A high incidence of vaginal adenosis-like lesions (ADL, 67%) was found in 30-day-old offspring of ICR/JCLmice given 4 daily subcutaneous injections of 2,000 µg diethylstilbestrol (DES) on days 15–18 of gestation. The prenatally DES-exposed mice and the controls ovariectomized at 10 days of age were given 5 daily subcutaneous injections of 10–4, 10–3, 10–2, 10–1, or 1 µg 17β-estradiol (E2), respectively, starting at 25 days of age. ADL was never observed in the age-matched controls given prepubertal injections of any E2 dose. In contrast, high incidences of ADL (63–100%) were found in the vaginal fornix and upper vagina of 30-day-old DES mice receiving prepubertal injections of 10–3–1 µg E2/day, whereas in DES mice given prepubertal injections of the vehicle alone and a lower daily dose of 10–4 µg E2, incidences were nil and 25%, respectively. Mitotic figures were found in the columnar epithelium with ADL, although the rate was lower than in the ADL-freed stratified epithelium. It is suggested, therefore, that an ovarian hormone (probably estrogen) participates in the prepubertal induction of ADL in DES mice.