The activity of the pyruvate carboxylase was determined in brains of newborn and adult mice as well as primary cultures of astrocytes, of cerebral cortex neurons, and of cerebellar granule cells. The activity was found to be 0.25 +/- 0.14, 1.24 +/- 0.07, and 1.75 +/- 0.13 nmol X min -1 X mg -1 protein in, respectively, neonatal brain, adult brain, and astrocytes. Neither of the two types of neurons showed any detectable enzyme activity (i.e., less than 0.05 nmol X min -1 X mg -1). It is therefore concluded that pyruvate carboxylase is an astrocytic enzyme.