Endocrinology, Vol 127, 272-277, Copyright © 1990 by Endocrine Society

ARTICLES

Characterization and distribution of binding sites for the hypothalamic peptide, pituitary adenylate cyclase-activating polypeptide

PE Gottschall, I Tatsuno, A Miyata and A Arimura
U.S.-Japan Biomedical Research Laboratories, Tulane University, Hebert Center, Belle Chasse, Louisiana 70037.

A novel bioactive peptide was recently isolated from ovine hypothalamus and was named PACAP (pituitary adenylate cyclase-activating polypeptide). PACAP was present in two bioactive, amidated forms, PACAP27 and PACAP38 (27 and 38 amino acids, respectively), and showed a 68% sequence homology with vasoactive intestinal peptide (VIP) in the N- terminal 28 residues. PACAP38 was at least 1000 times more potent than VIP in stimulating adenylate cyclase in pituitary cells, but both peptides exhibited comparable vasodepressor activity. Thus, we sought to determine whether PACAP acts on specific binding sites in the anterior pituitary or other tissues and whether these binding sites are different from those of VIP. Binding of [125I] PACAP27 to freshly prepared rat anterior pituitary membranes in the presence and absence of 212 nM unlabeled PACAP27 was specific, saturable, and more rapid at 22 C than at 4 C. Scatchard analysis of this binding site using increasing doses of unlabeled PACAP27 revealed a single high affinity site with a Kd of 446 +/- 141 pM and a maximum number of sites of 1312 +/- 182 fmol/mg protein. These results do not exclude the possibility of a second pituitary binding site with significantly lower affinity. Unlabeled PACAP38 and PACAP38OH exhibited significantly higher affinity binding (3- to 5-fold) than PACAP27 with a similar number of pituitary sites. A variable distribution of binding sites was observed between PACAP27 and VIP when binding to different tissue membranes was measured with 125I-labeled peptides. Very high specific binding of both PACAP27 and VIP was observed in lung membranes. An almost identical relative magnitude of binding was observed between PACAP27 and VIP in lung, liver, duodenum, ovary, and thymus. However, whereas PACAP27 binding to hypothalamic and pituitary membranes was great, VIP binding to these tissues was almost absent. To determine if VIP and PACAP might share a binding site in peripheral tissues, displacement curves were generated using [125I]PACAP27 binding to lung membranes and VIP, PACAP27, and PACAP38 as unlabeled ligands. VIP was highly potent in displacing [125I] PACAP27 binding in lung membrane, and the IC50 values for all three of these peptides were between 1-10 nM. These results suggest that 1) a saturable, high affinity binding site for PACAP is present on anterior pituitary membranes; 2) PACAP27 and PACAP38, but not VIP, share this binding site in the anterior pituitary and possibly the hypothalamus; and 3) PACAP27, PACAP38, and VIP share a similar or identical binding site on lung membranes and possibly other peripheral tissues.

This article has been cited by other articles:

				S. Lariviere, G. Garrel-Lazayres, V. Simon, N. Shintani, A. Baba, R. Counis, and J. Cohen-Tannoudji Gonadotropin-Releasing Hormone Inhibits Pituitary Adenylyl Cyclase-Activating Polypeptide Coupling to 3',5'-Cyclic Adenosine-5'-Monophosphate Pathway in L{beta}T2 Gonadotrope Cells through Novel Protein Kinase C Isoforms and Phosphorylation of Pituitary Adenylyl Cyclase-Activating Polypeptide Type I Receptor Endocrinology, December 1, 2008; 149(12): 6389 - 6398. [Abstract] [Full Text] [PDF]

				K. H. Sze, H. Zhou, Y. Yang, M. He, Y. Jiang, and A. O. L. Wong Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) as a Growth Hormone (GH)-Releasing Factor in Grass Carp: II. Solution Structure of a Brain-Specific PACAP by Nuclear Magnetic Resonance Spectroscopy and Functional Studies on GH Release and Gene Expression Endocrinology, October 1, 2007; 148(10): 5042 - 5059. [Abstract] [Full Text] [PDF]

				M. Barberi, B. Muciaccia, M. B. Morelli, M. Stefanini, S. Cecconi, and R. Canipari Expression localisation and functional activity of pituitary adenylate cyclase-activating polypeptide, vasoactive intestinal polypeptide and their receptors in mouse ovary Reproduction, August 1, 2007; 134(2): 281 - 292. [Abstract] [Full Text] [PDF]

				M. Ushiyama, R. Ikeda, H. Sugawara, M. Yoshida, K. Mori, K. Kangawa, K. Inoue, K. Yamada, and A. Miyata Differential Intracellular Signaling through PAC1 Isoforms as a Result of Alternative Splicing in the First Extracellular Domain and the Third Intracellular Loop Mol. Pharmacol., July 1, 2007; 72(1): 103 - 111. [Abstract] [Full Text] [PDF]

