Different NR2 subunits (NR2A-D) of NMDA receptors confer distinct properties on the receptors and the subunit composition of heteromeric NMDA receptor complex is tightly regulated. Here, we demonstrate that suppression of neuronal activity causes mRNA expression of the NR2B subunit to increase significantly, both in vitro and in vivo, and that this modulation of transcription is mediated by epigenetic mechanisms. Treating cortical neurons with TTX substantially increases the level of mRNAs for NMDA receptor subunits. Particularly, the NR2B expression increases over 2-fold, similar to the effects of dark-rearing. The increase of NR2B induced by TTX is occluded by inhibiting DNMTs. Furthermore, MeCP2 binds to NR2B and the association of MeCP2 with NR2B is reduced by TTX treatment. Together, these data indicate that DNA methylation as well as subsequent MeCP2 association mediates neuronal activity-dependent regulation of NR2B expressions.