ENHANCED CAMP PRODUCTION MEDIATES THE STIMULATORY ACTION OF PITUITARYADENYLATE-CYCLASE ACTIVATING POLYPEPTIDE (PACAP) ON IN-VITRO CATECHOLAMINE SECRETION FROM BOVINE ADRENAL CHROMAFFIN CELLS

The 38 amino acid peptide pituitary adenylate cyclase activating polyp eptide (PACAP) induced a dose dependent increase of catecholamine secr etion in cultures of bovine chromaffin cells. This secretagogue activi ty of PACAP was strictly dependent on the presence of calcium in the c ulture medium. If calcium was omitted from the medium no effect of PAC AP on catecholamine secretion could be detected during an incubation o f 20 min. Preincubation of cells with 1 nM PACAP for 5 min facilitated the subsequent nicotine stimulated catecholamine secretion during a 2 0 min incubation without addition of the peptide. PACAP induced catech olamine secretion was clearly accompanied by a dose dependent increase of intracellular cAMP concentrations. The percentage of cells respond ing to PACAP with increased catecholamine secretion was assessed by im munocytochemistry of the transient appearance of dopamine-beta-hydroxy lase, associated with the membranes of the chromaffin granules on the cell surface during the secretory process. About 70 % of adrenal medul lary cells responded to 100 nM PACAP with enhanced secretory activity. Though PACAP stimulated catecholamine secretion, we did not observe m ajor effects on intracellular free calcium concentrations ([Ca2+](i)) as determined with fura-2 by single cell fluorescence microscopy. In m aximally 20 % of the cells a rise in [Ca2+](i) in response to a challe nge with 500 nM PACAP was observed. Lower concentrations of PACAP were without an effect on [Ca2+](i). These data indicate that the stimulat ory action of PACAP on in vitro catecholamine secretion from bovine ch romaffin cells is linked to a rise of intracellular cAMP.