Protein kinase C and calcium regulation of adenylyl cyclase in isolated rat pancreatic islets

Rat islets express several isoforms of adenylyl cyclase (AC), and the regul ation of AC activity in isolated islets by Ca2+ and protein kinase C (PKC) was investigated. At basal 2.8 mmol/l glucose, the muscarinic receptor agon ist carbamylcholine chloride (CCh) evoked a concentration-dependent increas e in cAMP generation with a maximum increase at least 4.5-fold above contro l. In contrast, forskolin and glucagon-like peptide 1 fragment 7-36 amide i ncreased cAMP accumulation 23-fold and almost 10-fold, respectively. Cholec ystokinin 26-33 sulfated amide (CCK) also stimulated cAMP production by up to eightfold, as did the phorbol ester, phorbol 12,13-dibutyrate (PDBu). PD Bu and CCh or CCK responses were not additive. The effects of phorbol ester , CCh, and CCK were inhibited by as much as 75% by the PKC inhibitors GF 10 9203X and Ro-32-0432 and after PKC downregulation. In the absence of extrac ellular Ca2+, PDBu-, CCh-, and CCK-induced cAMP production was inhibited by similar to 50% in each case. Chelation of intracellular Ca2+ with 1,2-bis( o-amino-5-fluorophenoxy) ethane-N,N,N',N'-tetraacetic acid tetraacetoxymeth yl ester (BAPTA/AM) inhibited CCh- and CCK-stimulated cAMP generation by si milar to 50% but did not inhibit the stimulatory effect of PDBu. Stringent Ca2+ depletion by removal of extracellular Ca2+ and inclusion of BAPTA/ AM allowed for increased cAMP production in response to CCh and CCK; PKC inhib itors and PKC downregulation prevented this stimulation. Glucose stimulatio n also increased islet cAMP production, but PDBu did not potentiate the glu cose response. The results suggest that Ca2+ influx, Ca2+ mobilization, and PKC activation play important roles in the modulation of AC activity in pa ncreatic islets.