Earlier studies reported from this laboratory demonstrated the luteolytic effect of a 5-day sequential regimen of 3 days of estrogen overlapped with 3 days of PGF2 alpha administered during the midluteal phase to cynomolgus monkeys. In this study, a different regimen, containing estrogen, lower doses of PGF2 alpha, and methylergonovine maleate (MEM), was used. Individual components of this modified sequential regimen were not effective as luteolytic agents. When 3 days of estrogen (40 micrograms of depoestradiol cypionate [DEC] on the first day and 40 micrograms of estradiol benzoate [EB] on second and third day) was followed by 3 days of MEM (400 micrograms, twice daily) overlapping 1 day of estrogen treatment, there was a significant shortening of the length of the menstrual cycle (less than 26 days) in eight of 10 animals, with a concomitant decline in plasma levels of progesterone. When a low dose of MEM (100 micrograms, twice daily), which was completely ineffective in shortening cycle lengths with the same estrogen treatment, was administered alternately with PGF2 alpha (5 mg twice daily, 11 of 14 animals had shortened cycles, with an associated decline in levels of progesterone. PGF2 alpha and MEM without the estrogen pretreatment did not have a significant effect on cycle lengths or blood levels of progesterone, thus demonstrating a synergism between estrogen and the other compounds in inducing luteolysis. Since the agents used in these sequential regimens have potent effects on the uterus, the treatments were repeated in hysterectomized monkeys to determine whether the uterus was involved in the mediation of luteolysis. These treatments caused a significant drop in plasma levels of progesterone by the tenth day after the preovulatory estrogen peak, thus demonstrating that the uterus is not essential to the luteolytic action.