A culture system designed to examine events associated with maternal recognition of pregnancy in the mare was developed using trophoblastic vesicles and monolayers of uterine epithelial cells. Equine uterine epithelial cells were harvested from endometrial biopsy tissue obtained from pregnant and nonpregnant mares and a homogenous population of luminal and glandular epithelial cells was isolated by mechanical/enzymatic dissociation. To create polarized monolayers of uterine epithelial cells, isolated cells were plated on extracellular matrix-coated culture inserts. Polarity of the confluent monolayer two weeks after plating was demonstrated by ultrastructural analysis showing apical microvilli, tight junctions and desmosomes in the apico-lateral membranes of the cultured cells. Trophoblastic vesicles were generated for use in co-culture with the uterine cells and for the development of monolayers of trophectodermal cells. Functional studies using a fluorescence (carboxyfluorescein) recovery after photobleaching assay (gap FRAP) evaluated the degree of gap junction-mediated intercellular communication between monolayers of uterine cells. Endometrial cells isolated from mares either during pregnancy or diestrus were capable of cell-cell communication. Gap-junction mediated inter-cellular communication was also detected between trophectodermal cells in culture. The determination of the extent of cell-cell communication as well as the identification of specific signal pathways within endometrial cells can be monitored using the model in an effort to identify signalling events important to equine embryo recognition. The model may also have valuable potential as a research tool for the study of prostaglandin synthesis and protein synthesis during early pregnancy in the mare.