Further characteristics of the ATP-stimulated uptake of calcium into chromaffin granules.

Abstract

The ATP-stimulated uptake of 45Ca2+ [and [3H](-)-noradrenaline ([3H]NA)] into chromaffin granules and that into mitochondria are driven by a protonic gradient delta mu H+, composed of the components delta pH (concentration gradient of protons) and delta psi (electrical potential difference). The granular ATPase pumps protons into the matrix (delta pH inside acid, delta psi positive), but the mitochondrial ATPase ejects protons from the matrix (delta pH alkaline, delta psi negative inside). To show different driving forces of uptake, the rate of the ATP-stimulated uptake of 45Ca2+ (and [3H]NA) into chromaffin granules was compared with the rate of the ATP-stimulated uptake of 45Ca2+ into mitochondria (adrenomedullary or rat liver). In the presence of nitrate, the rate of the ATP-stimulated uptake of 45Ca2+ into chromaffin granules is higher than in the presence of acetate, because the lyotropic anion nitrate stimulates the granular ATPase and increases delta pH (acid inside). Compared with nitrate, the rate of the ATP-stimulated uptake of 45Ca2+ into mitochondria is higher in the presence of the proton-carrying anion acetate, which, after permeation, provides protons for ejection by the ATPase. In the absence of ATP, a valinomycin-mediated potassium influx (delta psi inside positive) stimulates the granular uptake of [3H]NA, which has an electrogenic component, but not the granular uptake of 45Ca2+, which is electroneutral. The electrogenic uptake of 45Ca2+ into mitochondria is stimulated by a valinomycin-mediated potassium efflux (delta psi negative inside). The ATP-stimulated uptake of 45Ca2+ into chromaffin granules is sensitive to ruthenium red, suggesting a carrier-mediated mechanism of uptake, and it is sensitive to atractyloside, indicating the simultaneous uptake of ATP. After collapse of delta pH by ammonia, the ATP-stimulated uptake of 45Ca2+ into chromaffin granules is abolished, but not that into mitochondria. In the presence of ammonia, the rate of the ATP-stimulated uptake of [3H]NA is very low, and an ATP-independent uptake of 45Ca2+ into chromaffin granules is observed which is similar to the ATP-independent Ca2+/Na+ exchange at the granular membrane.