DOSE-DEPENDENT EFFECTS OF ANGIOTENSIN-II, ACETYLCHOLINE AND VASOPRESSIN ON THE CYTOSOLIC CONCENTRATION OF CA2+ IN SUSPENSION PRIMARY CULTURES OF ZONA-FASCICULATA RETICULARIS CELLS FROM BOVINE ADRENAL-CORTEX

The effects of angiotensin II (AII), acetylcholine and vasopressin on the intracellular concentration of Ca2+ have been little studied in adrenocortical cells from the zona fasciculata/reticularis (ZFR). Primary cultures of bovine ZFR cells maintained in suspension culture for 72 h produce cortisol in response to AII (0.1-mu-M), acetylcholine (0.1 mM) and vasopressin (1-mu-M). This response is accompanied by a breakdown of membrane phosphoinositides from [H-3]inositol-prelabelled cells. Using cells loaded with the Ca2+ indicator fura-2, the intracellular concentration of Ca2+ was measured in response to increasing doses of all three agonists and found to increase in a graded fashion in each case. The basal intracellular concentration of Ca2+ was 75 +/- 3 nM (mean +/- S.E.M., n = 52), rising to a maximum 1.82 +/- 0.14-fold (n = 6) for AII (0.1-mu-M), 1.35 +/- 0.05-fold (n = 7) for acetylcholine (0.1 mM) and 1.27 +/- 0.10-fold (n = 6) for vasopressin (1-mu-M). In the case of AII and acetylcholine, agonists were added sequentially in medium of normal extracellular Ca2+ concentration (1.2 mM) or in medium in which the Ca2+ concentration was buffered to approximate to the intracellular concentration of Ca2+ (75-100 nM). Evidence was thereby obtained that both AII and acetylcholine mobilize a common intracellular pool of Ca2+. Our findings suggest that these three agonists, all of which stimulate phospholipase C, increase intracellular Ca2+ through a mechanism which depends, at least in part, on the release of Ca2+ from a common intracellular pool.