神経化学教育口演

神経化学教育口演 Neural Development・Neural Differentiation

2G1-01 The role of blood flow in neuronal turnover in the adult olfactory bulbs Ogino Takashi¹，Sawada Masato¹，Inada Hiroyuki²，Nabekura Junichi²，Sawamoto Kazunobu¹ ¹Department of Developmental and Regenerative Biology, Nagoya City University Graduate School of Medical Sciences，²Division of Homeostatic Development, Department of Developmental Physiology, National Institute for Physiological Sciences In the adult mouse olfactory bulbs（OBs）, new neurons generated from neural stem cells in the ventricular-subventricular zone are continuously supplied whereas old ones are eliminated, suggesting that olfactory interneurons are replaced throughout life. We have previously reported that olfactory input promotes reiterated use of the same positions by new neurons in the adult OB. However, mechanisms underlying the spatiotemporal regulation of neuronal turnover remain unknown. Here we show the relationship between neuronal turnover and blood flow in the adult OB. By performing in vivo two-photon imaging, we found that neuronal addition and elimination occur in the vicinity of blood vessels, suggesting that blood vessels provide preferable microenvironment for efficient neuronal turnover. To test the possibility that neuronal addition and elimination are correlated with the blood flow, we labeled blood plasma with fluorescent dye and measured capillary blood flow by in vivo two-photon line-scan imaging. Interestingly, newly added neurons were observed more frequently in the vicinity of blood vessels with high blood flow as compared with those with low flow. These data suggest that blood flow promotes neuronal turnover in the perivascular regions of adult OB.		2G1-02 Drebrin knockout mice show olfactory dysfunction by impairment of adult neurogenesis and cell survival Kajita Yuki¹，Koganezawa Noriko¹，Kojima Nobuhiko¹，Sakimura Kenji²，Sakurai Takashi³，Shirao Tomoaki¹ ¹Department of Neurobiology ＆ Behavior, Gunma University Graduate School of Medicine，²Department of Cellular Neurobiology, Brain Research Institute, Niigata University，³Electronics-Inspired Interdisciplinary Research Institute, Toyohashi University of Technology Drebrin is an F-actin-binding protein which plays an important role in regulation of spine morphology. Drebrin consists of two major isoforms, drebrin A and E. We have previously examined the functional role of drebrin using drebrin A specific knockout mice（DAKO）. DAKO shows behavioral abnormality in context dependent fear conditioning test（Kojima et al., 2010）. To better understanding of the role of drebrin for brain function, we generated drebrin null-knockout mice（DXKO）. DXKO showed abnormal behavior in buried food test and three-chamber social interaction test which were olfactory bulb（OB）related behavior. In Golgi staining, we observed normal dendritic spines in the OB of DXKO, suggesting that the olfaction disorder was not caused by spine abnormality. We then immunohistochemically analyzed the number of dying cells and mature neurons in the OB. In the OB of DXKO, the number of cell death decreased compared to that of wild-type mice（WT）, whereas the number of mature neurons did not change. Since newly generated neurons migrate from subventricular zone（SVZ）to OB, we next examined the number of arriving newly generated neurons in the OB. One day after injection of BrdU, the number of newly generated neurons in the OB of DXKO was smaller compared to that of WT. At 1 week, however, there was no difference in the cell number, and the cell number was larger in DXKO at 7 weeks. These results suggest that the adult neurogenesis decreases whereas the neuronal survival in OB increases in DXKO. These abnormalities might cause olfaction disorder in DXKO.

