学習・長期記憶 Learning and Long-term Memory
P3-2-79 恐怖条件付けパラダイムを用いた健常者における情動学習の検証 Learning process in association with emotion in healthy people, using fear conditioning paradigm ○多田光宏1, 内田裕之1, 前田貴記1, 小西海香1, 梅田聡2, 寺澤悠理3, 大谷愛1, 水野裕也1, 吉田和生1, 坪井貴嗣1, 北畑亮輔1, 三村將1, 高橋琢哉4 ○Mitsuhiro Tada1, Hiroyuki Uchida1, Takaki Maeda1, Mika Konishi1, Satoshi Umeda2, Yuri Terasawa3, Ai Otani1, Yuya Mizuno1, Kazunari Yoshida1, Takashi Tsuboi1, Ryosuke Kitahata1, Mimura Masaru11, Takuya Takahashi4 慶應義塾大学　精神・神経科学教室1, 慶應義塾大学文学部　心理学研究室2, (独) 国立精神・神経医療研究センター　精神生理研究部3, 横浜市立大学大学院・医学研究科　生理学教室4 Department of Neuropsychiatry, Keio University School of Medicine, Tokyo, Japan1, Department of Psychology, Keio University, Tokyo, Japan2, Department of Psychophysiology, National Institute of Mental Health, NCNP, Tokyo, Japan3, Department of Physiology, Yokohama City University Graduate School of Medicine, Yokohama, Japan4 Background Fear is one of the indispensable conditions for survival and this condition is caused by a variety of stimuli. Since fear is also associated with a number of psychiatric illnesses, it is critically important to elucidate how this condition is acquired and extinguished in humans in response to various modalities of stressors. Method In this preliminary feasibility study, 11 healthy people underwent fear conditioning and extinction procedures in response to the following three types of stimuli: (1) a simple aversive sound, (2) disgusting pictures, and (3) a picture of actor's face with unpleasant message that was designed to cause an interpersonal conflict. A magnitude in the skin conductance response (SCR) was used as an outcome of conditioned responses. Results During the fear acquisition phase, nine participants (81.8%) showed an autonomic response to at least one of the stimuli and got conditioned. The proportion of the participants who demonstrated an autonomic response to a picture of actor's face with unpleasant message was greater than those in response to simple aversive sound, and disgusting pictures (N=8, 72.7%; N=5, 45.5%; N=5, 45.5%, respectively). Moreover, the interpersonal conflict stimuli tended to be associated with a greater degree of arousal than the simple aversive sound and the disgusting pictures during the fear acquisition phase. During the extinction phase, all of the participants showed attenuation and extinction of an autonomic response to the conditioned fear memory. Conclusions These results suggest that the use of those three types of stimuli, especially the interpersonal conflict stimuli, is feasible in the fear conditioning paradigm. This paradigm could contribute to further understanding of pathoetiology of psychiatric conditions, including mood disorders, with a focus on fear.	P3-2-80 遺伝子改変鳴禽類のさえずり発達 Song development of the transgenic songbird ○安部健太郎1,2, 渡邉大1,3 ○Kentaro Abe1,2, Dai Watanabe1,3 京都大学大学院　生命科学研究科　高次脳機能学1, 独立行政法人科学技術振興機構、さきがけ2, 京都大学　医学部　生体情報科学3 Grad School of Biostudies, Kyoto Univ, Kyoto, Japan1, JST, PRESTO, Kawaguchi, Saitama, Japan2, Faculty of Medicine, Kyoto Univ, Kyoto, Japan3 Songbirds have been a promising experimental animal to study how one's ability develops according to postnatal experiences. However, the lack of efficient methods to manipulate the genome of songbirds has hampered the research of revealing the contribution of genetic factors on the vocal learning. Recently, genetic modification of songbird became possible by virus-mediated transgenesis. Here, we use the transgenic technology in songbirds to achieve specific manipulation of the genome of zebra finch (Taeniopygia guttata). By injecting lentiviral vectors to the early embryos, we obtained several lines of transgenic zebra finches. Through analyzing the acoustic quality of songs uttered by those transgenic animals, we revealed that a specific molecular signaling pathway was involved in the motor learning of songs. In addition, by performing classical fear conditioning in songbirds, we found that these transgenic birds also showed a deficiency in auditory memory formation, although the basal hearing ability of such animals was not changed. Specific brain nucleus was involved in this fear conditioning, and infusion of antagonists hindered the formation of such memory. Collectively, our data demonstrate the molecular mechanism of how genetic factor affects motor and auditory aspects of the postnatal development of the communication ability of songbirds.
P3-2-81 発達段階の違いによる恐怖記憶の再燃への影響 Effect of brain development during adolescence on spontaneous recovery of contextual fear memory ○石井大典1, 松澤大輔1,2, 松田真悟1, 富澤はるな1, 須藤千尋2, 清水栄司1,2 ○Daisuke Ishii1, Daisuke Matsuzawa1,2, Shingo Matsuda1, Haruna Tomizawa1, Chihiro Sutoh2, Eiji Shimizu1,2 千葉大院・医・認知行動生理学1, 子どものこころの発達研究センター2 Dept Cognitive Behav Physiol, Univ of Chiba, Chiba1, Research center for child mental development2 The paradigm of fear conditioning and extinction in rodents has been considered as a valuable animal model for studying anxiety disorders. Fear conditioning to a context represents a form of associative learning involving the formation of linkages between a neutral stimulus and the innate behavioral significance. As a conditioned stimulus (CS) such as context and an aversive unconditioned stimulus (US) such as an electric footshock are repeatedly and consistently paired, CS alone begins to elicit a fear response in anticipation of the US presentation. Fear extinction refers to the repeated presentations of the CS in the absence of the US previously paired. It has been hypothesized that extinction does not erase the original fear memory but forms a new memory of safety which inhibits fear expression. Previous studies using young adult rodents revealed that a reduced fear response could be recovered spontaneously after the passage of time (spontaneous recovery). Understanding the mechanisms of spontaneous recovery is important for the prevention of recurrence of anxiety in patients. However, it is still unclear whether spontaneous recovery occurs among all age groups. Thus, we examined behavioral experiments to investigate whether young or adult mice recover fear response reduced by extinction training. We found that young and adult mice did not recover fear response, but young adult mice only recovered fear response. Moreover, enhanced fear acquisition and retrieval were observed when mice received fear conditioning in early development. These results suggest that there exists a unique period in which spontaneous recovery occurs and may prove informative for understanding mechanisms to erase fear memory.	P3-2-82 成獣期での卵巣摘出手術が恐怖の消去と再発に与える影響 Effects of adult ovariectomy on fear extinction and recovery ○松田真悟1,2, 松澤大輔1,3, 石井大典1, 富澤はるな1, 清水栄司1,3 ○Shingo Matsuda1,2, Daisuke Matsuzawa1,3, Daisuke Ishii1, Haruna Tomizawa1, Eiji Shimizu1,3 千葉大院・医・認知行動生理1, 日本学術振興会特別研究員2, 千葉大院・子どものこころの発達研究センター3 Dept Cognitive behavioral physiology, Univ of Chiba, Chiba1, JSPS Research Fellow, Tokyo, JAPAN2, Research Center for Child Mental Development, Univ of Chiba, Chiba3 Introduction Stress-related disorders such as major depression and anxiety disorder are disproportionately prevalent in women. However, the biological mechanism of the sex difference in the prevalence rate remains unclear. Fear conditioning and extinction paradigm is a widely used model in researching the biological basis of stress-related disorders. It is known that even after the fear extinction, the conditioned fear responses can recover with a passage of time (spontaneous recovery). One clinical interpretation of the spontaneous recovery is the relapse or return of fear/anxiety in stress-related disorders. We reported that female mice required longer days for fear extinction to inhibit the spontaneous recovery than male mice (Matsuda et al., 35th Annual Meeting of JNS). Considering that gonadal hormones affect fear extinction, to study whether gonadal hormones affect consecutive fear extinction and spontaneous recovery, we used ovariectmized-mice in consecutive fear extinction paradigm. Results & Discussion As like intact females, ovariectmized-mice required 15 days of fear extinction to inhibit spontaneous recovery. Freezing curves in early days of fear extinction differed between ovariectmized- and sham-mice but the curves in the late days of fear extinction did not. These results suggest that gonadal hormones may affect process of fear extinction but not spontaneous recovery of the conditioned fear.
P3-2-83 注意や学習におけるマウス束傍核の遺伝学的機能解析 Genetic analysis of the roles of NMDA receptors in the parafascicular nucleus in attention and learning ○安田光佑1,2, 林悠1, 田中三佳1, 糸原重美1 ○Kosuke Yasuda1,2, Yu Hayashi1, Mika Tanaka1, Shigeyoshi Itohara1 理研・BSI・行動遺伝学技術開発チーム1, 東大院・農生・比較病態生理2 Laboratory for Behavioral Genetics, RIKEN, BSI, Saitama1, Laboratory of Comparative Pathophysiology, Graduate School of agricultural and life science, The University of Tokyo, Tokyo2 Animals selectively and rapidly redirect their attention to salient or unfamiliar stimuli, allowing rapid response to factors that threaten or promote survival of the species. The molecular and neuronal mechanisms underlying this attentional shift, however, remain largely unknown.The parafascicular nucleus (PFN) of the thalamus may play an important role in attentional shift. Anatomically, the PFN is classified as a part of the reticular activating system, which is critical for attention and arousal. In vivo recordings and pharmacological studies suggest that the PFN detects behaviorally significant stimuli. Neurons of the PFN are selectively activated by unfamiliar stimuli, and these firing responses often habituate rapidly when the same stimuli are presented repeatedly. These characteristics likely allow the PFN to act as a detector of salient or unfamiliar stimuli.NMDA receptors, members of the ionotropic glutamate receptor family, allow entry of Ca2+, which serves as a second messenger to activate intracellular signaling cascades, and act as a coincidence detector underlying synaptic plasticity. Thus, we postulated that NMDA receptors are involved in the rapid habituation of the PFN neurons, enabling the PFN neurons to respond selectively to novel stimuli. To test this hypothesis, we generated PFN-specific NR1 knockout mice using the cre-loxP system. We developed a transgenic mouse line expressing Cre selectively in this brain area and crossed these mice with mice in which the NMDA receptor subunit gene NR1 is flanked by loxP. In addition, to address the effect of acute disruption of the PFN, we generated adeno-associated viral (AAV) vectors by which, in combination with Cre mice, neural firing or transmission can be blocked specifically in the PFN. We will report the behavioral phenotypes of these genetically engineered mice.	P3-2-84 ゼブラフィッシュ手綱核-縫線核経路は逃避行動学習を制御する The Habenula-Raphe pathway Regulates Active Avoidance Learning in Zebrafish ○天羽龍之介1, 揚妻正和1,2, 木下雅恵1, 白木利幸1, 青木田鶴1, 山崎昌子1, 東島眞一3, 松田勝4川上浩一5, 大島登志男6, 相澤秀紀1,7, 岡本仁1 ○Ryunosuke Amo1, Masakazu Agetsuma1,2, Masae Kinoshita1, Toshiyuki Shiraki1, Tazu Aoki1, Masako Yamazaki1, Shin-ichi Higashijima3, Masaru Matsuda4, Maximiliano L Suster5, Koichi Kawakami5, Toshio Ohshima6, Hidenori Aizawa1,71,7, Hitoshi Okamoto1 理研BSI発生遺伝子制御1, コロンビア大2, 岡崎統合バイオサイエンスセンター生理研3, 宇都宮大学バイオサイエンス教育研究センター4, 国立遺伝研個体遺伝初期発生5, 早稲田大院先進理工生医6, 東京医科歯科大分子神経科学研究室7 RIKEN BSI, Saitama1, Dept Biological Sciences, Columbia Univ, New York2, NIPS, Okazaki Institute for Integrative Bioscience, Aichi3, Center for Bioscience Research and Education, Utsunomiya Univ, Tochigi4, Division of Molecular and Developmental Biology, NIG, Shizuoka5, Dept Life Science and Medical Bioscience, Waseda Univ, Tokyo6, Dept Molecular Neurosciences, Tokyo Medical and Dental Univ, Tokyo7 Appropriate choice of voluntary behavior such as avoiding predictable danger is critical for animals to survive. Monoaminergic modulation of neural circuits including the cortico-basal ganglia circuit is one of the key mechanisms regulating this behavior. Recent studies have suggested that the lateral habenula conveys negative emotional information. Interestingly, anatomical and electrophysiological studies showed that the lateral habenula regulates the activity of midbrain dopaminergic neurons and serotonergic neurons in the raphe. Although the lateral habenula is a major afferent of the raphe, function of the habenula-raphe pathway is poorly understood. We previously identified the homolog of the mammalian lateral habenula in zebrafish, the ventral habenula. Notably, the ventral habenula projects exclusively to the raphe but not to the dopaminergic nuclei in zebrafish. In this study, we have successfully established a transgenic fish line expressing tetanus toxin to inhibit neural transmission in the ventral habenula-raphe pathway. We found that Inhibition of the ventral habenula impaired active avoidance learning, a type of operant conditioning, while learning of classical fear conditioning was unaffected. Furthermore, ventral habenula neurons were phasically excited by an aversive stimulus and optogenetic stimulation of ventral habenula axons increased the firing of serotonergic neurons in the raphe. Taken together, our study suggests that the evolutionarily conserved lateral habenula-raphe pathway conveys negative emotional information to modify behavior to avoid danger through the regulation of serotonergic neurons.
