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P3-1-241 菱脳唇由来神経細胞特異的P/Q型カルシウムチャネル欠損マウスは平行線維－プルキンエ細胞間のシナプス伝達に変異を生じ、運動失調、運動発作、欠神発作のCacna1a遺伝子変異に見られる表現型を再現する Postnatal loss of P/Q-type channels confined to rhombic lip derived neurons alters synaptic transmission at the parallel fiber to purkinje cell synapse and replicates genomic Cacna1a mutation phenotype of ataxia and seizures in mice ○前島隆司1 ○Takashi Maejima1, Patric Wollenweber1, Lena U.C. Teusner1, Jeffrey L. Noebels2, Stefan Herlitze1, Melanie D. Mark1 ルール大学ボーフム・神経生物1, ベイラー医大・神経学2 Dept Zoology and Neurobiology, Ruhr-University Bochum, Bochum, Germany1, Dept Neurology, Baylor College of Medicine, Houston, USA2 Ataxia, episodic dyskinesia and thalamocortical seizures are associated with an inherited loss of P/Q-type voltage-gated Ca2+ channel function. P/Q-type channels are widely expressed throughout the neuraxis, obscuring identification of the critical networks underlying these complex neurological disorders. We recently showed that the conditional postnatal loss of P/Q-type channels in cerebellar Purkinje cells (PCs) in mice leads to these aberrant phenotypes, suggesting that intrinsic alteration in PC output is a sufficient pathogenic factor for disease initiation. The question arises whether P/Q-type channel deletion confined to a single upstream cerebellar synapse might induce the pathophysiological abnormality of genomically inherited P/Q-type channel disorders. PCs integrate two excitatory inputs, climbing fibers from inferior olive and parallel fibers (PFs) from granule cells (GCs) that receive mossy fiber (MF) input derived from precerebellar nuclei. Here, we introduce a new mouse model with a selective knock-out of P/Q-type channels in rhombic lip derived neurons including PF- and MF-pathways (quirky). We found that in quirky mice, PF-PC synaptic transmission is reduced during low-frequency stimulation. Using focal light stimulation of GCs that express optogenetic light-sensitive channels, channelrhodopsin-2, we found that modulation of PC firing via GC input is reduced in quirky. Phenotypic analysis revealed that quirky mice display ataxia, motor seizures and absence epilepsy. These results suggest that developmental alteration of patterned input confined to only one of the main afferent cerebellar excitatory synaptic pathways has a significant role in generating the neurological phenotype associated with the global genomic loss of P/Q-type channel function.	P3-1-242 脳卒中片麻痺患者の継時的MRI計測による高齢健常者との比較検討：事例報告 A case report of chronic stroke recovery by using longitudinal neuroimaging approach ○磯谷悠子1, 倉重宏樹1, 大高洋平2,3, 大須理英子1 ○Yuko Isogaya1, Hiroki Kurashige1, Yohei Otaka2,3, Rieko Osu1 CNS, ATR1, 慶應義塾大学2, 東京湾岸リハビリテーション病院3 ATR, Kyoto, Japan1, Keio University, Tokyo, Japan2, Tokyo Bay Rehabilitation Hospital, Chiba, Japan3 Now we are constructing a brain image database of stroke patients instages from sub-acute to chronic, because there is little informationabout change of brain image during their chronic rehabilitation periodin Japan. In this study, we report some cases of partial recovery afterstroke in stages from sub-acute to chronic state. The aim of this studyis to determine the initial abnormality and the longitudinal changes ofthe brain in both morphologic and functional aspects for the sameindividuals after stroke. All participants are registrants of thisdatabase.A multimodal neuroimaging approach based on resting state functionalmagnetic resonance image (fMRI) and was used to investigate plasticityof damaged brain system.We investigated a patient (Male, 76 years old) after stroke and 14age-matched healthy volunteers by using jackknife group comparison. Thestroke patient underwent MRI set (T1-weighted structural MRI, DTI andresting state) and physical functional assessments 2 times at intervalsof approximately 3 months (at the time of admission and discharge).The healthy volunteers underwent same MRI set just one time.We compared the gray matter volume segmented from MRI and the bloodoxygenation level dependent (BOLD) measured by fMRI of the strokepatient was compared with those of healthy volunteers using statisticalparametric mapping, which is a voxel-based analysis in stereotacticspace. We will examine these brain changes are able to predictfunctional recovery.As a proof of concept investigation, the present findings encouragefurther research utilizing neuro-imaging technique to realize a tailoredrehabilitation for each patient.
