MECHANISM OF CYCLIC-AMP FACILITATION OF STIMULATION-EVOKED CATECHOLAMINE RELEASE IN ADRENAL CHROMAFFIN CELLS .1. EVIDENCE FOR ENHANCEMENT OF EVOKED INCREASE IN CYTOSOLIC FREE NA+ CONCENTRATION BY CYCLIC-AMP ELEVATION

The mechanism by which cyclic AMP facilitates the secretagogue-stimulated catecholamine (CA) release from cultured bovine adrenal chromaffin cells was investigated. Both 8-Br cyclic AMP and forskolin alone showed no detectable change in cytosolic free Ca2+ concentration ([Ca2+]i) but markedly potentiated acetylcholine (ACh)-, nicotine- or excess KCl-induced rise in [Ca2+]i. Verapamil, nicardipine and diltiazem antagonized ACh-evoked CA release. 8-Br cyclic AMP still enhanced CA release elicited by ACh even in the presence of Ca2+ channel blockers. Potentiation of stimulation-evoked CA release and Ca-45 uptake induced by cyclic AMP or forskolin was antagonized by a Na+ channel blocker, tetrodotoxin (TTX), or by the removal of Na+ from the medium, or by amiloride. Enhancement of secretagogue-stimulated CA release and of Ca-45 uptake induced by ouabain was also antagonized by TTX. Ouabain and veratrine mimicked the effect of cyclic AMP and these agents counteracted each other. Forskolin alone did not increase Na-22 uptake but enhanced Na-22 uptake induced by ACh. TTX markedly reduced Na-22 uptake induced by ACh with forskolin or by ACh with ouabain. The present study suggests that cyclic AMP induced potentiation of stimulation-evoked CA release from cultured bovine chromaffin cells primarily by enhancing the accumulation of intracellular free Na+. From the analogous results obtained using ouabain and veratrine, and from the counteraction of the effects of cyclic AMP by treatments which elevate the intracellular Na+, the possibility that cyclic AMP may inhibit Na+, K+-ATPase in plasma membrane and produce the accumulation of Na+, thereby increasing [Ca2+]i through the mechanism of Na+/Ca2+ exchange is discussed.