				K. Sayasith, K. A Brown, and J. Sirois Gonadotropin-dependent regulation of bovine pituitary adenylate cyclase-activating polypeptide in ovarian follicles prior to ovulation Reproduction, February 1, 2007; 133(2): 441 - 453. [Abstract] [Full Text] [PDF]

				S. Vaccari, S. Latini, M. Barberi, A. Teti, M. Stefanini, and R. Canipari Characterization and expression of different pituitary adenylate cyclase-activating polypeptide/vasoactive intestinal polypeptide receptors in rat ovarian follicles. J. Endocrinol., October 1, 2006; 191(1): 287 - 299. [Abstract] [Full Text] [PDF]

				S.-H. Lee and C. L. Cox Excitatory Actions of Vasoactive Intestinal Peptide on Mouse Thalamocortical Neurons Are Mediated by VPAC2 Receptors J Neurophysiol, August 1, 2006; 96(2): 858 - 871. [Abstract] [Full Text] [PDF]

				J. P. Moore Jr, L. L. Burger, A. C. Dalkin, and S. J. Winters Pituitary Adenylate Cyclase Activating Polypeptide Messenger RNA in the Paraventricular Nucleus and Anterior Pituitary During the Rat Estrous Cycle Biol Reprod, September 1, 2005; 73(3): 491 - 499. [Abstract] [Full Text] [PDF]

				W. I. DeHaven and J. Cuevas VPAC Receptor Modulation of Neuroexcitability in Intracardiac Neurons: DEPENDENCE ON INTRACELLULAR CALCIUM MOBILIZATION AND SYNERGISTIC ENHANCEMENT BY PAC1 RECEPTOR ACTIVATION J. Biol. Chem., September 24, 2004; 279(39): 40609 - 40621. [Abstract] [Full Text] [PDF]

				M. Delgado, D. Pozo, and D. Ganea The Significance of Vasoactive Intestinal Peptide in Immunomodulation Pharmacol. Rev., June 1, 2004; 56(2): 249 - 290. [Abstract] [Full Text] [PDF]

				S. Cecconi, G. Rossi, M. Barberi, L. Scaldaferri, and R. Canipari Effect of Pituitary Adenylate Cyclase-Activating Polypeptide and Vasoactive Intestinal Polypeptide on Mouse Preantral Follicle Development in Vitro Endocrinology, April 1, 2004; 145(4): 2071 - 2079. [Abstract] [Full Text] [PDF]

				E. M. Apostolakis, R. Lanz, and B. W. O'Malley Pituitary Adenylate Cyclase-Activating Peptide: A Pivotal Modulator of Steroid-Induced Reproductive Behavior in Female Rodents Mol. Endocrinol., January 1, 2004; 18(1): 173 - 183. [Abstract] [Full Text] [PDF]

				D. Alexandre, H. Vaudry, L. Grumolato, V. Turquier, A. Fournier, S. Jegou, and Y. Anouar Novel Splice Variants of Type I Pituitary Adenylate Cyclase-Activating Polypeptide Receptor in Frog Exhibit Altered Adenylate Cyclase Stimulation and Differential Relative Abundance Endocrinology, July 1, 2002; 143(7): 2680 - 2692. [Abstract] [Full Text] [PDF]

				R. Apa, A. Lanzone, F. Miceli, S. Vaccari, E. Macchione, M. Stefanini, and R. Canipari Pituitary Adenylate Cyclase-Activating Polypeptide Modulates Plasminogen Activator Expression in Rat Granulosa Cell Biol Reprod, March 1, 2002; 66(3): 830 - 835. [Abstract] [Full Text] [PDF]

				M. Roberto, R. Scuri, and M. Brunelli Differential Effects of PACAP-38 on Synaptic Responses in Rat Hippocampal CA1 Region Learn. Mem., September 1, 2001; 8(5): 265 - 271. [Abstract] [Full Text] [PDF]

				N. M. Sherwood, S. L. Krueckl, and J. E. McRory The Origin and Function of the Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP)/Glucagon Superfamily Endocr. Rev., December 1, 2000; 21(6): 619 - 670. [Abstract] [Full Text]

				M. E. Freeman, B. Kanyicska, A. Lerant, and G. Nagy Prolactin: Structure, Function, and Regulation of Secretion Physiol Rev, October 1, 2000; 80(4): 1523 - 1631. [Abstract] [Full Text] [PDF]

				W. Savino and M. Dardenne Neuroendocrine Control of Thymus Physiology Endocr. Rev., August 1, 2000; 21(4): 412 - 443. [Abstract] [Full Text] [PDF]

				S. Gräs, C. Hedetoft, S. H. Pedersen, and J. Fahrenkrug Pituitary Adenylate Cyclase-Activating Peptide Stimulates Acute Progesterone Production in Rat Granulosa/Lutein Cells via Two Receptor Subtypes Biol Reprod, July 1, 2000; 63(1): 206 - 212. [Abstract] [Full Text]

