Pituitary adenylyl cyclase-activating peptide activates multiple intracellular signaling pathways to regulate ion channels in PC12 cells

Pituitary adenylyl cyclase-activating peptide (PACAP) stimulates calcium transients and catecholamine secretion in adrenal chromaffin and PC12 cells. The PACAP type 1 receptor in these cells couples to both adenylyl cyclase and phospolipase C pathways, but although phospolipase C has been implicated in the response to PACAP, the role of adenylyl cyclase is unclear. In this study, we show that PACAP38 stimulates Ca2+ influx in PC12 cells by activating a cation current that depends upon the dual activation of both the PLC and adenylyl cyclase signaling pathways but does not involve protein kinase C. In activating the current, PACAP38 has to overcome an inhibitory effect of Ras. Thus, in cells expressing a dominant negative form of Ras (PC12asn17-W7), PACAP38 induced larger, more rapidly activating currents. This effect of Ras could be overidden by intracellular guanosine-5'-O-3-(thio)triphosphate (GTP gamma S), suggesting that it was mediated by inhibition of down-stream G proteins. Ras may also inhibit the current through a G protein-independent mechanism, because cAMP analogues activated the current in PC12asn17-W7 cells, provided GTP gamma S was present, but not in PC12 cells expressing wild type Ras. We conclude that coupling of PACAP to both adenylyl cyclase and phospholipase C is required to activate Ca2+ influx in PC12 cells and that tonic inhibition by Ras delays and Limits the response.