Neuronal Per Arnt Sim domain 4 (Npas4), a brain-specific helix-loop-helix transcription factor, was recently shown to regulate the development of GABAergic inhibitory neurons. Npas4 mRNA expression levels were decreased in the hippocampus of mice exposed to stress, which was accompanied by brain dysfunction. We have suggested that transient stress reduced Npas4 transcription through the glucocorticoid receptor. In the present report, we investigated the potential contribution of epigenetic modifications induced by stress on Npas4 gene expression. The Npas4 promoter region contains two CpG islands; in the hippocampus, chronic restraint stress increases the DNA methylation levels of both of these CpG islands. In the Neuro2a cell line, treatment with a DNA methyltransferase inhibitor, 5-aza-2'-deoxycytidine, increased Npas4 mRNA levels and markedly reduced the DNA methylation levels of CpG island 2 in the Npas4 promoter. The DNA methylation sites in CpG island 2 overlap with two cyclic adenosine monophosphate response element (CRE) sequences. Mutation of these CRE sequences reduced Npas4 promoter activity. These results suggest that transcription of the Npas4 gene is downregulated by stress through DNA methylation of its promoter.