There is ample evidence that glucose metabolism in the pregenual anterior cingulate cortex (PACC) is increased in major depressive disorder (MDD), whereas it is still unknown whether glucose levels per se are also elevated. Elevated cerebrospinal fluid (CSF) lactate concentrations in MDD patients might indicate that increased glycolytical metabolization of glucose to lactate in astrocytes either alone or in conjunction with mitochondrial dysfunction results in an accumulation of lactate and contributes to pathophysiological mechanisms of MDD. However, until now, no study investigated in vivo PACC glucose and lactate levels in MDD. Proton magnetic resonance spectroscopy was therefore used to test the hypothesis that patients with MDD have increased PACC glucose and lactate levels. In 40 healthy and depressed participants, spectra were acquired from the PACC using a maximum echo J-resolved spectroscopy protocol. Results show significant increases of glucose and lactate in patients, which are also associated with depression severity. These findings indicate impaired brain energy metabolism in MDD with increased fraction of energy utilization via glycolysis and reduced mitochondrial oxidative clearance of lactate. Targeting these metabolic disturbances might affect the balance of metabolic pathways regulating neuronal energetics and result in an attenuation of the elevated basal activity of brain regions within the neural circuitry of depression.