Importance of epigenetic mechanisms in visceral pain induced by chronic water avoidance stress

Abstract

Epigenetic molecular mechanisms, which include DNA methylation and histone deacetylation, are implicated in the dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis. Previously, we demonstrated that repeated water avoidance stress (WAS), a validated model of chronic psychological stress, induces heightened visceral pain behaviors in rodents that resemble irritable bowel syndrome (IBS) sequelae. However, the involvement of epigenetic molecular mechanisms in the pathophysiology of stress-induced visceral pain has not been explored. Our hypothesis is that epigenetic mechanisms within the central nervous system (CNS) are important to chronic stress-induced visceral hypersensitivity. Adult male F-344 rats with intracerebroventricular (i.c.v.) cannulae were exposed to 7 days of repeated WAS. Controls received a SHAM stress. Following the daily 1h stressor, trichostatin A (TSA; 100 ng/ml), a potent histone deacetylase inhibitor, or vehicle (VEH; 0.1% DMSO/saline,) as control was administered via the i.c.v. cannula. Visceral sensitivity was assessed 24h after the final WAS and quantified the visceromotor response (VMR) by recording the number of abdominal contractions in response to graded pressures (20-60 mmHg) of colorectal distensions (CRD). From a separate group of rats that were exposed to repeated WAS or SHAM stress, the amygdala was isolated to assess the methylation status of glucocorticoid receptor (GR) and corticotropin releasing-factor (CRF) genes via bisulfite sequencing and verified by pyrosequencing. GR and CRF gene expression was quantified via qRT-PCR. Stressed rats exhibited visceral hypersensitivity that was significantly attenuated by TSA. Compared to SHAM controls, methylation of the GR gene was increased following WAS while expression of the GR gene was decreased. Methylation of the CRF promoter was decreased with WAS with a concomitant increase in CRF expression. This study demonstrates the involvement of central epigenetic mechanisms in regulating stress-induced visceral hypersensitivity and provides a foundation for exploring the epigenetic mechanisms that may contribute to IBS-like symptomatology.