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. 2022 Jan 12;15:757628. doi: 10.3389/fnins.2021.757628

FIGURE 2.

FIGURE 2

Specific activation of the rACCGlu-vlPAG neural circuit leads to hyperalgesia and anxiety-like behaviors in sham mice. (A) Schematic diagram of the experimental process. (B) Schematic diagram of chemical genetic virus injection (left) and representative image of the mCherry-labeled cells in the rACC after virus injection (right). Scale bar, 200 μm. (C) The mCherry (red) and glutamatergic neuron marker VGLUTs 1/2 (green) were co-localized in the rACC. Scale bar, 20 μm. (D) Percentage of mCherry and VGLUTs 1/2 co-localization (n = 15 slices from three mice). (E) Schematic diagram of chemogenetics (left) and representative image of viral expression within the right rACC (right). Scale bar, 100 μm. (F) Representative images of the glutamatergic neurons (red) in the rACC that co-localized with c-Fos. Scale bar, 20 μm. (G) Percentage of co-localization of glutamate neurons and c-Fos in each group (n = 15 slices from three mice, *P < 0.05). (H) Time course of the changes in the PWTs. (I) Time spent in the open arms in the EPM test. (J) There is no significant difference between the entries in the open arms covered by the mice among the three groups. (K) Representative movement trajectory diagram and activity heatmap of the sham + mCherry + CNO, sham + hM3D + saline, and sham + hM3D + CNO groups in the EPM test. (L) Time spent in the center area in the OFT. (M) Center distance covered by the mice in the OFT. (N) There is no significant difference between the total distance covered by the mice in the OFT among the three groups. (O) Representative movement trajectory diagrams and activity heatmaps in the sham + mCherry + CNO, sham + hM3D + saline, and sham + hM3D + CNO groups in the OFT. Data are presented as means ± standard errors of the means. *P < 0.05 vs. the sham + mCherry + CNO group; #P < 0.05 vs. the sham + hM3D + saline group; NS, not significant. n = 7–13 mice per group in (H–N). PWT, paw withdrawal threshold; EPM, elevated plus maze; OFT, open field test; CNO, clozapine-N-oxide; SNI, spared nerve injury.