In previous experiments it was found that PGF2 alpha originating from the uterine endometrium plays a physiologically important role in corpus luteum regression. The present study was undertaken to investigate the regulation of PGF2 alpha release from the uterus in luteolysis. The effect of ovariectomy on PGF2 alpha release was examined. Bilateral ovariectomy was performed on day 14 of pseudopregnancy. PGF2 alpha urinary metabolite excretion was almost constant from day 15 to 22, although a significant increase of PGF2 alpha UM excretion could be seen in the control rabbits. Ovariectomy was performed on day 17. Levels of PGF2 alpha UM excretion increased from day 15 to 17, however, an abrupt decrease occurred after ovariectomy. This indicates that the ovaries are necessary for PGF2 alpha release from the uterus. The effect of steroids on PGF2 alpha release was examined. No significant increase in PGF2 alpha UM excretion could be seen in the ovariectomized pseudopregnant rabbits by the administration of estradiol and/or progesterone, although they are reported to be involved in other species. The effect of the ligation of the vessels between the uterus and ovaries on PGF2 alpha release was investigated. The ligation was performed on day 9. No increase in the production of PGF2 alpha occurred after ligation. Luteal life span was prolonged by 6 days. Endometrial perifusion was carried out to investigate PGF2 alpha production activity on day 18 of the pseudo-pregnant rabbits after the ligation of the vessels between the uterus and ovaries. PGF2 alpha production activity in the treated rabbits was lower than in the controls. These results indicate that a non-steroidal regulator may regulate PGF2 alpha release via the circulation of the uterus and ovaries in the pseudopregnant rabbit.