The purpose of this study was to examine the possible mechanism through which RU486 induces luteolysis during the late-luteal phase in pseudopregnant (PSP) rats. PSP rats received a subcutaneous injection of RU486 in sesame oil (5 mg/kg body weight) or sesame oil alone once a day between day 9 and day 11 of pseudopregnancy. Serial blood samples were collected on days 5, 9, 10, 11 and 12 and assayed for progesterone content. To examine the possible action of RU486 through a uterine and/or a pituitary (prolactin-dependent) mechanism, PSP rats and chronic hysterectomized PSP rats which had been hysterectomized before PSP induction received a subcutaneous injection of RU486 in sesame oil (5 mg/kg body weight), sesame oil alone, prolactin in 50% polyvinylpyrrolidone (15 IU/day), or RU486 and prolactin once a day between day 9 and day 11 of pseudopregnancy. Serial blood samples were collected on days 5, 9, 10 and 11 and assayed for progesterone content. Blood samples were also collected at 0400 h on day 12 and used for prolactin and progesterone determinations. To examine the direct effect of RU486 on corpus luteum and/or pituitary, hysterectomized rats underwent hypophysectomy and pituitary autotransplantation on dioestrus 1 and received a subcutaneous injection of RU486 in sesame oil or sesame oil alone for 3 days between day 21 and day 23 after surgery. Serial blood samples were collected on days 10, 21, 22, 23 and 24 and assayed for progesterone and prolactin contents. In ordinary PSP rats, serum progesterone levels were significantly (P < 0.01) lower in the RU486-treated group than in the control group (9 +/- 1 vs 53 +/- 7 ng/ml; mean +/- S.E.M.) on day 11. Serum prolactin levels at 0400 h on day 12 of pseudopregnancy were significantly (P < 0.05) lower in the RU486-treated group than in the control group (16 +/- 4 vs 154 +/- 44 ng/ml; mean +/- S.E.M.). The concomitant prolactin treatment reversed the luteolytic effects of RU486 on day 11 of pseudopregnancy. In hysterectomized PSP rats, RU486 also suppressed serum prolactin levels, and the concomitant prolactin treatment again reversed the luteolytic effects of RU486. In hysterectomized rats which were hypophysectomized and pituitary autotransplanted, RU486 treatment did not induce any significant changes in serum progesterone and prolactin levels. These results indicated that RU486 induced luteolysis during the late-luteal phase in PSP rats by suppressing prolactin secretion via a hypothalamic mechanism.