Summary/Abstract Neurons and local CNS-resident immune cells engage in complex interactions to regulate brain function. In the mediobasal hypothalamus (MBH), a critical integrator of energy balance, diets rich in fat rapidly induce the inflammatory activation and accumulation of a heterogeneous population of myeloid cells, broadly termed microglia. We showed that microglia activated in this context are sufficient to stimulate food intake and body weight gain, however the metabolic factors initiating this response are unknown. The goal of this proposal is to determine the role of the gut microbiota in regulating the function of MBH myeloid cells. The objective is to identify specific myeloid populations responsive to gut microbiota-derived signals and determine how they transduce this response to influence hypothalamic regulation of metabolic function. We will use this information to design novel myeloid cell-based therapeutic interventions to limit diet-induced obesity and its metabolic consequences. To reach our objective, we will leverage our expertise in single-cell profiling of MBH myeloid cells to define specific populations under gut microbial regulation. Diet is a major factor affecting the composition of the gut microbiota, and diets rich in saturated fat induce unfavorable alterations in the type and numbers of commensal microorganisms in the gut. These changes may impact MBH myeloid cell function. Our aim is to determine the contribution of the gut microbiota in determining hypothalamic myeloid responses to dietary saturated fat. The following sub-aims are proposed: (1) Determine how the interaction between dietary fat composition and gut microbiota influences hypothalamic myeloid responses; (2) Determine the specific contribution of gut microbiota on hypothalamic myeloid function. Completing the proposed work will provide novel insights into how the dietary fat consumption and the gut microbiota interact to regulate the functional diversity of diet-responsive MBH myeloid cells and the hypothalamic control of energy metabolism.