P3-2-85 長期間のメチルドナー欠乏は長期記憶を阻害する Long-term methyl donors deficit impairs long-term memory ○富澤はるな1, 松澤大輔1,2, 石井大典1, 松田真悟1,3, 須藤千尋2, 清水栄司1,2 ○Haruna Tomizawa1, Daisuke Matsuzawa1,2, Daisuke Ishii1, Shingo Matsuda1,3, Chihiro Sutoh2, Eiji Shimizu1,2 千葉大院・医・認知行動生理学1, 千葉大院・医・子どものこころの発達研究センター2, 日本学術振興会特別研究員3 Dept Cogn Behav Physiol, Univ of Chiba, Chiba, Japan1, Research Center for Child Mental Development, Univ of Chiba, Chiba, Japan2, JSPS Research Fellow, Tokyo, Japan3 DNA methylation is known as one of the epigenetic process. Both of hyper- and hypo-methylated phenomena are related to the improper regulation of gene function. Accumulating evidence from patients with neurodegenerative and neurodevelopmental disorders such as Alzheimer's disease, schizophrenia and autism indicated that epigenetic mechanism is related to the pathogenesis. Nutrients related to one-carbon metabolism are known to play important roles on the maintenance of genomic DNA methylation. Previous studies have shown that the long-term administration of a diet lacking essential one-carbon nutrients such as methionine, choline and folic acid (folate/methyl donor: FMD) resulted in the rearrangement of global DNA methylation in the brain. Taken together, we hypothesized that the long-term feeding of methyl-donor deficient diet during developmental period might affect the brain functioning related to specific cognitive process even at later adult stage. In the current study, we used FMD diet (lacking choline and folic acid with very low methionine). One experimental group was fed with the diet from postnatal 3 to 6 weeks (FMD-6) and the other was fed to 12 weeks (FMD-12). The comparison groups were fed with normal diet (CONT-6, and -12). Open-field, object recognition, social recognition, and social interaction tests were employed as behavioral tests, and the behaviors were evaluated at 6 weeks (FMD/CONT-6), or at 12 weeks (FMD/CONT-12). We found that the FMD-6 mice had impairment in the novel object recognition and the social recognition. Decreased locomotor activity in the open-field test was observed in FMD-12 mice. In conclusion, our data suggest that the long-term administration of a diet lacking methyl-donor impairs long-term memory and locomotion. The long-term methyl-donor deficient might have influenced the brain development.	P3-2-86 線虫における個体群密度依存的嗅覚可塑性の分子機構の解析 Analysis of the molecular mechanism of population density dependent olfactory plasticity in C. elegans ○土屋純一1, 山田康嗣1, 飯野雄一1 ○Jun-ichi Tsuchiya1, Koji Yamada1, Yuichi Iino1 東京大院・理・生物化学1 Dept. of Biophys. and Biochem., Grad. Sch. of Sci., Univ. of Tokyo1 Among various information in the environment, stimuli from other individuals of th same spiecies can largely influence the behaviors of animals. a previous study showed that olfactory plasticity of the nematode C. elegans is dependent on population density of animals on the cultivation plates. C. elegans is attracted to a series of odors such as benzaldehyde. However, after preexposure to benzaldehyde in the absence of food, worms show reduced attraction to it and rather avoid of the odorant. Interestingly, avoidance of benzaldehyde after preexposure to it becomes stronger, as the population density becomes higher. Population density information is transmitted via the pheromone, and the pheromonal signal down-regulates SNET-1, a neuropeptide which suppresses olfactory plasticity. In addition, it has been suggested that SNET-1 is negatively regulated by the extracellular peptidase encoded by nep-2. However, we still do not know where in the nervous system SNET-1 is received and how SNET-1 regulates olfactory plasticity. The severe defect in olfactory plasticity in the nep-2 mutant probably results from overe-accumulated SNET-1. Therefore, we performed a forward genetic screen for mutations that suppress the olfactory plasticity defect of the nep-2 mutant, in order to identify molecules that act downstream of SNET-1. As a result, we acquired two suppressor mutants of nep-2 , and one of them suppressed nep-2 mutant phenotype to the same extent as the nep-2; snet-1 double mutant. Genetic mapping with single nucleotide polymorphisms between two strains of C. elegans suggested that this mutation resides on the left arm of X chromosome. It is probable that further analysis of this mutation will enable us to identify a molecule that functions downstream of SNET-1, and provide insights into the mechanisms of how interactions between individuals affect learning.
P3-2-87 メマンチン投与による海馬依存的記憶の破壊 Erasure of hippocampus-dependent memory by memantine ○石川理絵1, 喜田聡1,2 ○Rie Ishikawa1, Satoshi Kida1,2 東京農大・農・バイオ1 Dep. of Bioscience, Tokyo Univ. of Agriculture1, JST, CREST2 Facilitation to erase fear memory is thought to be a candidate for reducing acquired fear that leads to emotional disorders such as post-traumatic stress disorders. Memantine (MEM) is a noncompetitive N-methyl-D-aspartate (NMDA) glutamate receptor antagonist. Recent study displayed that high-dose of MEM treatment promoted adult hippocampal neurogenesis. In this study, we tried to examine effects of MEM treatment on memory retention in contextual fear conditioning and Morris water maze tasks, both of which are hippocampus-dependent. In contextual fear conditioning test, mice were trained with an electrical footshock and tested 24 hrs after the training (test 1). After that mice received systemic injections of MEM (50 mg/kg body weight) every week for 4 weeks and then assessed contextual fear memory 1 week after the last MEM treatment (test 2). We found that MEM-treated group displayed an impaired contextual fear memory at the test 2, but not test 1, while control group treated with saline displayed contextual fear memory at test 1 and 2. Importantly, MEM-treated group did not show spontaneous recovery of fear memory 4 weeks after test 2. These observations suggest that systemic injections of MEM disrupted contextual fear memory. Moreover, we observed similar effects of MEM treatment on spatial memory in Morris water maze. Thus we found the possibility that MEM treatment disrupts hippocampus-dependent memory recently generated through the enhancement of neurogenesis. Now we try to confirm whether our results generalize to other fear memory task.	P3-2-88 リン酸化CREBとErkの発現レベルを指標とした恐怖記憶強化と消去制御に関わるニューロン集団の同定 Distinct time-dependent roles of Erk activation in the reconsolidation and extinction memory phases ○張悦1,2, 福島穂高1,2, 喜田聡1,2 ○Yue Zhang1,2, Hotaka Fukushima1,2, Satoshi Kida1,2 東京農大院・農・バイオ1, 日本科学振興機構2 Dept. of Bioscience, Tokyo University of Agriculture1, JST, CREST2 Memory retrieval induces two opposite memory processes; reconsolidation and extinction. We have characterized reconsolidation and extinction phases in the inhibitory avoidance (IA) task because this task is thought to allow to discriminate reconsolidation and extinction phases at the time point when mice enter into dark box from light box. We previously found that the re-exposure to the light box leads to reconsolidation and enhancement of reactivated IA memory through new gene expression in amygdala (AMY), medial prefrontal cortex (mPFC) and hippocampus (HP). Additionally, we also revealed that the re-exposure to the dark box leads to long-term extinction through new gene expression in the mPFC and AMY, but not HP. To compare molecular signatures of enhancement/reconsolidation and extinction phases, we measured time-course of phosphorylations of cAMP-responsive element-binding protein (pCREB) and extracellular signal-regulated kinase (pErk) following the re-exposure to light or dark box using immunohistochemistry. Significant increases in pErk level in the mPFC, AMY and HP were observed 5 min after the re-exposure to light and dark box (in both enhancement/reconsolidation and extinction phases). In contrast, increases in pErk level in these regions were observed 30 min after the re-exposure to the dark box, but not light box, suggesting that Erk is activated in the late extinction phase. On the other hand, significant increases in pCREB levels were observed in the HP, mPFC and AMY 30 min after the re-exposure to the light box (reconsolidation phase), while these increases were observed in the mPFC and AMY, but not the HP, in the extinction phase. Consistently, micro-injection of MEK inhibitor U0126 into the mPFC 30 min after the re-exposure blocked long-term extinction, but not enhancement/reconsolidation. These observations suggest that Erk displayed distinct time-dependent roles in reconsolidation/enhancement and extinction phases.
