Previous studies have shown that chronic ethanol treatment up-regulates the expression of the N-methyl-D-aspartate (NMDA) receptor number and function both in vitro and in vivo. In vitro chronic ethanol treatment specifically augments mRNA levels of the R2B subunit without altering R1 subunit mRNA levels, although similar treatment results in increased levels of both R1 and R2B polypeptides. To further delineate the molecular mechanisms involved in differential regulation of NMDA receptor subunits by chronic ethanol treatment (50 mM, 5 days), we have determined the mRNA stability of the NMDA R1 and R2B subunits and the transcription rate of the respective genes using mouse fetal cortical neurons. Our observations demonstrated that ethanol stabilized the NMDA R1 mRNA over the time period examined (24 h) without altering the stability of the R2B mRNA. Chronic exposure of fetal cortical neurons to ethanol enhanced the rate of R2B gene transcription approximately twofold. Taken together, these results suggest that up-regulation of the NMDA receptor number seen in cultured cortical neurons after chronic ethanol treatment is due to the regulation of the NMDA R2B receptor subunit at the transcriptional level and that of the NMDA R1 subunit mainly at the posttranscriptional level.