Recently, the rat ovary was shown to contain significant levels of a GnRH-like protein, but no detectable GnRH. In the present studies, extracts of bovine ovaries and ovine corpora lutea were examined for their content of both GnRH-like protein and GnRH. The GnRH-like proteins were detected with a rat ovarian membrane radioreceptor assay, and GnRH was detected with a specific GnRH RIA. The biological activity of the GnRH-like protein was evaluated with a rat luteal cell assay. GnRH-like activity, but not GnRH, was clearly present in extracts of the entire bovine ovary, bovine corpus luteum, bovine granulosa cells, and ovine corpus luteum. The highest levels of GnRH-like activity were present in granulosa cells. Neither GnRH-like activity nor GnRH was detected in extracts of bovine follicular fluid or bovine jugular plasma. Fractionation of the bovine and ovine GnRH-like proteins by reverse phase HPLC resulted in retention times similar to that of the rat ovarian GnRH-like protein, but distinctly different from that of authentic GnRH. The bovine and ovine GnRH-like fractions, like those in the rat, were sensitive to protease and heat. The bovine GnRH-like protein, obtained by preparative reverse phase HPLC, evoked a dose-dependent inhibition of LH-stimulated cAMP accumulation in rat luteal cells similar to that caused by GnRH. Based on these results, we suggest that the bovine ovary, granulosa cells, and corpus luteum and the ovine corpus luteum contain an antigonadotropic GnRH-like protein similar to the GnRH-like protein of the rat ovary. The presence of a similar GnRH-like protein (but the absence of GnRH) in ovaries of domestic species and the rat raises the possibility that this substance may play a paracrine antigonadotropic role in the ovary of diverse species.