一般口演（Oral）

Oral Synaptogenesis and Activity-dependent Development 一般口演シナプス形成と活動依存的発達

7月27日（土）16:30～16:45　第9会場（朱鷺メッセ　3F　306+307） 3O-09e1-1 Mitochondrial fission regulates presynaptic function and axon branching by limiting axonal mitochondrial size Seok-Kyu Kwon（Kwon Seok-Kyu）¹，Tommy Lewis（Lewis Tommy）^2,3，Annie Lee（Lee Annie）³，Reuben Shaw（Shaw Reuben）⁴，Franck Polleux（Polleux Franck）³ ¹Korea Institute of Science and Technology, Seoul, South Korea ²Department of Neuroscience, Columbia University, New York, USA ³Oklahoma Medical Research Foundation, Oklahoma City, USA ⁴Salk Institute for Biological Studies, La Jolla, USA Neurons have highly polarized structures comprising axons and dendrites. More interestingly, mitochondria also display extreme degrees of compartment-specific morphology. In cortical pyramidal neurons, dendritic mitochondria are long and tubular whereas axonal mitochondria have uniformly short length in vitro and in vivo. Mitochondrial morphology is dynamically regulated by fusion and fission, however compartment-dependent control mechanism is still not known. In addition, the functional significance of maintaining small axonal mitochondria has not been suggested yet. Here we demonstrate that Mitochondrial Fission Factor (MFF), one of the Drp1 receptors, is required for determining the mitochondrial size entering the axon, and then maintaining their small shapes. In MFF-deficient neurons, axonal mitochondria are elongated, but normally trafficked along the axon and well targeted to presynaptic sites. In addition, their membrane potential and ability to generate ATP are not significantly altered. More strikingly, elongated presynaptic mitochondria in MFF-deficient condition augment their Ca2+ uptake capacity thereby decrease presynaptic Ca2+ and synaptic vesicle exocytosis. Presynaptic release is essential for normal axon development, therefore elongated mitochondria-dependent changes lead reduced terminal axon branching in vivo. Our results uncover a novel mechanism controlling neurotransmitter release and axon branching through fission-dependent regulation of presynaptic mitochondrial size.		7月27日（土）16:45～17:00　第9会場（朱鷺メッセ　3F　306+307） 3O-09e1-2 The specialised synaptic development of layer 5 corticothalamic projections in posterior thalamic nucleus depends on regulated vesicular release. Shuichi Hayashi（林周一）¹，Anna Hoerder-Suabedissen（Hoerder-Suabedissen Anna）¹，Emi Kiyokage（清蔭恵美）²，Kazunori Toida（樋田一徳）³，Graham Knott（Knott Graham）⁴，Zoltan Molnar（Molnár Zoltán）¹ ¹Department of Physiology, Anatomy and Genetics, University of Oxford, Oxford, United Kingdom ²医療技術学部、川崎医療福祉大、岡山、日本 ³解剖学教室、川崎医科大、岡山、日本 ⁴Centre of Electron Microscopy, EPFL, Lausanne, Switzerland Cortical layer 5 provides strong feedforward inputs to higher-order (association) nuclei of the thalamus. The synaptic development of those layer 5 corticothalamic projections is not fully understood. We used the Rbp4-Cre; Rosa26-lsl-tdTomato mouse line to label a subset of cortical layer 5 projections and examined Rbp4-Cre-positive (Rbp4+) boutons in the posterior thalamic nucleus (Po) by pre- and post-immunoelectron microscopy against tdTomato during postnatal development. Rbp4+ boutons emerged in Po during the first postnatal week, and they formed synapses with the shaft of Po dendrites. Rbp4+ boutons became larger during the second and third postnatal weeks and formed multiple synapses with excrescences protruded from Po dendrites. The 3D reconstruction from serial block face electron microscopy (SBF-SEM, for methods see Maclachlan et al., 2018) of Rbp4+ boutons and connecting Po dendritic excrescences at P18 revealed their unique features: multiple excrescences from one Po dendrite connect with the same single bouton that completely encloses them. We also examined Rbp4+ boutons in SNAP25 conditional knockout (cKO) mice (Rbp4-cre; Rosa26-lsl-tdTomato; SNAP25-f/f), in which regulated vesicular release at synapses of Rbp4+ layer 5 neurons was removed (Hoerder-Suabedissen et al., 2018). SNAP25-cKO axons projected normally to thalamic nuclei, and there was no significant difference in the initial synapse formation between SNAP25-cKO and control boutons in Po at P8. However, the size of the SNAP25-cKO boutons were significantly smaller than that of control by P18. Although the SNAP-cKO boutons retained synapses with the shaft of Po dendrites, no dendritic excrescences protruded into the boutons and this caused a decrease in the total number of synapses on each SNAP25-cKO bouton. Our results suggest that regulated vesicular release at presynapses is required for the development of mature specialised connections between cortical layer 5 projections and thalamic dendrites in Po. References 1. Maclachlan C, Sahlender DA, Hayashi S, Molnar Z, Knott G. Block Face Scanning Electron Microscopy of Fluorescently Labeled Axons Without Using Near Infra-Red Branding. Front. Neuroanat. 2018;12:88. 2. Hoerder-Suabedissen A, Korrell KV, Hayashi S, Jeans A, Ramirez DMO, Grant E, et al. Cell-Specific Loss of SNAP25 from Cortical Projection Neurons Allows Normal Development but Causes Subsequent Neurodegeneration. Cereb. Cortex. 2018.

