Previous studies have implicated arginine vasopressin (AVP) as a negative intratesticular modulator of androgen production. The present studies were undertaken to further define the testicular AVP system by investigating potential influences of pituitary hormones on testicular receptors for AVP. Binding of [3H]AVP to testicular membranes was time, temperature, and pH dependent, with optimum binding at 90 min, 22 C, and pH 8.0. Addition of divalent cations to the incubation mixture significantly increased binding by 4-, 10-, 15-, and 35-fold with Ca2+, Zn2+, Mg2+, and Mn2+, respectively. The influence of pituitary hormones on testicular AVP receptors was initially assessed by comparing binding parameters obtained by Scatchard analysis of saturation binding data between membranes prepared from intact or hypophysectomized rats. Testicular membranes from intact animals bound 66 +/- 2 fmol AVP/mg protein, whereas membranes from animals hypophysectomized 10 days previously showed a significantly lower binding capacity of 40 +/- 3 fmol AVP/mg protein; however, the affinity of the receptors for AVP did not differ between these two groups. A similar decrease was observed when binding was compared between purified Leydig cells isolated from intact and hypophysectomized animals by Metrizamide density gradients. Time-course analysis of this effect showed a continuous decline in total testicular AVP-binding capacity for the first 2 weeks after hypophysectomy and a slight increase at 3 weeks. Treatment of hypophysectomized animals with pituitary hormones beginning on the third day after hypophysectomy and continuing for either 4 or 7 days showed that LH and GH were able to dose-dependently restore testicular AVP-binding capacity to levels found in intact animals, whereas FSH and PRL were ineffective. These data indicate that testicular AVP receptors are under the control of LH and GH from the pituitary and suggest that pituitary hormones may impact on the efficacy of the testicular AVP system by regulating AVP receptor levels.