The objective of this study was to investigate the role of androgens and nonandrogenic Leydig cell products in maintaining Leydig cell and macrophage numbers in the testis of the adult rat. Adult male Sprague-Dawley rats received Silastic implants containing testosterone and estradiol (T-E) in order to suppress endogenous luteinizing hormone (LH) for 9 weeks. After T-E treatment, Leydig cell and macrophage numbers, quantified using the optical disector approach, were reduced by 40 and 60%, respectively, compared with controls. Administration of human chorionic gonadotropin (hCG) for a period of 10 days restored Leydig cell numbers to control levels, and macrophage numbers were partially restored. Administration of the antiandrogen, flutamide, in combination with hCG treatment in T-E implanted animals prevented the restoration of Leydig cell numbers but did not prevent the recovery of macrophage numbers. In the T-E-implanted animals, there was a decrease in testicular macrophage nuclear size, which was not restored by either hCG or hCG plus flutamide treatment. The results of this study support the hypothesis that LH is the main pituitary regulator of both Leydig cell and macrophage number in the adult rat testis and further indicate that androgens are responsible for maintaining Leydig cell numbers and/or differentiation, but nonandrogenic Leydig cell factors are primarily responsible for controlling macrophage numbers. Testicular macrophage function, as indicated by nuclear size, does not appear to be influenced by LH or testosterone in the adult rat.