P3-1-243 低刺激強度の経頭蓋磁気刺激が神経障害性疼痛モデルマウスに与える影響 Effects of low-intensity repetitive transcranial magnetic stimulation (rTMS) in the neuropathic pain model mice ○由比友顕1, 辻貞俊1 ○Tomoaki Yuhi1, Sadatoshi Tsuji1 産業医科大学神経内科1 Dept Neurol, Univ of Occupation Environment Hlth, Fukuoka1 Previously we reported high-intensity (120% motor threshold: MT) repetitive transcranial magnetic stimulation (rTMS) had the antinociceptive effects in the neuropathic pain model mice. In this study, we studied whether low-intensity (90% MT) rTMS would have the antinociceptive effect same as high-intensity rTMS. Partial sciatic nerve injury model (Seltzer model) was used as neuropathic pain model. Left side sciatic nerve of 5 weeks old ddY mice (n=10) were ligated with 9-0 silk suture around approximately 1/3 to 1/2 the diameter. From 7 to 10 weeks old, nerve-ligated mice were divided into two groups. High-intensity rTMS group (n=5) have received rTMS (1Hz, 120% MT, 1000 stimuli/day) with a round coil (diameter=8 cm) using magnetic stimulator (Nihon Koden, Japan) for 5 days/week. Low-intensity rTMS group (n=5) also received the same condition of rTMS except low-intensity (90% MT). Withdrawal threshold to heat was measured with a plantar test (Ugo Basile, Italy) at 7, 12 and 15 weeks old. The latency of hind paw withdrawal from radiation heat stimuli was measured five times at 5-min intervals and its average value was used as the latency of the response. At 7 weeks (before rTMS), withdrawal latency of left and right hind paw were 2.8±0.5 sec and 7.4±0.9 sec respectively. Operated left hind paw showed the hypersensitivity to heat. At 12 and 15 weeks, withdrawal latency of right hind paw didn't change in both groups. At 12 weeks, withdrawal latency of left hind paw in high-intensity and low-intensity rTMS mice were 3.4±0.5 and 3.8±0.4 sec respectively. At 15 weeks, withdrawal latency of left hind paw in high-intensity and low-intensity rTMS mice were 4.6±0.9 and 4.6±0.3 sec respectively. In both group rTMS prolonged the withdrawal latency of operated hind paw. Our results show that not only high-intensity but also low-intensity rTMS have the antinociceptive effects.	P3-1-244 神経堤由来細胞を蛍光標識したマウスを用いた薬剤性ヒルシュスプルング病モデルの有用性に関する検討 Evaluation of the drug-induced Hirschsprung's disease model with the genetic labeling for the neural crest derivatives ○藤村匠1,2, 芝田晋介1, 下島直樹2, 黒田達夫2, 岡野栄之1 ○Takumi Fujimura1,2, Shinsuke Shibata1, Shimojima Naoki2, Tatsuo Kuroda2, Hideyuki Okano1 慶應大・医・生理学1, 慶應大・医・小児外科学2 Dept Physiol, Sch of Med, Keio Univ, Tokyo1, Dept Ped Surg, Sch of Med, Keio Univ, Tokyo2 The absence of the enteric ganglion cell originated from neural crest is the main feature of the Hirschsprung's disease. The drug-induced Hirschsprung's disease using Benzalkonium chloride (BAC) is well established mouse model, whose ganglion cells are known to be ablated selectively. This model reflects the pathophysiological change of the patient suffering from Hirschsprung's disease. The purpose of this study is to analyze the aganglionic segment in the alimentary tract of this model in detail. In order to visualize the ganglion cells in vivo, the transgenic mouse strain was used to label the neural crest derivatives with Green Fluorescence Protein (GFP). The accumulation of feces was observed at the oral side of the constricted portion by the treatment of BAC. The labeled enteric ganglion cells in the myenteric plexus were evaluated immunohistochemically at two to eight weeks after surgery. The control and aganglionic segment from the colon were stained with the specific markers for ganglion cell, glial cell, muscle cell and interstitial cell along with the GFP antibody. The deletion of the ganglionic cells was confirmed with the disappearance of the markers including GFP and neuronal markers. Addition to the neuronal cell damages, the other types of the cells were also influenced in the BAC-treated gut. The detailed analysis of the drug-induced Hirschsprung's disease model revealed not only the usefulness for the future study but also the discrepancy from the original disease onset.
