Developmental and regional alternative splicing of the NMDAR1 subunit gene transcript was examined by in situ hybridization in the developing and adult rat brain. NMDAR1 mRNA, barely detectable at embryonic day 14, increased gradually during development until the third postnatal week, after which it declined slightly to adult levels, when it was detected in every examined neuronal type. Each splice form of the primary NMDAR1 gene transcript was found to follow a parallel profile of abundance in the brain, but marked regional differences were observed in splicing at both 5′ and 3′ sequences. The individual regional distributions of splice forms appeared to be established around birth, with little change thereafter, except in the overall abundance. The NMDAR1-a and NMDAR1–2 splice forms occurred extensively and approximately homogeneously throughout brain gray matter. The NMDAR1-b variant was found primarily in the sensorimotor cortex, neonatal lateral caudate, thalamus, hippocampal CA3 field, and cerebellar granule cells, but was absent from adult caudate. The NMDAR1–1 and -4 splice forms were detected in almost complementary patterns; the former was concentrated in more rostral structures such as cortex, caudate, and hippocampus, while the latter was principally in more caudal regions such as thalamus, colliculi, and cerebellum. These two splice forms accounted for a greater proportion of the adult NMDAR1 mRNA than that of the neonate. The NMDAR1–3 mRNA variant was scarce, being detected only at very low levels in postnatal cortex and hippocampus. The different splice forms may generate regional differences in NMDA receptor properties during development and in the adult CNS.