The distribution of glucosephosphate isomerase (GPI, D-glucose-6-phosphate ketol isomerase) in mouse nervous tissue has been determined at the light microscopic level by immunofluorescence and histochemical procedures. The fluorescence procedure, which utilizes anti-GPI antibodies, detected lower levels of GPI than the histochemical procedure, which relies upon the catalytic activity of the enzyme. The distribution of GPI in nervous tissue is very similar to that of hexokinase. High levels of GPI were found in the Purkinje cells, the molecular layer, and the glomeruli of the granular layer in the cerebellar cortex; the pontine nuclei and the inferior olivary nuclei of the pons and medulla; the neurons of the thalamus and hypothalamus; the pyramidal cells, the dentate nuclei, and Ammons' horn of the cerebral cortex; the ventral horn cells of the spinal cord; and ventricular cells, choroid plexus cells, and the leptomeninges. The neuropil throughout the central nervous system (CNS) stained uniformly with moderately high levels of GPI. No GPI was observed in the myelin sheaths of the CNS.