Transient colitis leads to long-term postinflammatory GI dysmotility and structural remodeling of the myenteric plexus. (A) Experimental design of transient 3×DSS colitis (3.5% DSS in drinking water). (B) Percentage of body weight change in 3×DSS- or water-treated WT mice (n = 10 per group). (C) Fecal lipocalin-2 in 3×DSS- or water-treated WT mice, measured by ELISA (control, n = 4; DSS, n = 8). (D) Percentage of total viable cells (left) and absolute cell counts (right) of colonic mucosal macrophages in 3×DSS- or water-treated mice, quantified by FC (control, n = 5–7; DSS, n = 4–7). (E) Percentage of total viable cells (left) and absolute cell counts (right) of colonic mucosal neutrophils in 3×DSS- or water-treated mice, quantified by FC (control, n = 5–7; DSS, n = 4–7). (F) GI transit in 3×DSS- or water-treated mice at week 1 and week 10 after starting DSS (control, n = 10; DSS, n = 9–10). (G) Confocal images of colonic myenteric plexus isolated from 3×DSS- or water-treated mice and stained with Hu (neuronal somas), βIII-tubulin (nerve fibers), and MHCII (MMs, channel turned off) antibodies. Images are representative of control (n = 5–10) and DSS (n = 4–13) for each time point. Scale bars, 100 μm. (H–J) Changes in colonic myenteric ganglia displayed in G of mice described in A showing (H) total counts of Hu⁺ neurons, (I) counts of neuron clusters, and (J) counts of Hu⁺ neurons per cluster, quantified by confocal microscopy. Data are shown as the mean value per mouse (left, control, n = 5–10; DSS, n = 4–13) or as distribution of pooled values generated from analysis of 4–9 fields of view per mouse (right). (K and L) Changes in nerve fiber architecture of colonic myenteric plexus displayed in G of mice described in A showing (K) counts of IGFT regions and (L) area of IGFT regions, quantified by confocal microscopy. Highlighted area in G is an example of IGFT regions identified as spaces between Hu⁺ myenteric ganglia and βIII-tubulin⁺ fiber tracts (see Materials and methods). Data are shown as the mean value per mouse (left, control, n = 5–10; DSS, n = 4–13) or as distribution of pooled values generated from analysis of 4–9 fields of view per mouse (right). All graphs show the mean ± SEM, n—mice. Data are representative of at least three independent experiments with similar results. Statistical analyses: unpaired Student’s t test (F), multiple t tests (C–E), and two-way ANOVA for multiple comparisons (H–L), *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, and ****P ≤ 0.0001. # above pooled data in violin plots (right, H–L) indicates empirical P values calculated by the Nested Leave-One-Out Jackknife strategy, where, ###P < 0.001. ANOVA, analysis of variance; FC, flow cytometry.