Technical Abstract: Gonadotropin-inhibitory hormone (GnIH) was identified as an RFamide-related peptide (RFRP) in avian species. Mammalian orthologs (RFRP-1 and RFRP-3) have been reported in the human, rodents, and recently in sheep, but the role of RFRPs in the domestic pig is not established. We hypothesize that a GnIH neuronal system is present in the domestic pig. Hypothalami were collected from prepubertal gilts (PIC composite; 160 d of age, n=2) and total RNA purified for reverse transcription. The cDNA was used in PCR with primers based on sequence conservation of known RFRP genes. The resulting sequence revealed a reading frame that coded for at least two porcine RFamide-related peptides (RFRP-1 and RFRP-3) with a high degree of homology to the human orthologs (92 and 79%, respectively). Prepubertal gilts (n=5) were perfused transcardially with 4% paraformaldehyde. Hypothalami were dissected and processed for immunoflouorscence histochemistry. The numbers of RFRP-ir neurons were evaluated in every fourth section (20 -M) using the Axio Vision 4.6 Imaging system. RFRP-ir neurons (5-15 per section) were observed to be solitarily distributed in the posterior hypothalamic, lateral mammillary and dorsomedial nuclei. Numerous ir-perikarya (16-25 per section) were found in the ventromedial, lateral hypothalamic, suprachiasmatic and lateral preoptic nuclei. The paraventricular nucleus had densely organized RFRP-ir neurons found in clusters of 26-50 neurons near the 3rd ventricle. These results indicate that a GnIH neuronal system is present in the pig. Currently, we seek to elucidate the role these neuropeptides may have in regulating gonadotropin secretion in swine.