growth factor beta-induced expression of chondroitin sulphate proteoglycans in reactive astrocytes: roles of non-Smad signaling pathways and autophagy
*S. S. HANNILA, N. JAHAN, S. GHAVAMI;
Human Anat. and Cell Sci., Univ. of Manitoba, Winnipeg, MB, Canada
Following spinal cord injury, reactive astrocytes express high levels of chondroitin sulphate proteoglycans (CSPGs), which have been shown to inhibit axonal regeneration both in vitro and in vivo. The factors responsible for inducing CSPG expression have not been fully defined, but there is strong evidence that transforming growth factor β (TGFβ) plays an important role in this process. Administration of anti-TGFβ antibodies reduces glial scarring following spinal cord injury, and expression of CSPGs is significantly increased when astrocytes are treated with TGFβ. Inhibiting TGFβ signaling may therefore be an effective way to reduce CSPG levels in the injured spinal cord. Canonical TGFβ signaling is initiated by binding to the TGFβ receptor, which in turn leads to activation of the Smad2/3 signaling pathway. We have recently shown that levels of Smad2 are significantly reduced in neurons in response to elevation of intracellular cyclic AMP (cAMP), and so, we initially hypothesized that expression of the CSPG neurocan would be reduced following treatment with dibutyryl cAMP (dbcAMP). Surprisingly, while Smad2 levels were significantly reduced in astrocytes treated with dbcAMP and TGFβ, neurocan expression remained elevated, suggesting that Smad signaling is not required for CSPG expression. Subsequent experiments confirmed that activation of the TGFβ receptor is necessary for expression of neurocan, brevican, and aggrecan, but when astrocytes were incubated with either Smad2 or Smad4 siRNA prior to TGFβ treatment, CSPG levels were still significantly increased, which further suggests that TGFβ-induced CSPG expression is Smad-independent. In addition to the Smad signaling pathway, TGFβ also activates several non-Smad signaling pathways, and to identify which of these pathways is involved in CSPG expression, astrocytes were treated with inhibitors of the PI3K-Akt-mTOR or Ras-Erk pathways. No significant reductions in CSPG levels were observed when astrocytes were treated with the Erk inhibitor U0126, but levels of neurocan, brevican, and aggrecan were all significantly reduced following treatment with two inhibitors of the PI3K-Akt-mTOR pathway: LY294002 and rapamycin, which suggests that CSPG expression is mediated primarily through this pathway. In addition, TGFβ induces several markers of autophagy, and CSPG expression was reduced in response to treatment with choloroquine, which indicates that autophagy may also play a role in this process. Targeting these pathways could potentially prevent CSPG deposition within the spinal cord and thereby create a more favorable environment for axonal regeneration.



621.10/BB23. Aftertaste sans taste: ageusia with palinageusia
*A. R. HIRSCH1, K. V. GAFTANYUK2;
1Smell & Taste Treatment and Res. Fndn., Chicago, IL; 2Intl. Univ. of the Hlth. Sci. Sch. of Med., Winnipeg, MB, Canada
Introduction: A case study with vivid and perceptually correct aftertaste in the absence of taste is described. Case Study: A 55 y/o male two years prior to presentation was exposed to Scott´s Crabgrass Preventer herbicide. Within one week there was a gradual decline in sweet and salt taste perception and dysgeusia where potato chips taste like potatoes, apple pie tastes like apples, Pepsi tastes like chemicals, chocolate tastes bitter and corn flakes have a salty taste. For the last one year, he has intermittent salty phantageusia of his entire mouth, whereby regular water tastes like seawater. Six months ago he noted onset of palinageusia, a strong aftertaste of food that was consumed earlier in the day, the way it should taste, but of 50% intensity. Cheese, has no taste, but 8 hours later he can taste cheese flavor. Cinnamon bun is flavorless, but half-hour after eating, the taste appears and lasts for 5 minutes. Sausage has a slight taste at the time of consumption, but two hours later he experiences a much more flavorful and savory taste of sausage, which lasts for half an hour. The aftertaste is always of the last food he has eaten. Rather than being repulsed by these aftertastes, he "savors them" because it provides he only taste experience. Since onset, he has lost of pleasure in eating and has lost 60 pounds. He complains of mild dysphagia, halitosis and omeprazole responsive reflux. Neuropsychiatric exam: decreased short-term memory, sadness, and irrational fears of others out to do him harm. Beck Depression Inventory and Zung Anxiety Scale: normal. Neurologic exam: decreased blink frequency, bilateral palmar erythema and 1+ bilateral pedal edema. Motor: R pronator drift. Reflexes: absent. Candidiasis cultures - negative. Chemosensory tests: see table. Discussion: Potential causes for his condition include: Zenker´s diverticulum, esophageal dysmotility, Gastroesophageal Reflux, herbicide (Pendimethalin) toxicity with phantom aftertaste, prolonged chemosensory deprivation phantageusia and palinageusia (chemosensory equivalent of Phantom Eye Syndrome), illusion of taste due to autosuggestion, somatoform delusion or somatic manifestation of depression or psychosis. Aftertaste warrants exploration in those with chemosensory dysfunction.
Olfactory Functions
Name of TestInterpretation
Sniffin´ Sticks Threshold TestHyposmia
Sniffin´ Sticks Discrimination TestHyposmia
Sniffin´ Sticks Identification TestHyposmia
Name of Test
Olfactometer Threshold Test
Olfactometer Identification Test
Name of Test
Quick Smell Identification Test
Sniff Magnitude Test
Alcohol Sniff Test
Odor Memory Test
Suprathreshold Amyl Acetate Odor Intensity Taste
Suprathreshold Amyl Acetate Odor Hedonic Taste
Jelly Bean Retronasal Smell Test
Gustatory Functions
Name of Test
Taste Threshold Test
Quadrant Taste Test
Propylthiouracil
Piesesthesiometry Test
Name of TestLocation
Electrogustometry TestPalate
RLAnterior
2824RR
>34>34
Name of TestResult
Saxon Test12 grams




