Oligonucleotide and cRNA probes were used for nonradioactivein situ hybridizations carried out to identify the neural cell types expressing the glutamate transporter GLAST mRNA in the rat CNS. Additionally, the regional distribution of GLAST mRNA-expressing cells was studied, and the results were complemented by immunocytochemical investigations using an antibody against a synthetic GLAST peptide. The findings documented that GLAST is expressed by Bergmann glia and by astrocytes throughout the CNS. The glial localization of GLAST mRNA was verified unequivocally by double-labeling with an astrocytic marker protein. Additionally, GLAST mRNA reactivity and GLAST immunoreactivity were found in ependymal cells. In other neural cell types of the CNS, GLAST expression was not detectable. A high level of astrocytic immunolabeling was observed in the entire gray matter of the brain, with variations in intensity in different regions. Those brain areas that are known to possess high glutamatergic activity and astrocytic glutamate metabolism stained intensely for both GLAST mRNA and GLAST protein. The latter observation suggests that the GLAST glutamate transporter participates in the regulation of extracellular glutamate concentrations, especially in brain areas receiving an intense glutamatergic innervation.