Coumestrol (Cou) is a plant-derived phytoestrogen that induces various pathologies in the female reproductive tract. Although effects of phytoestrogens on reproductive function in other species are well documented, their influence on progesterone (P4) and prostaglandin (PG) secretion in the mare is unknown. The aim of this study was to determine if Cou directly affects P4and PG concentrations (in vivo) and endometrial PG secretion (in vitro) in the mare. In experiment 1, the mares (n = 4) were fed for 14 days on a diet containing increasing proportions of alfalfa pellets (250 g-1 kg/day). An additional 4 mares were fed a standard diet (control group). Sequential blood samples were obtained for 8 h after feeding on Days 13 and 14 (1 kg/day alfalfa pellets). Feeding the mares alfalfa pellets up-regulated PGE2and 13,14-dihydro-15-ketoprostaglandin F2alpha(PGFM) and down-regulated P4in the blood plasma compared to those in the control group (P< 0.05). In experiment 2, epithelial and stromal cells were exposed to E2(10-9M) or Cou (10-8M) for 24 h. In the in vitro study, Cou increased PG secretion in epithelial and stromal cells (P< 0.05). In both types of endometrial cells, Cou up-regulated PTGS-2 protein expression (P< 0.05). Moreover, PGES and PGFS proteins were up-regulated by Cou in epithelial cells (P< 0.01). These results indicate that Cou can disturb reproductive function by affecting reproductive hormone secretion and altering the endometrial milieu through PG stimulation. Coumestrol therefore may impair physiologic regulation of the estrous cycle and early pregnancy.