一般演題（ポスター）

一般演題（ポスター） Neurotransmission I

1P-10 CAPS1 stabilizes SVs docking state in hippocampal CA3-CA1 synapses Ishii Yuuki¹，Ishii Chiaki¹，Shinoda Yo^1,2，Fukazawa Yugo³，Sadakata Tetsushi⁴，Iwasato Takuji^5,6，Itohara Shigeyoshi²，Furuichi Teiichi^1,2 ¹Department of Applied Biological Science, Tokyo University of Science，²Laboratory for Memory Mechanisms, RIKEN Brain Science Institute，³Division of Cell Biology and Neuroscience, Faculty of Medical Sciences, University of Fukui，⁴Advanced Scientific Research Leaders Development Unit, Gunma University，⁵Division of Neurogenetics, National Institute of Genetics，⁶Department of Genetics, SOKENDAI AbstractCalcium-dependent activator protein for secretion 1（CAPS1）is a cytosolic protein, which associates with dense-core vesicle secretion in endocrine cells, however, their neuronal function is still largely unknown because of Caps1 knock-out（KO）results in prenatal death. Here we show that CAPS1 stabilizes the docking state of synaptic vesicle（SV）to presynaptic active zone using forebrain specific Caps1 conditional KO（cKO）mice. The synaptic transmission is strongly reduced and paired-pulse facilitation shows significant alteration in Caps1 cKO so that the impairment of SV release is expected. Morphological analysis shows accumulation of SVs in presynapse without any morphological changing. Interestingly, even though SV accumulation is occurred, the percentage of presynaptic button contained docked vesicle is markedly reduced in Caps1 cKO. Finally, SV release experiment revealed by time-lapse imaging indicates the decreased SV release in the absence of CAPS1. These data suggest that CAPS1 stabilizes SV docking state to enhance SV release.		1P-11 Conditional knockout and optogenetic study on the involvement of the secretion-related protein CAPS1 in oxytocin-associated social and maternal behavior Minami Haruka¹，Yamaga Ryosuke¹，Shinoda Yo¹，Sakimura Kenji²，abe Manabu²，Furuichi teiichi¹ ¹Dept. of Appl. Biol. Sci., Fac. of Sci. and Tech. Tokyo Univ. of Sci.，²Dept of Cellular Neurobiology BRI, Niigata Univ Calcium-dependent activator protein for secretion 1（CAPS1）plays a regulatory role in dense-core vesicle（DCV）exocytosis pathway. CAPS1 is widely expressed in the mouse brain including paraventricular nucleus（PVN）and supraoptic nucleus（SON）that contain oxytocin（OXT）-producing neurons. OXT is known as a neuropeptide, which is associated with social and maternal behavior. Thus, we hypothesized that CAPS1 potentially regulates social and maternal behavior through regulating OXT secretion. In this study, we generated conditional knock-out（cKO）mice lacking Caps1 in OXT neurons and tried to compare WT and cKO mice by optogenetic control of PVN or SON via microinjection of adeno-associated virus vectors containing channelrhodopsin 2（ChR2）. The effect of CAPS1 deficiency in OXT neurons was analyzed in terms of activity-dependent OXT secretion, social and maternal behavior. We will show the recent progress in testing the implication of CAPS1 in OXT-related brain function and behavior.