P3-1-245 TRAP1によるミトコンドリア形態制御とDrp1、Mff発現変化 TRAP1 controls mitochondrial morphology through Drp1 and Mff expression ○高村明孝1,2, 小山佳久3, 松崎伸介1,2,3, 山田浩平1,2, 服部剛志4, 宮田信吾1,3,5, 嶽本香菜3, 遠山正彌1,2,3,5, 片山泰一2 ○Hironori Takamura1,2, Yoshihisa Koyama3, Shinsuke Matsuzaki1,2,3, Kohei Yamada1,2, Tsuyoshi Hattori4, Shingo Miyata1,3,5, Kana Takemoto3, Masaya Tohyama1,2,3,5, Taiichi Katayama2 大阪大院・連合小児発達・附属子どものこころの分子統御機構研究センター1, 大阪大院・連合小児発達・分子生物遺伝学2, 大阪大院・医・神経機能形態学3, 大阪大院・医・分子精神神経学4, 近畿大・東洋医学研究所・分子脳科学5 Mol. Res. Ctr. for Children's Mental Develop., United Grad. Sch. Child Develop., Osaka Univ., Osaka, Japan1, Dept. Child Develop. and Mol. Brain Science, United Grad. Sch. Child Develop., Osaka Univ., Osaka, Japan2, Dept. Anat. and Neuroscience, Grad. Sch. Med., Osaka Univ., Osaka, Japan3, Dept. Mol. Neuropsychiatry, Grad. Sch. Med., Osaka Univ., Osaka, Japan4, Div. Mol. Brain Science, Res. Inst. Asian Oriental Med., Kinki Univ., Sayama, Osaka, Japan5 Mitochondria are dynamic organelles that change in response to extracellular stimuli. These changes are essential for normal mitochondrial/cellular function and are controlled by a tight balance between two antagonistic pathways that promote fusion and fission. Although some molecules have been identified to mediate the mitochondrial fusion and fission process, the underlying mechanisms remain unclear. Tumor necrosis factor receptor-associated protein 1 (TRAP1) is a mitochondrial molecule that regulates a variety of mitochondrial functions. Here, we examined the role of TRAP1 in the regulation of morphology. Stable TRAP1 knockdown cells showed abnormal mitochondrial morphology, and we observed significant decreases in dynamin-related protein 1 (Drp1) and mitochondrial fission factor (Mff), mitochondrial fission proteins. Similar results were obtained by transient knockdown of TRAP1 in two different cell lines, SH-SY5Y neuroblastoma cells and KNS-42 glioma cells. However, TRAP1 knockdown did not affect expression levels of fusion proteins. The reduction in Drp1 and Mff protein levels was rescued following treatment with the proteasome inhibitor MG132. These results suggest that TRAP1 regulates the expression of fission proteins and controls mitochondrial fusion/fission, which affects mitochondrial/cellular function.	P3-1-246 デンドリマーナノ粒子のニューロン内分布（in vitro研究） Intraneuronal distribution of dendrimer nanoparticles: an in vitro study ○黒河佳香1, , 平野靖史郎1, 曽根秀子1 ○Yoshika Kurokawa1, Tin-Tin Win-Shwe1, Seishiro Hirano1, Hideko Sone1 （独）国立環境研究所　環境リスク研究センター1 National Institute for Environmental Studies, Tsukuba, Ibaraki, Japan1 Dendrimer is a composite nanomaterial with a diameter below 10 nm as a primary particle. This nanoparticle has unique properties such as sphere shape, wide variation in number of branches or surface radicals, and pocket-like inner structure, and has been expected to be applied to drug delivery, gene therapy and biological imaging. The particle, if administrated by nasal inhalation, is supposed to be easily absorbed via nasal mucosa, circulate with blood flow, pass through blood-brain barrier, and reach the surface of cerebral neuronal cells. In this study, we prepared particles of polyamidoamine (PAMAM) dendrimer conjugated with molecules of Alexa Fluor 488 or 546, and added the fluoresce particles into media culturing ES cell-derived neuronal cells. We examined under confocal laser microscopy whether the particles show intracellular transport and localization, and we observed the fluorescence signals within lysosomal vesicles around nuclei in the cells one day after adding the particles.