631.07/II16. Theta frequency oscillations predominate in the mesencephalic locomotor region during voluntary treadmill locomotion
*B. R. NOGA1, F. J. SANCHEZ1, C. O'TOOLE1, L. VILLAMIL1, S. STASIENKO2, S. KASICKI2, U. SLAWINSKA2, L. M. JORDAN3;
1Miami Project To Cure Paralysis, Univ. of Miami Sch. of Med., MIAMI, FL; 2Dept. of Neurophysiol., Nencki Inst. of Exptl. Biol., Warsaw, Poland; 3Dept. of Physiol., Univ. of Manitoba, Winnipeg, MB, Canada
Oscillatory rhythms in local field potentials (LFPs) are thought to coherently bind cooperating neuronal ensembles to produce behaviours, including locomotion. A variety of frequencies occur at LFPs recorded during locomotion, and have been used as a basis for identification of appropriate targets for deep brain stimulation (DBS) to enhance locomotor recovery in patients with gait disorders. The pedunculopontine nucleus (PPN) has been targeted to improve locomotor activity in Parkinson´s disease (PD), for example, and the presence of gamma band activity (20-100 Hz) has been a basis for identification of this part of the presumed mesencephalic locomotor region (MLR). It is also evident that theta band activity (6-12 Hz) in the hippocampus as well as in the locomotor nuclei of the hypothalamus is associated with locomotor activity. There is sparse information available, however, about the LFPs that occur in the MLR during locomotion, the primary targets for DBS to improve locomotion in PD and after spinal cord injury. Here we use electrodes implanted in the MLR of rats to induce locomotion, and then to monitor the oscillatory activity that is associated with spontaneous locomotion. Thresholds for inducing locomotion were determined in awake rats sitting motionless in an open field. LFPs were then recorded from the same electrodes during treadmill locomotion. Here we show that the predominant oscillatory rhythms recorded from the most effective MLR stimulus sites (thresholds: 15-40 µA) are within the theta range of frequencies (n=5). Theta activity was minimal at rest, but it appeared at the onset of locomotion, and the amplitude was correlated with the speed of locomotion. In animals (n=5) with higher thresholds (70-300 µA), the correlations between locomotor speed and theta LFP oscillations were less robust. Changes in the gamma band were rather small and inconsistent. These results support the suggestion that the oscillatory rhythm responsible for the selection of locomotor output through the MLR is within the theta range of frequencies, with other frequencies, including the gamma band, being less important. These results indicate that theta and not gamma oscillations are suitable characteristics for identifying the functional MLR for DBS.



