In the rat central nervous system, the mRNA encoding the N-methyl-D-aspartate receptor subunit R1 is the most ubiquitously distributed among the cloned subunit mRNAs of this glutamate receptor subtype. The N-methyl-D-aspartate R1 mRNA is very abundantly expressed and N-methyl-D-aspartate R1 coexpression is necessary for functional expression of all other cloned N-methyl-D-aspartate receptor subunits. Therefore, the R1 subunit is likely to be an essential component of all known N-methyl-D-aspartate receptors in rat brain. By employing sequence specific polyclonal antibodies, we demonstrate that rat brain N-methyl-D-aspartate R1, as well as recombinantly expressed receptor protein, has an apparent molecular mass of 116 kDa in sodium dodecyl sulfate polyacrylamide gel electrophoresis. The receptor protein is heavily glycosylated. It is specifically localized to the central nervous system, and it co-enriches with synaptic membranes upon subcellular fractionation of the cerebral cortex. Chemical cross-linking of synaptic membrane proteins shows that the N-methyl-D-aspartate R1 protein is part of a receptor protein complex with a molecular mass of 730 kDa. By using immunocytochemical methods, we demonstrate a widespread but distinct distribution of N-methyl-D-aspartate R1 in neurons of the rat brain, with prominent immunostaining in certain layers of the cerebral cortex, in the hippocampus and dentate gyrus, as well as in the cerebellum.