Fig 2: pGBP reduces the excitability of neurons in intermediate gray matter below the level of SCI(A and A’) Representative grayscale (top row) and inverted binarized (A’) images of FosB-immunostained T8 spinal cord sections. Scale bar, 200 µm.(B) Basal levels of FosB+ neurons in laminae I–IV were unchanged after SCI with or without pGBP.(C) In laminae V–X, SCI + saline mice had significantly more FosB+ neurons relative to sham + saline mice; this effect was blocked by pGBP.(B and C) Kruskal-Wallis one-way ANOVA with Dunn’s multiple comparisons, n = 8 mice/group, ap < 0.0001 versus sham + saline, *p < 0.05 for SCI + saline versus SCI + GBP, mean ± SEM.

Fig 3: Effects of nonretrieval intra-AI BLF on the protein levels of ?FosB using Western blot and immunofluorescence analyses. (A) Western blot analysis of the relative protein content of ?FosB and FosB in different groups. (B, C) Semiquantitative analysis of the relative levels of ?FosB and FosB by densitometric analysis (n = 5 mice per group, one-way ANOVA). (D) Semiquantitative analysis of the relative levels of ?FosB by densitometric analysis in the NR + BLF and NR + Veh groups (n = 5 per group). (E) Representative immunofluorescence images of ?FosB in AI from the NR + BLF and NR + Veh groups in the left panel. The corresponding scope of the magnified images right is marked by red squares. Data are presented as the mean ± SEM analyzed by one-way ANOVA. ** represents p < 0.01, compared with the saline group. ns represents no significant difference.

Fig 4: Effects of intra-AI 0.06 nmol/side BLF and 0.2 µl/side vehicle on the conditioned response 6 h after reward memory retrieval. (A) Timeline of the experimental procedure. (B, C) Intra-AI BLF 6 h after reward memory retrieval has no effect on the conditioned response and locomotor activities. (D) Western blot analysis of the relative protein content of ?FosB and FosB in different groups. (E, F) Semiquantitative analysis of the relative levels of ?FosB and FosB by densitometric analysis (n = 5 mice per group, one-way ANOVA). Data are presented as the mean ± SEM. 6hR + BLF = Intra-AI 0.06 nmol/ side BLF 6 h after retrieval; 6hR + Veh = Intra-AI 0.2 µl/side 0.9% sterile saline 6 h after retrieval. ** represents p < 0.01. ns represents no significant difference.

Fig 5: Stress effects on FosB in the hippocampus are microbe dependent. Brain hippocampal FosB and IBA-1 expression were analyzed by immunohistochemistry following behavior testing. Representative sagittal brain hippocampal sections from experimental groups stained with either (A) FosB or (C) Iba1. Stress-exposed mice had increased (B) FosB expression in the hippocampus when compared to control mice (*p < 0.05 vs all other groups). The FosB expression increase was ameliorated in stressed mice gavaged with antibiotics (stress x antibiotic interaction, p < 0.05). Similar to FosB, (D) Iba1 expression was increased in stress-exposed mice gavaged with water when compared to all other groups (*p < 0.05 vs all other groups). n = 5–6 per group from 2 replicate experiments.