We have characterized opioid binding sites in the Sertoli cells of adult and 18-day-old rat testes. Maximal specific etorphine binding was attained after 30 min at 4 C. The binding was reversible, with association and dissociation rate constants of 0.98 X 10(5) M-1 min-1 and 0.33 min-1, respectively. Scatchard analyses and saturation curves revealed a single class of high-affinity, low-capacity binding sites. No opioid binding was observed in Leydig cell cultures. Exposure to opioids for 3 days caused a significant increase in [3H]etorphine specifically bound to the Sertoli cells that was completely prevented by naloxone, demonstrating opioid up-regulation of its own receptor. Chronic opioid treatment of the cultures significantly inhibited androgen-binding protein production, and this effect was prevented by naloxone. Since the circulating concentrations of endorphins (10(-12) M) are lower than the Kd of testis opiate receptors, it is conceivable that opioids of Leydig cell origin act on the specific high-affinity receptors of the Sertoli cells, and may play a role in modulating their function.