P3-2-89 社会的認知記憶に基づく新規性認知に関わる脳領域の組織学的解析 Identification of neural network recognizing "novelty" in the social recognition task in mice ○谷水俊之1, 門間和音1, 岡野絵美子1, 張悦1,2, 福島穂高1,2, 喜田聡1,2 ○Toshiyuki Tanimizu1, Kazune Kadoma1, Emiko Okano1, Yue Zhang1,2, Hotaka Fukushima1,2, Satoshi Kida1,2 東京農大院・農・バイオ1, 科学技術振興機構2 Dep. of Bioscience, Tokyo Univ. of Agriculture,1, CREST, JST2 Memory consolidation is a process in which long-term memory (LTM) is generated by stabilizing short-term labile memory. A critical biochemical feature of memory consolidation is a requirement of new gene expression. In the social recognition (SR) task, mice form a SR memory of a juvenile mouse. Our studies have shown that mice form a SR memory following the exposure to a juvenile mouse for 3 min, but not 1 min. In this study, we have tried to understand mechanisms underlying formation and regulation of SR memory at the anatomical level. To do this, we measured expression levels of c-fos and Arc, markers of gene expression during memory consolidation, following the exposure to a juvenile mouse for 0, 1 or 3 min using immunohistochemistry. We found significant inductions of c-fos and Arc in the medial prefrontal cortex (mPFC), anterior cingulate cortex (ACC), hippocampus (HP) and basolateral amygdala (BLA) following the exposure to a juvenile mouse for 3 min. Consistently, the inhibition of protein synthesis in the mPFC, ACC and HP impaired long-term SR memory, suggesting that protein synthesis in these regions is required for consolidation of SR memory. Moreover, we found that the number of brain regions displaying positive correlations of c-fos expression levels was increased with time of exposure to a juvenile mouse. We next compared c-fos expression when mice retrieved SR memory or formed a new SR memory; mice were exposed to the same juvenile mouse or a novel juvenile mouse. We observed the c-fos inductions in the mPFC, ACC and BLA, but not HP, following the retrieval of SR memory. This observation indicated that HP was not activated when mice were re-exposed to familiar juvenile mouse, suggesting that HP plays a critical role in discrimination of novel and familiar mice. Thus we will report brain regions that play critical roles in formation and regulation of SR memory.	P3-2-90 前脳領域におけるCREB情報伝達経路活性化が時間的関連付け学習に与える影響 Effects of up-regulation of CREB activity on learning ○芹田龍郎1, 福島穂高1,2, 喜田聡1,2 ○Tatsurou Serita1, Hotaka Fukushima1,2, Satoshi Kida1,2 東京農業大学大学院　農学研究科　バイオサイエンス専攻1 Dep.of Bioscience, Tokyo Univ.of Agriculture, Tokyo1, JST・CREST2 Previous our findings indicated that loss-of-CREB function leads to an impairment of long-term memory (LTM). To further understand roles of CREB in memory formation, we have examined effects of gain-of-CREB function on memory formation by generating and analyzing transgenic mice expressing a constitutively active CREB mutant (CREB DIEDML or CREB Y134F) in the forebrain. Interestingly, these transgenic mice displayed enhancements of short-term memory (STM) and LTM, suggesting that CREB positively regulates the formation of STM and LTM. In this study, to further understand effects of gain-of-CREB function, we first examined the ability of DIEDML and Y134F mice in temporal association learning using trace fear conditioning task, where mice learn an association between the conditioned stimulus (CS; tone) and the unconditioned stimulus (US; footshock) separated by the trace interval. During the training that consists of 8 CS-trace-US-intertrial interval (ITI), mice were assessed freezing responses during each trace interval. Twenty-four hours after training, mice were assessed freezing scores again for each ITI during test that consists of 8 CS-ITI. DIEDML mice learned the trace fear conditioning significantly faster and better and also displayed significantly stronger trace fear memory during the test, compared to wild-type (WT) mice although Y134F mice showing weak up-regulation of CREB activity displayed normal learning and memory. These observations suggest that DIEDML mice exhibit enhanced temporal association learning in trace fear conditioning. Furthermore, DIEDML mice displayed enhanced within-session extinction of contextual fear memory. These our observations suggest that constitutive up-regulation of CREB activity enhances learning. We are now investigating the ability of DIEDML mice in spatial temporal association learning using T-maze and working memory version of water maze task.
P3-2-91 恐怖条件付け文脈記憶形成時の海馬における糖鎖修飾の役割の解析 Roles of glycosylation in the hippocampus in formation of contextual fear memory ○稲葉洋芳1, 甲斐大輔1, 喜田聡1,2 ○Hiroyoshi Inaba1, Daisuke Kai1, Satoshi Kida1,2 東京農大院・農・バイオ1 Dept. of Bioscience, Tokyo Univ. of Agriculture, Tokyo1, JST・CREST, Saitama2 Memory consolidation is a process to stabilize short-term labile memory, generating long-term memory. A critical biochemical feature of memory consolidation is a requirement of new gene expression. Protein glycosylation is one of important post-translational modifications and plays critical roles in the protein function. Previous studies using chicks showed that novel glycoproteins were generated in the cell surface after learning and that the inhibition of protein glycosylation led to an impairment of avoidance memory (Rose SP., 1995). However, roles of protein glycosylation in memory formation have not been well examined in mammals. In this study, we have tried to understand roles of glycosylation in formation of hippocampus-dependent memory in mice. To do this, we examined effects of a micro-infusion of tunicamycin, N-linked glycosylation inhibitor, into the hippocampus on contextual fear memory. Mice were micro-infused tunicamycin (0.25 or 0.5 μg /side) into the dorsal hippocampus 30 min before the training with a single footshock (0.4 mA, 2 sec), and assessed freezing at 2 or 24 hrs after the training. Mice micro-infused tunicamycin showed significantly less freezing at the test 24 hrs, but not 2 hrs, after the training compared to control groups in a dose-dependent manner, suggesting that the inhibition of glycosylation in the hippocampus impaired long-term contextual fear memory. Furthermore, mice micro-infused tunicamycin showed normal locomotor activity in the open field test, suggesting that the impaired contextual fear memory by inhibition of glycosylation was not due to abnormal locomotor activity. These observations suggest that hippocampal glycosylation is required for the consolidation of contextual fear memory.	P3-2-92 海馬ガンマ波は動物の経験依存的に増強するが、それは左右非対称に生じる Lateralized enhancement of experience-induced hippocampal gamma oscillations ○篠原良章1, 細谷亜季1, 平瀬肇1 ○Yoshiaki Shinohara1, Aki Hosoya1, Hajime Hirase1 理化学研究所神経グリア回路研究チーム1 Neuron Glia Circuit Team, RIKEN BSI1 Gamma oscillations have gathered attention because they are implicated in higher order brain functions such as sensory binding and memory. But, how an animal's experience organizes these gamma activities remains largely elusive. We find that the power of hippocampal theta-associated gamma oscillations tends to be greater in rats reared in an enriched environment than those reared in an isolated condition. This experience-dependent gamma enhancement is consistently larger in the right hippocampus across subjects, coinciding with a lateralized increase of synaptic density in the right hippocampus. Moreover, interhemispheric coherence in the enriched environment group is significantly elevated at the gamma frequency. These results suggest that enriched rearing sculpts the functional left-right asymmetry of hippocampal circuits by reorganization of synapses.
P3-2-93 ラットの逆転弁別学習における前部帯状皮質と海馬のERP A study of event-related potentials of rats elicited by reversal discrimination learning in the anterior cingulate cortex and hippocampus ○坂田省吾1, 武田梢1, 岡田佳奈1, 服部稔2 ○Shogo Sakata1, Kozue Takeda1, Kana Okada1, Minoru Hattori2 広島大院・総合科学・行動科学1, 広島大院・医歯薬保・トレーニングセンター2 Dept Behav Sci, Grad Sch of Inte Arts & Sci, Hiroshima Univ, Japan1, Advan Med Skill Training C, Inst Biomed & Health Sci, Hiroshima Univ, Japan2 The purpose of this study was to investigate whether the rat P2 and P3-like potential reflects 'attention processes'. Twelve rats were trained to discriminate stimuli between light and tone with food reinforcements or not. One group with six rats was reinforced for light (S+) stimulus and tone (S-) stimulus without reinforcement. The other group with six rats was reinforced with tone (S+) stimulus and light (S-) with no food. After reached the learning criterion for the performance of the first stimulus discrimination task, then S+ and S- were inversed. This discriminative reversal procedure did three times after reached the learning criterion. The event-related potentials (ERPs) of the rats were recorded at the anterior cingulate cortex and hippocampal CA1 during performing tasks. The amplitude of the P2 and P3-like potentials with S+ during the discrimination task was greater than S- for both sites. These results provide evidence that the rat P2 and P3-like component reflects 'attention processes'. Rats can provide a useful model for investigation of the neural mechanisms of the P2 and P3-like potentials in learning performance.	P3-2-94 線虫C. elegansにおいて、記憶の忘却を制御するTIR-1/JNK-1経路の下流因子の探索 Downstream signaling factors of TIR-1/JNK-1 pathway for forgetting in C. elegans ○北園智弘1, 井上明俊1, 石原健1,2 ○Tomohiro Kitazono1, Akitoshi Inoue1, Takeshi Ishihara1,2 九大・シス生・シス生1, 九大・院理・生物2 Dept. of Sys. Life Sci., Kyushu Univ., Fukuoka1, Fac. of Sci., Dept. of Biol., Kyushu Univ., Fukuoka2 Animals can properly respond to continuously changing environments by erasing past memories that are inconsistent with present circumstances. To elucidate the regulatory mechanism of forgetting, we used the olfactory adaptation in C. elegans as a simple model. By forward genetic analyses, we found that TIR-1, a p38 MAPK adaptor protein, and its downstream signaling pathway (TIR-1/JNK-1 pathway) mutants exhibited prolonged retention of the olfactory adaptation, and that the TIR-1/JNK-1 pathway accelerates forgetting by releasing forgetting signals from AWC neurons to other sensory neurons. However, the molecular mechanisms underlying this forgetting remain largely unknown. To identify downstream components of the TIR-1/JNK-1 pathway, we carried out a suppressor screening of the gain of function mutants of tir-1, which shows a weak adaptation phenotype probably because forgetting signals are always released. By the screening, we isolated 71 independent lines, which showed prolonged retention of olfactory adaptation. These mutants are classified into 2 classes. One class shows prolonged retention of olfactory adaptation to both diacetyl, which is sensed by AWA sensory neurons, and isoamylalcohol, which is sensed by AWC sensory neurons. The other class shows prolonged retention of olfactory adaptation to diacetyl, but not of that to isoamylalcohol. This result may suggest that the distinct downstream signaling pathways regulate forgetting of olfactory adaptation. By the whole genome sequencing and genetic mapping, we identified a gene responsible for one of the mutants, qj143 as maco-1, which is conserved across the species and is orthologous to human macoilin. In C. elegans, macoilin, which is a membrane protein expressed in many neurons, is involved in assembly or traffic of ion channels or ion channel regulators and necessary for normal thermotaxis. This study enables us to provide new insights on the regulatory mechanisms of forgetting.
