Objectives: Polyethylene Glycol 3350 (PEG3350) is a laxative commonly used to treat constipation in children. The Food and Drug Administration has received reports of increased anxiety, aggression, and obsessive-compulsive behaviors in children administered PEG3350. Thus, we assessed whether daily administration of PEG3350 leads to anxiety-like behavior in mice. Methods: Outbred CD-1 IGS mice were administered either a high or a low dose of PEG3350 via daily oral gavage for 2 weeks. As a laxative comparison and control, additional mice were given a high or low dose of magnesium citrate or vehicle (water). Weight and stool consistency were assessed after each gavage to determine laxative effectiveness. Anxiety-like behaviors were assessed using light/dark, open field, and elevated plus maze (EPM) tests at baseline, after 2 weeks of daily gavage, and after a 2 week washout in experiment 1, and after 2 weeks of daily gavage in experiment 2. Stool samples were collected for microbiome analysis in experiment 2 at baseline, after 2 weeks of daily gavage, and after 2 weeks washout. Results: PEG3350 and magnesium citrate significantly changed stool consistency, as well as microbiome alpha and beta diversity. Anxietylike behaviors were not, however, different in mice administered low or high doses of PEG3350 or magnesium citrate. Conclusions: Although changes in stool consistency and the gut microbiome occurred, administration of PEG3350 did not alter anxiety-like behaviors.

Stressor exposure increases colonic inflammation. Because inflammation leads to anxiety-like behavior, we tested whether stressor exposure in mice recovering from dextran-sulfate-sodium (DSS)-induced colitis enhances anxiety-like behavior. Mice received 2% DSS for five consecutive days prior to being exposed to a social-disruption (SDR) stressor (or being left undisturbed). After stressor exposure, their behavior was tested and colitis was assessed via histopathology and via inflammatory-cytokine measurement in the serum and colon. Cytokine and chemokine mRNA levels in the colon, mesenteric lymph nodes (MLNs), hippocampus, and amygdala were measured with RT-PCR. SDR increased anxiety-like behaviors, which correlated with serum and hippocampal IL-17A. The stressor also reduced IL-1β, CCL2, and iNOS in the colonic tissue, but increased iNOS, IFNγ, IL-17A, and TNFα in the MLNs. A network analysis indicated that reductions in colonic iNOS were related to elevated MLN iNOS and IFNγ. These inflammatory markers were related to serum and hippocampal IL-17A and associated with anxiety-like behavior. Our data suggest that iNOS may protect against extra-colonic inflammation, and when suppressed during stress it is associated with elevated MLN IFNγ, which may coordinate gut-to-brain inflammation. Our data point to hippocampal IL-17A as a key correlate of anxiety-like behavior.