Pituitary adenylate cyclase-activating polypeptide (PACAP) is a member of the glucagon/secretin peptide family, and its molecular structure is highly conserved in vertebrates. In this study, the functional role of PACAP in regulating GH release in the goldfish was investigated. Using immunohistochemical staining, nerve fibers with PACAP immunoreactivity were identified in the vicinity of goldfish somatotrophs, suggesting that this neuropeptide may influence GH release in the goldfish. The direct regulatory action of PACAP on GH secretion was demonstrated in vitro in perifused goldfish pituitary cells. PACAPs (0.01 nM to 1 M) from different species, including ovine PACAP 27 , ovine PACAP 38 , frog PACAP 38 , zebra fish PACAP 27 , and zebra fish PACAP 38 , were all effective in stimulating GH release with ED 50 values of 8.9 Ϯ 3.5, 3.3 Ϯ 1.6, 14.4 Ϯ 3.5, 15.4 Ϯ 4.1, and 1.4 Ϯ 0.2 nM, respectively. Similar concentrations of vasoactive intestinal polypeptide (VIP), a peptide related to PACAP, was not effective in this respect. In addition, the GH-releasing action of ovine PACAP 38 (10 nM) was inhibited by the PACAP antagonist PACAP 6 -38 (10 M), but not by the VIP antagonist [4-Cl-D-Phe 6 ,Leu 17 ]VIP (10 M). The pharmacology of these GH responses is consistent with the mammalian type I PACAP receptors, suggesting that a similar receptor subtype is present in the goldfish pituitary and mediates the GH-releasing action of PACAP. To establish the structural identity of this goldfish PACAP receptor, a complementary DNA (cDNA) clone sharing a high degree of sequence homology with mammalian type I PACAP receptors was isolated from a goldfish pituitary cDNA library. This cDNA was 5.2 kb in size with a 1.4-kb open reading frame and encoded a 465-amino acid protein with the typical structure of a 7-transmembrane domain-containing, G protein-coupled receptor. Functional expression of this cDNA in COS-7 cells revealed that this fish type I PACAP receptor could be activated by ovine PACAP 27 and PACAP 38 to increase cAMP synthesis with ED 50 values of 2.4 Ϯ 0.8 and 4.2 Ϯ 1.2 nM, respectively. Other structurally related peptides, including VIP (100 nM), GH-releasing hormone (100 nM), glucagon (100 nM), secretin (100 nM), gastric inhibitory polypeptide (100 nM), and PTH (100 nM), were not effective in altering cAMP production. Using Northern blot and RT-PCR, messenger RNA transcripts of this PACAP receptor were identified in the brain, heart, and pituitary of the goldfish. These results, taken together, support the hypothesis that PACAP functions as a novel GH-releasing factor in the goldfish through activation of type I PACAP receptors. (Endocrinology 139: [3465][3466][3467][3468][3469][3470][3471][3472][3473][3474][3475][3476][3477][3478][3479] 1998) P ITUITARY adenylate cyclase-activating polypeptide (PACAP) is a member of the secretin/glucagon/vasoactive intestinal polypeptide (VIP) family (1). In mammals, two biologically active forms of PACAP have been purified, namely PACAP 38 (2) and PACAP 27 (3). PACAP 38 is ...