P ituitary adenylate cyclase activating polypeptide (PACAP) is present in gastric nerves, and PACAP receptors (PAC1) are found on gastric enterochromaffin-like (ECL) cells. Expression of PAC1 splice variants in purified ECL cells was determined by RT-PCR. PACAP effects on ECL cells were analyzed by video imaging of [Ca²⁺]_i and histamine release; its effects on gastric glands were examined by confocal microscopy of [Ca²⁺]_i in ECL and parietal cells. PACAP action on D cells was measured by [Ca²⁺]_i and radioimmunoassay. PACAP effects on acid secretion were determined in fistula rats with or without neutralizing anti-somatostatin antibodies. All splice variants of PAC1 were found, but vasoactive intestinal polypeptide (VIP) receptor (VPAC) products were absent. PACAP-27 and -38 dose-dependently raise [Ca²⁺]_i in ECL cells, and stimulated histamine release. VIP had a much lower affinity, which demonstrates the presence of PAC1 but not VPAC. PACAP elevated [Ca²⁺]_i in ECL and parietal cells of superfused gastric glands, but only the parietal cell signal was inhibited by ranitidine, showing the absence of PAC1 on parietal cells, and demonstrating functional coupling between the cell types. PACAP and VIP stimulated calcium signaling and somatostatin release from D cells with almost equal efficacy. Acid secretion was stimulated after intravenous injection of PACAP into rats treated with somatostatin antibody. PACAP is a candidate as a mediator of neural regulation of acid secretion.J. Clin. Invest. 104:1383–1391 (1999).