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

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.