PROJECT SUMMARY The recent increase in obesity prevalence is a major public health concern in the United States. Since energy balance regulation is rooted in the brain, understanding the neural feeding circuits that impede weight loss and maintenance is necessary for the development of novel obesity treatments. Agouti-related protein-expressing neurons in the hypothalamic arcuate nucleus (ARCAGRP) are both necessary and sufficient for food intake. Indeed, acute stimulation of ARCAGRP neurons drives food intake, and chronic stimulation leads to dramatic hyperphagia and weight gain. ARCAGRP neurons can be divided into distinct subpopulations that project to one of several target regions. Independent stimulation of four of these distinct projection subpopulations [ARCAGRP- bed nucleus of the stria terminalis (BNST), -paraventricular hypothalamic nucleus (PVH), -lateral hypothalamic area (LHA) and paraventricular thalamic nucleus (PVT)] is sufficient to drive food intake. We have also shown that ARCAGRP neurons transmit a negative valence that may be likened to the negative affect that is associated with hunger. Here, we take advantage of the one-to-one neuron connectivity to test the affective, functional and physiological relevance of ARCAGRP neuron subpopulations. The main goal of this proposal is to test the hypothesis that ARCAGRP neurons contribute to diet-induced obesity by driving food intake and negative valence through distinct neuron subpopulations. Specific Aim I experiments will examine the valence of the four ARCAGRP neuron subpopulations that increase feeding upon stimulation to provide insight into the specific neural projections that mediate negative affect associated with dieting for weight loss. Specific Aim II examines the neural activity dynamics of feeding-sufficient ARCAGRP neuron subpopulations during the gradual onset of hunger and feeding in lean and obese mice to better understand the endogenous role of feeding-sufficient ARCAGRP subpopulations in hunger and weight gain. Finally, given that persistent ARCAGRP neuron firing is associated with obesity, Specific Aim III experiments will directly examine the role of ARCAGRP neurons in the development and treatment of diet-induced obesity. Overall, results from these experiments will identify ARCAGRP neuron subpopulations, target regions, and mechanisms that can be leveraged to develop novel treatments for obesity.