Behavioral and Oxidative Stress Changes in Mice Subjected to Combinations of Multiple Stressors Relevant to Irritable Bowel Syndrome

Abstract

Background and Objectives: Irritable bowel syndrome (IBS) is a well-known functional gastrointestinal (GI) disorder exhibiting a wide range of symptoms due to individual variability and multifactorial etiology. Stress exposure is a major risk factor for the development of IBS. Here, we investigate the differential effects of psychological stress exposures on behavior and oxidative status in mice by using increasingly complex combinations of etiologic IBS-relevant stressors (maternal separation and chronic unpredictable mild stress combinations). Materials and Methods: Mice were subjected to three different combinations of psychological stress factors and subsequent behavioral cognitive and affective parameters and oxidative status markers (superoxide dismutase and glutathione peroxidase antioxidant activity and malondialdehyde level) in the brain and bowel tissues of the animals were analyzed. Results: GI transit modifications reflected by decreased fecal output, cognitive and affective behavioral deficits were observed in all stress exposed groups, but were more evident for the more complex combinations of stressors. Behavioral deficits were accompanied by mild oxidative stress occurring in the bowel and to a greater extent in brain tissue. Conclusions: The presented data depict the effect of various associations in mimicking IBS symptoms and comorbidities and suggest that an all-inclusive combination of early and adult-life psychological stressors is more effective in IBS symptoms modulation. Oxidative stress in both brain and bowel, suggestive for brain-gut molecular connectivity, may play an important role in IBS mechanistic.

Keywords: behavioral tasks; chronic unpredictable mild stress; irritable bowel syndrome; mice; neonatal maternal separation; oxidative stress; restraint stress.

Figure 1

Experimental design of maternal separation and chronic unpredictable mild stress combination in a complex IBS mice model (PD1-PD111 = postnatal days 1-111; U.M.S = unpredictable mild stressors).

Figure 2

The effect of various combinations of stress factors on gastrointestinal tract habits, as given by fecal pellets count per 24 h. The values are mean ± S.E.M (n = 10 per group, ** p < 0.01, *** p < 0.001, MS = maternal separation, MF = multifactorial stress.

Figure 3

The effects of various combinations of stress factors in the Y-maze test, as showed by the spontaneous alternation parameter. The values are mean ± S.E.M (n = 10 per group, * p < 0.05 vs. control, MS = maternal separation, MF = multifactorial stress.

Figure 4

The effects of various combinations of stress factors on the parameters evaluated in elevated plus maze: (a) open arms entries, (b) open arms time, (c) closed arms entries and (d) grooming time. The values are mean ± S.E.M (n = 10 per group, * p < 0.05 vs. control, MS = maternal separation, MF = multifactorial stress.

Figure 5

Effect of various combinations of stress factors on the parameters evaluated in the forced swim test: (a) swimming, (b) floating and (c) struggling time (seconds). The values are mean ± S.E.M (n = 10 per group, * p < 0.05 vs. control and # p < 0.05 vs. MS group (MS = maternal separation, MF = multifactorial stress).

Figure 6

The effect of various combinations of stress factors on oxidative stress markers in the mice brain: (a) SOD activity, (b) GPx activity and (c) MDA levels. The values are mean ± S.E.M. (n = 10 per group, * p < 0.05, and ** p < 0.01, MS = maternal separation, MF = multifactorial stress).

Figure 7

The effect of various combinations of stress factors on oxidative stress markers in the mice bowel tissue: (a) SOD activity, (b) GPx activity and (c) MDA levels. The values are mean ± S.E.M. (n = 3 per group), * p < 0.05 vs. control, MS = maternal separation, MF = multifactorial stress.

Figure 7

The effect of various combinations of stress factors on oxidative stress markers in the mice bowel tissue: (a) SOD activity, (b) GPx activity and (c) MDA levels. The values are mean ± S.E.M. (n = 3 per group), * p < 0.05 vs. control, MS = maternal separation, MF = multifactorial stress.

Figure 8

Statistical correlations between behavioral parameters: (a) swimming vs. frequency of open arms entries, (b) floating duration vs. frequency of open arms entries, (c) swimming vs. open arms time (n = 40, OAE = open arms entries, OAT = open arms time).

Figure 9

Statistical correlations between behavioral parameters: (a) floating duration vs. MDA brain levels, (b) open arms time vs. MDA brain levels, (c) floating vs. brain GPx activity, (d) spontaneous alternation vs. brain SOD activity, (e) open arms entries vs. brain GPx activity, (f) floating and bowel MDA levels (n = 40, OAT = open arms time, OAE = open arms entries, MDA-B = MDA brain level, SOD-B = brain activity of SOD, GPx-B = activity of brain GPx, MDA-C = colon MDA level).

Figure 10

Statistical correlations between oxidative stress parameters in brain and colon: (a) brain GPx activity vs. brain MDA levels, (b) bowel GPx activity vs. bowel MDA levels, (c) brain GPx activity vs. bowel MDA levels, (d) brain GPx and bowel GPx activity, (e) brain MDA and bowel MDA levels (n = 40, MDA-B = MDA brain level, GPx-B = brain GPx activity, GPx-C = colon GPx activity, MDA-C = colon MDA level).