Characterisation of Distinct Inositol 1,4,5‐Trisphosphate‐Sensitive and Caffeine‐Sensitive Calcium Stores in Digitonin‐Permeabilised Adrenal Chromaffin Cells

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Abstract
The effect of inositol 1,4,5‐trisphosphate [Ins‐(1,4,5)P3] and caffeine on Ca2+ release from digitonin‐per‐meabilised bovine adrenal chromaffin cells was examined by using the Ca2+ indicator fura‐2 to monitor [Ca2+]. Permea‐bilised 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(l,4,5)P3 had no effect on the magnitude of the response to a subsequent addition of caffeine. The response to Ins(l,4,5)P3 was prevented by prior addition of caffeine or CaCl2, indicating that the Ins(l,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(l,4,5)P3‐ or caffeine‐sensitive stores. Ca2+ release from the caffeine‐sensitive store was selectively blocked by ryano‐dine. The Ins(l,4,5)P3‐sensitive store was emptied by thap‐sigargin, 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.