Plasma and pituitary PRL levels are significantly greater in adult female than in male rats. This difference is thought to be brought about by ovarian steroids. We found a similar sex difference in pituitary PRL content in normal mice from 30 days of age. Ovariectomy reduced pituitary PRL content and this reduction was prevented by sc implants of estradiol-17 beta (E2). Implants of E2 also increased the pituitary PRL content in normal male mice to a level approaching that in normal females. The hypogonadal (hpg) mice did not show a sex difference in pituitary PRL content. Implantation of E2 into mutants of both sexes raised the pituitary content of PRL to the level in normal females. Treatment of intact but not ovariectomized hpg females with two injections/day of 6 micrograms FSH (NIH-FSH-S15) produced uterine growth and an increase in pituitary PRL content. Estrogen implants significantly increased plasma PRL concentrations in ovariectomized normal female and normal male mice but not in adult hpg male or female animals. These results show that elevation of the plasma E2 concentration increased significantly the synthesis of PRL in both normal and hpg mice of both sexes, and in the normal, but not the hpg mice, also increased the plasma concentration of PRL.