Effects of monovalent and divalent cations on Ca2+ fluxes across chromaffin secretory membrane vesicles.

Abstract

Bovine chromaffin secretory vesicle ghosts loaded with Na+ were found to take up Ca2+ when incubated in K+ media or in sucrose media containing micromolar concentrations of free Ca2+. Li+- or choline+-loaded ghosts did not take up Ca2+. The Ca2+ accumulated by Na+-loaded ghosts could be released by the Ca2+ ionophore A23187, but not by EGTA. Ca2+ uptake was inhibited by external Sr2+, Na+, Li+, or choline+. All the 45Ca2+ accumulated by Na+-dependent Ca2+ uptake could be released by external Na+, indicating that both Ca2+ influx and efflux occur in a Na+-dependent manner. Na+-dependent Ca2+ uptake and release were only slightly inhibited by Mg2+. In the presence of the Na+ ionophore Monensin the Ca2+ uptake by Na+-loaded ghosts was reduced. Ca2+ sequestered by the Na+-dependent mechanism could also be released by external Ca2+ or Sr2+ but not by Mg2+, indicating the presence of a Ca2+/Ca2+ exchange activity in secretory membrane vesicles. This Ca2+/Ca2+ exchange system is inhibited by Mg2+, but not by Sr2+. The Na+-dependent Ca2+ uptake system in the presence of Mg2+ is a saturable process with an apparent Km of 0.28 microM and a Vmax = 14.5 nmol X min-1 X mg protein-1. Ruthenium red inhibited neither the Na+/Ca2+ nor the Ca2+/Ca2+ exchange, even at high concentrations.