P3-2-95 サルにおける瞬目反射条件づけ：筋電位測定法と高速イメージング法を用いた瞬目反射条件づけ方法の開発とその評価 Eyeblink conditioning in monkeys: Development of an evaluation system for conditioned eyeblink response by using electromyographic measurements and high-speed image processing techniques ○桐野豊1, 山本茂幸2, 鈴木一隆2, 豊田晴義2, 狩野方伸3, 塚田秀夫2, 岸本泰司1 ○Yutaka Kirino1, Shigeyuki Yamamoto2, Kazutaka Suzuki2, Haruyoshi Toyoda2, Masanobu Kano3, Hideo Tsukada2, Yasushi Kishimoto1 徳島文理大・香川薬・生物物理学1, 浜松ホトニクス・中央研2, 東京大院・医・神経生理3 Lab of Neurobiopysics, Tokushima Bunri Univ, Sanuki, Japan1, Central Res Lab, Hamamatsu Photonics, Hamamatsu, Japan2, Dept Neurophysiol, Univ of Tokyo, Tokyo3 Delay eyeblink conditioning, in which a 1-kHz tone (conditioned stimulus, CS) is paired with an air puff (0.02 MPa; unconditioned stimulus, US), is a cerebellum-dependent learning paradigm supposedly unrelated to human awareness. This learning paradigm has been applied to various mammalian species; however, delay eyeblink conditioning in monkeys has not yet been reported. We developed a system that we believe is the first for delay eyeblink conditioning in a monkey species (Macaca mulatta). The eye blinking of a monkey was monitored and evaluated simultaneously by using orbicularis oculi electromyographic (OO-EMG) measurements and an eyelid-state detection system comprising a 1-kHz high-speed image processor (Intelligent Vision Sensor, C8201; Hamamatsu Photonics). EMG signal analysis indicated that monkeys (n = 5) exhibited a conditioned response (CR) frequency of more than 60% during the 5-day acquisition session and an extinguished CR during the 2-day extinction session. High-speed image processing analysis also yielded similar results for CRs in both the acquisition and extinction sessions. Hence, we concluded that both analytical methods are effective in evaluating monkey eyeblink conditioning. This multi-measuring system, incorporating different measuring principles, has enabled us to elucidate the relationship between the actual state of eyelid closure and the OO-EMG activities during the experiment. An interesting finding of this study was that the monkeys often elicited the obvious CRs even when their eyes were closed. Our quantitative analysis confirmed that the probability of CR incidence is not greatly influenced by whether the monkeys closed their eyelids just before CS onset. This novel multi-measuring system for eyeblink CRs in monkeys will serve as a useful tool for elucidating the neural mechanism underlying implicit/unconscious learning in non-human primates.	P3-2-96 想起時のNR2Bサブユニット含有NMDA受容体の活性化が恐怖記憶に必要な2段階のArcタンパク質合成を誘導する Activation of NR2B-containing NMDA receptors determines the requirement of two-phase Arc synthesis for reactivated fear memory ○中山大輔1, 山崎良子1, 岩田浩一1, 松木則夫1, 野村洋1 ○Daisuke Nakayama1, Yoshiko Yamasaki1, Hirokazu Iwata1, Norio Matsuki1, Hiroshi Nomura1 東京大学大学院薬学系研究科薬品作用学教室1 Lab Chem Pharmacol, Univ of Tokyo, Tokyo, Japan1 Memory retention requires long-term maintenance. Retrieved memory undergoes destabilization via activation of NR2B-containing NMDA receptors (NMDAR) and stabilization via de novo protein synthesis around the time of retrieval. However, the mechanisms underlying stabilization at longer delayed time and the relevant destabilization have not been explored. The activity-regulated cytoskeletal-associated protein (Arc/Arg3.1) is an immediate early gene that has been widely believed to play a role in synaptic plasticity. Here, we show two-phase Arc synthesis contributing to reactivated fear memory and its induction mechanisms. Arc expression in the basolateral amygdala (BLA) was examined at several time points after retrieval of contextual fear memory. The western blotting showed biphasic increase in Arc expression. Arc expression increased at 2 and 12 hr after retrieval. To examine the role of late Arc expression in memory retention, Arc antisense oligodeoxynucleotide, an Arc translation selective inhibitor, was infused into the basolateral amygdala. Inhibition of late Arc synthesis attenuated fear memory 7 days later, but not 2 days later, suggesting that late Arc synthesis is required for memory persistence. Next, infeprodil, a NR2B subunit-selective NMDAR antagonist, was infused into the BLA before retrieval to investigate the relationship between the requirement of Arc synthesis and NR2B-containing NMDAR. Pre-retrieval infusion of ifenprodil prevented the memory impairment by inhibiting immediate or late Arc synthesis. These results indicate that activation of NR2B-containing NMDAR during retrieval induces the requirement of two-phase Arc synthesis for fear memory and that late-phase Arc synthesis contributes to the persistence of fear memory.
P3-2-97 新規物体認知記憶固定化を制御する脳領域の同定 Analysis of brain regions regulating consolidation of novel object recognition memory ○河野恭平1, 張悦1,2, 谷水俊之1, 岡野絵美子1, 喜田聡1,2 ○Kyohei Kono1, Yue Zhang1,2, Toshiyuki Tanimizu1, Emiko Okano1, Satoshi Kida1,2 東京農大院・農・バイオ1, 科学技術振興機構2 Dept. of Bioscience, Tokyo Unive. of Agriculture1, CREST, JST2 Memory consolidation is a process to stabilize short-term labile memory, generating long-term memory (LTM). A critical biochemical feature of memory consolidation is a requirement of gene expression. Importantly, previous studies have shown that memory consolidation requires the activation of cAMP responsive element binding protein (CREB)-mediated transcription. Although novel object recognition task has been used as a benchmark task to examine learning and memory in rodents, mechanisms by which novel object recognition memory is consolidated still remain unclear at the anatomical level. In this study, to understand these mechanisms, we identified brain regions where CREB-mediated gene expression is activated following the training of novel object recognition task by analysing the expression levels of c-fos, a target gene of CREB, using immunohistochemistry. Mice were exposed to novel two objects (A and B) in the open field for 15 min at 4th day following the exposure to this field in the absence of objects for 3 days (habituation session). As control groups, mice were exposed to the objects in the open field at 4th day without the habituation session, and mice were exposed to the open field in the absence of objects at 4th day with or without the habituation session. Similarly with previous studies indicating that perirhinal cortex (PRh) and insular cortex (IC) play critical roles in novel object recognition learning and memory, we observed increases in c-fos expression in these two brain regions only when mice were exposed to the novel objects following habituation session. Furthermore, similar induction of c-fos expression was also observed in the medial prefrontal cortex (mPFC), anterior cingulate cortex (ACC) and the hippocampus (HP). These results suggest that gene expression in the mPFC, ACC, HP, IC and PRh regions play essential roles in the consolidation of novel object recognition memory.	P3-2-98 海馬依存的マウス瞬目反射条件付けの記憶固定過程におよぼす抗ヒスタミン薬の効果 Effect of ketotifen on memory consolidation after trace eyeblink conditioning in mice ○北川浩規1, 臼井弘児1, 池田真行2, 川原茂敬1 ○Hiroki Kitagawa1, Koji Usui1, Masayuki Ikeda2, Shigenori Kawahara1 富山大・工・生命工1, 富山大・理・生物2 Dpt. Life Sciences and Bioengineering, Fac. Engineering, Univ. Toyama1, Dpt. Biol., Fac. Sciences, Univ. Toyama2 Sleep is considered to play an important role in memory consolidation after learning. During a sleep, two characteristic phases, rapid-eye-movement (REM) and non-REM phases, are known to alternate. Among them, the REM sleep is thought to be involved in memory consolidation. Recently, it was found that a H1 histamine receptor antagonist ketotifen decreased REM sleep and increased non-REM sleep in rats. Therefore, in the present study, we investigated the effect of ketotifen on the memory consolidation which would take place during a sleep after daily learning. We used a hippocampus-dependent type of eyeblink conditioning, the trace paradigm, in mice.Male C57BL/6 mice (8-10 weeks old) were maintained on a 12:12h light:dark cycle and conditioned for 7 days. The daily conditioning, which took about 30 min, started immediately after the onset of light period. During the conditioning, mice were trained with a 350-ms tone and a 100-ms periorbital electrical shock with a 500-ms stimulus-free interval between them. There were two kinds of groups which differed in the timing of daily injection. Immediate groups received an intraperitoneal injection of ketotifen (30 mg/kg) or saline immediately after the conditioning and brought back to their home cage, while 3-hrs groups received the injection after a rest of 3 hrs.We found that there were no significant differences between the saline and ketotifen groups receiving the injection immediately after the conditioning. However, the mice that received ketotifen-injection 3 hrs after conditioning showed a significantly lower rate of memory formation compared to the saline-injected mice. There were no significant differences in the temporal pattern of the acquired conditioned responses. These data suggest that histamine receptors play an important role in the memory consolidation after the daily trace eyeblink conditioning and that this histamine-dependent memory consolidation occurs around at least 3 hrs after the conditioning.
