The melanocortin receptors, melanocortin-3-receptor (MC3-R) and melanocortin-4-receptor (MC4-R), are expressed in many discrete medial hypothalamic nuclei implicated in feeding regulation. The pro-opiomelanocortin product α-melanocyte stimulating hormone (α-MSH), an MC3/4-R agonist, decreases food intake following intracerebroventricular (ICV) injection in rats. MC4-R’s involvement in feeding has been established although a function for the MC3-R is unclear. We investigated endogenous melanocortin ligand binding and activation at the MC3-R and MC4-R and their effects on feeding. We have shown that α-MSH, desacetyl-α-MSH and β-MSH bound to the MC3-R and MC4-R with similar affinity and stimulated cAMP with similar potency in HEK 293 cells transfected with MC3-R and MC4-R. In contrast γ₂-MSH showed selectivity for the MC3-R over the MC4-R both in binding affinity and cAMP stimulation. α-MSH and β-MSH injected ICV into fasted rats at doses of 1, 3 and 6 nmol resulted in a decrease in food intake, (2 h food intake: α-MSH 6 nmol, 1.7±0.3 g; β-MSH 6 nmol, 1.5±0.3 g vs. saline 6.0±0.5 g, P<0.001). Desacetyl α-MSH did not reduce food intake at low doses but was significant at 25 nmol (2 h food intake: desacetyl-α-MSH 6.1±1.0 g vs. saline 9.5±1.4 g, P<0.05). In contrast, γ₂-MSH had no effect on food intake when administered ICV to fasted rats. We were unable to establish a role for the MC3-R in feeding regulation. Our evidence, however, strengthens the hypothesis that the melanocortin’s effects on food intake are mediated via the MC4-R.