| Budget Amount *help |
¥6,600,000 (Direct Cost: ¥6,600,000)
Fiscal Year 1995: ¥900,000 (Direct Cost: ¥900,000)
Fiscal Year 1994: ¥5,700,000 (Direct Cost: ¥5,700,000)
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| Research Abstract |
We have monitored changes in membrane capacitance (DELTAC) and conductance (DELTAG) induced by muscarinic acetylcholine stimulation in single rat pancreatic acinar cells. Acetylcholine (ACh, 500nM) induced simultaneous increases of DELTAC and DELTAG.In contrast, a low concentration (50nM) of ACh exclusively induced DELTAC-increases without DELTAG.These reponses were abolished by the internal perfusion of heparin. It indicates that inositol 1,4,5-trisphosphate-mediated internal Ca^<2+>-mobilization either simultaneously activates exocytosis and ion channels or exclusively initiates exocytosis. In comparison, a low concentration of A23187 selectively activated ion channels but a high concentration exocytosis and ion channels simultaneously. These selective response patterns of DELTAC and DELTAG depend on the choice of agonist and the internal EGTA concentration. From this, we postulated two possible explanation for the selective action of muscarnic ACh stimulation on exocytosis. First, a
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n area of high [Ca^<2+>] i, spatially close to secretory granules, activates exocytosis. Second, an as yet unknown signalling factor sensitizes the Ca^<2+>-affinity of the exocytotic apparatus. Applying a phase-sensitive capacitance measurement to the ER preparation, the time course of the changes in membrane capacitance were monitored. At the local patch-membrane level, the changes occurred at random as discrete on-and off-steps, which can be interpreted as reversible vesicle formation (membrane transformation). The size distribution of the steps were fitted by a negative exponential curve, suggesting the membrane transformation energy seems to have a single energy peak. Increasing the lumenal Ca^<2+>-concentration at the whole-ER level, a rise in the membrane capacitance was found. This indicates that the vesicles are reversibly transformed to caps with a connection between the vesicle interior and the ER lumen. The result suggests the involvement of the ER lumenal Ca^<2+> in the vesicular transport. Less
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