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Characterization and Messenger Ribonucleic Acid Distribution of a Cloned Pituitary Adenylate Cyclase-Activating Polypeptide Type I Receptor in the Frog Xenopus laevis Brain¹

Department of Psychiatry, Mental Retardation Research Center, University of California at Los Angeles, Los Angeles, California 90024-1759

Address all correspondence and requests for reprints to: James A. Waschek, 68–225 NPI, Department of Psychiatry, University of California at Los Angeles, 760 Westwood Plaza, Los Angeles, California 90024. E-mail: jwaschek{at}mednet.ucla.edu

Pituitary adenylate cyclase-activating polypeptide (PACAP) has been found to modulate neuroendocrine functions in the frog brain and pituitary, but the nucleotide sequence and brain distribution of messenger RNA (mRNA) for the selective type I receptor for PACAP (PAC₁) in the frog are still unknown. Here, we report the isolation and characterization of a PAC₁ receptor complementary DNA (cDNA) clone from a frog (Xenopus laevis) tadpole brain cDNA library. This cDNA encoded a 466-amino acid protein that has 74% homology with human PAC₁ receptor and 48% homology to the frog vasoactive intestinal peptide/PACAP receptor. Injection of in vitro synthesized mRNA of the cloned cDNA into Xenopus oocytes resulted in expression of selective high affinity PACAP receptors (K_d = 47 pM). IC₅₀ values for PACAP-38, PACAP-27, and VIP were 27 pM, 110 pM and >1 µM, respectively. These results indicated that the cloned cDNA represents a Xenopus PACAP-preferring PAC₁ receptor. Northern hybridization revealed that PAC₁ receptor mRNA was present at high levels in the brain. In situ hybridization showed that the PAC₁ receptor gene was expressed highly in the pallium, preoptic nucleus, and nucleus of cerebellum, and moderately in the Purkinje cell layer of the cerebellum. Moderate PAC₁ receptor mRNA signals were detected in the distal lobe of the pituitary. A knowledge of the molecular structure and expression pattern of the PAC₁ receptor will facilitate further investigation of the physiological roles of PAC₁ receptor in the frog brain.

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