7月27日（土）17:00～17:15　第9会場（朱鷺メッセ　3F　306+307） 3O-09e1-3 発達期大脳皮質ニューロンにおけるBDNFのプロモーター活性を制御する発火活動パターン Yumi Miyasaka（宮阪優美），Nobuhiko Yamamoto（山本亘彦）大阪大院生命機能細胞分子神経生物学 During development, neuronal activity is known to modify cortical circuits. Activity-dependent gene expression plays a critical role in this process. Previous studies have demonstrated that brain-derived neurotrophic factor (BDNF) is expressed in cortical cells in an activity-dependent manner and contributes to axonal and dendritic development. However, it is unknown what patterned activity efficiently induces its expression in cortical cells. To address this issue, we studied BDNF promoter activity in individual cortical cells by applying various patterns of stimulation. For this, live imaging using luciferase assay was performed in combination with an optogenetic technique. A channelrodopsin-2 (ChR2) vector was transfected into the mouse cortex by in utero electroporation at embryonic day 15, when upper layer neurons are born. After giving a birth, slice cultures were prepared from the cortical region containing ChR2-positive cells. To visualize BDNF promoter activity, a luciferase (luc) vector under control of BDNF exon4 promoter was transfected sparsely into upper layer neurons around ChR2-expressing axons. After 2 weeks in culture when cortical circuits are formed with synaptic connections, the luc signals in individual cells were observed by an EMCCD camera every 30 minutes for up to 24 hr before and after photostimulation with various frequency (0.1~10 Hz). As a result, 2 and 10 Hz stimulation significantly increased the luc signal. In particular, 2 Hz stimulation produced more rapid elevation of the luc activity. In both cases, the signals began to increase just after the stimulation, peaked around 6 hr thereafter, and then gradually decreased. The luc signals were elevated in a fraction of cells by 5 Hz stimulation, but the increase was not significantly different from the control. The infrequent stimulation (0.1, 1Hz) did not change the signals. Moreover, the increase in the luc activity by high frequency stimulation was substantially suppressed in the presence of glutamate receptor blockers. Thus, the present result suggests that BDNF expression in individual cortical cells is up-regulated efficiently by presynaptic firing with preferential frequency and may contribute to neuronal circuit remodeling in the developing cortex.		7月27日（土）17:15～17:30　第9会場（朱鷺メッセ　3F　306+307） 3O-09e1-4 インテグリンα5β1を介したヒトiPS細胞由来ドパミン神経前駆細胞移植におけるシナプス形成の促進 Kaneyasu Nishimura（西村周泰）^1,2，Jun Takahashi（高橋淳）² ¹京都薬大統合薬科 ²京都大iPS細胞研 Cell transplantation therapies have great promise for treatment against neurodegenerative disorders such as Parkinson's disease (PD). Accordingly, several groups have reported methods to induce midbrain dopaminergic (DA) neurons from human pluripotent stem cells, such as embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs), and a robust and efficient induction method of midbrain DA progenitors was established toward clinical application. To realize cell transplantation therapy for PD, the grafted neurons should be integrated into the host neuronal circuit in order to restore the lost neuronal function. Here, we show that integrin α5β1 is selectively expressed in striatal neurons that are innervated by midbrain DA neurons using wheat germ agglutinin-based trans-synaptic tracing. Additionally, we found that integrin α5β1 was activated by the administration of estradiol-2-benzoate (E2B) in striatal neurons of adult female rats. Then, we observed that the systemic administration of E2B into hemi-parkinsonian rat models facilitates the functional integration of grafted DA neurons derived from human induced pluripotent stem cells into the host striatal neuronal circuit via the activation of integrin α5β1. Finally, methamphetamine-induced abnormal rotation was recovered earlier in E2B-administrated rats than in rats that received other regimens. Our results suggest that the simultaneous administration of E2B with stem cell-derived DA progenitors can enhance the efficacy of cell transplantation therapy for PD.