Previous studies indicated that there is a separate hypothalamic control of follicle-stimulating hormone (FSH) release distinct from that of luteinizing hormone (LH), An FSH-releasing factor (FSHRF) was purified from rat and sheep hypothalami, but has not been isolated, We hypothesized that FSHRF might be an analogue of mammalian luteinizing hormone-releasing hormone (m-LHRH) and evaluated the activity of many analogues of m-LHRH and of the known LHRHs found in lower forms, Here we demonstrate that lamprey (1) LHRH-III has a potent, dose-related FSH-but not LH-releasing action on incubated hemipituitaries of male rats, 1-LHRH-I on the other hand, had little activity to release either FSH or LH, m-LHRH was equipotent to 1-LHRH-III to release FSH, but also had a high potency to release LH in contrast to 1-LHRH-III that selectively released FSH, Chicken LHRH-II had considerable potency to release both LH and FSH, but no selectivity in its action, Salmon LHRH had much less potency than the others tested, except for 1-LHRH-I, and no selectivity in its action, Because ovariectomized, estrogen, progesterone-treated rats are a sensitive in vivo assay for FSH- and LH-releasing activity, we evaluated 1-LHRH-III in this assay and found that it had a completely selective stimulatory effect on FSH release at the two doses tested (10 and 100 pmols), Therefore, 1-LHRH-III is a highly potent and specific FSH-releasing peptide that may enhance fertility in animals and humans, It may be the long sought after m-FSHRF.