1P-12 IRBIT suppresses CaMKII-alpha activity and contributes to catecholamine homeostasis through tyrosine hydroxylase phosphorylation Kawaai Katsuhiro¹，Mizutani Akihiro²，Shoji Hirotaka³，Ogawa Naoko¹，Ebisui Etsuko¹，Kuroda Yukiko⁴，Wakana Shigeharu⁵，Miyakawa Tsuyoshi³，Hisatsune Chihiro¹，Mikoshiba Katsuhiko¹ ¹Laboratory for Developmental Neurobiology, Brain Science Institute, RIKEN，²Department of Pharmacotherapeutics, Showa Pharmaceutical University，³Fujita Health University，⁴Laboratory of Cell and Tissue Biology, Keio University School of Medicine，⁵Technology and Development Team for Mouse Phenotype Analysis, BioResource Center, RIKEN Inositol 1,4,5-trisphosphate receptor（IP₃R）binding protein released with IP₃（IRBIT）contributes to various physiological events（electrolyte transport, mRNA polyadenylation, and the maintenance of genomic integrity）through its interaction with multiple targets. However, little is known about the physiological role of IRBIT in the brain. In this report, we identified calcium calmodulin-dependent kinase II alpha（CaMKIIα）as an IRBIT-interacting molecule in the central nervous system. IRBIT binds to and suppresses CaMKIIα kinase activity by inhibiting the binding of calmodulin to CaMKIIα. In addition, we show that IRBIT knockout mice show elevated catecholamine levels, increased locomotor activity, and social abnormalities. The level of tyrosine hydroxylase（TH）phosphorylation by CaMKIIα, which affects TH activity, was significantly increased in the ventral tegmental area of IRBIT deficient mice. We concluded that IRBIT suppresses CaMKIIα activity and contributes to catecholamine homeostasis through TH phosphorylation.		1P-13 Generation of GAD65 knockout rats using TALEN-mediated genome editing Yanagawa Yuchio¹，Kakizaki Toshikazu¹，Watanabe Masahiko² ¹Dept. of Genet. ＆ Behav. Neurosci., Gunma Univ. Grad. Sch. of Med.，²Dept. of Anat., Hokkaido Univ. Grad. Sch. of Med. GABA is the major inhibitory neurotransmitter in the adult mammalian CNS. Glutamate decarboxylase（GAD）is the rate-limiting enzyme that catalyzes the production of GABA from glutamate. There are two isoforms of GAD, GAD65 and GAD67 according to their molecular masses, and they are encoded by independent genes. GAD65 knockout mice showed an increase in susceptibility to seizures and changes in emotional behavior such as anxiety and aggression, whereas GAD67 knockout mice were shown to die of cleft palate. However, the size of the mouse is a potential limitation for some types of physiological monitoring, behavioral testing, brain mapping and repeated blood sampling. To overcome the problem, we have generated GAD65 knockout rats using TALEN genome editing. mRNAs encoding TALENs targeted to exon 1 of GAD65 were microinjected into single-cell rat embryos and transferred to pseudopregnant recipients. Three founders with monoallelic or biallelic mutations were backcrossed to wild-type rats, and then heterozygous mutants were obtained. These heterozygous rats possessed both wild-type GAD65 allele and GAD65 mutant allele containing either 8 bp, 314 bp or 490 bp deletions. Homozygous GAD65 mutant rats with 8 bp or 314 bp deletions were generated by crossing their respective heterozygous mutant rats each other. Western blot analysis demonstrated that GAD65 protein was not detected in the homozygous mutant brain. This rat model provides a new experimental tool for investigating the pathophysiology of GABAergic transmission.

1P-14 Functional characterization of oxytocin/vasopressin-like peptide in social insect, ants Tahara Hiroki¹，Miura Masayuki^1,2，Koto Akiko^1,2 ¹Department of Genetics, Graduate School of Pharmaceutical Sciences, The University of Tokyo，²CREST Oxytocin（OXT）and arginine-vasopressin（AVP）are neurohypophysial hormones composed of 9 amino acids and known to function as neuromodulators. OXT/AVP-like peptides are evolutionarily conserved among wide range of animals, from worms to humans. Their function is well studied in highly social mammals to regulate social behaviors. In addition, recent studies also suggest that OXT/AVP-like peptides function to regulate social behaviors such as courtship and mating in birds, fishes and some invertebrates such as molluscs and nematodes. However, the evolutionary origin of their molecular function is still unrevealed and we aim to study the neuronal function of their homologous nonapeptide, inotocin（INT）in social insects, ants. Ants exhibit sophisticated social organization within their colonies which is characterized by the reproductive caste differentiation and the division of labor. Ant colonies have a reproductive caste which consists of queens and males, and a non-reproductive caste, workers. Workers show the division of labor, that is, each worker specializes in one job such as foraging, nurturing and nest construction. They can flexibly change their jobs according to surrounding environment and colony demands. From this, we hypothesize that INT signaling would be involved in the regulation of the unique social behaviors in ants. We firstly established the in vitro assay system to measure the INT receptor activity by utilizing cultured cells, and confirmed that the putative INT receptor is activated by INT peptide. We also found that OXT peptide can activate the INT receptor at high dose, which suggests the similarity of INT and OXT signaling pathways. We are now testing the effect of OXT/AVP signaling agonists/antagonists in INT signaling, for the further in vivo study to manipulate INT signaling pathway and examine whether it is involved in the regulation of social behaviors in ants.