In two different enriched populations of Leydig cells (called FI and FII) obtained from human testes (young patients: mean age 36 +/- 3 years, n = 6; aged men: mean age 73 +/- 2 years, n = 5), the dehydroepiandrosterone and testosterone in vitro outputs were increased in a dose- and time-related manners by hCG. Similar results were obtained when the Leydig cells were incubated in presence of either dbcAMP or 22R-hydroxycholesterol. In presence of either hCG or dbcAMP, the coefficient of stimulation (in terms of steroid outputs) was higher in FII when compared to FI. Conversely, the basal production of steroids was greater in FI than in FII, mainly for testosterone. The addition of increasing amounts of seminiferous tubule culture medium (STM) to the Leydig cell incubation medium led to a dose-related enhancement of the steroid production in both enriched-Leydig cell fractions under basal and hCG-stimulated conditions. Similar results were obtained in presence of increased seminiferous tubules length. Additional experiments realized with either concentrated STM or the coculture of seminiferous tubules with purified Leydig cells have confirmed the existence of a paracrine control of Leydig cell steroidogenesis by seminiferous secreted factor(s). A paracrine factor (or factors) from seminiferous tubular origin influences positively and with a high efficiency the Leydig cell function in humans, whatever the age.