In vitro perifusion was employed to compare the potencies of mammalian, avian, salmon, and lamprey gonadotropin-releasing hormones (GnRHs) on the release of luteinizing hormone (LH) from the pituitaries of an amphibian (Rana pipiens) and a reptile (Chrysemys picta). The chicken-I and salmon GnRH variants were equipotent with mammalian GnRH in both the frog and the turtle glands. By contrast, the lamprey GnRH was inactive (less than 1% as potent as the others). Lamprey GnRH also failed to stimulate LH release or to induce GnRH priming when administered chronically to the frog gland. These results support the hypothesis that the GnRH receptors on nonmammalian pituitary cells are much less specific than those of the mammal with regard to the amino acid at position 8 of the GnRH molecule. These data suggest that the native GnRH variant or the one most like that found in the brain of a species is not necessarily the most potent biologically in that species. However, the nonmammalian pituitary does show some specificity with regard to the structure of natural GnRHs in that none of the tetrapod species studied is responsive to lamprey GnRH.