The neuromodulatory effect of pyroglutamylated RFamide peptide 43 (QRFP43) on the hypothalamic appetite regulation centre in sheep has not yet been investigated. The present work focuses on the role of QRFP43 in modulating mRNA expression of pyroglutamylated RFamide peptide receptor (QRFPR), neuropeptide Y (NPY), agouti related neuropeptide (AGRP), cocaine- and amphetamine-regulated transcript prepropeptide (CARTPT), proopiomelanocortin (POMC), peptidyglycine alpha-amidating monooxygenase (PAM) and NPY protein expression in the hypothalamic arcuate nucleus in sheep. The aim of this study was to investigate whether QRFP43 could affect gene expression of the appetite-regulating centre in the hypothalamus. The experiment was conducted from September to December and included forty-eight female Polish Merino sheep randomly assigned to three groups. The control group received an intracerebrovenctricular infusion of Ringer-Locke solution (480 μl/day), whereas the experimental groups were administered QRFP43 in two doses: 10 or 50 μg/480 μl/day (referred to as the RFa10 and RFa50 groups). Selected brain structures were collected from animals for immunohistochemical and real-time PCR analyses. Central infusions of QRFP43 induced changes in mRNA expression of the NPY, CARTPT, POMC and QRFPR genes. A decrease in NPY and QRFPR mRNA expression, and an increase in CARTPT mRNA expression were detected. Furthermore, infusion of QRFP43 in the RFa50 group resulted in decreased POMC mRNA expression. Our findings suggest that QRFP43 may exert an inhibitory effect on the hypothalamic neuronal network responsible for appetite regulation in sheep. Moreover, it appears that the effect of QRFP43 may vary in different animal species, thus further research is required, especially involving hormonal proteomic analyses.