Reduced metabolic rate may contribute to weight gain in leptin-deficient (ob/ob) mice; however, available studies have been criticized for referencing O₂ consumption (V˙o ₂) to estimated rather than true lean body mass. To evaluate whether leptin deficiency reduces energy expenditure, four separate experiments were performed:1) NMR spectroscopy was used to measure fat and nonfat mass, permittingV˙o ₂ to be referenced to true nonfat mass; 2) dietary manipulation was used in an attempt to eliminate differences in body weight and composition between ob/ob and C57BL/6J mice; 3) short-term effects of exogenous leptin (0.3 mg ⋅ kg⁻¹ ⋅ day⁻¹) on V˙o ₂ were examined; and4) body weight and composition were compared in leptin-repleted and pair-fedob/ob animals.ob/ob animals had greater mass, less lean body mass, and a 10% higher metabolic rate whenV˙o ₂ was referenced to lean mass. Dietary manipulation achieved identical body weight inob/ob and C57BL/6J animals; however, despite weight gain in C57BL/6J animals, percent fat mass remained higher in ob/ob animals (55 vs. 30%). Exogenous leptin increasedV˙o ₂ inob/ob but not control animals. Weight loss in leptin-repleted ob/ob mice was greater than in pair-fed animals (45 vs. 17%). We conclude, on the basis of the observed increase inV˙o ₂ and accelerated weight loss seen with leptin repletion, that leptin deficiency causes a reduction in metabolic rate in ob/obmice. In contrast, these physiological studies suggest that comparison of V˙o ₂ in obese and lean animals does not produce useful information on the contribution of leptin to metabolism.