Gonadotropin-specific primary antisera and gold-conjugated secondary antibodies were used to immunocytochemically localize gonadotropins in the anterior pituitary of intact pony mares. Electron microscopy was then used to characterize the ultrastructure and immunoreactive staining characteristics of equine gonadotropes. Cells containing LH were morphologically indistinguishable from those containing FSH. Gonadotropes were relatively large and commonly had eccentric nuclei. The rough endoplasmic reticulum was well developed and dilated. Secretory granules were present in two morphologically distinct forms. Large polymorphic granules were generally located in perinuclear cytoplasmic areas, whereas small and uniformly shaped granules were in the peripheral cytoplasm, close to the cell membrane. Double-labeling revealed cells with granules that stained for both LH and FSH as well as cells that stained for either LH or FSH. Gonadotropes constituted 15 to 32% of all pituitary cells in the anterior pituitaries from the three mares included in this study. Cells that stained for only LH constituted 2 to 16% of all pituitary cells, cells that stained for only FSH ranged from 1 to 4.5%, and cells staining for both hormones constituted 6.2 to 24% of the pituitary cells. These results indicate that there are in fact three distinct subclasses of gonadotropes in the equine anterior pituitary based on immunocytochemical staining, which is similar to the situation described for several other mammalian species.