Introduction: Bile acids (BA) are cholesterol-derived molecules mostly known for their role in digesting lipids. By activating the Takeda G protein coupled receptor 5 (TGR5) in peripheral organs, they can also act as signaling molecules to reduce body weight and improve glucose homeostasis. Notably, TGR5 activation can increase energy expenditure in brown adipocytes, although the metabolic pathways involved in these effects are not yet clear. These outcomes imply an anti-obesity function for TGR5. However, all studies investigating BA in energy balance have exclusively focused on peripheral tissues. Since the major center of convergence of nutrient, hormonal, and environmental cues is the brain, particularly the hypothalamus, we hypothesized a role for TGR5 in this brain structure, suggesting that hypothalamic TGR5 activity may participate in energy balance, specifically under dietinduced obesity. Objective: To demonstrate the function of the BA – TGR5 system in hypothalamic populations known to control energy homeostasis, and disentangle its relevance for the treatment of diet-induced obesity. Methods: C57Bl6/J male mice that were either lean (standard chow) or diet-induced obese (60% high-fat diet; HFD) were implanted with an intra-cerebroventricular (ICV) cannula for the pharmacological delivery of TGR5 agonists. TGR5flox/flox mice were used to target the sitespecific deletion of the receptor within the mediobasal hypothalamus (MBH), through the stereotaxic delivery of AAV-Cre. The following metabolic outputs were measured: body weight, food intake, body composition (EchoMRI analyzer), insulin sensitivity, serum and hypothalamic BA (liquid mass spectrometry), and energy expenditure (TSE Phenomaster system). To block sympathetic signaling, we exposed mice to thermoneutrality (30°C) or performed chemical sympathectomy (6-hydroxydopamine; 80mg/kg i.p.). Markers of lipolysis, thermogenesis, and thyroid metabolism were measured in the liver, adipose and hypothalamic tissues by qPCR or western blots. All studies received the approval from the animal ethical committee of the University of Bordeaux. Results: We demonstrate that TGR5 and BA transporters are expressed in the MBH and that diet-induced obese mice have decreased circulating and hypothalamic BA. Acute ICV or intra-MBH administration of TGR5 agonists reduced food intake and body weight in dietinduced obese mice only, and improved insulin sensitivity. Accordingly, chronic ICV administration of the TGR5 agonist in obese mice reduced their body weight and adiposity, while increasing energy expenditure and mRNA markers of sympathetic activity in the adipose tissue. Indeed, experiments conducted at thermoneutrality or chemical sympathectomy blunted these effects, demonstrating that central TGR5 effects require an enhanced sympathetic tone. By using TGR5flox/flox mice coupled with the delivery of an AAVCre, we observed that the deletion of TGR5 in the MBH had no effect in chow-fed mice. However, a HFD switch rapidly increased their body weight, food intake and adiposity. When exposed to the cold (4 h at 4°C), protein levels of lipolysis and thermogenesis markers in the adipose tissue were blunted, implying an interruption in sympathetic signaling to the periphery due to hypothalamic downregulation of TGR5. Lastly, Cre-dependent deletion of TGR5 in the MBH of already obese mice rapidly increased adiposity by inducing hyperphagia, worsening their obese phenotype. Conclusions: Our work proves the existence of a functional hypothalamic BA – TGR5 receptor system. We show for the first time that the activation of TGR5 in the MBH decreases body weight and adiposity, while increasing energy expenditure through recruitment of the sympathetic nervous system. Taken together, these results expose a new mechanism of action for potential anti-obesity therapies.