Correlation of real-time catecholamine release and cytosolic Ca2+ at single bovine chromaffin cells

Abstract

Previous investigations of the role of Ca2+ in stimulus-secretion coupling have been undertaken in populations of adrenal chromaffin cells. In the present study, the simultaneous detection of intracellular Ca2+, with the fluorescent probe fura-2, and catecholamine release, using a carbon-fiber microelectrode, are examined at single chromaffin cells in culture. Results from classic depolarizing stimuli, high potassium (30-140 mM) and 1,1-dimethyl-4-phenylpiperazinium (3-50 μM), show a dependence of peak cytosolic Ca2+ concentration and catecholamine release on secretagogue concentration. Catecholamine release induced by transient high K+ stimulation increases logarithmically with K+ concentration. Continuous exposure to veratridine (50 μM) induces oscillations in intracellular Ca2+ and at higher concentrations (100 μM) concomitant fluctuation of cytosolic Ca2+ and catecholamine secretion. Mobilization of both caffeine- and inositol trisphosphate-sensitive intracellular Ca2+ stores is found to elicit secretion with or without extracellular Ca2+. Caffeine-sensitive intracellular Ca2+ stores can be depleted, refilled, and cause exocytosis in medium without Ca2+. Single cell measurement of exocytosis and the increase in cytosolic Ca2+ induced by bradykinin-activated intracellular stores reveal cell to cell variability in exocytotic responses which is masked in populations of cells. Taken together, these results show that exocytosis of catecholamines can be induced by an increase in cytosolic Ca2+ either as a result of transmembrane entry or by release of internal stores.