CHARACTERIZATION OF PACAP RECEPTORS AND SIGNALING PATHWAYS IN RABBIT GASTRIC MUSCLE-CELLS

Pituitary adenylate cyclase-activating peptide (PACAP) receptors and t heir signaling pathways were characterized in dispersed rabbit gastric muscle cells. I-125-PACAP-27 and I-I25-vasoactive intestinal peptide (VIP) binding to muscle cells were inhibited equally by PACAP and VIP (mean inhibitory concentration 0.8 to 1.3 nM) and desensitized to the same extent (70-80%) by exposure to either peptide. PACAP, like VIP, i ncreased cytosolic free Ca2+ and the formation of L-[H-3]citrulline, N O3-/NO2-, guanosine 3',5'-cyclic monophosphate (cGMP), and adenosine 3 ',5'-cyclic monophosphate (cAMP) and induced relaxation (mean effectiv e concentration 1.8 +/- 0.1 nM) that was partly inhibited by N-G-nitro -L-arginine (L-NNA), VIP-(10-28), and PACAP 6-38. L-[H-3]citrulline an d cGMP formation were blocked by nifedipine, L-NNA, and pertussis toxi n (PTx), implying activation of a G protein-coupled, Ca2+-calmodulin-d ependent nitric oxide (NO) synthase. PACAP-induced relaxation was inhi bited to the same extent (46-49%) by nifedipine, L-NNA, PTx, and the p rotein kinase G inhibitor KT-5823; the inhibition reflected the compon ent of relaxation mediated by the NO-cGMP pathway. The residual relaxa tion was abolished by the protein kinase A inhibitor 11-89. The patter n of inhibition of all responses was identical to that observed with V IP. Desensitization with VIP or PACAP abolished cAMP formation but had no effect on L-[H-3]citrulline and cGMP formation induced by either p eptide. Receptor protection with VIP or PACAP preserved fully all resp onses (L-[H-3]citrulline, cGMP, and cAMP formation and relaxation) to either peptide. The complete cross-competition, cross-desensitization, cross-antagonism, and cross-protection of receptors by either VIP or PACAP are consistent with interaction of both peptides with the same r eceptors; the receptors consist of two classes, each coupled to a dist inct signaling pathway.