1989 Fiscal Year Final Research Report Summary
Mechanisms of 'run-down' of calcium channels
| Project/Area Number |
63570044
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| Research Category |
Grant-in-Aid for General Scientific Research (C)
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| Allocation Type | Single-year Grants |
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| Research Field |
General physiology
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| Research Institution | Okazaki National Research Institutes, National Inst. for Physiol. Sci. |
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Principal Investigator |
KAMEYAMA Masaki Natl. Inst. Physiol. Sci., Res. Associate, 生理学研究所, 助手 (60150059)
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| Project Period (FY) |
1988 – 1989
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| Keywords | calcium channel / run-down / cardiac myocyte / patch clamp |
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| Research Abstract |
A cytoplasmic protein that activates L-type Ca^<2+> channel was investigated by using both electrophysiological and biochemical methods. First, we attempted to establish a purification procedure using lipid vesicles into which bovine cardiac Ca^<2+> channels were reconstituted. The Ca^<2+> channel activating protein (CCAP) was eluted from a gel-filtration column as a mass having an apparent molecular weight (M'_r) of 200-300 x 10^3. On DEAE-sepharose chromatography, CCAP was eluted at 100-150 mM KCl. The partially Purified CCAP had M'_r of about 100 x 10^3 on SDS-polyacrylamid gel electrophoresis. These biochemical properties were clearly different from the cAMP- dependent protein kinase or any subunit of the Ca^<2+> channel. Second, CCAP could restore the activity of Ca^<2+> channel in inside-out patches of cardiac myocytes. Neither number of channels nor single channel conductance was affected by CCAP, but open-state probability of the channel was dramatically increased by CCAP. Third, we investigated tissue distribution of CCAP. A supernatant fraction of the homogenate from brain, heart, skeletal muscle, liver or kidney was applied to cardiac Ca^<2+> channels which were previously led to 'run-down' in inside-out patches. The tissue extracts from brain, heart, skeletal muscle and liver, but not kidney, recovered the channels from the run-down.
These results suggest that the activity of the L-type Ca^<2+> channel is maintained by the cyto- plasmic protein CCAP. It would be important. in future studies to characterize this protein further and to clarify possible regulatory mechanisms in which CCAP is involved.
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