NMDA receptor subunit mRNA and protein expression in ethanol-withdrawal seizure-prone and -resistant mice

Background: Ethanol withdrawal seizure-prone (WSP) and -resistant (WSR) mic e have been genetically selected for differences in handling-induced convul sion severity during withdrawal from chronic ethanol administration. Import antly, drug-naive mice from these selected lines also differ in handling-in duced convulsion severity. Different N-methyl-D-aspartate (NMDA) receptor s ubunit and splice variant associations confer varying sensitivities to etha nol, and may play a role in the different behavioral responses of the WSP a nd WSR mice. Methods: In situ hybridization of riboprobes was used to characterize NMDA receptor subunit and splice variant mRNA expression in cortex and hippocamp us from WSP and WSR mice. In addition, immunoblotting and immunohistochemis try were used to examine the expression of specific NMDA receptor subunits and splice variants in hippocampus and cortex from the selected mouse lines . Results: In situ hybridization of riboprobes indicated that, in brain secti ons from both WSP and WSR mice, there was a differential regional distribut ion of mRNA for the mouse NR1, NR2A, NR2B, and NR2C NMDA receptor subunits. However, there were no differences between the selected lines in the hybri dization of riboprobes to hippocampal subfields or cortical layers. In addi tion, hybridization of the probe for a 63-base N1-terminal cassette of etha nol-sensitive NR1 splice variants labeled both cortex and hippocampus. The level of hybridization did not differ across subfields of the hippocampus. Results from Western blot and immunohistochemical experiments also indicate d that there were no differences between selected lines in NMDA receptor su bunit protein expression. However, there was a correlation between mRNA and protein expression in hippocampus and cortex for each NMDA receptor subuni t that was examined. Conclusions: The data suggest that at the level of both mRNA and protein, N MDA receptor subunit and splice variant expression can be uncoupled from co nvulsion severity in mice that have been selectively bred for symptoms of e thanol withdrawal.