P3-1-247 神経障害性疼痛下における脳内 miRNA-200b/429 クラスター発現低下 Reduction of miRNA-200b/429 cluster expression in the brain is critical for neuropathic pain ○山下哲1, 浜田麻美1, 須原佑樹1, 池上大悟1, 成田道子1, 津山淳2, 葛巻直子2, 岡野栄之2, 成田年1 ○Akira Yamashita1, Asami Hamada1, Yuki Suhara1, Daigo Ikegami1, Michiko Narita1, Jun Tsuyama2, Naoko Kuzumaki2, Hideyuki Okano2, Minoru Narita1 星薬大・薬・薬理1, 慶應大・医・生理2 Dept. Pharmacol., Hoshi Univ., Sch. Pharm. Pharmaceut. Sci., Toyko, Japan1, Dept. Physiol., Keio Univ. Sch. Med., Tokyo, Japan2 Neuropathic pain is the most difficult type of pain to control, and patients lose their motivation for the purposive pursuit with a decrease in their quality of life. It has been widely recognized that the ascending anatomical dopamine projection from the ventral tegmental area (VTA) to the nucleus accumbens (N.Acc.) is mostly related to motivational functions and the reinforcing effects of opioids. MicroRNAs are small, noncoding RNA molecules that direct the post-transcriptional suppression of gene expression, and play an important role in regulating synaptic plasticity. In this study, we found that sciatic nerve ligation induced a drastic decrease in the expression of miR200b and miR429 in N.Acc. neurons. The expression of DNA methyltransferase 3a (DNMT3a), which is the one of the predicted targets of miR200b/429, was significantly increased in the N.Acc. at 7 days after sciatic nerve ligation. Next, we microinjected the miR200b/429-expressing lentivirus into the N.Acc. to evaluate pain behaviors by sciatic nerve ligation. We found that its microinjection partially but significantly reduced the thermal hyperalgesia and mechanical allodynia by sciatic nerve ligation. The results of these analyses provide new insight into an epigenetic modification that is accompanied by a dramatic decrease in miR200b and miR429 along with the dysfunction of "mesolimbic motivation/valuation circuitry" under a neuropathic pain-like state. These phenomena may result in an increase in DNMT3a in neurons of the N.Acc. under neuropathic pain, which leads to the long-term transcription-silencing of several genes.	P3-1-248 自閉性障害患者に見出したGPR85変異についての解析 GPR85, a G-protein coupled receptor located in the AUTS9 (7q31) locus, is associated with autism spectrum disorder ○神保恵理子1, ゆじりん1, 山形崇倫1, 小島華林1, 松本歩1, 田辺裕子2, 桃井隆2, 桃井真里子1 ○Eriko Jimbo1, Zhi-Ling Yu1, Takanori Yamagata1, Karin Kojima1, Ayumi Matsumoto1, Yuko Tanabe2, Takashi Momoi2, Mariko Momoi1 自治医科大学　医学部1, 国際医療福祉大学　基礎医学センター2 Department of Pediatrics, Jichi Medical University1, Medical Research Center, International University of Welfare and Health2 Autism spectrum disorder (ASD) is a neurodevelopmental disorder with complex genetic etiology. Previous studies have shown that the molecular pathogenesis of ASD includes G-protein coupled-receptor (GPCR, GPR) signaling through neurotransmitters GABA, NMDA, and peptide hormones such as oxytocin and secretin. We aimed to identify ASD-associated gene mutations in Caucasian and Japanese patients by screening GPRs on chromosome 7q31, an ASD linkage locus. We identified ASD-related alleles in GPR85 that encodes a GPCR being expressed abundantly in the brain. We found two missense mutations, T1033C (M152T) and G1239T (V221F), at conserved sites in males with ASD from independent Japanese families. In the hippocampus, and not in the cortex, neurons of Gpr85-deficient mice showed significantly reduced expression of the gamma-aminobutyric acid type B receptor subunit 2 receptor (GABA(B)R2) protein but no change of GABA(B)R2 mRNA expression. The Gpr85 deficiency did not alter expression of GAD65/67, PSD-95, or synapsin I in the hippocampus. Thus, GABA(B)R2 down-regulation in hippocampal neurons was specifically related to Gpr85 deficiency. These results raised the possibility that the mutation caused an imbalance in excitatory and inhibitory synaptic transmission, consistent with the functional deficiency reported to be the basis of ASD pathology in neurons with a GPR85 deletion.