654.06/UU10. Changes in the orexin (hypocretin) system in rats following footshock exposure
H. WANG1, S. LI1, G. LIU1,2, *G. J. KIROUAC3;
1Oral Biol., Univ. of Manitoba, Winnipeg, MB, Canada; 2Intl. School, Guangdong Food and Drug Vocational Col., Guangzhou, China; 3Oral Biol. and Psychiatry, Fac. of Dent., Winnipeg, MB, Canada
Orexins (hypocretins) are peptides produced by neurons located exclusively in the hypothalamus. Orexin neurons project to many areas of the brain known to be important for regulating arousal levels. A number of studies have also shown that orexin neurons are activated by stressful conditions and that these neurons play a role in the physiological and behavioral responses to stress. Post-traumatic stress disorder (PTSD) is an anxiety disorder triggered by traumatizing and stressful events. Similarly, rats exposed to an acute episode of moderately intense footshocks display long lasting fear and anxiety. Work in our laboratory has demonstrated that a subset of shocked rats which showed a high level of acute fear to a novel tone the day after the footshock experience (high responders; HR), showed high levels of anxiety that last for several weeks. Previous work in our laboratory has also shown that systemic treatment of HR with an orexin antagonist has an anxiolytic effect in these rats. The present study examined if the mRNA levels for prepro-orexin (ppOX), orexin 1 receptor (OX1R) and orexin 2 receptor (OX2R) are increased in rats exposed to footshocks. On day 1, shocked rats were exposed to footshocks (5 × 2 sec episodes of 1.5 mA with an inter shock period of 10-50 s presented randomly over 3 min), whereas nonshocked (NS) rats were placed in the shock chamber for the same amount of time. On day 2, rats were placed in a novel chamber and the time that each rat spent immobile during the presentation of a novel tone was scored. Based on the immobility to the novel tone, shocked rats were assigned to the HR (rats spent > 60% of the time immobile) or low responders (LR; rats spent < 40% of the time immobile) groups. On day 14, the rats were anesthetized and the posterior hypothalamus, dorsal midline thalamus and locus coeruleus/parabrachial region were removed for analysis using real-time RT-PCR. The level of ppOX mRNA was increased in the hypothalamus of HR compared to the LR and NS. In addition, the level of OX1R mRNA was found to be increased in the hypothalamus of HR compared to NS. No difference was observed in OX2R mRNA levels in the hypothalamus between the 3 groups. There was no difference between groups in the level of OX1R and OX2R in samples of the midline thalamus and locus coeruleus/parabrachial region. Protein expression levels for the OX1R and OX2R are being quantified using Western blot and will be presented at the meeting. The results of the present study indicate that changes in orexin and OX1R synthesis occur in a subset of rats exposed to footshocks. These results may provide a better understanding of the brain mechanisms contributing to hyperarousal and anxiety in PTSD.



687.11/B14. Localization of equilibrative nucleoside transporter 3 (ENT3) in mouse brain
L. ROBERTS, W. XIONG, N. LEVINE, M. F. JACKSON, *F. E. PARKINSON;
Univ. Manitoba, Winnipeg, MB, Canada
Adenosine is a signaling molecule acting via cell surface G-protein coupled receptors. In brain, adenosine has neuroprotective, anticonvulsant and sedative properties; however, the mechanisms which regulate adenosine concentrations are poorly understood. Four members of the equilibrative nucleoside transporter family have been identified (ENT1-4) where ENT1 and 2 have been best characterized to date. The current study was initiated to explore the role of ENT3 in regulating brain adenosine levels. Mouse ENT3 gene sequence was inserted into pIRES puro-flag plasmid. HEK293T cells were transiently transfected then used for western blot analysis with a commercial polyclonal ENT3 specific antibody and a monoclonal antibody specific for the flag epitope. Mouse brain was dissected and mRNA was isolated from cortex, cerebellum, striatum and hippocampus. Reverse transcriptase polymerase chain reaction (RT-PCR) was performed using ENT3 specific primers designed to amplify a 245 bp product. The ENT3-specific antibody was used in western blot analysis of proteins isolated from the dissected regions of mouse brain. Cortical astrocytes were cultured and used for RT-PCR or immunocytochemistry of ENT3. Western blot analysis performed using ENT3-transfected and wild type HEK293T cells showed a 40kD band in transfected cells only that was detected with either the ENT3 specific or the anti-flag antibody. RT-PCR analysis was positive for expression of ENT3 in all brain regions tested as well as in cortical astrocytes. Western blot analysis of dissected mouse brain regions, using the ENT3-specific antibody, showed a major band at 52kD and a minor band at 30kD. Immunocytochemistry images indicate intracellular localization of ENT3 in astrocytes. Our results indicate that ENT3 is widely expressed in mouse brain and in cultured cortical astrocytes.



