Ketone bodies are important alternate brain fuels, but their capacity to replace glucose and support neural function is unclear. In this study, the contributions of ketone bodies and glucose to cerebral cortical metabolism were measured in vivo in halothane-anesthetized rats fasted for 36 hours (n=6) and receiving intravenous [2,4-(13)C(2)]-D-β-hydroxybutyrate (BHB). Time courses of (13)C-enriched brain amino acids (glutamate-C4, glutamine-C4, and glutamate and glutamine-C3) were measured at 9.4 Tesla using spatially localized (1)H-[(13)C]-nuclear magnetic resonance spectroscopy. Metabolic rates were estimated by fitting a constrained, two-compartment (neuron-astrocyte) metabolic model to the (13)C time-course data. We found that ketone body oxidation was substantial, accounting for 40% of total substrate oxidation (glucose plus ketone bodies) by neurons and astrocytes. D-β-Hydroxybutyrate was oxidized to a greater extent in neurons than in astrocytes (≈ 70:30), and followed a pattern closely similar to the metabolism of [1-(13)C]glucose reported in previous studies. Total neuronal tricarboxylic acid cycle (TCA) flux in hyperketonemic rats was similar to values reported for normal (nonketotic) anesthetized rats infused with [1-(13)C]glucose, but neuronal glucose oxidation was 40% to 50% lower, indicating that ketone bodies had compensated for the reduction in glucose use.