P3-1-249 Combined central and peripheral stimulation for treatment of chronic tinnitus, randomized controlled pilot study ○Mohamed Thabit1, Fouad Naglaa2, Shahat Badawy3, Hashim Ramadan3 Department of Neurology, Sohag University Hospital, Sohag, Egypt1, Depatment of Physical therapy, Sohag General Hospital, Sohag, Egypt2, Depatrement of Otolaryngology, Sohag University Hospital, Sohag, Egypt3 Background and objective: Tinnitus is common untreatable distressing symptom. Chronic tinnitus originates from peripheral cochlear or auditory nerve lesions that induce central maladaptive plasticity in auditory and non-auditory brain centers. Repetitive transcranial magnetic stimulation (rTMS) targeting auditory area was tried for treatment of chronic tinnitus with clinically unsatisfactory effect. Accordingly many studies aimed to augment the effect of rTMS by combined stimulation of auditory and non-auditory brain areas. Moreover, trans-meatal cochlear low level laser (TCL) therapy for treatment of tinnitus has induced minimal subjective improvement. However, to our knowledge, no studies targeted both peripheral and central areas implicated in tinnitus pathophysiology. We tested the effect of combined rTMS and TLC therapy targeting both peripheral and central auditory areas as a new therapeutic strategy for tinnitus. Methods: 30 patients were randomized to three equal groups received three different interventions; rTMS treatment (1Hz, 110% suprathreshold, 2000 pulse) to left auditory cortex, TCL therapy (200mW, 808 nm) to affected ear(s) and combination of both treatments. The changes in tinnitus handicap inventory (THI) and visual analogue scale (VAS) were used as an outcome measures. Those outcome measures were taken before, at end, two and four weeks after 10 consecutive daily sessions for each intervention type. Results: Repeated measures ANOVA showed significant effect of time × intervention interaction for THI and VAS scores (P=0.003 and, P<0.0001). One-way ANOVA showed significant main effect of time for THI scores in combined stimulation intervention only (P<0.0001). For VAS scores, there was significant effect of time for both combined and TCL interventions (P<0.0001 and, P<0.001). Conclusion: combined central rTMS and peripheral TCL stimulation of auditory pathway is more beneficial new therapeutic strategy for management of chronic tinnitus.	P3-1-250 Withdrawn
P3-1-251 Individual differences in trait anxiety in common marmosets (Callithrix jacchus) are associated with different types of cognitive flexibility: evidence for two independent neurobiological dimensions ○Yoshiro Shiba1, Katrin Braesicke1, Carmen Agustin-Pavon2, Andrea M. Santangelo1, Angela C. Roberts1 Department of Physiology, Development and Neuroscience, University of Cambridge, UK1, Center for Genomic Regulation, Spain2 Individuals with high trait anxiety are vulnerable to emotional trauma and prone to mood and anxiety disorders. An understanding of the neurobiological basis of these individual differences will therefore provide important clues as to the aetiology and treatment of such disorders. In human and rodents, individuals with high trait anxiety are poor at discriminating safety from danger cues. Thus, in Experiment 1, we tested marmosets on their ability to develop discriminative autonomic and behavioural conditioned responses in an aversive Pavlovian discrimination paradigm, in which one of two auditory cues predicted a burst of aversive loud noise. The results showed 26% of the subjects failed to discriminate. However, their successful discriminative performance on an appetitive Pavlovian discrimination paradigm ruled out a general learning deficit. Instead, the finding that their aversive discrimination failure was predicted by enhanced responsivity to the safety cue, and/or to the context (as shown by baseline measures), early on in training, suggested increased anxiety. Consistent with this proposal was their enhanced emotionality upon exposure to a model snake, a traditional test of anxiety in primates. Neurobiological studies of trait anxiety have implicated enhanced amygdala responsivity and reduced prefrontal control in the aetiology of anxiety. To investigate the former, in Experiment 2 we investigated the performance of the same group of animals on two tests of cognitive flexibility, dependent upon orbitofrontal and ventrolateral prefrontal cortex, respectively. We found that increased perseveration on the two tests was correlated differentially with the enhanced cue and baseline anxiety measures on the aversive discrimination task. Together these findings suggest that different prefrontal circuits contribute to different aspects of anxiety. We will discuss our results in the light of the hypothesis that enhanced responsivity to cues and contexts may lead to heightened anxiety in aversive settings but also improved cognitive processing in non-aversive settings.

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