The control of cytosolic Ca2+ concentration: studies of high affinity Ca2+ transport in permeabilized acini of rat submandibular glands

Active Ca2+ transport was studied in acini that had been permeabilized by incubation in buffer nominally free of Ca2+ in order to avoid any contribution to measured Ca2+ uptake by elements of the plasma membrane. In most experiments, ruthenium red was used to inhibit mitochondrial Ca2+ uptake. Non-mitochondrial Ca2+ transport was greatest at pH 7.0-7.5 and required Mg2+-ATP at concentrations typical of the cytosol (K0.5 = 1.36 +/- 0.53 mM). Other substrates were much less effective than ATP. Ca2+ uptake was stimulated by Ca2+ at concentrations near those measured in intact cells (K0.5 = 0.43 +/- 0.17 microM). Hill coefficients at subsaturating concentrations of Ca2+ (1.08 +/- 0.27) and Mg2+-ATP (0.81 +/- 0.22) indicated that cooperative interactions are not characteristic of the major cationic regulators of Ca2+ transporting activity. Na+ did not release Ca2+ from acinar mitochondria, but consistently reduced non-mitochondrial Ca2+ uptake by about 20% as compared to uptake in the presence of an equimolar K+ concentration. The properties of non-mitochondrial Ca2+ uptake in permeabilized acini are similar to those of high affinity Ca2+ uptake which, in broken cell preparations, has been found distributed in parallel with elements of the endoplasmic reticulum. The Ca2+-sequestering properties of a non-mitochondrial organelle in permeabilized submandibular gland acini are those expected of a principal regulator of cytosolic Ca2+ and could account for the ionized Ca2+ concentration measured in resting salivary acinar cells.