Non-Alcoholic Fatty Liver Disease (NAFLD) is the most common cause of chronic liver disease worldwide, affecting 70-90% of obese individuals. In humans, a lower NAFLD incidence is reported in pre-menopausal women, although the mechanisms affording this protection remain under-investigated. Here, we tested the hypothesis that the constitutive androstane nuclear receptor (CAR) plays a role in the pathogenesis of experimental NAFLD. Male and female wild-type (WT) and CAR knock-out (CAR-/-) mice were subjected to a high-fat diet (HFD) for 16 weeks. We examined the metabolic phenotype of mice through body weight follow-up, glucose tolerance tests, analysis of plasmatic metabolic markers, hepatic lipid accumulation, and hepatic transcriptome. Finally, we examined the potential impact of HFD and CAR deletion on specific brain regions, focusing on glial cells. HFD-induced weight gain and hepatic steatosis are more pronounced in WT males than females. CAR-/- females present a NASH-like hepatic transcriptomic signature suggesting a potential NAFLD to NASH transition. Transcriptomic correlation analysis highlighted a possible cross-talk between CAR and ERα receptors. The peripheral effects of CAR deletion in female mice were associated with astrogliosis in the hypothalamus. These findings prove that nuclear receptor CAR may be a potential mechanism entry-point and a therapeutic target for treating NAFLD/NASH.