The localization of adenosine receptors in rat testicular cells was investigated by evaluating the binding of cyclohexyladenosine (CHA) to somatic and germ cells and by assessing the effect of purine nucleosides on the function of testicular cells. Most of the specific binding of CHA in the testicular homogenate was found in the seminiferous tubule fraction, with little specific binding present in the interstitial compartment. Specific binding was also detected in a total-cell suspension prepared by dispersion of the seminiferous tubules. However, the specific activity of the binding did not increase when such a suspension was fractionated to produce germ cells at different stages of spermatogenesis. Adenosine receptors were present in the crude particulate fraction prepared from testes of rats from 10 to 90 days of age. While the affinity of these receptors did not vary as the testis matured, the number of binding sites per testis increased markedly up to 60 days after birth. Conversely, when binding activity was expressed per mg of testicular protein, no major changes in the specific activity of the binding were detected in testes up to 90 days of age. Sertoli cell-enriched cultures possessed a large number of binding sites, with a receptor density of 722 +/- 147 fmoles bound/mg protein (mean +/- SE; n = 5) and an affinity constant (Kd) of 1-2 nM. Among the characteristics of this binding site were kinetics constants and specificity similar to those measured in total particulate fractions of the testis and in the brain. Specific binding of CHA was not detected in cultures of testicular peritubular cells.(ABSTRACT TRUNCATED AT 250 WORDS)