Chronic Ultrasound (‘Emotional’) Stress in Mice Exacerbates Colitis Through GSK3β-Mediated Neuroimmune Dysregulation and Is Ameliorated by Probiotic Intervention

Abstract

Background: Growing evidence supports a bidirectional relationship between ‘emotional’ (psychological) stress and intestinal inflammation. ‘Emotional stress’, a form of stress that is triggered by negative information and experiences of non-physical nature, is an established risk factor for the development of subclinical intestinal inflammation, called irritable bowel syndrome (IBS), to inflammatory bowel disease (IBD). However, the underlying mechanisms of this transition remain unclear, owing to the limited availability of appropriate animal models. Glycogen synthase kinase-3β (GSK3β), a molecular hub of ‘emotional stress’ response, as shown particularly in the ultrasound stress (US) model, and an important player in gut inflammation, may underlie this transition. Here, we propose a mouse paradigm for the progression from IBS to IBD-like syndrome by combining the US with dextran sulfate sodium (DSS)-induced colitis paradigm in mice and investigate the role of GSK3β signaling under these conditions. Methods: C57BL/6 mice were subjected to a 21-day US consisting of irradiation of alternating ‘negative’ and ‘neutral’ frequencies, followed by 5 days of 0.5% DSS exposure via drinking water. A standardized anti-inflammatory probiotic VSL#3 whose efficacy was shown in DSS-based models was administered to a subgroup of mice. Behavioral, histological, and molecular outcomes previously established in US and IBS models were evaluated. This included parameters of anxiety, anhedonia, time of intestinal transit, fecal water content, and brain and gut neutrophil infiltration, as well as gene expression of pro-inflammatory cytokines and of GSK3β, and serum levels of pro-inflammatory molecules. Given the important role of the nervus vagus in brain–gut interactions, molecular changes were studied in the nervus vagus as well. Results: Mice exposed to the combined US and DSS challenge had impaired intestinal transit and increased fecal water content, neutrophil infiltration, the upregulation of pro-inflammatory mediators and GSK3β in the gut that together manifested the IBD-like state. In this group, upregulated gene expression of these markers was found in the prefrontal cortex and the hippocampus, as well as in the vagus. The combined US and DSS challenge exacerbated anxiety- and anhedonia-like behaviors and elevated blood cytokine and chemokine levels. In comparison to these changes, solitary exposures to US or DSS exerted smaller effects on the majority of measured parameters. Probiotic treatment attenuated most of the physiological, inflammatory, and behavioral abnormalities induced by the combined US and DSS intervention. Conclusions: The mouse model employed here using both US and DSS recapitulated the progression of IBS to the IBD associated with ‘emotional stress’ and demonstrated upregulated gut and brain Gsk3β expression accompanying this transition. The described model provides a useful paradigm for investigating the mechanisms underlying the disease development from IBS- to IBD-like pathology.