756.06/VV5. Sources of dopamine fibers in the paraventricular nucleus of the thalamus
*S. LI1, G. J. KIROUAC2;
1Oral Biol., 2Oral Biol. and Psychiatry, Univ. of Manitoba, Winnipeg, MB, Canada
The paraventricular nucleus of the thalamus (PVT) sends a dense projection to the nucleus accumbens, bed nucleus of stria terminalis, central nucleus of the amygdala, and the prefrontal cortex. These areas of the brain play a key role in regulating reward and motivation. Recent experimental evidence indicates that the PVT is involved in the modulation of drug seeking behavior and reward behavior in rats. It is well known that dopamine (DA) neurons in the ventral tegmental area (VTA; A10 group) through its projections to the nucleus accumbens are a key regulator of reward and motivation. Dopamine fibers have also been localized in the PVT and it is possible that DA from the VTA could influence behavior by acting at PVT. Previous studies that have attempted to identify the source of DA fibers in the PVT in the rat have been inconsistent in terms of whether the VTA is a source of these fibers. The PVT is a small nucleus which is bordered by the mediodorsal nucleus and the habenula, both of which receive DA projections from the VTA. The present study was done to re-examine this important question. Small iontophoretic injections of cholera toxin B (CTB) that were restricted to the PVT were used to retrogradely labeled tyrosine hydroxylase positive neurons that project to the PVT. Dopamine (tyrosine hydroxylase positive) neurons that were found to project to the PVT were scattered through different regions of the hypothalamus (A11, A13, A15 DA cell groups) and the periaqueductal gray. Double-labeled neurons were sometimes found in the VTA in cases with larger injections that spread outside the PVT, indicating that the areas outside the PVT may receive DA fibers from the VTA. We can conclude from these experiments that DA fibers in the PVT do not originate from VTA but from a heterogeneous population of DA neurons located in the hypothalamus and periaqueductal gray. The significance and function of DA in the PVT remains to be determined.



800.18/U24. BNIP3 interacting with LC3 triggers excessive mitophagy
*R. SHI1, S. ZHU2, V. LI4, S. B. GIBSON3, X. XU5, J. KONG1;
1Dept. of Human Anat. and Cell Sci., 2Dept. of Pharmacol. & Therapeut., 3Dept. of Biochem. and Med. Genet., Univ. of Manitoba, Winnipeg, MB, Canada; 4Fac. of Med., Univ. of British Columbia, Vancouver, BC, Canada; 5Inst. of Neurosci., Soochow Univ., Suzhou, Jiangsu Province, China
A basal level of mitophagy is essential in mitochondrial quality control in physiological conditions while excessive mitophagy contributes to cell death in a number of diseases including ischemic stroke. Signals regulating this process remain unknown. BNIP3, a pro-apoptotic BH3-only protein, has been implicated as a regulator of mitophagy. Here, using in vivo and in vitro models of stroke, we show that BNIP3 and its homologue BNIP3L (NIX) are highly expressed in a `delayed´ manner and contribute to delayed neuronal loss following stroke. Deficiency in BNIP3 significantly decreases both neuronal mitophagy and apoptosis but increases non-selective autophagy following ischemic/hypoxic insults. The mitochondria-localized BNIP3 interacts with the autophagosome-localized LC3, suggesting that BNIP3, similar to NIX, functions as a LC3-binding receptor on mitochondria. Although NIX expression is upregulated when BNIP3 is silenced, upregulation of NIX cannot functionally compensate for the loss of BNIP3 in activating excessive mitophagy. Therefore, NIX primarily regulates basal level of mitophagy in physiological conditions, whereas BNIP3 exclusively activates excessive mitophagy leading to cell death.



809.17/Z30. Chronic treatment with mood stabilizer lithium inhibits amphetamine-increased risk-taking behaviour in rats
Z. ZHOU1, Y. WANG1, H. TAN1, Y. SUN1, Y. CHE2, *J.-F. WANG1;
1Dept. of Pharmacol. & Therapeutics, Fac. of Medicine, Univ. of Manitoba, Winnipeg, MB, Canada; 2Med. Col. of Soochow Univ., Suzhou, China
In humans frequent use of amphetamine causes symptoms that resemble psychotic mania such as hyperactivity, elevated energy, risk-taking, enhanced mood and racing thoughts. The drug lithium is commonly used to treat mania. In this study, we analyzed the effects of lithium on amphetamine-induced risk-taking behaviour in rats. Risk-taking behaviours were measured by elevated plus maze test. The number of stretch-attend postures, the open-arm duration and the open-arm entrance frequency were each recorded for 5 min. We found that treatment with amphetamine at 2 mg/kg for 10 and 14 days significantly increased the number of stretch-attend postures, the open-arm duration and the centre entrance frequency, but treatment with amphetamine for 1, 3 and 7 day has no effect on these behaviours. We also found that chronic treatment with lithium significantly inhibited amphetamine- increased open arm duration and entrance frequency. In addition, treatment with amphetamine for 14 days increased protein carbonylation in rat frontal cortex, and chronic treatment with lithium inhibited amphetamine treatment-increased protein carbonylation. These results suggest that repeated amphetamine treatment in rats displays risk-taking manic-like behaviors in rats, and this risk-taking behaviour can be inhibited by chronic lithium treatment. Our findings also indicate that the process of oxidative stress may be targeted by lithium for its anti-manic treatment.