Effect of gestational ethanol exposure on the NMDA receptor complex in rat forebrain: from gene transcription to cell...

Effects of gestational ethanol exposure on the trafficking of the NMDA receptor complex were investigated. Studies focused on three distinct processes in NMDA receptor translocation: (1) the level of gene transcription (2) nascent NMDA receptor subunits (NR) associated with the endoplasmic reticulum bound chaperone protein calnexin and (3) NMDA receptors associated with the cell surface anchoring protein PSD-95. Forebrain mRNA and membrane proteins were isolated from postnatal day 1 rat pups from prenatally ethanol exposed, pair-fed and ad libitum experimental groups. Ribonuclease protection assays were carried out to determine the levels of NR2A, NR2B, and NR2C mRNA within the treatment groups determined. Results indicated that gestational ethanol exposure did not affect the gene transcription of the NR2 subunits. Immunoprecipitation experiments were conducted with an anti-calnexin antibody or an anti-PSD-95 antibody and the immunoprecipitates probed for NR1 and NR2 subunits. Within the anti-calnexin immunoprecipitates, no NR2A, NR2B or NR2C subunits were detectable, but a significant pool of NR1 subunits was identified. These findings suggest that NR1 subunits but not NR2 subunits are associated with calnexin within the endoplasmic reticulum. Further, gestational ethanol exposure significantly increased the NR1 polypeptide levels in the anti-calnexin immunoprecipitate. Anti-PSD-95 immunoprecipitates revealed an abundance of NR1 and NR2B subunits, and these complexes were unaffected by gestational ethanol exposure. No NR2A or NR2C subunits were detected. These results suggest that gestational ethanol exposure significantly affects the assembly and transport of NMDA receptors. Gestational ethanol exposure may not alter the composition of the PSD-95 associated NMDA receptor complex.