AJP - Heart and Circulatory Physiology, Vol 261, Issue 6 1927-H1936, Copyright © 1991 by American Physiological Society

ARTICLES

Vasopressin (V1) receptor characteristics in rat aortic smooth muscle cells

V. Gopalakrishnan, Y. J. Xu, P. V. Sulakhe, C. R. Triggle and J. R. McNeill
Department of Pharmacology, College of Medicine, University of Saskatchewan, Saskatoon, Canada.

We report saturable, high-affinity, specific, reversible binding sites for both [3H]arginine vasopressin ([3H]AVP) and d(CH2)5Tyr(Me)-[3H]AVP, a V1-selective antagonist, in cultured smooth muscle cells obtained from rat aorta (RA) and rat mesenteric artery (RMA). Specific binding of [3H]AVP had the following characteristics in adherent monolayers of RA and RMA smooth muscle cells: dissociation constant (KD) = 1.42 and 1.23 nM and maximal binding capacity (Bmax) = 9,500 and 29,910 sites/cell, respectively. Lower KD and higher Bmax values were detected for 3H-labeled V1 antagonist binding to both types of cells. Complete dissociation of [3H]-AVP binding to RA cells occurred in a biphasic manner with two rate constants of dissociation, suggesting high- and low-affinity states of the binding site for the agonist interaction. Partial dissociation of the antagonist-specific binding occurred, and it was monophasic, suggesting interaction of the 3H-labeled V1 antagonist radioligand to the high-affinity binding state. Inhibition constant (Ki) values determined by competitive inhibition of [3H]AVP binding to RA cells by a series of AVP-related peptide analogues and antagonists were consistent with the saturation data. AVP in a concentration-dependent manner induced the accumulation of inositol phosphates [mean effective concentration (EC50) 1 nM] in the adherent RA cells. The free cytosolic Ca2+ level in the dispersed RA smooth muscle cells was increased by AVP (EC50 8.1 nM). Pretreatment with the V1 antagonist abolished these AVP-evoked responses. The data support the conclusion that the agonist binding occurs at a homogeneous population of V1-subtype receptors in the high-affinity (KD = approximately 1 nM) state and that these receptors are functionally coupled to phospholipase C.

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