KINETICS OF CALCIUM ION FLUXES ASSOCIATED WITH HISTAMINE RELEASE IN THE RAT MAST CELL

The involvement of Ca('2+) in secretion is not completely understood. It has been proposed that stimulation leads to an uptake of extracellular Ca('2+) into the cytosol, thus triggering Ca('2+)-sensitive, intracellular processes that lead to exocytosis. In the mast cell, one of the difficulties in testing this hypothesis has been the inability to measure with sufficient precision and accuracy the rapid Ca('2+) movements and histamine secretion that occur after stimulation. This study describes a new method of rapid sampling capable of distinguishing the sequences of histamine release, Ca('2+) uptake, and Ca('2+) efflux in the stimulated mast cell. Measurements of Ca('2+) efflux and histamine release during stimulation show that the two events occur simultaneously. Ca('2+) may therefore be released by the same mechanism that releases histamine from the cell. The bulk of the Ca('2+) uptake from the extracellular medium is shown to occur after histamine release in compound 48/80- and ionophore A23187-stimulated cells. This indicates Ca('2+) uptake is a late event in the secretion process and not the trigger for secretion. Cell-associated calcium pools must therefore provide the Ca('2+) that increases the cytosolic concentration of this ion. The involvement of these calcium pools was probed using the Ca('2+)-chelating agent, EGTA, and the Ca('2+)-binding protein, calmodulin. There are several EGTA-sensitive Ca('2+) pools in the cell, but the identity of these stores remains a mystery. A novel, Ca('2+)-dependent inhibition of histamine release by extracellular calmodulin has been identified, suggesting there are Ca('2+)-binding control sites at or near the outer surface of the plasma membrane. The results presented here are consistent with recent reports that stimulation of membrane-bound phosphatidylinositol metabolism may be the critical event in mast cell stimulation which triggers the release of endogenous stores of Ca('2+) into the cytosol.