Rapid down‐regulation of glucocorticoid receptor gene expression in the dentate gyrus after acute stress in vivo: Role of DNA Methylation and microRNA Activity

Background: Although glucocorticoid receptors (GRs) in the hippocampus play a vital role in the regulation of physiological and behavioural responses to stress, the regulation of receptor expression remains unclear. This work investigates the molecular mechanisms underpinning stress-induced changes in hippocampal GR mRNA levels in vivo.

Methods: Male Wistar rats were killed either under baseline conditions or after forced swim stress (FSS; 15 min in 25°C water). Rat hippocampi were micro-dissected (for mRNA, microRNA, and DNA methylation analysis) or frozen whole (for chromatin immunoprecipitation (ChIP). In an additional experiment, rats were pre-treated with RU486 (a GR antagonist) or vehicle.

Results: FSS evoked a dentate gyrus-specific reduction in GR mRNA levels. This was associated with increased DNMT3a protein association with a discreet region of the Nr3C1 (GR gene) promoter, shown here to undergo increased DNA methylation after FSS. FSS also caused a time-dependent increase in the expression of miR-124a, a microRNA known to reduce GR mRNA expression, which was inversely correlated with a reduction in GR mRNA levels at 30 min post-FSS. FSS did not affect GR binding to a putative negative glucocorticoid response element (GRE) within the Nr3c1 gene.

Conclusions: Acute stress results in decreased GR mRNA expression specifically in the dentate gyrus. Our results indicate that a complex interplay of multiple molecular mechanisms including increased DNA methylation of discrete CpG residues within the Nr3c1 gene, most likely facilitated by DNMT3a, and increased expression of miR-124a could be responsible for these changes.