P3-2-99 条件づけ恐怖の復元はNMDA受容体と蛋白質合成によって制御される Reinstatement of conditioned fear is controlled by NMDA receptors and protein synthesis ○五十嵐ひかる1, 沈慧蓮1, 松木則夫1, 野村洋1 ○Hikaru Igarashi1, Huilian Shen1, Norio Matsuki1, Hiroshi Nomura1 東京大学大学院 薬学系研究科1 Lab of Chem Pharm, Grad School of Pharm Sci, Univ of Tokyo, Tokyo1 Conditioned fear induces diverse anxiety diseases such as phobia and PTSD. The extinguished response can be reinstated with a weak stimulus, not enough to induce conditioned fear. While extensive studies on the neuronal circuitry and neurochemical mechanisms leading to fear acquisition and extinction have been conducted, few studies have focused on reinstatement. In this study, we investigated the effects of N-methyl-D-aspartic acid receptor (NMDAR) antagonists, protein synthesis inhibitors, cannabinoid receptor type 1 (CB1R) antagonists, and benzodiazepine on reinstatement of conditioned fear. Conditioned mice were re-exposed to the conditioning chamber for 40 min to induce fear extinction. Then, they received a weak footshock (reminder shock). They showed freezing behavior in the original conditioning chamber the next day. Intraperitoneal injection of the NMDAR antagonist MK-801 or the protein synthesis inhibitor anisomycin 30 min before the reminder shock attenuated fear reinstatement tested the next day. However, anisomycin had no effect on fear reinstatement tested 2 h after the reminder shock. CB1R antagonists, SR141716A, and the benzodiazepine diazepam had no effect on fear reinstatement. These results suggested that NMDAR and protein synthesis-dependent plasticity contributed to the reinstatement of conditioned fear and that protein synthesis was involved in consolidation of reinstated fear.	P3-2-100 C. elegansの塩走性学習を制御するインスリン／PI3K経路との相互作用因子の同定 Identification of molecules interacting with the insulin/PI3K pathway that regulates salt chemotaxis learning in C. elegans ○酒井奈緒子1, 富岡征大2, 姜涛1, 安達健3, 飯野雄一1,2 ○Naoko Sakai1, Masahiro Tomioka2, Tao Jiang1, Takeshi Adachi3, Yuichi Iino1,2 東京大学大学院　理学系研究科　生物化学専攻1, 東京大学遺伝子実験施設2, 神奈川大学　理学部　生物科学科3 Dept. Biophys. Biochem., Grad. Schl. of Sci., Univ. of Tokyo, Tokyo1, Molecular Genetics Research Lab., Grad. Schl. of Sci., Univ. of Tokyo, Tokyo2, Dept. of Biological sci., Faculty of Sci., Kanagawa Univ., Kanagawa3 The insulin/PI 3-Kinase (PI3K) signaling pathway is widely conserved among many species including mammals, and regulates a variety of phenomena such as tumor progression and metabolism. Recently, this pathway has been reported to be involved also in learning and memory. However, so far, a mechanism by which this pathway regulates neuronal plasticity is poorly understood. We have found that the nematode C. elegans changes responses to external salt concentrations depending on food conditions. When worms are cultivated on a medium that contains sodium chloride (NaCl) and bacterial food, they are attracted to the NaCl concentration at which they were grown. In contrast, after exposure to NaCl under starvation conditions, they learn to avoid the NaCl concentration. This behavioral change is called "salt chemotaxis learning". Mutants of the insulin/PI3K pathway components show strong defects in salt chemotaxis learning. Although the DAF-16/FOXO-dependent transcription is a major output of the insulin/PI3K pathway in the control of developmental decision and aging, other machinery downstream of the insulin/PI3K pathway are predicted in salt chemotaxis learning . Mutants of daf-18 encoding PTEN, in which the PI3K signaling is abnormally increased, show avoidance of NaCl regardless of food conditions. To understand a molecular mechanism downstream of the PI3K pathway we performed a screen for suppressors of the abnormal behavior in daf-18. As a result, we obtained 4 suppressors and found that the three of them have point mutations in the goa-1 locus. GOA-1 encodes an alpha subunit of Go. Therefore communication between the Go protein signaling and the PI3K signaling may be important for salt chemotaxis learning.
P3-2-101 報酬条件下で進行が早められたフィードフォワード学習によって、報酬による学習定着効果の改善が予測される Faster progression of feedforward learning induced under rewarded training conditions predicted greater improved long-term retention of newly acquired motor memories ○阿部十也1,2, 花川隆1,4, 本田学1 ○Mitsunari Abe1,2, Nicholas Schweighofer3, Leonardo Cohen2, Takashi Hanakawa1,4, Manabu Honda1 独立行政法人　国立精神・神経医療研究センター 神経研究所 疾病研究第七部1, 米国国立衛生研究所、脳梗塞・神経疾患研究所、大脳生理学・神経リハビリテーション部門2, 南カリフォルニア大学、運動学・理学療法学部門3, （独)国立精神・神経医療研究センター　脳統合病態イメージングセンター　分子イメージング部4 Dept of Functional Brain Research, National Institute of Neuroscience, NCNP, Kodaira, JAPAN1, NINDS, NIH, Bethesda, USA2, Dept of Biokinesiology and Physical Therapy, University of Southern California, Los Angeles, USA3, Dept of Molecular Imaging, IBIC, NCNP, Kodaira, JAPAN4 Training in which good performance is rewarded or bad performance is punished results in transient behavioral improvements. We demonstrated that reward is a more effective reinforcer than punishment in eliciting improved long-term retention of newly acquired memories through posttraining offline gains. However, the neural substrate underlying improved long-term retention gains in the rewarded conditions were unknown. Here, we examined if faster progression of feedforward learning would predict induction of greater offline gains in the rewarded conditions. We studied healthy subjects who trained on a motor task under rewarded or non-rewarded neutral conditions. Subject's goal was to maintain a cursor within a target, which moved along a consistently repeated trajectory across trials. In training trials, feedforward movements of the cursor that preceded trajectory of the target were identified among periods of time in which the cursor was maintained within the target. Error was also tested before, and immediately and 30 days after the training period in the absence of reward. The two groups had similar error improvements across the training period. The rewarded group increased amount of the successful feedforward movements faster than the neutral group during training trials, although the two groups ultimately reached the comparable amount of time at the end of training. Furthermore, faster increase in the feedforward movements that were successful for the first time, but not more repetitive successful experiences of the same feedforward movements, predicted greater error improvements at 30days after the end of training in the rewarded conditions. We conclude that faster progression of feedforward learning induced under rewarded training conditions predicted greater improved long-term retention of newly acquired motor memories.	P3-2-102 記憶の維持・想起における海馬歯状回シナプス亜鉛の関与 Involvement of Zn2+ signal in dentate granule cells in LTP maintenance and memory recall ○南埜達也1, 藤井洋昭1, 中村仁総1, 高田俊介1, 奥直人1, 武田厚司1 ○Tatsuya Minamino1, Hiroaki Fujii1, Masatoshi Nakamura1, Shunsuke Takada1, Naoto Oku1, Atsushi Takeda1 静岡県大・薬・生物薬品化学1 Dept Bioorganic Chem, Univ of Shizuoka, Shizuoka1 We have demonstrated that the loss of synaptic Zn2+ by zinc chelators transiently affect dentate gyrus long term-potentiation (LTP) and object recognition memory at the same time in young rats, suggesting that synaptic Zn2+ signal in the dentate gyrus is required for memory acquisition. On the other hand, the role of Zn2+ signal in the retention and recall of memory is entirely unknown. In the present study, the real-time relationship between dentate gyrus LTP maintenance under conscious condition and recognition memory acquired was examined focused on the loss of synaptic Zn2+ by lipophilic zinc chelators. The maintenance of dentate gyrus LTP induced previously was almost abolished after intraperitoneal injection of clioquinol (30 mg/kg) or TEPN (10 mg/kg). This abolishment was not rescued even when Zn2+ signals were returned to the basal levels. To examine whether the transient loss of Zn2+ signal in the dentate gyrus affects the maintenance of dentate gyrus LTP, clioquinol was locally injected into the dentate gyrus. LTP maintenance was affected by the injection of clioquinol (200 nmol). In contrast, LTP was maintained when clioquinol-induced loss of Zn2+ signal in the dentate gyrus was ameliorated by co-injection of ZnCl2. Furthermore, the pretreatment with jasplakinolide, a stabilizer of F-actin, rescued the impairment of LTP maintenance induced by the loss of Zn2+ signal. It is likely that Zn2+ signal in dentate granule cells participates in the formation of F-actin, which is required for LTP maintenance. Simultaneously, acquired recognition memory was affected by the loss of Zn2+ signal in the dentate gyrus, but not by the pretreatment with Jasplakinolide. The present study indicates that LTP maintenance and acquired memory are erased by transient loss of Zn2+ signal in the dentate gyrus and that dentate gyrus LTP maintenance under conscious condition may be real-timely linked to the retention of recognition memory.
P3-2-103 アルツハイマー病における参照記憶障害が場所の見当識障害を引き起こすかもしれない：βアミロイド注入ラットにおける居住型迷路行動の検討 Reference memory deficit in AD may induce disorientation to place: analysis of residential maze behavior in β-amyloid protein infused rats ○角正美1,2, 山田一夫2, 一谷幸男2, 永田博司1 ○Masami Kaku1,2, Kazuo Yamada2, Yukio Ichitani2, Hiroshi Nagata1 茨城県立医療大学　保健医療学部　医科学センター1, 筑波大学大学院　感性認知納科学専攻　行動神経科学分野2 Center for Med Sci, Ibaraki Pref Univ of Health Sciences, Ibaraki1, Dept Behav Neurosci, kansei Bahav Brain sci, Univ of Tsukuba, Ibaraki2 Alzheimer's disease (AD) is characterized by a progressive impairment of cognitive function. People with AD sometimes get lost in familiar places. Impairment of working and reference memory is thought to cause confusion with places, which can lead to wandering. However, previous animal studies have focused mainly on working memory, while reference memory deficit had not completely understood. We have demonstrated that Aβ infused rats caused impairment of reference memory using "residential maze task (RMT)". RMT has some characteristics that can complement insufficient evidence of other task. Rats can acquire RMT without receiving explicit training, pain and fear. Rats receive a daily repeated training and pain in the acquisition of many behavioral tasks, however human memory is not only formed by intensive training and fear. RMT is similar to the natural feeding and drinking behavior for rats. Water cup is put in the start box, and powder-food is placed in the goal box, thus moderately hungry rats can learn the maze by coming and going between start and goal boxes in residence period. Rats can retain the acquired information in RMT over a month. Briefly, RMT is thought to be the most suitable task to evaluate reference memory in AD model rats. The aim of this study was to examine reference memory in Aβ-infused rats using RMT. In the test trial conducted everyday before the residence period, rats showed a significant decrease both of error response and running time to reach the goal box for 5 days. After the acquisition of RMT, rats were given surgery. Immediately and three weeks after the cessation of Aβ or vehicle infusion, rats were tested reference memory in RMT. Aβ-infused rats have increased the number of errors and running time compared to control rats. The results suggest that a continuous infusion of Aβ induces impairment of memory retention. This is the first report that reference memory deficit in AD animal model is concerned with disorientation to place.	P3-2-104 線虫C. elegansの塩濃度記憶を制御する神経回路の解析 A gustatory neural circuit for salt concentration memory in C. elegans ○佐藤博文1, 國友博文1, 小田茂和2, 飯野雄一1 ○Hirofumi Sato1, Hirofumi Kunitomo1, Shigekazu Oda2, Yuichi Iino1 東京大学大学院　理学系研究科　生物化学専攻1, MRC分子生物学研究所2 Department of Biophysics and Biochemistry, Graduate School of Science, the University of Tokyo, Tokyo, Japan1, Medical Research Council Laboratory of Molecular Biology, Cambridge, UK.2 Caenorhabditis elegans has a simple neurvous system and is often used for studying learning and memory. Our group has recently found that C. elegans is able to memorize a particular salt concentration. I therefore, in this study, we searched for the neural circuit required for the memory of salt concentration. First, we monitored the activity of the salt-sensing chemosensory neuron ASER. We found that ASER changed the magnitude of its responses depending on previously exposed salt concentrations. Then, we investigated the response of ASER in two kinds of mutants, which are defective in the release of either synaptic vesicles or dense core vesicles. In both mutants, the amplitude of ASER response changed similar to that of the wild type. These results suggest that the plasticity of ASER response is independent of inputs from other neurons. Next, we investigated the activity of three interneurons; AIA, AIB, and AIY. These neurons are postsynaptic neurons of ASER. We found that AIB and AIY change their responses depending on the previously experienced salt concentrations, but the response of AIA was constant regardless of the salt concentration at which the worms had been cultivated. To determine whether inputs from ASER are required for the change of AIB response, we used cell-specific rescue of sensory functions, and the results indicate that ASER contributes to the change of AIB response. According to these results, ASER, AIB, and AIY are likely to participate in memorizing salt concentrations. To assess the contribution of the three interneurons to behavior, we monitored behavioral responses of worms whose interneurons are genetically ablated individually. Although reversal frequency of the worms was different from that of the wild type, indicating successful ablation, the worms showed normal salt chemotaxis. The result indicates that there are redundancies in the neural circuit that participates in salt perception.
