Nelivaptan was prepared as a solution in physiological saline containing 0.1% Tween80 or 5% DMSO and 5% Cremphor EL. Drugs were delivered in a constant volume of 2 mL/kg (rat) and containing 3, 10 or 20 mg/kg of Nelivaptan. Rats were evaluated for stress response in a Punished Drinking Test, in an elevated Plus-Maze and a forced swim test. Doses of 3 and 10 mg/kg significantly increased punished drinking. A dose of 10 and 30 mg/kg increased the boldness of rats in the elevated maze test. In the forced swim test, nelivaptan had a significant effect on the immobility time at doses of 10 mg/kg and up.

CHO-dhFr- cells were transfected with an expression vector derived from plasmid 7055 containing the cDNA encoding the human V1b receptor. Stabley transfected cells were grown in 10 mM HEPES, pH 7.4, minimal essential medium supplemented with 5% fetal calf serum and 8 g/L sodium bicarbonate and 300 micro-G.mL geneticin at 37 deg-C and under a 5% CO2 atmosphere. Binding assays on membranes of CHO cells transfected with the human or rat V1b receptor were performed in an incubation medium containing 50 mM Tris-HCl, pH 7.4; 3 mM MgSO4; 0.1% BSA; 0.1% bacitracin; [3H]AVP; and increasing amounts of Nelivaptan (0.9, 1.8, 3.7, 7.5, and 15 nM). Nonspecific binding was determined in the presence of 1 μM unlabeled AVP. Nelivaptan dose-dependently antagonized [3H]AVP binding to various membrane preparations from CHO cell lines transfected with the rat and human V1b receptors. Nelivaptan had an affinity for human V1b receptors close to that of the natural hormone, AVP (Ki values of 1.54 ± 0.82 and 0.80 ± 0.25 nM, respectively)