Figure 4. Lactobacillus supplementation improves behavior by moderating kynurenine metabolism.

(a) Representation of tryptophan-kynurenine pathway, depicting H₂O₂ inhibition of pathway-initiating enzyme IDO1. (b) Production of H₂O₂ by E. coli and L. reuteri (n = 3 wells per group; representative of 2 independent experiments; two-tailed t-test, ***p < 0.001; mean ± s.e.m.). (c) Production of H₂O₂ by endogenous Lactobacillus species compared to L. reuteri (n = 7–8 colonies per group; one-way ANOVA with Dunnet’s multiple comparisons, ***p < 0.001; mean ± s.e.m.). (d) Fecal H₂O₂ levels in naïve, stressed and L. reuteri treated stressed mice (n = 10 naïve, 11 stressed and 7 stressed + L.reuteri mice per group; one-way ANOVA with Dunnet’s multiple comparisons, *p < 0.05, ***p < 0.001; mean ± s.e.m.). (e) Correlation between Lactobacillus and H₂O₂ levels in naïve and stressed mice (n = 5 mice per group; Spearman r test, **p = 0.01). (f) qRT-PCR quantification of ido1 expression in the intestines of naïve, stressed mice and L. reuteri treated stressed mice, relative to GAPDH. (n = 4 naïve, 7 stressed and stressed + L.reuteri mice per group, 1-way ANOVA followed by Dunnett’s post-hoc, p < 0.01; mean ± s.e.m.). (g) Experimental design of L. reuteri and/or kynurenine administration. (h) Forced swim test quantification of escape behavior of naïve mice treated with L-kynurenine or saline control (n = 10 saline and 8 L-kynurenine mice per group; representative of 2 independent experiments; two-tailed t test with Welch’s correction, *p < 0.05; mean ± s.e.m.). (i) Forced swim test quantification of escape behavior of stressed mice treated with either L. reuteri alone or L. reuteri and L-kynurenine (n = 7 per group; one-way ANOVA followed by Dunnett’s post-hoc, **p < 0.01; mean ± s.e.m.).