The central neuropeptide Y (NPY) Y1 receptor (Y1-R) system has been implicated in feeding, endocrine, and autonomic regulation. In the present study, we systematically examined the brain distribution of Y1-R mRNA in rodents by using radioisotopic in situ hybridization histochemistry (ISHH) with a novel sensitive cRNA probe. Within the rat hypothalamus, Y1-R–specific hybridization was observed in the anteroventral periventricular, ventromedial preoptic, suprachiasmatic, paraventricular (PVH), dorsomedial, ventromedial, arcuate, and mamillary nuclei. In the rat, Y1-R mRNA expression was also seen in the subfornical organ, anterior hypothalamic area, dorsal hypothalamic area, and in the lateral hypothalamic area. In addition, Y1-R hybridization was evident in several extrahypothalamic forebrain and hindbrain sites involved in feeding and/or autonomic regulation in the rat. A similar distribution pattern of Y1-R mRNA was observed in the mouse brain. Moreover, by using a transgenic mouse line expressing green fluorescent protein under the control of the melanocortin-4 receptor (MC4-R) promoter, we observed Y1-R mRNA expression in MC4-R–positive cells in several brain sites such as the PVH and central nucleus of the amygdala. Additionally, dual-label ISHH demonstrated that hypophysiotropic PVH cells coexpress Y1-R and pro–thyrotropin-releasing hormone mRNAs in the rat. These observations are consistent with the proposed roles of the central NPY/Y1-R system in energy homeostasis. J. Comp. Neurol. 482:217–243, 2005. © 2004 Wiley-Liss, Inc.