P3-2-105 行動タグとしてのニューロプシン Neuropsin as a behavioral tag ○石川保幸1, 塩坂貞夫1 ○Yasuyuki Ishikawa1, Sadao Shiosaka1 奈良先端大・神経機能科学1 Laboratory of Functional Neuroscience, Nara Institute of Science and Technology, Nara, Japan1 Activity-dependent synaptic plasticity is widely accepted to provide a cellular basis for learning and memory. Synaptic associativity could be involved in activity-dependent synaptic plasticity, because it distinguishes between local mechanisms of synaptic tags and cell-wide mechanisms that are responsible for the synthesis of plasticity-related proteins. An attractive hypothesis for synapse specificity of long-term memory is synaptic tagging: synaptic activity generates a tag, which captures the PRPs (plasticity-related proteins) derived outside of synapses. Previously we have been reported that neuropsin, a plasticity-related extracellular protease, was involved in synaptic tag setting. In the present study, we tested the hypothesis that neuropsin was engaged in behavioral tag setting for LTM in vivo. Behaviorally, weak training, which induces short-term memory (STM) but not LTM, can be consolidated into LTM by exposing animals to novel but not familiar environment 1 h before training. We found that neuropsin deficient mouse impaired such transformation short-term into long-term memory. These results suggest neuropsin as a tag setting for synaptic plasticity and memory.	P3-2-106 記憶想起はサーカディアン制御される Circadian regulation of memory retrieval ○長谷川俊介1,2, 細田浩司1, 張悦1,2, 太田美穂1, 谷水俊之1喜田聡1,2 ○Shunsuke Hasegawa1,2, Hiroshi Hosoda1, Yue Zhang1,2, Miho Ohta1, Toshiyuki Tanimizu1, Paul W. Frankland3, Sheena A. Josselyn3, Satoshi Kida1,2 東農大・応生科・バイオ1, 科学技術振興機構・CREST2 Dept Bio, Univ of Tokyo Agri, Tokyo, Japan1, JST, CREST, Tokyo, Japan2, Institute of Medical Sciences, University of Toronto, Toronto, Ontario, Canada3 Cognitive performance in people varies according to time-of-day, with memory retrieval declining in the later afternoon-early evening. Here we show that mice exhibit a similar time-of-day retrieval deficit following weak hippocampus-dependent learning that correlates with low forebrain levels of a key circadian transcription factor, BMAL1. To test whether BMAL1 function regulates the efficiency of memory retrieval, we inducibly expressed a dominant negative BMAL1 (BMAL1 R91A; dnBMAL1; Hosoda et al, 2004) exclusively in the forebrain. Biochemical analyses showed that dnBMAL1 mice exhibit disruptions of circadian expression cycle of BMAL1/CLOCK-target genes in the forebrain, but not in the hypothalamus. In addition, dnBMAL1 mice displayed normal circadian rhythms at the behavioral level. These results indicated that inhibition of BMAL1 activity forebrain-specifically impairs circadian transcription rhythms without affecting behavioral circadian rhythms. Behavioral analyses using social recognition, novel object recognition and contextual fear conditioning tasks showed that these mutant mice exhibited impairments of memory retrieval tested around ZT10 in a dnBMAL1 expression-dependent manner, while these mutant mice displayed normal memory retrieval tested at ZT4, 16, or 22. Furthermore, similar impairments of memory retrieval in these mutant mice were also observed in constant dark (CT) conditions. Importantly, dnBMAL1 mice also displays a deficit in c-Fos induction in the hippocampus when contextual fear memory was tested at ZT10, suggesting that hippocampal activation associated with memory retrieval is impaired at ZT10 in these mutant mice. These findings indicate that CLOCK/BMAL1 signaling pathway in the forebrain contributes to circadian regulation of memory retrieval.
P3-2-107 恐怖学習強度は嫌悪的体験の引き起こす扁桃体外側核錐体細胞の活性化によって決定される Aversive experiences set the strength of fear learning through activation of amygdala pyramidal neurons ○小澤貴明1 ○Takaaki Ozawa1, Edgar A Ycu1, Jenny Koivumma1, Joshua P Johansen1 理研BSI記憶神経回路1 Lab. for Neural Circuitry of Memory, BSI, RIKEN, Saitama, Japan1 During auditory fear conditioning, animals learn that a neutral auditory tone (conditioned stimulus, CS) predicts an aversive event (unconditioned stimulus, US). Learning finally reaches a steady state at a certain memory strength (termed the learning asymptote) beyond which further training is ineffective at producing learning. Fear memory storage occurs in the lateral nucleus of the amygdala (LA) where enhancement of auditory CS inputs onto LA pyramidal neurons may result when these cells are co-activated by the shock US. We previously found a learning dependent, gradual reduction in the US-evoked response in LA neurons (also termed prediction error coding) that is negatively correlated with the development of fear learning . This suggests the hypothesis that prediction error coding in LA neurons may set the strength of fear memories. To test this idea, we examined whether optogenetic manipulations of aversive responding in LA neurons at learning asymptote regulates fear memory strength. We first developed a 4-day fear conditioning paradigm in which rats were trained (days 1, 3) and tested (days 2, 4) twice. We found that rats reached learning asymptote after the initial 8 CS-US pairings (day 1) as learning was not enhanced by overtraining (day 3). However, learning strength was increased if a higher US intensity was used during overtraining. Combining this behavioral paradigm and optogenetic techniques, we found that: (1) additive activation of US-evoked responses in LA pyramidal cells during overtraining enhanced fear learning on the 2nd test compared to 1st test (2) Inhibition of US-evoked LA neuronal activity during overtraining abolished the increase in fear learning induced by a higher shock intensity. These results suggest that fear memory strength is set by a learning induced reduction in aversive US-evoked activation of LA pyramidal neurons, supporting a functional role for LA prediction error coding in regulating the strength of fear memories.	P3-2-108 ビタミンB1欠乏は海馬依存的学習・記憶形成の障害を誘導する Thiamine deficiency induces deficits in hippocampus-dependent learning and memory formation ○岸本拓也1, 大石諭1, 永田幹1, 長谷川俊介1,2, 喜田聡1,2 ○Takuya Kishimoto1, Satoru Ohishi1, Kan Nagata1, Shunsuke Hasegawa1,2, Satoshi Kida1,2 東農大・応生科・バイオ1, 科学技術振興機構・CREST2 Dep. of Bio, Tokyo Univ. of Agri, Japan1, JST, CREST, Tokyo, Japan2 Thiamine deficiency (TD) induces cognitive dysfunction including deficits in learning and memory known as Wernicke-Korsakoff's syndrome in human. Previous studies have shown that TD reduces basal brain-derived neurotrophic factor (BDNF) levels in the hippocampus and frontal cortex of rats with pyrithiamine-induced thiamine deficiency (PTD) (Ryan et al., 2011). However, the molecular mechanisms by which TD leads to deficits in learning and memory still remain unclear. In this study, to understand these mechanisms, we first examined effects of PTD on hippocampus-dependent memory performances in mice. PTD-treated mice displayed bodyweight loss and motor ataxia just after the PTD treatment with dietary thiamine deprivation for 10 days together with daily injections of thiamine antagonist. However, these PTD mice displayed comparable body weight and motor performance with control groups by feeding normal diet for at least 3 weeks together with the treatment with a single injection of thiamine after the PTD treatment, suggesting that these PTD-treated mice recovered from PTD. However, these PTD mice recovered from the PTD displayed impairments in social recognition memory, contextual fear memory and spatial memory. Interestingly, biochemical analyses showed that PTD mice displayed decreases in expressions of GluA1 in the hippocampus. Taken together, these observations indicated that PTD leads to severe deficits in hippocampus-dependent memory performances.
P3-2-109 条件づけ恐怖の復元における内側前頭前皮質の関与 The involvement of medial prefrontal cortex in the spontaneous recovery and reinstatement of conditioned fear ○手代木知恵1, 今村菜津子1, 沈慧蓮1, 松木則夫1, 野村洋1 ○Chie Teshirogi1, Natsuko Imamura1, Huilian Shen1, Norio Matsuki1, Hiroshi Nomura1 東京大学大学院薬学系研究科　薬品作用学教室1 Laboratory of Chemical Pharmacology, Graduate School of Pharmaceutical Sciences, The University of Tokyo, Tokyo, Japan1 Anxiety disorder is a blanket term covering several different forms of mental illness of abnormal fear and anxiety. In some cases, despite successful reduction of fear through exposure psychotherapy, inappropriate fear responses reappear with passage of time (spontaneous recovery) or by re-exposure to stimuli or stress (reinstatement). In contrast to extensive studies on fear acquisition and extinction, few studies have focused on spontaneous recovery or reinstatement. We investigated what region of the brain is involved in spontaneous recovery or reinstatement. Mice were conditioned with footshocks in a chamber, and re-exposed to the chamber without footshocks for 40 min on the next day. This long-term re-exposure extinguished conditioned fear responses. Then, they were left in their home cages for 4 weeks or received a weak immediate shock in a different chamber. As a result, re-exposure to the original conditioning chamber induced fear responses. We investigated the neuronal activity during the expression of recovered fear by measuring the density of c-Fos positive neurons. In the prelimbic region (PL) of medial prefrontal cortex (mPFC), c-Fos expression was elevated with fear conditioning and during spontaneous recovery as well, and in the infralimbic region (IL) of mPFC, c-Fos expression was lower with reinstatement than that with extinction, indicating that exposure to the conditioning chamber activated PL with spontaneous recovery but not IL with reinstatement. When mice received microinjection of anisomycin, a protein synthesis inhibitor, into mPFC before the weak immediate shock, the reinstatement was impaired. This suggests that de novo protein synthesis in mPFC is required for reinstatement. These findings indicate that an important role of mPFC in spontaneous recovery and reinstatement of conditioned fear.	P3-2-110 順序および運動学習のマウスモデルとしての3レバー・オペラント課題 The three-lever operant task as mouse model of motor sequence and skill learning ○米田貢1, 田端佑樹1, 菊池ゆひ1, 少作隆子1 ○Mitsugu Yoneda1, Yuki Tabata1, Yui Kikuchi1, Takako Ohno-Shosaku1 金沢大学　医薬保健研究域　保健学系　リハビリテーション科学領域1 Fac. Health Sci. Kanazawa Univ., Kanazawa, Japan1 To elucidate the neural basis of motor sequence and skill learning, we have established the three-lever operant task in rats. In this task, rats were trained to press three levers in a given sequence. We previously reported that the performance of this task was impaired in Parkinson's model rats, indicating its dependence on the basal ganglia. We also found that rats can perform the three-lever task either quickly (lever press interval <0.5 s) or slowly (>2 s). In this study, we examined whether this task is applicable to mice, which have a great advantage in the manipulation of genetic properties. Three C57BL/6NCr male mice were used. Operant boxes were modified for mice. Three levers (1.8 × 1.5 cm) were protruded 1.8 cm into the chamber, and the left, center and right levers were positioned 2, 5.5 and 2 cm above the floor, respectively. One training session lasting 60 min was given once a day and five times a week. The total amount of food per day was set to 3 g, but decreased by 0.5-1.5 g when the number of lever press was small. The weight was maintained between 20-25g. At the age of eight weeks, mice were trained to press any one of the three levers for a food reward (single-lever task) as shaping (9-19 sessions), and then trained to press three levers in a given sequence (three-lever task) with or without time limitation. In the three-lever task with time limitation, mice were required to press the second (or third) lever within a given time (T) after the onset of the first (or second) lever press. Although the performance was markedly impaired at T= 0.5 s, the mice showed good performance at T= 1 s and obtained more than 100 pellets in one session, which is comparable to rats. The present study suggests that the three-lever operant task might be useful for studying the neural basis of motor sequence and skill learning in mice.
