To identify regions in the hypothalamus involved in refeeding and their regulation by -melanocyte-stimulating hormone (-MSH), adult rats were subjected to a 3-day fast and 2 h after refeeding, the distribution of cfos-immunoreactive neurons was elucidated. Compared with fed and fasted animals, a significant increase (P < 0.001) in the number of cfos-immunoreactive cells was identified in refed animals in the supraoptic nucleus, magnocellular and ventral parvocellular subdivision (PVNv) of the hypothalamic paraventricular nucleus, and the dorsal (DMNd) and ventral (DMNv) subdivisions of the dorsomedial nucleus. Refeeding shifted the location of cfos-labeled neurons from the medial to lateral arcuate where cfos was induced in 88.7 ± 2.2% of -MSH-containing neurons. -MSH-containing axons densely innervated the PVNv, DMNd and DMNv and organized in close apposition to the majority of refeeding-activated cfos-positive neurons. To test whether the melanocortin system is involved in induction of cfos in these regions, the melanocortin 3/4 receptor antagonist, agouti related protein (AGRP 83-132), was administered to fasting animals just before refeeding. Compared with aCSF, a single icv bolus of AGRP (5 µg/5 µl) not only significantly increased the total amount of food consumed within 2 h, but also nearly abolished refeeding-induced cfos expression in the PVNv and DMNd, and partially reduced cfos-immunoreactivity in the DMNv. We conclude that refeeding activates a subset of neurons in the PVN and DMN as a result of increased melanocortin signaling, and propose that one or more of these neuronal populations mediate the potent anorexic actions of -MSH.