We have investigated the role of cAMP in nitric oxide (NO)- and prostanoid-independent vascular relaxations evoked by acetylcholine (ACh) in isolated arteries and perfused ear preparations from the rabbit. These EDHF-type responses are shown to be associated with elevated cAMP levels specifically in smooth muscle and are attenuated by blocking adenylyl cyclase or protein kinase A (PKA). Relaxations are amplified by 3-isobutyl-1-methylxanthine, which prevents cAMP hydrolysis, while remaining susceptible to inhibition by the combination of two K-Ca channel blockers, apamin and charybdotoxin. Analogous endothelium- and cAMP-dependent relaxations were evoked by cyclopiazonic acid (CPA) which stimulates Ca2+ influx via channels linked to the depletion of Ca2+ stores. Responses to ACh and CPA were both inhibited by interrupting cell-to-cell coupling via gap junctions with 18 alpha -glycyrrhetinic acid and a connexin-specific Gap 27 peptide. The findings suggest that EDHF-type responses are initiated by capacitative Ca2+ influx into the endothelium and propagated by direct intercellular communication to effect relaxation via cAMP/PKA-dependent phosphorylation events in smooth muscle.