P3-2-111 カルシニューリンを起点とした想起後の受動回避記憶の増強 Enhancement of fear memory after retrieval through the activation of calcineurin ○福島穂高1,2, 張悦1,2, 金亮1,2, 喜田聡1,2 ○Hotaka Fukushima1,2, Yue Zhang1,2, Ryang Kim1,2, Satoshi Kida1,2 東京農大・応生科・バイオ1 Dep. of Bioscience, Tokyo Univ. of Agriculture1, JST, CREST2 Recent studies have shown that retrieved memory becomes labile through proteasome-dependent protein degradation and then re-stabilized through a gene expression-dependent reconsolidation, whereas prolonged retrieval induces memory extinction that is also a gene expression-dependent process. We have shown using light-dark inhibitory avoidance (IA) task that the re-exposure to the light box enhanced IA memory through the activations of proteasome-dependent protein degradation and protein synthesis in amygdala (AMY), hippocampus (HP) and medial prefrontal cortex (mPFC). In this study, we have tried to identify upstream signal transduction pathway to activate proteasome-dependent protein degradation. Calcium/calmodulin-dependent phosphatase 2B, calcineurin (CaN) in HP is required for extinction of contextual fear memory (de la Fuente et al., 2011). However, the role of CaN activation on memory reconsolidation and enhancement still remains unclear. Therefore, we examined effects of micro-infusion of CaN inhibitor, FK506. First, our immunohistochemical analyses showed a systemic injection of FK506 blocked significant increases in poly-ubiquitinated proteins in AMY, HP and mPFC regions following the re-exposure to the light box. Second, a micro-infusion of FK506 into AMY blocked the disruption of IA memory by protein synthesis inhibition and the enhancement of IA memory, suggesting that the activation of CaN in AMY is required for both destabilization and enhancement of IA memory. Furthermore, the inhibition of CaN in HP or mPFC also blocked the enhancement of IA memory, suggesting that CaN activation in HP/mPFC is required for memory enhancement. Taken together, our observations suggest that the activation of CaN enhances reactivated IA memory though proteasome-dependent protein degradation and new gene expression.	P3-2-112 遅発的なArc発現は神経活動非依存的であるが恐怖記憶の持続に寄与する Delayed Arc expression is independent of neuronal activity but, contributes to persistence of fear memory ○岩田浩一1, 中山大輔1, 松木則夫1, 野村洋1 ○Hirokazu Iwata1, Daisuke Nakayama1, Norio Matsuki1, Hiroshi Nomura1 東京大院　薬　薬品作用学1 Dept of Pharm, Univ of Tokyo, Tokyo1 Memory consolidation requires de novo protein synthesis. Arc (activity-regulated cytoskeleton protein, Arg3.1) is an immediate early gene and involves in AMPA receptor endocytosis. The expression is transiently up-regulated by robust patterned synaptic activities related to associative learning, and it contributes to consolidation of synaptic plasticity and long-term memory. However, the detailed temporal pattern of Arc expression following learning remains unclear. Here, we show 2 peaks of Arc expression accompanying fear conditioning. Mice received contextual fear conditioning and were sacrificed 2, 6, 12, 24 h later. Arc expression in the dorsal hippocampus (DH) was increased at 2 and 12 h after conditioning. To test whether the delayed Arc expression depends on neuronal activity, mice received microinjection of tetrodotoxin (TTX) into DH 9 h after conditioning and Arc expression 12 h after conditioning was examined. TTX had no effect on the delayed Arc expression, suggesting that delayed Arc expression is independent of neuronal activity. To examine the role of delayed Arc expression, mice underwent microinjection of Arc-antisense oligodeoxynucleotide (ODN) into DH after conditioning. Arc-antisense ODN attenuated fear memory after 7 days, but not after 2 days. The result indicates that the delayed Arc expression contributes to the persistence of contextual conditioned fear. These findings suggest that the expression mechanisms and contribution to memory consolidation are different between immediate and delayed Arc expression.
P3-2-113 マウスにおける行動柔軟性：包括的解析のための新規ハイスループット試験系の開発 Behavioral flexibility in mice: Development of a novel high-throughput assay for comprehensive analysis ○遠藤俊裕1, ベナー聖子1, 遠藤のぞみ1, 古瀬民生2, 若菜茂晴2, 梶井靖3遠山千春1, 掛山正心1 ○Toshihiro Endo1, Seico Benner1, Nozomi Endo1, Tamio Furuse2, Shigeharu Wakana2, Yasushi Kajii3, David P. Wolfer4, Hans-Peter Lipp4, Chiharu Tohyama1, Masaki Kakeyama1 東京大院・医・疾生セ・健康環境医工学1, 理研　バイオリソースセンター　マウス表現型解析開発チーム2 Lab of Environ Health Sci, CDBIM, Univ of Tokyo, Tokyo1, Tech. Dev. Team for Mouse Phenotype Analysis, RIKEN BRC, Tsukuba2, Abbott Japan, Tokyo3, Inst of Anat, Univ of Zurich, Zurich, Switzerland4 Behavioral flexibility is ability that humans and animals are endowed to adapt to an ever-changing environment. Although development of behavioral flexibility assay for mice as human models has been challenged by many researchers, the standardized test has not been available. To overcome this problem, we aimed to develop a high-throughput and comprehensive assay of behavioral flexibility applicable to psychiatry research using mouse models. The present behavioral flexibility test is based on a behavioral sequencing task using a fully computerized testing apparatus for group-housed mice, IntelliCage (Endo et al., Behav. Brain Res., 2011). The validity and efficiency of the behavioral sequencing task (i.e., reward retrieval by shuttling between distantly-positioned chambers) and its serial reversals on assaying behavioral flexibility was evaluated through the time-course analyses under different timescales (i.e., hours, days, and weeks). We also employed mouse models of mental disorders (i.e., NR1/Grin1 mutant mice, Tuba1 mutant mice, and mice perinatally exposed to dioxin) for the validation. The time-course analyses successfully delineated the processes of behavioral flexibility in mice. Intriguingly, mice became adept at behavioral shift through the serial reversal learning, suggesting that they are able to learn the task rule itself. In contrast, each mouse model exhibited the signs of delayed adaptation to the new rule in the time-course analyses, compared to the wild type mice, suggesting that behavioral flexibility is impaired. The present results demonstrated that our newly developed method based on a behavioral sequencing task is valid and efficient for characterizing the complex processes of mouse behavioral flexibility. Since the signs of behavioral inflexibility were shown robustly in both genetic and environmental mouse models of mental disorders, the method developed in this study was also validated to be applicable to psychiatric research.	P3-2-114 3秒間の聴覚刺激を用いた恐怖記憶消去学習の特性 Characterization of fear extinction using 3-sec auditory conditioned stimulus in male rats ○三上香織1, 清川泰志1, 武内ゆかり1, 森裕司1 ○Kaori Mikami1, Yasushi Kiyokawa1, Yukari Takeuchi1, Yuji Mori1 東京大院・農・獣医動物行動1 Laboratory of Veterinary Ethology, The University of Tokyo, Tokyo, Japan1 In fear conditioning, an animal shows fear responses to a conditioned stimulus (CS) if it is previously paired with an aversive unconditioned stimulus (US). Recently, we found in male rats that a 3-sec tone can be used as the auditory CS and induces fear responses including freezing behavior when it is paired with foot shocks as the US beforehand. Because 3-sec is an unusually short duration for the auditory CS, characters of the fear responses to 3-sec CS are still unknown. Here we assessed the efficacy of extinction training using 3-sec CS. On day 1, all the subjects received 7 repetitions of a 3-sec tone (8kHz, 70dB) that terminated concurrently with a 0.5-sec foot shock (0.55mA) during the conditioning procedure. On day 2, half of the subjects underwent extinction training in which only the CS was presented 24 times at randomly varied intervals from 1 to 2-min (training group). The other half of the subjects was placed in the test box during the same period without CS presentation (non-training group). On day 3, both groups were again placed in the test box and presented CS twice. The duration of freezing during 20-sec after the onset of each CS was defined as the response to each CS and its intensity was expressed as a ratio. We evaluated the efficacy of the extinction training by comparing the average of two freezing intensity on day 3. The conditioning procedure increased the intensity of freezing to the CS in both groups. At the end of the extinction training, the intensity of freezing was decreased as compared to the initial one. However, on day 3, the training group showed the same intensity of freezing as compared to the non-training group. To verify our procedure, we also prepared the subjects with the same procedures but using 20-sec tone as the CS. In these subjects, the extinction training decreased the intensity of freezing on day 3 as seen in many literatures. These results suggest that the extinction training is less effective when we use 3-sec CS.
