We investigated the effect of pituitary adenylate cyclase activating peptide (PACAP) on the colon-inferior mesenteric ganglion (IMG) reflex loop in vitro. PACAP27 and PACAP38 applied to the IMG caused a prolonged depolarization and intense generation of fast EPSPs and action potentials in IMG neurons. Activation of PACAP- preferring receptors (PAC1-Rs) with the selective agonist maxadilan or VIP/PACAP (VPAC) receptors with VIP produced similar effects whereas prior incubation of the IMG with selective PAC1-R antagonists PACAP6-38 and M65 inhibited the effects of PACAP. Colonic distension evoked a slow EPSP in IMG neurons that was reduced in amplitude by prolonged superfusion of the IMG with either PACAP27, maxidilan, PACAP6-38, M65 or VIP. Activation of IMG neurons by PACAP27 or maxadilan resulted in an inhibition of ongoing spontaneous colonic contractions. PACAP-LI was detected in nerve trunks attached to the IMG and in varicosities surrounding IMG neurons. Cell bodies with PACAP-LI were present in lumbar 2-3 dorsal root ganglia and in colonic myenteric ganglia. Colonic distension evoked release of PACAP peptides in the IMG as measured by radioimmunoassay. Volume reconstructed images showed that a majority of PACAP-LI, VIP-LI and VAChT-LI nerve endings making putative synaptic contact onto IMG neurons and a majority of putative receptor sites containing PAC1-R-LI and nAChR-LI on the neurons were distributed along secondary and tertiary dendrites. These results suggest involvement of a PACAP-ergic pathway, operated through PAC1-Rs, in controlling the colon-IMG reflex.