The Ca2+-dependent, exocytotic release of catecholamine from adrenal medullary chromaffin cells is fundamental to the function of the adrenal medulla and has important cardiovascular and metabolic effects. It also serves as a model for exocytosis of catecholamine, other neurotransmitters and hormones from nerve terminals and cells. In bovine adrenal chromaffin cells nicotinic receptor-induced increase in cytosolic Ca2+ caused by Ca2+ influx is the physiological trigger for exocytosis. In this proposal three subjects concerning the metabolism and effects of Ca2+ that are possibly important in the control of exocytosis or other chromaffin cell functions will be investigated: 1) Ca2+ sequestration - Ca2+ entry induced by secretagogue raises cytosolic Ca2+ which in turn triggers exocytosis. Subsequent sequestration of Ca2+ probably contributes to the termination of secretion. Experimentally, secretagogue-induced 45Ca2+ uptake is rapidly sequestered. The site of short-term Ca2+ sequestration will be investigated. 2) Phosphatidylinositol metabolism and the release of inositol phosphates - Pilot experiments indicate the nicotinic agonists and elevated K+ which are secretagogues as well as muscarinic agonists which are not secretagogues, stimulate the release of inositol phosphates from chromaffin cells. The possibility will be investigated that secretagogue-induced increase in cytosolic Ca2+ induces the release of inositol phosphates, perhaps by activation of the Ca2+-dependent enzyme phospholipase C. The ability of inositol 1,4,5 trisphosphate to release Ca2+ from digitonin-permeabilized cells will also be investigated. 3) Ca2+-stimulated adenylate cyclase - The amount of cellular cyclic AMP is increased in a Ca2+-dependent manner by nicotinic agonist stimulation. It is unlikely that the increase is caused by direct nicotinic receptor stimulation of adenylate cyclase. Ca2+ and calmodulin stimulate adenylate cyclase in membranes from some tissues. The possibility that increases in cytosolic Ca2+ activates adenylate cyclase in situ in chromaffin cells will be investigated.
