| Co-Investigator(Kenkyū-buntansha) |
TAKAGAKI Yotaro Kitasato University, School of Medicine, Professor, 医学部, 教授 (50281324)
NAKAMURA Kazuo Kitasato University, School of Medicine, Associate Professor, 医学部, 助教授 (40189030)
AZUMA Sadahiro Kitasato University, School of Medicine, Assistant Professor, 医学部, 助手 (80348507)
KATAOKA Masakazu Shinshu University, Deapartment of Technology, Associate Professor, 学部, 助教授 (90332676)
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| Research Abstract |
Neurotrophins and various growth factors are expressed in brain and play important roles both in developing and in adult brains. Recent studies indicate that various trophic factors involve in the regulation of synaptic plasticity underlying learning and memory, however, the precise mechanisms are still remained obscure. In order to elucidate molecular mechanisms of trophic factor-mediated regulation of presynaptic functions, effects of epidermal growth factor (EGF) and insulin-like growth factor-1 (IGF-1) on neurotransmitter release were studied in clonal rat pheochromocytoma PC12 cells. A brief treatment of EGF and IGF-1 enhanced Ca^<2+> dependent dopamine (DA) release from PC12 cells in concentration-dependent manners. EGF activated both mitogen-activated protein kinase (MAPK) and phosphatidylinositol 3-kinase (PI3-kinase) pathways, and EGF-dependent enhancement of DA release was suppressed by a MAPK kinase inhibitor as well as by PI3-kinase inhibitors. By the striking contrast, IGF
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-1 activated PI3-kinase pathway but not MAPK pathway, and IGF-1-dependent enhancement was suppressed by PI3-kinase inhibitor but not by MAPK kinase inhibitor. EGFP-tagged PH domain of protein kinase B (PKB), which selectively bind to phosphatidylinositol 3,4-bisphosphate and phosphatidylinositol 3,4,5-triphosphate, expressed in PC12 cells was translocated to plasma membrane after treatments with either EGF or nerve growth factor (NGF), and the translocations were abolished by wortmannin, a potent inhibitor of PI3-kinase. By the contrast, no significant redistribution of EGFP-PKB-PH was induced by IGF-1. EGF as well as NGF induced reorganization of actin cytoskeleton especially in the tip of cell process, however, only slight change was induced by IGF-1. More than nine cellular proteins were phosphorylated by PKB, a downstream enzyme of PI3-kinase, after a treatment with either EGF or NGF, however different set of proteins were phosphorylated by IGF-1. These results indicate that PI3-kinase participates in the enhancement of neurotransmitter release by two distinct mechanisms. Less
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