P3-2-115 恐怖記憶形成におけるオレキシンの役割 Orexin system plays an important role in fear memory formation ○征矢晋吾1, 長谷川恵美1, 昌子浩孝2, 宮川剛2, 三枝理博1, 桜井武1 ○Shingo Soya1, Emi Hasegawa1, Hirotaka Shoji2, Tsuyoshi Miyakawa2, Michihiro Mieda1, Takeshi Sakurai1 金沢大学大学院　医学系研究科　分子神経科学・統合生理学1, 藤田保健衛生大・総合医科学研究所・システム医科学研究部門2 Departmet of molecular neuroscience and integrative physiology, Kanazawa Univ. Kanazawa , Japan1, Division of Systems Medical Science., Institute for Comprehensive Medical Science, Fujita Health Univ., Aichi, Japan2 Orexin A and B acts on two G-protein coupled receptors, orexin receptor 1 (OX1R) and receptor 2 (OX2R) and have been implicated in regulating sleep and wakefulness, energy homeostasis, feeding behavior, and reward system. Orexin system may be also implicated in the regulation of stress and arousal response and the formation of emotional memory, since orexins densely innervate the monoaminergic nuclei such as locus coeruleus (LC), dorsal raphe, tuberomammillary nucleus (TMN), and their neurons strongly involves in fear, anxiety, and stress response. Moreover, orexin neurons receive abundant inputs from the limbic system, and the importance of this connection is apparent in the defense response. These observations suggest that orexin neurons are activated during emotional event and play an important role in fear memory formation. To address this hypothesis, we examined OX1R deficient mice (Ox1r-/- ), OX2R deficient mice (Ox2r-/-), orexin receptor double knockout mice (OXDKO), and prepro-orexin knockout mice (ORXKO) in a classic cued and contextual fear-conditioning test. We found that these mutant mice showed significant impairment in conditioning and test period of contextual fear conditioning test. Also, in cud fear test, mutant mice showed significant decrease in freezing behavior and especially OXDKO and ORXKO presented larger decreases in freezing time during conditioning period. Similar pattern was seen in the test period except Ox2r-/-showed normal fear response against auditory CS. These observations strongly suggest that orexin system involves in establishing fear memory via OX1R and OX2R signaling. Also, cue-dependent fear memory formation may require specifically OX1R contribution. Restoring OX1R in noradrenergic neurons in the LC increase the freezing time during the test period comparable to wild type, supporting this hypothesis. These findings suggest that lateral hypothalamus plays a crucial role in the development of fear memory through orexin system.	P3-2-116 恐怖条件づけ時の前頭連合野の神経活動 Neuronal activity in the frontal association cortex during fear conditioning ○野中綾子1, 松木則夫1, 野村洋1 ○Ayako Nonaka1, Norio Matsuki1, Hiroshi Nomura1 東京大院・薬・薬品作用1 Lab. Chem. Pharmacol., Grad. Sch. Pharm. Sci., Univ. Tokyo, Tokyo1 Fear conditioning is a representative associative learning task in which subjects learn the relationship between a neutral conditioned stimulus (CS; e.g. tone or context) and an aversive unconditioned stimulus (US; e.g. footshock), and after the conditioning, the subjects become to express fear responses. Frontal association cortex (FrA) is one of the important regions for fear conditioning. It is known that inactivation of the FrA impairs memory consolidation, and spine elimination in the FrA is induced by fear conditioning. However, how memories are controlled in the FrA is still unclear. Observation of neuronal activity at individual neuron level is required to understand the memory processing because memories are controlled specific neuronal populations. To reveal the neuronal activity in the FrA at individual neuron level, we used in vivo two-photon calcium imaging. We set up the fear conditioning system in which we can carry out calcium imaging during the conditioning and retrieval. Male C57BL/6J mice were anesthetized with urethane (1.5 g/kg, i.p.) and FrA neurons were labeled with Oregon Green BAPTA-1 (OGB-1). Conditioning tones were presented via earphones fixed at the mice side, and electrical shocks were delivered via 26-gauge needles inserted into the footpad. Under this fear conditioning system, we could observe calcium transients in FrA during CS and US presentation. Elucidation of neuronal activity in the FrA during fear conditioning and retrieval can be possible with this system.
P3-2-117 フタホシコオロギの感覚的事前条件付け Sensory preconditioning in the cricket Gryllus bimaculatus ○松本幸久1,2, 平島大輔3, 水波誠2 ○Yukihisa Matsumoto1,2, Daisuke Hirashima3, Makoto Mizunami2 東京医科歯科大・教養・生物1, 北大・理・生命2, 東北大・理・生命3 Dept Biol, Coll Lib Art Sci, Tokyo Med Dent Univ, Chiba1, Grad Sch Life Sci, Hokkaido Univ, Sapporo, Japan2, Grad Sch Life Sci, Tohoku Univ, Sendai, Japan3 Sensory preconditioning is a variation of high-order conditioning, which forms an association between two neutral stimuli without an apparent reinforcement. The purpose of this study is to establish the sensory preconditioning paradigm in the cricket Gryllus bimaculatus and to investigate the mechanism of learning between neutral stimuli. In the first phase of training, crickets received pairing trials of odor and visual pattern, and in the second phase, they received pairing trials of the visual pattern and water reward (or saline punishment). As a result, crickets showed conditioned response to the odor used in the training over another odor which was only used in the odor preference tests, suggesting that crickets are capable of the sensory preconditioning. In our recent study, we demonstrated that appetitive reinforcement requires octopaminergic transmission and aversive reinforcement requires dopaminergic transmission in classical conditioning of crickets. Therefore we investigated whether the association between neutral stimuli in the first phase of sensory preconditioning is octopaminergic or dopaminergic transmission-dependent. As a result, blockade of neither octopaminergic nor dopaminergic signaling impaired association between neutral stimuli. We conclude that the mechanisms for association between neutral stimuli differ from those for reward or punishment learning.	P3-2-118 Persistent activity at perirhinal and lateral entorhinal cortices supporting long-term temporal conditioning ○Eugenie Suter1, Craig Weiss1, John F. Disterhoft1 Northwestern University1 Perirhinal (PR) and lateral entorhinal (latEC) cortices, major bidirectional connecting nodes between higher-order cortex and hippocampus, exhibit persistent activity in vitro following a brief stimulus. PR is essential for acquisition, but latEC only for retention, of trace eyeblink conditioning (tEBC), suggesting that these parahippocampal areas may possess mechanisms for bridging time between stimuli in vivo. We recorded multiple single neurons from PR, latEC and CA1 in the awake rabbit during tEBC, in which a 500ms gap ("trace period") separated conditioned (CS; whisker vibration) and unconditioned stimuli (US; corneal airpuff). Among 408 cells recorded in PR and latEC, 46% were significantly modulated during task performance, showing either an increase or decrease in firing rate during CS presentation or in the trace interval between CS and US. One month after acquisition, rabbits were retrained on tEBC, and results suggest that rate-decreasing cells are more prevalent post-consolidation than during acquisition in PR and latEC, possibly representing an increased inhibitory component in the circuit. We conclude that persistent post-stimulus activity may contribute to binding stimuli together across time, and in latEC, may support long-term (hippocampal-independent) tEBC. Further data are needed to conclusively compare roles of PR, latEC and hippocampus pre- and post-consolidation and draw conclusions for network function in long-term memory.
P3-2-119 Sensory memory forms in the auditory-associated cortical area in zebra finch song learning ○Christian Flecke1, Yoko Yazaki-Sugiyama1 Neuronal Mechanism for Critical Period Research Unit, Okinawa Institute of Science and Technology (OIST) Graduate University, Okinawa, Japan1 During the sensory phase of song learning (10-30 post-hatch days [PHD]), juvenile male zebra finches (Taeniopygia guttata) memorize a tutor song (TUT) based upon their auditory experiences. This memory template is then used during the sensorimotor phase (35-90 PHD) by an unknown auditory feedback comparator circuit, to render the birds' own vocalizations similar to those of the TUT. Increasing evidence suggests that the neural substrate for the TUT memory is located in the caudomedial nidopallium (NCM), the area homologous to the mammalian auditory-associated cortex. However, to date, no single neuronal units showing clear TUT-biased auditory responses have been identified in this brain area. In this study we found that neurons in the NCM showed strong TUT-biased responses after only a short period of TUT exposure. Male juvenile zebra finches were isolated from TUT together with their mothers and siblings, starting at 10-12 PHD, before they could form a memory of it. With this protocol, it is possible to delay closure of the sensory phase, and to more easily observe singing behavior and neuronal responses. Then, starting at 55 PHD, they were tutored by a male zebra finch for five days. Although this short period of tutor exposure was long enough to form a memory for subsequent motor mimicking, it was not long enough for motor copying. Subsequent to tutoring, we probed neuronal units throughout the entire NCM with TUT, using familiar and unfamiliar songs. Unlike neurons in the anterior forebrain pathway (homologous to the mammalian basal ganglion thalamocortical pathway), which start to show a selective response to TUT or to a birds' own song only after they start mimicking TUT (ca. 60 PHD during normal development), neurons in the subarea of NCM acquired and showed selective responses to TUT after only 5 days of exposure. These results suggest that sensory memory forms within a few days in the auditory-associated cortical area in zebra finches during song learning.	P3-2-120 Differential Responses of the CA1, CA2 and CA3 Regions to Gradual Contextual Change ○Marie Wintzer1, Roman Boehringer1, Thomas J. McHugh1 Riken Brain Science Institute1 The hippocampus is necessary for the learning and mnemonic association of context. In rodents, changes in the context stimulate exploration and concomitant activation of neuronal ensembles representing the explored space and the cues contained within it. We have subjected mice to various degrees of cue and contextual change and using Arc/H1a+ catFISH we demonstrate that the hippocampal CA subregions (CA3/CA2/CA1) exhibit differential responses to shifts in objects and contexts. The distribution of the joint CA3/CA2/CA1 response across the protocols indicates there may be unique network states representing varying degrees of environmental change. Further, our results suggest that ensemble activity in CA2 may be highly responsive to any change in the context and support the hypothesis that encoding in each field is unique.
P3-2-121 Withdrawn	P3-2-122 Dopaminergic Polymorphisms and Memory ○Chun-Yen Chang1, Pei-Jung Lin1,2, Ying-Chun Cho1, Ting-Kuang Yeh1, Chung-Yi Hu3 Science education center, national taiwan normal university1, Graduate Institute of Clinical Medicine, National Taiwan University2 This study investigates the association between memory and SNPs in genes involved in the dopaminergic pathway and serotoninergic pathway in a sample of healthy individuals. The sample includes one hundred and thirty-four Taiwanese undergraduate volunteers of similar cognitive ability. The Chinese versions of the Wechsler Memory Scale (WMS-III) was employed. The genes involved in the dopaminergic and serotoninergic pathway selected for analysis included TH, DDC, SLC6A3, SLC6A2, SLC18A1, SLC18A2, MAOA, MAOB, COMT,DRD1, DRD2, DRD3, DRD4, DRD5, SLC6A4, HTR1B, HTR2A, HTR2C, HTR3A, HTR3C, HTR3D, and HTR3E. Sixty nonsynonymous SNPs among all the candidate genes involved in the dopaminergic and serotoninergic pathway were identified on the basis of the information available in the Entrez Gene, HapMap, and Ensembl databases. We performed a pilot study in which we genotyped DNA samples from 20 healthy participants. SNPs with a minor allele frequency greater than 5% in the samples analyzed were identified as the most promising candidates, having representative common variants. Our findings indicate that the dopamine receptor D3 (DRD3) Ser9Gly polymorphism are associated significantly with long-term auditory memory. Further analysis detects no significant associations in the other polymorphisms and indices. Future replicated studies with larger sample sizes, and studies that consider different ethnic groups, are encouraged.

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