The neuropeptide FF receptor 2 (NPFFR2) is highly expressed in the hypothalamus where it is activated by a set of RFamide peptides. However, its function is unknown. Here we show that lack of NPFFR2 in female mice results in significant reduction in body fat mass and increase in lean mass, that is associated with increase in energy expenditure during the light phase, increases in body temperature and physical activity. Interestingly, female NPFFR2^-/- mice exhibit equal glucose tolerance to wild type (WT) at the expense of enhanced glucose-stimulated insulin secretion, suggesting an impaired insulin action. Importantly, female NPFFR2^-/- mice on a high fat diet (HFD) for 8 weeks show greater weight and fat gains compared to those of WT mice, which are associated with significant decreases in energy expenditure and physical activity. In addition, whereas HFD increases energy expenditure in both genotypes, the increase is less pronounced in NPFFR2^-/- compared to that of WT mice, suggesting an impaired diet-induced adaptive thermogenesis by lack of NPFFR2 signaling. In support of this, HFD significantly increased the protein levels of UCP-1 and PGC-1a in the brown adipose tissue of WT but failed to do so in NPFFR2^-/- mice. Finally, NPFFR2 signaling may regulate energy expenditure and adaptive thermogenesis via hypothalamic neuropeptide Y (NPY) pathway, since the HFD-induced decrease in hypothalamic NPY expression observed in WT mice is absent in NPFFR2^-/- mice. Taken together, these data demonstrate that NPFFR2 signalling plays important roles in the regulation of energy metabolism and glucose homeostasis. The regulation of NPFFR2 signaling in diet-adaptive thermogenesis may involve hypothalamic NPY pathway and down-stream brown adipose tissue thermogenesis.