| Project/Area Number |
62480119
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| Research Category |
Grant-in-Aid for General Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Research Field |
General pharmacology
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| Research Institution | Nagoya University School of Medicine |
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Principal Investigator |
HIDAKA Hiroyoshi Nagoya University School of Medicine, Professor, 医学部, 教授 (80100171)
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| Co-Investigator(Kenkyū-buntansha) |
NAKA Michiko Mie University School of Medicine, Staff, 医学部, 助手 (10093139)
TANAKA Toshio Mie University School of Medicine, Professor, 医学部, 教授 (00135443)
ISHIKAWA Naohisa Nagoya University School of Medicine, Assistant professor, 医学部, 助教授 (80109321)
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| Project Period (FY) |
1987 – 1988
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| Project Status |
Completed (Fiscal Year 1988)
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| Budget Amount *help |
¥6,000,000 (Direct Cost: ¥6,000,000)
Fiscal Year 1988: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1987: ¥4,000,000 (Direct Cost: ¥4,000,000)
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| Keywords | Intracellular Calcium Requlation / Myosin Light Chain Phosphorylation / Myosin Light Chain Kinase Vascular Smooth Muscle Contraction / Aggregation and Secretion Response of Platelets / Myosin Light Chain Kinase Inhibitor / ML-9 / ミオシン軽鎖キナーゼ阻害剤 / MLー9 / 分子薬理学 / ML-9 / カルシウム依存性リン酸化反応 / 平滑筋収縮反応 / 血小板放出反応 |
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| Research Abstract |
The purpose of this research is investigate the role of myosin light chain (MLC) phosphorylation in vascular contraction and platelet function with direct pharmacological manipulation of MLC-kinase, since we have developed a series of novel inhibitors of intracellular Ca^<2+> messenger system.
1. We found that a newly synthesized compound, 1-(5-chrolonaphthalene-1-sulfonyl)-1H-hexahydro-1,4-diazepine (ML-9) is a direct and selective inhibitor of MLC-kinase,with Ki value 3.8 M, and its inhibition was of the competitive type with respect to ATP Superprecipitation and Mg-ATPase activity of actomyosin from bovine aorta was inhibited by the addition of ML-9 in a dose-dependent manner. In chemically skinned smooth muscle cells of rabbit mesenteric artery, ML-9 inhibited the both Ca^<2+>- and Ca^<2+>-independent MLC-kinase-induced contraction. In the intact vascular strips, ML-9 suppressed the KCl-induced contraction concomitant with the inhibition 20-kDa MLC phosphorylation.
2. Monophosphorylated and diphosphorylated 20-kDa MLC were demonstrated in thrombin-stimulated human platelets by two different gel electrophoretic methods. The more rapid monophosphorylation was catalyzed by MLC-kinase while the slower and additional phosphorylation was catalyzed mainly by protein kinase C.
3. The rate of phosphorylation and daphosphorylation of myosin was closely related to the change of myosin conformation.
4. These results suggest the significance of Ca^<2+>, calmodulin-depenpent MLC phosphorylation in the regulation of vascular contractile activity and platelet function.
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