| Project/Area Number |
63065003
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| Research Category |
Grant-in-Aid for Specially Promoted Research
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| Allocation Type | Single-year Grants |
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| Research Institution | Kyoto University |
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Principal Investigator |
NUMA Shosaku Kyoto University Professor Faculty of Medicine, 医学部, 教授 (50025516)
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| Co-Investigator(Kenkyū-buntansha) |
FUJITA Yoshihiko Kyoto University Faculty of Medicine Assistant, 医学部, 助手 (80192730)
FUKUDA Kazuhiko Kyoto University Faculty of Medicine Assistant, 医学部, 助手 (90199224)
TAKESHIMA Hiroshi Kyoto University Faculty of Medicine Assistant, 医学部, 助手 (70212024)
TANABE Tsutomu Kyoto University Faculty of Medicine Assistant, 医学部, 助手 (70183069)
IMOTO Keiji Kyoto University Faculty of Medicine Associate Professor, 医学部, 助教授 (00176512)
久保 泰 京都大学, 医学部, 助手 (10178030)
野田 昌晴 京都大学, 医学部, 助教授 (60172798)
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| Project Period (FY) |
1988 – 1991
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| Project Status |
Completed (Fiscal Year 1991)
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| Budget Amount *help |
¥316,000,000 (Direct Cost: ¥316,000,000)
Fiscal Year 1991: ¥72,000,000 (Direct Cost: ¥72,000,000)
Fiscal Year 1990: ¥72,000,000 (Direct Cost: ¥72,000,000)
Fiscal Year 1989: ¥72,000,000 (Direct Cost: ¥72,000,000)
Fiscal Year 1988: ¥100,000,000 (Direct Cost: ¥100,000,000)
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| Keywords | Nicotinic acetylcholine receptor / Na^+ channel / Ca^<2+> channel / Ca^<2+> release channel / Muscarinic acetylcholine receptor / cDNA expression / Site-directed mutagenesis / Excitation-contraction coupling / 部位特異的変異 / グルタミン酸受容体 / Ca^<2+> releaseチャンネル / cGMP-gatedチャンネル / K^+チャンネル / 電位感受機構 / パッチクランプ法 / E-C coupling / ムスカリン性アセチルコリン受容体サブタイプ |
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| Research Abstract |
(A) By analyzing site-specifically mutated nicotinic acetylcholine receptor, we identified the regions which form the inner wall of the channel, the region which forms a channel constriction and and the region which determines ion selectivity. (B) By functional analysis of mutated Na^+ channels, we identified structural parts involved in voltage-sensitive activation or inactivation. Furthermore, we identified the region which is speculated to form the external mouth or the inner wall of the channel. (C) The primary structure of the cardiac dihydropyridine (DHP) receptor was determined by cDNA cloning and expression of the cDNA yielded a L-type Ca^<2+> channel. We elucidated the primary structure of a brain Ca^<2+> channel by cDNA cloning and showed that this channel is a high-threshold channel, insensitive to nifedipine and omega-conotoxin, and that it is predominantly expressed in cerebellum. We provided evidence that the skeletal muscle DHP receptor functions not only as an indispens
… More
able component in excitation-contraction (E-C) coupling but also as a slow Ca^<2+> channel and that it has a close relation to intramembrane charge movement. The region in the DHP receptor which determines the type of E-C coupling (skeletal muscle type or cardiac type) and the region which determines activation kinetics were identified. The primary structures of the skeletal muscle and the cardiac ryanodine receptor were determined by cDNA cloning and functional Ca^<2+> release channels were expressed from the cDNAs. (D) We showed that mAChR subtypes selectively couple to different effector systems. By functional analysis of chimaeric mAChRs, we identified the region involved in selective effector coupling and the region involved in antagonist binding. cDNA cloning and expression of two K^+ channels from NG108-15 neuroblastoma-glioma hybrid cells and a cGMP-gated channel from retina photoreceptor cells. (E) We expressed ionic channel proteins in CHO cells or cultured insect cells. Using purified proteins, we investigated conditions for crystallization of the proteins. Less
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