Structural determination of agonist binding site of ADPbS-sensitive P2
| Project/Area Number |
09670057
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| Research Category |
Grant-in-Aid for Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
General physiology
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| Research Institution | NIHON UNIVERSITY |
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Principal Investigator |
KOKUBUN Shinichiro Nihon Univ. Sch. Of Med., Professor, 医学部, 教授 (20153520)
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| Co-Investigator(Kenkyū-buntansha) |
TAKAO Kyoichi Nihon Univ. Sch. Of Med., Assistant, 医学部, 助手 (90187922)
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| Project Period (FY) |
1997 – 1998
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| Project Status |
Completed (Fiscal Year 1998)
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| Budget Amount *help |
¥3,000,000 (Direct Cost: ¥3,000,000)
Fiscal Year 1998: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1997: ¥1,500,000 (Direct Cost: ¥1,500,000)
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| Keywords | P2X1 / P2Y2 / P2Y4 / Urinary bladder / RT-PCR / ntracellular Calcium / ラット / 膀胱平滑筋細胞 / ヌクレオチド受容体 / クローニング / 細胞内Ca / Polymerase Chain Reaction |
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| Research Abstract |
We investigated subtypes of P2-purinoceptors in rat urinary bladder smooth muscle (UBSM) cells. In the experiments using isolated single UBSM cells, in the presence of extracellular 1.8 mM Ca2+ (10ィイD1-8ィエD1 M to 10ィイD1-4ィエD1 M) and P2X1 agonist, α,β-methylene adenosine 5'-triphosphate (α,β-Me ATP) (10ィイD1-9ィエD1 M to 10ィイD1-4ィエD1 M), increased intracellular CaィイD12+ィエD1 ([CaィイD12+ィエD1]ィイD2iィエD2) monitored with 1-[2-(5-carboxyoxazol-2-yl)-6-aminobenz-furan-5-oxy]-2-(a-amino-5-methylphenoxy)-ethane-N,N,N,N-tetraacetic acid (fura-2). In the absence of extracellular CaィイD12+ィエD1, α,β-Me ATP (10ィイD1-9ィエD1 M to 10ィイD1-4ィエD1 M) didnot increase [CaィイD12+ィエD1]ィイD2iィエD2, whereas ATP (10ィイD1-8ィエD1 M to 10ィイD1-4ィエD1 M) increased it. The potency order of P2Y-purinoceptor agonists in the increase in [CaィイD12+ィエD1]ィイD2iィエD2 in the absence of extracellular CaィイD12+ィエD1 was ATP>UTP>ADP>UDP. Reverse transcription polymerase chain reaction (RT-PCR) showed P2X1, P2Y2, P2Y4 and P2Y1 transcripts in these cells, and the relative quantity of mRNAs determined by ABI PRISM 7700 was P2X1>>P2Y2>>P2Y4>>>>P2Y1. These results suggest that the major purinoceptors expressed in rat UBSM cells are P2X1 and P2Y2.
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