All androgen-sensitive peripheral tissues and cells, including the prostate, are commonly believed to possess the ability to metabolize testosterone. We report on the in vitro metabolism of tritiated testosterone performed in immortalized human adult normal prostatic epithelial cells and in human adult normal prostate fibroblastic cells (stromal cells). These two cell types were incubated separately with increasing testosterone concentrations (1 to 50 nM.) for 2 and 4 hours, after which the testosterone metabolic profile was analyzed. Data analysis provided evidence, for the first time, of the existence of two different 5 alpha-reduced metabolic pathways. Stromal cells preferred the androsterone pathway via the oxidative androstenedione formation, whereas epithelial cells preferred the reductive 5 alpha-dihydrotestosterone pathway. These two 5 alpha-reduced metabolites were produced in nearly equal quantities regardless of testosterone concentration or time of incubation. Since interactions between epithelial and stromal cells are involved in the development of the prostate, the availability of defined epithelial and stromal cells suitable for in vitro experiments provides a useful tool for the study of the contribution of androgens to these interactions. The model presented in this study would permit a better evaluation of the intraprostatic regulation of androgen metabolism and the contribution of the anti-5-alpha-reductase drugs to the management of benign prostate hyperplasia.