Recently, lactate has been receiving great attention as an energy substrate in the brain. In this study, the role of lactate was evaluated by "bioradiography" system with 2-deoxy-2-[(18)F]fluoro-d-glucose ([(18)F]FDG), which is a positron emitting radiotracer for glucose uptake quantification. "Bioradiography" is the dynamic living tissue slice imaging system for positron-emitter labeled compounds. We investigated the brain energy metabolism under resting state and neural activated conditions induced by KCl addition. The monocarboxylate transporter inhibitor, alpha-cyano-4-hydroxycinnamate (4-CIN), had no effect on [(18)F]FDG uptake rate in rat brain slices before KCl addition. On the other hand, addition of 4-CIN induced larger [(18)F]FDG uptake rates under the activated condition in comparison with the control condition. Because neurons cannot utilize lactate under the 4-CIN loaded conditions, this indicates that activated neurons consume lactate as an energy substrate. The lactate concentration in the incubation medium was increased with KCl treatment in both groups and the extent was slightly greater in 4-CIN group. These results suggested that: (1) the brain mainly uses glucose, not lactate, as an energy substrate in resting state; (2) when neuron is stimulated, excess amounts of lactate might be produced in astrocytes and the lactate is mobilized as an energy substrate.