2G1-03 Modeling and analysis of intercellular adhesion between cells from the developing cerebral cortex Matsunaga Yuki¹，Noda Mariko¹，Murakawa Hideki²，Miura Takashi³，Kubo Ken-ichiro¹，Nakajima Kazunori¹ ¹Department of Anatomy, Keio University School of Medicine，²Faculty of Mathematics, Kyushu University，³Department of Anatomy and Cell Biology, Kyushu University Graduate School of Medical Sciences The mammalian neocortex has a highly organized 6-layered structure of neurons. Cortical neurons are generated within the ventricular zone（VZ）or subventricular zone（SVZ）, and migrate along radial fibers toward the pial surface. Newly born excitatory neurons migrate radially into the cortical plate（CP）past the earlier-born neurons, resulting in the birth-date-dependent“inside-out”alignment of neurons in the CP. Although the Reelin-deficient mouse, reeler, has been studied for more than 60 years and Reelin is indispensable for the establishment of the“inside-out”neuronal layers, cellular and molecular functions of Reelin for layer formation are still largely unknown. Reaggregation culture is a tool for studying intercellular adhesion. In the previous study, several clusters of MAP2-positive neurons were abnormally observed in the reaggregates of the reeler cerebral cortical cells. This result suggests the possibility that intercellular adhesion is altered in the reeler cerebral cortex. In the present study, to uncover how Reelin controls the intercellular adhesion among cortical cells, we performed Reelin stimulation experiments using reaggregation culture of the cells from the reeler cerebral cortex. We transfected an expression vector for Reelin into part of the cortical cells in the reeler reaggregates. Overexpression of Reelin unexpectedly caused clustering of nestin-positive cells in the inner part of the reeler reaggregates. To understand the mechanism of this cell clustering, we made mathematical models of cell aggregation, and examined the factors important for recapitulating the cell clustering patterns in the presence or absence of Reelin. The reaggregation culture and the mathematical model of cell sorting suggest that transient increase in neuronal adhesion is required for nestin-positive cluster formation in the inner part of the reeler reaggregates. Transient but not persistent increase in cell-cell adhesion might be necessary for the highly organized layered structure of neurons in the mammalian neocortex.		2G1-04 Ergothioneine promotes neuronal differentiation via induction of neurotrophin 5 in cultured neural stem cells. Ishimoto Takahiro，Nakamichi Noritaka，Masuo Yusuke，Kato Yukio Fac. Pharm., Kanazawa Univ. Ergothioneine（ERGO）is a food-derived hydrophilic antioxidant, distributed to the brain, and taken up into neural stem cells（NSCs）via carnitine/organic transporter OCTN1/SLC22A4. OCTN1-mediated ERGO uptake in mouse NSCs promoted neuronal differentiation accompanied with induction of Math1, one of the basic helix-loop-helix（bHLH）transcription factors, via unidentified mechanisms different from antioxidant action（Ishimoto et al., PLOS ONE 9, e89434, 2014）. In the present study, we focused on neurotrophins（NTs）, which promote neuronal differentiation by induction of bHLH transcription factors, as one of the candidate mechanisms. Some NTs are known as important factors related with pathogenesis of neuropsychiatric disorders. Since NTs are not distributed to the brain across the blood-brain barrier unlike ERGO, induction of NTs by ERGO may be a novel therapy for neuropsychiatric disorders. Exposure of NSCs to ERGO at 500 μM for 9 days significantly increased mRNA expression of Math1 and neurotrophin 5（NT-5）, and tended to increase expression of brain-derived neurotrophic factor（BDNF）and neurotrophin 3（NT-3）. Short term exposure of NSCs to ERGO also increased mRNA expression of NT-5, BDNF and NT-3 depending on the exposure time of ERGO until 12 hours, followed by increase in expression of Math1 at 24 hours. NT-5, BDNF and NT-3 activate neurotrophic tyrosine kinase receptor type2（TrkB）. To clarify the intracellular signaling pathway related with induction of Math1 by ERGO exposure, NSCs were incubated with ERGO in either the presence or absence of the inhibitor of TrkB or its three downstream signaling pathways, PI3K/Akt, PLCγ and MAPK/ERK signaling, and expression of Math1 was examined. As a result, all inhibitors of TrkB, Akt, PLCγ or Erk suppress induction of Math1 by ERGO. These results suggest that ERGO may promote neuronal differentiation at least partially by activation of TrkB signaling via autocrine/paracrine action of NT-5. Further studies are required in order to clarify upstream mechanisms underlying induction of NT-5 by ERGO.