The natural history of benign prostatic hyperplasia (BPH) in the dog is characterized by a slow progression through two phases. The early phase of the disease process is characterized by glandular hyperplasia and occurs as early as 2.5 yr of age. The late phase of the disease is characterized by cystic hyperplasia and occurs after 4 yr. The results of the present study are the first to compare Leydig cell structure and steroidogenic function in dogs with BPH with those in age-matched controls. It was discovered that glandular BPH can occur in young dogs (2.5 yr) in which Leydig cell mass and ultrastructure and maximally stimulated androgen secretion are indistinguishable from those in age-matched controls. These results support the concept that the early phase of BPH (glandular hyperplasia) is not related temporally to some defect in the Leydig cell. In contrast, the late phase of BPH (cystic hyperplasia) in beagles 6 yr of age is associated with diminished smooth endoplasmic reticulum in the Leydig cell and with diminished production of androgens by perfused testes in vitro. During the course of these studies, we discovered that the testes of young beagles with BPH, but not age-matched controls or old beagles with BPH, secrete an unidentified molecule (putative estrogen). This molecule was characterized partially in that it is extractable from testicular venous effluent with diethyl ether, elutes in a discrete fraction in several different high performance liquid chromatographic systems, reacts with an antibody that recognizes 17 beta-estradiol, estrone, and estriol, and competes with 17 beta-[3H]estradiol for the rat uterine estrogen receptor. Based on the elution volume from high performance liquid chromatography, the unknown molecule (putative estrogen) is not estriol, estradiol, or estrone.