Linear V1-vascular vasopressin antagonists suitable for radioiodination, biotinylation, and fluorescent labeling

We modified several linear V1-vascular arginine vasopressin (AVP) antagonists to obtain compounds suitable for radioiodination, biotinylation, and fluorescent labeling. In binding competition experiments with human platelet V1-vascular AVP receptors, the linear V1 antagonist phenylacetyl-D-Tyr(Et)-Phe-Gln-Asn-Lys-Pro-Arg-NH2 (PhaaGln) displayed the greatest affinity [dissociation constant (Kd) = 0.05 +/- 0.01 nM]. The radioiodinated compound phenylacetyl-D-Tyr(Et)-Phe-Val-Asn-Lys-Pro-125I-labeled Tyr-NH2 (125I-labeled TyrPhaa) was characterized by a high affinity (Kd = 1.42 +/- 0.19 nM), a low nonspecific binding, and good stability. PhaaGln coupled to dodecabiotin retained a good affinity for V1-vascular AVP receptors (Kd = 1.41 +/- 0.20 nM). The complex PhaaGln-dodecabiotin-avidin is bifunctional, since an avidin-agarose column specifically bound V1-vascular AVP receptors labeled with 125I-TyrPhaa-dodecabiotin. In A7r5 vascular smooth muscle cells loaded with fura-2, PhaaGln and PhaaGln-dodecabiotin were pure antagonists as they blocked AVP-induced calcium mobilization but did not elicit a calcium signal by themselves. V1-vascular AVP receptors of A7r5 vascular smooth muscle cells were visualized by bound PhaaGln-dodecabiotin made fluorescent by labeling with fluorescein avidin. Thus linear V1-vascular AVP antagonists can be used as high affinity and specificity radioiodinated, biotinylated, and fluorescent probes to explore V1-vascular AVP receptors of human and animal origin.