Store-operated influx of Ca(2+) in pancreatic beta-cells exhibits graded dependence on the filling of the endoplasmic reticulum.
The store-operated pathway for Ca(2+) entry was studied in individual mouse pancreatic beta-cells by measuring the cytoplasmic concentrations of Ca(2+) ([Ca(2+)](i)) and Mn(2+) ([Mn(2+)](i)) with the fluorescent indicator fura-2. Influx through the store-operated pathway was initially shut off by pre-exposure to 20 mM glucose, which maximally stimulates intracellular Ca(2+) sequestration. To avoid interference with voltage-dependent Ca(2+) entry the cells were hyperpolarized with diazoxide and the channel blocker methoxyverapamil was present. Activation of the store-operated pathway in response to Ca(2+) depletion of the endoplasmic reticulum was estimated from the sustained elevation of [Ca(2+)](i) or from the rate of increase in [Mn(2+)](i) due to influx of these extracellular ions. Increasing concentrations of the inositol 1,4,5-trisphosphate-generating agonist carbachol or the sarco(endo)plasmatic reticulum Ca(2+)-ATPase inhibitor cyclopiazonic acid (CPA) cause gradual activation of the store-operated pathway. In addition, the carbachol- and CPA-induced influx of Mn(2+) depended on store filling in a graded manner. The store-operated influx of Ca(2+)/Mn(2+) was inhibited by Gd(3+) and 2-aminoethoxydiphenyl borate but neither of these agents discriminated between store-operated and voltage-dependent entry. The finely tuned regulation of the store-operated mechanisms in the beta-cell has direct implications for the control of membrane potential and insulin secretion.
Dyachok
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O
Department of Medical Cell Biology, Uppsala University, Biomedicum, Box 571, SE-751 23 Uppsala, Sweden.
Gylfe
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D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels / Q000378:metabolism; D002217:Carbachol / Q000494:pharmacology; D002478:Cells, Cultured; D004721:Endoplasmic Reticulum / Q000187:drug effects / Q000378:metabolism*; D005453:Fluorescence; D005682:Gadolinium / Q000494:pharmacology; D005947:Glucose / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007328:Insulin / Q000378:metabolism; D000078790:Insulin Secretion; D017136:Ion Transport / Q000187:drug effects; D007515:Islets of Langerhans / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D008345:Manganese / Q000378:metabolism; D051379:Mice; D018160:Receptors, Cytoplasmic and Nuclear / Q000037:antagonists & inhibitors / Q000378:metabolism
D015220:Calcium Channels; D007211:Indoles; D053496:Inositol 1,4,5-Trisphosphate Receptors; D007328:Insulin; D018160:Receptors, Cytoplasmic and Nuclear; D008345:Manganese; D002217:Carbachol; D005682:Gadolinium; D005947:Glucose; D002118:Calcium; C000543:cyclopiazonic acid