The effects of FSH on the testes of immature hypophysectomized rats were investigated by comparing functional changes in Leydig cells with changes in their number and morphological appearance. Rats were treated twice daily for 7 days with 0.5 ml saline vehicle, 10 micrograms rat FSH, or 20 ng ovine LH (an equivalent amount of LH known to contaminate the FSH preparation). FSH treatment caused a significant increase in testis weight and stimulated more advanced spermatogenic activity compared to saline or LH treatment. Morphometric analysis of glutaraldehyde perfusion-fixed testes showed no significant increase in Leydig cell number after LH treatment [saline, 4.63 +/- 0.14 million cells; LH, 6.38 +/- 0.55 million mean +/- SE)], but a significant (P less than 0.001) increase after FSH treatment (11.55 +/- 0.79 million). FSH, but not LH or saline, treatment resulted in Leydig cell hypertrophy and ultrastructural features identical to those of adult Leydig cells, these changes being reflected by enhanced hCG- and LHRH agonist-stimulated testosterone production in vitro by whole testes or dispersed interstitial cells. FSH and LH treatment caused minor but significant decreases in LH receptor numbers on dispersed interstitial cells compared to saline treatment. LHRH receptor numbers on interstitial cells were significantly increased only by FSH treatment. It is suggested that since FSH acts only on the seminiferous epithelium, then the hypertrophy, hyperplasia, and functional enhancement of Leydig cells after FSH treatment may be mediated by the secretion of one or more factors from the seminiferous tubules, providing additional evidence to support the view that gonadotropic regulation of testicular function is modulated by local interactions between the seminiferous tubules and the interstitial tissue.