Non-alcoholic fatty liver disease (NAFLD) is a common and morbid metabolic disease that independently predicts development of type 2 diabetes mellitus and its cardiovascular, renal and retinal complications. More than one billion individuals have NAFLD, making this the most common chronic liver disease worldwide. A barrier to effectively treating and preventing this disease and its complications is the lack of effective NAFLD treatments. Our long-term goal is to understand the underlying glucose metabolic pathways that lead to NAFLD, so that we can precisely modulate these pathways to treat or prevent its progression. We identified the hepatic carbohydrate carrier, GLUT8, as a therapeutic target because: (i.) GLUT8 promotes diet-induced hepatic steatosis and metabolic syndrome and (ii.) the disaccharide and glucose mimetic trehalose inhibits GLUT8 and activates an adaptive fasting response (e.g. AMP kinase and autophagy), to reverse hepatic steatosis. The objective here is to define the interaction between hepatocyte glucose transport, trehalose, and adaptive hepatocyte fasting responses. Our preliminary data suggest that acute hepatocyte-selective GLUT8 knockdown and hepatocyte- specific germline GLUT8 gene deletion (GLUT8 LKO mice) each induce hepatocyte fasting signals and peripheral thermogenesis. We therefore hypothesize that genetic and pharmacological hepatocyte GLUT8 blockade induces thermogenesis and confers resistance to hepatic steatosis. The two Specific Aims are to 1) Identify mechanisms by which GLUT8 regulates hepatic and extrahepatic metabolism, and 2) Define mechanisms mediating trehalose-induced thermogenesis and protection from NAFLD. We will accomplish these aims by leveraging novel experimental mouse models and unique in vivo imaging techniques. Completing these aims informs how to optimally target hepatocyte glucose transport to augment hepatocyte and whole-organism energy homeostasis.