Characterization of VIP receptor-effector system antagonists in rat and mouse peritoneal macrophages.
In the present study we show that the synthetic peptides [4-Cl-D-Phe6,Leu17]VIP and the growth hormone releasing factor (GRF) analog [Ac-Tyr1,D-Phe2]GRF-(1-29)-NH2 inhibit in a competitive manner the specific [125I]VIP binding to both rat and mouse peritoneal macrophages. In rat peritoneal macrophages, the order of potency of the different peptides, as expressed by the IC50 values was: VIP (IC50 = 1.90 +/- 0.16 nM) > [4-Cl-D-Phe6,Leu17]VIP (IC50 = 125.8 +/- 13.2 nM) > [Ac-Tyr1,D-Phe2]GRF-(1-29)-NH2 (IC50 = 354.8 +/- 21.2 nM). In mouse peritoneal macrophages a similar pattern of potency was observed: VIP (IC50 = 1.58 +/- 0.12 nM) > [4-Cl-D-Phe6,Leu17]VIP (IC50 = 110.8 +/- 10.7 nM) > [Ac-Tyr1,D-Phe2]GRF-(1-29)-NH2 (IC50 = 251 +/- 19.2 nM). The behavior as VIP receptor antagonists of both [4-Cl-D-Phe6,Leu17]VIP and [Ac-Tyr1,D-Phe2]GRF-(1-29)-NH2 in rat and mouse peritoneal macrophages was confirmed by: (a) the shift to the right of VIP dose-stimulated cyclic AMP production curves in the presence of the two antagonists; (b) the agreement between the order of efficacy of the two peptides in competition experiments with the corresponding inhibition of cyclic AMP production; (c) the inefficiency of the two antagonists on the stimulation of cyclic AMP production by the beta-adrenoceptor agonist isoproterenol, which indicates the specificity of the interaction; (d) the synergic effect of VIP on isoproterenol-stimulated cyclic AMP production was completely abolished by [4-Cl-D-Phe6,Leu17]VIP or [Ac-Tyr1,D-Phe2]GRF-(1-29)-NH2, suggesting that both antagonists acted via specific VIP receptors. Moreover, propranolol, a beta-adrenoceptor antagonist, did not affect the VIP-stimulated cyclic AMP production and the antagonist role of [4-Cl-D-Phe6,Leu17]VIP or [Ac-Tyr1,D-Phe2]GRF-(1-29)-NH2; (e) in cross-linking experiments, the intensity of the labeling of the [125I]VIP/receptor complexes was significantly lower with the antagonists than in the control experimental situation in both mouse and rat peritoneal macrophage membranes.
Pozo
D
D
Department of Medical Biochemistry and Molecular Biology, University of Seville School of Medicine, Spain.
Montilla
M L
ML
Guerrero
J M
JM
Calvo
J R
JR
D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D000319:Adrenergic beta-Antagonists / Q000494:pharmacology; D000818:Animals; D001667:Binding, Competitive; D002462:Cell Membrane / Q000737:chemistry; D003432:Cross-Linking Reagents; D000242:Cyclic AMP / Q000096:biosynthesis; D066298:In Vitro Techniques; D007545:Isoproterenol / Q000494:pharmacology; D017737:Macrophages, Peritoneal / Q000187:drug effects* / Q000378:metabolism; D051379:Mice; D011433:Propranolol / Q000494:pharmacology; D051381:Rats; D018005:Receptors, Vasoactive Intestinal Peptide / Q000037:antagonists & inhibitors*; D017337:Sermorelin / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D014660:Vasoactive Intestinal Peptide / Q000031:analogs & derivatives / Q000737:chemistry / Q000378:metabolism / Q000494:pharmacology
D000318:Adrenergic beta-Agonists; D000319:Adrenergic beta-Antagonists; D003432:Cross-Linking Reagents; D018005:Receptors, Vasoactive Intestinal Peptide; C048507:vasoactive intestinal peptide, 4-chloro-Phe(6)-Leu(17)-; D014660:Vasoactive Intestinal Peptide; D017337:Sermorelin; C045616:somatotropin releasing hormone (1-29)-amide, N-Ac-Tyr(1)-Phe(2)-; D011433:Propranolol; D000242:Cyclic AMP; D007545:Isoproterenol