Fig. 4: ACC^Glu modulates chronic visceral pain-induced change in food preference.

a Representative images of c-Fos expression following CRD stimulation in the ACC and the total number of c-Fos-expressing neurons (n = 3 mice, p < 0.0001). Scale bar:20 µm. b Representative images showning the coexpression of c-Fos neurons (green) and glutamatergic neurons (red) in the ACC, along with the percentage of coexpressed neurons (n = 3 mice). Scale bar:20 µm. c Schematic representation of the optical fiber photometry assay and representative images of EGFP expression in the ACC. Scale bar:20 µm. d Heatmaps and average calcium activity showing Ca²⁺ signals time-locked to the CRD stimuli in NCI and CON mice. e Area under the curve of Ca²⁺ activity in Glu-positive ACC neurons in CON and NCI mice receiving CRD stimulation (n = 7 mice, p = 0.0435). f Schematic of viral injection (top) and experimental protocol of chemogenetic (bottom) and representative images of mCherry expression in the ACC. Scale bar:20 µm. g Spatial location heatmaps from free-feeding tests. h Colorectal distention (CRD) thresholds before and after hM3Dq activation of ACC^Glu neurons in CON mice (n = 9 mice/group, p < 0.0001). i Percentage of time spent in each food zone and intake of each food type before and after hM3Dq activation of ACC^Glu neurons in CON mice (n = 9 mice/group, Food intake percentage: p < 0.0001, Food zone time: p = 0.0173 in IL and p = 0.0107 in S). j Correlation between the percentage of food zone time and percentage of food intake (p = 0.001, r = 0.5254). k CRD threshold before and after hM4Di inhibition of ACC^Glu neurons in NCI mice (n = 9 mice, p = 0.0005). l The percentage of food zone time and food intake for each food before and after hM4Di inhibition of ACC^Glu neurons in NCI mice (n = 9 mice/group, Food intake percentage: p < 0.0001, Food zone time: p = 0.0172 in IL and p = 0.0356 in S). m Correlation between the percentage of food zone time and percentage of food intake (p < 0.0001, r = 0.6115). All the data are presented as the means ± SEMs. Two-sided student’s t test (a, e). Two-way ANOVA with Sidak’s multiple comparisons test (i, l). Ordinary one-way ANOVA with Dunnett’s multiple comparisons test (h, k). Linear regression and Correlation (j, m). *p < 0.05; **p < 0.01; ***p < 0.001.