Characterisation of distinct inositol 1,4,5-trisphosphate-sensitive and caffeine-sensitive calcium stores in digitonin-permeabilised adrenal chromaffin cells.
The effect of inositol 1,4,5-trisphosphate [Ins-(1,4,5)P3] and caffeine on Ca2+ release from digitonin-permeabilised bovine adrenal chromaffin cells was examined by using the Ca2+ indicator fura-2 to monitor [Ca2+]. Permeabilised cells accumulated Ca2+ in the presence of ATP and addition of either Ins(1,4,5)P3 or caffeine released 17% or 40-50%, respectively, of the accumulated Ca2+, indicated by sustained rises in [Ca2+] in the cell suspension. Prior addition of Ins(1,4,5)P3 had no effect on the magnitude of the response to a subsequent addition of caffeine. The response to Ins(1,4,5)P3 was prevented by prior addition of caffeine or CaCl2, indicating that the Ins(1,4,5)P3 response was blocked by elevated [Ca2+]. The responses were essentially identical in the presence of the proton ionophore carbonyl cyanide m-chlorophenylhydrazone, indicating that the Ca2+ release was not from mitochondria or secretory granules and that a proton gradient was not required for Ca2+ accumulation into the Ins(1,4,5)P3- or caffeine-sensitive stores. Ca2+ release from the caffeine-sensitive store was selectively blocked by ryanodine. The Ins(1,4,5)P3-sensitive store was emptied by thapsigargin, which had no effect on caffeine responses. These data suggest that permeabilised chromaffin cells possess two distinct nonoverlapping Ca2+ stores sensitive to either Ins(1,4,5)P3 or caffeine and support previous conclusions that these stores possess different Ca2(+)-ATPases.
Robinson
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IM
MRC Secretory Control Research Group, Department of Physiology, University of Liverpool, England.
Burgoyne
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RD
D000311:Adrenal Glands / Q000166:cytology / Q000378:metabolism*; D000818:Animals; D002110:Caffeine / Q000494:pharmacology*; D002118:Calcium / Q000378:metabolism*; D000252:Calcium-Transporting ATPases / Q000037:antagonists & inhibitors; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone / Q000494:pharmacology; D002463:Cell Membrane Permeability / Q000187:drug effects; D002838:Chromaffin System / Q000166:cytology / Q000378:metabolism*; D004072:Digitonin / Q000494:pharmacology*; D016257:Fura-2; D015544:Inositol 1,4,5-Trisphosphate / Q000494:pharmacology*; D012433:Ryanodine / Q000494:pharmacology; D013729:Terpenes / Q000494:pharmacology; D019284:Thapsigargin
D013729:Terpenes; D012433:Ryanodine; D002110:Caffeine; D002258:Carbonyl Cyanide m-Chlorophenyl Hydrazone; D019284:Thapsigargin; D015544:Inositol 1,4,5-Trisphosphate; D000252:Calcium-Transporting ATPases; D004072:Digitonin; D002118:Calcium; D016257:Fura-2