				D. Vaudry, B. J. Gonzalez, M. Basille, L. Yon, A. Fournier, and H. Vaudry Pituitary Adenylate Cyclase-Activating Polypeptide and Its Receptors: From Structure to Functions Pharmacol. Rev., June 1, 2000; 52(2): 269 - 324. [Abstract] [Full Text] [PDF]

				M. L. Scaldaferri, A. Modesti, C. Palumbo, S. Ulisse, A. Fabbri, E. Piccione, G. Frajese, and C. Moretti Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) and PACAP-Receptor Type 1 Expression in Rat and Human Placenta Endocrinology, March 1, 2000; 141(3): 1158 - 1167. [Abstract] [Full Text] [PDF]

				H.-J. Park, J. Lee, L. Wang, J.-H. Park, H.-B. Kwon, A. Arimura, and S.-Y. Chun Stage-Specific Expression of Pituitary Adenylate Cyclase-Activating Polypeptide Type I Receptor Messenger Ribonucleic Acid During Ovarian Follicle Development in the Rat Endocrinology, February 1, 2000; 141(2): 702 - 709. [Abstract] [Full Text] [PDF]

				J. J. Evans Modulation of Gonadotropin Levels by Peptides Acting at the Anterior Pituitary Gland Endocr. Rev., February 1, 1999; 20(1): 46 - 67. [Abstract] [Full Text]

				J. Lee, H.-J. Park, H.-S. Choi, H.-B. Kwon, A. Arimura, B.-J. Lee, W.-S. Choi, and S.-Y. Chun Gonadotropin Stimulation of Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) Messenger Ribonucleic Acid in the Rat Ovary and the Role of PACAP as a Follicle Survival Factor Endocrinology, February 1, 1999; 140(2): 818 - 826. [Abstract] [Full Text]

				U. Pagotto, T. Arzberger, E. Ciani, F. Lezoualc’h, C. Pilon, L. Journot, D. Spengler, and G. K. Stalla Inhibition of Zac1, a New Gene Differentially Expressed in the Anterior Pituitary, Increases Cell Proliferation Endocrinology, February 1, 1999; 140(2): 987 - 996. [Abstract] [Full Text]

				S. Lamouche, D. Martineau, and N. Yamaguchi Modulation of adrenal catecholamine release by PACAP in vivo Am J Physiol Regulatory Integrative Comp Physiol, January 1, 1999; 276(1): R162 - R170. [Abstract] [Full Text] [PDF]

				H. Oka, L. Jin, J. C. Reubi, X. Qian, B. W. Scheithauer, K. Fujii, T. Kameya, and R. V. Lloyd Pituitary Adenylate-Cyclase-Activating Polypeptide (PACAP) Binding Sites and PACAP/Vasoactive Intestinal Polypeptide Receptor Expression in Human Pituitary Adenomas Am. J. Pathol., December 1, 1998; 153(6): 1787 - 1796. [Abstract] [Full Text] [PDF]

				J. A. Waschek, R. A. Casillas, T. B. Nguyen, E. M. DiCicco-Bloom, E. M. Carpenter, and W. I. Rodriguez Neural tube expression of pituitary adenylate cyclase-activating peptide (PACAP) and receptor: Potential role in patterning and neurogenesis PNAS, August 4, 1998; 95(16): 9602 - 9607. [Abstract] [Full Text] [PDF]

				S. J. Winters, A. C. Dalkin, and T. Tsujii Evidence That Pituitary Adenylate Cyclase Activating Polypeptide Suppresses Follicle-Stimulating Hormone-{beta} Messenger Ribonucleic Acid Levels by Stimulating Follistatin Gene Transcription Endocrinology, October 1, 1997; 138(10): 4324 - 4329. [Abstract] [Full Text] [PDF]

				G. Geng, R. Gaspo, F. Trabelsi, and N. Yamaguchi Role of L-type Ca2+ channel in PACAP-induced adrenal catecholamine release in vivo Am J Physiol Regulatory Integrative Comp Physiol, October 1, 1997; 273(4): R1339 - R1345. [Abstract] [Full Text] [PDF]

				Y. Aoki, Y. Iwasaki, M. Katahira, Y. Oiso, and H. Saito Regulation of the Rat Proopiomelanocortin Gene Expression in AtT-20 cells. II: Effects of the Pituitary Adenylate Cyclase-Activating Polypeptide and Vasoactive Intestinal Polypeptide Endocrinology, May 1, 1997; 138(5): 1930 - 1934. [Abstract] [Full Text] [PDF]

				D. A. Przywara, X. Guo, M. L. Angelilli, T. D. Wakade, and A. R. Wakade A Non-cholinergic Transmitter, Pituitary Adenylate Cyclase-activating Polypeptide, Utilizes a Novel Mechanism to Evoke Catecholamine Secretion in Rat Adrenal Chromaffin Cells J. Biol. Chem., May 3, 1996; 271(18): 10545 - 10550. [Abstract] [Full Text] [PDF]