Vesicular Ca2+ participates in the catalysis of exocytosis

Effects of vesicular monoamine transporter inhibitors on catecholamine rele ase from bovine chromaffin cells have been examined at the level of individ ual exocytotic events. As expected for a depletion of vesicular stores, rel ease evoked by depolarizing agents was decreased following 15-min incubatio ns with reserpine and tetrabenazine, as evidenced by a decrease in exocytot ic frequency and amount released per event. In contrast, two reserpine deri vatives, methyl reserpate and reserpic acid, were much less effective. Surp risingly, the incubations also decreased the accompanying rise in intracell ular Ca2+ evoked by depolarizing agents. Subcellular studies revealed that reserpine and tetrabenazine at concentrations near their Ri values not only could increase cytoplasmic catecholamines but also could displace Ca2+ fro m vesicles. Furthermore, transient exposure to tetrabenazine and reserpine, but not methyl reserpate and reserpic acid, induced exocytotic release of catecholamines, Reserpine induced a rise in intracellular Ca2+, as detected by whole-cell measurements with Fura-2, It could induce exocytosis, albeit at a lower frequency, in Ca2+-free solutions, supporting an internal Ca2source. Depletion of endoplasmic reticulum and mitochondrial Ca2+ pools did not eliminate the reserpine-activated release. These results indicate that vesicular Ca2+ can play an important role in exocytosis and under some con ditions may be involved in initiating this process.