The hypothalamic neuropeptide neurons play the pivotal role in energy balance. The arcuate nucleus (ARC) neurons respond to peripheral hormones and nutrients through neuropeptide Y/agouti-related peptide (NPY/AgRP) and pro-opiomelanocortin/cocaine-amphethamine regulated transcript (POMC/CART). These neurons project to the paraventricular nucleus (PVN) and lateral area of the hypothalamus (LHA) where activate the second order of neurons producing e.g. orexins and melanin-concentrating hormone (MCH). The aim of this study was to investigate the effect of feeding with different fats on neuropeptide expression in the ARC and LHA, and melanocortin recepror-4 (MC4R) in the PVN. Three groups of rats were fed six weeks a low-fat diet (10% energy from fat), next three a high-fat diet (40% energy from fat) prepared with the same fat; lard – source of saturated fatty acids, sunflower oil – source of n-6 polyunsaturated fatty acids (PUFA), or fish oil rich in n-3 PUFA (8 rats/group). Throughout the experiment body weight and food intake were monitored. Before decapitation and brain removal rats were i.v. anaesthetized with ketamine (20 mg/kg) plus xylazine (10 mg/kg) and blood was drawn by heart puncture. Plasma level of glucose, leptin and insulin were measured. The ARC, PVN and LHA were punctured from frozen brain sections. NPY, galanin-like peptide (GALP), POMC, pre-orexin, MCH, MC4R expression was measured by RT-PCR. The whole hypothalamus was assayed for NPY and POMC expression by in situ hybridization (4 rats/group). The lard diets fed rats gained the highest body weight and ate the most energy vs. rats fed the PUFA diets. The NPY expression in the ARC and compact zone of the dorsomedial nucleus in obese high-lard fed rats was lower than in low-lard, and high-PUFA fed rats. The GALP expression increased by about 50% and 40% in the ARC in high-sunflower oil and high-fish oil groups, respectively vs. rats fed the corresponding low-fat diet. POMC expression in the ARC was increased in both high-PUFA fed rats vs. that fed the corresponding low-PUFA and high-lard diets. The POMC expression in the whole hypothalamus was increased, and the pre-orexin and MCH mRNA transcript levels in the LHA were decreased exclusively in high-fish oil fed rats. The expression of MC4R in the PVN was lower in high-lard than in low-lard fed rats. The high-fat fed rats had a significantly increased leptin, insulin and glucose levels independently of the kind of dietary fat. Summarizing, high ingestion of fish oil originate PUFA favoured anorexigenic POMC expression in the hypothalamus and down-regulated orexigenic peptides in the LHA. The high-saturated feeding failed to up-regulate POMC expression and down-regulated MC4R expression in the PVN. Our study indicates that hyperphagia caused by diet high in saturated fat may impair melanocortin transmission in the hypothalamus.