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Abstract

To identify and characterize V₁ vasopressin receptors, photoreactive antagonists of the glycogenolytic and vasoconstrictor activity of vasopressin have been synthesized. The following analogues with 3‐mercapto‐ 3,3‐cyclopentamethylene‐propionic acid (Mca) and N‐methylalanine (MeAla) in position 1 and 7 of vasopressin (VP) were effective V₁ antagonists: [Mca¹,D‐Tyr²,MeAla⁷,Lys⁸]VP (l), [Mca¹,MeAla⁷,Arg⁸,Lys⁹]VP (2), [Mca¹,MeAla⁷,Arg⁸,d‐Lys⁹]VP (3). Introduction of the photoreactive 4‐azidophenylamidino group into the side‐ chain of Lyss⁸ in analogue 1 or into Lys⁹ in analogues 2 and 3 increased the potency (for analogue 1 a tenfold increase in the antiglycogenolytic effect and a fivefold increase in the antivasopressor effect) and binding affinity for the rat hepatic V₁ receptor. Mono‐iodination at Tyr² with ¹²⁵I resulted in photoreactive antagonists of high specific radioactivity, which had roughly the same binding affinity as vasopressin for the rat hepatic V₁ receptor (K_d= 0.9—1.8 nM).

In photoaffinity labelling experiments with purified rat liver membranes, containing 2–3 pmol V₁ receptor/ mg protein, the analogues labelled specifically two proteins with the relative molecular masses (M_r) of 30000 and 38 000. These results and the results of a recent study using ³H‐labelled photoreactive vasopressin agonists [Boer, R. and Fahrenholz, F. (1985) J. Bid. Chem. 260, 15051 ‐ 150541 provide evidence that both vasopressin agonists and antagonists can interact with the same two subunits of the heterodimeric hepatic V₁ receptor. Furthermore the radioiodinated photoreactive V₁ antagonists should be helpful to identify V₁ receptor proteins in membranes of other cell types.