Fig. 6: Down-regulation of mGluR5 in ACC astrocytes decreases astrocytic Ca2+ signals in CCI mice. | Communications Biology

Fig. 6: Down-regulation of mGluR5 in ACC astrocytes decreases astrocytic Ca2+ signals in CCI mice.

From: mGluR5-mediated astrocytes hyperactivity in the anterior cingulate cortex contributes to neuropathic pain in male mice

Fig. 6: Down-regulation of mGluR5 in ACC astrocytes decreases astrocytic Ca2+ signals in CCI mice.

a Schematic representation of the experimental design. b Immunohistochemical images of the astrocytes in the ACC of CCI mice. Upper panel: colocalization of mCherry with the astrocytic-specific marker s100β (green) in the ACC (b1, 90.21 ± 4.904%, n = 6 slices from 3 mice); middle and below panel: immunohistochemical staining of astrocyte shRNA (red) and mGluR5 (green) in the ACC. Quantification of mGluR5 fluorescence intensity in shRNA mCherry-expressing astrocytes relative to those expressing scramble-shRNA mCherry (b2, n = 7 slices from 3 scramble-shRNA mice, n = 6 slices from 3 shRNA mice, scramble-shRNA (pink) = 50.10 ± 1.599 a.u., shRNA (blue) = 30.05 ± 7.04 a.u., p = 0.0122, unpaired t-test). c Summary plots illustrating that Ca2+ signals elicited by CHPG are attenuated by expression of shRNA (n = 10 cells from 3 shRNA mice; n = 8 cells from 3 scramble-shRNA mice; Baseline_shRNA (blue circle) = 0.003342 ± 0.002943▵F/F, CHPG_shRNA (blue circle) = 0.07953 ± 0.02701 ▵F/F, baseline_scramble-shRNA (pink circle) = 0.05103 ± 0.1469 ▵F/F, CHPG_scramble-shRNA (pink circle) = 0.3477 ± 0.1157 ▵F/F, p = 0.5003, two-way ANOVA (F (1, 16) = 12.79, p = 0.0025), Fisher’s LSD post hoc test). d Representative images displaying all AQuA-detected events from a 1-minute ex vivo astrocytic GCaMP7b Ca2+ imaging experiment from different groups. Upper: representative image in CCI mice; middle: representative image in shRNA-expressing mice; below: representative imaging in scramble-shRNA-expressing mice; Colors indicate detected events. Scale bar: 10 μm. e Graph depicting the frequency of astrocytic Ca2+ signals in the ACC of CCI, shRNA and scramble-shRNA mice (n = 8 cells from 3 CCI mice; n = 8 cells from 3 shRNA mice; n = 6 cells from 3 scramble-shRNA mice; CCI (red) = 1.324 ± 0.09899 min/100 μm2, shRNA (blue) = 0.5763 ± 0.07242 min/100 μm2, scramble-shRNA (pink) = 1.240 ± 0.2585 min/100 μm2, p = 0.0021, F = 8.690, one-way ANOVA followed by Dunnett’s test). f Graph showing the amplitude of astrocytic Ca2+ signals in the ACC of CCI, shRNA and scramble-shRNA mice (CCI (red) = 1.556 ± 0.2733 ▵F/F, shRNA (blue) = 0.7032 ± 0.09216 ▵F/F, scramble-shRNA (pink) = 1.144 ± 0.06367 ▵F/F, p = 0.0109, F = 5.781, one-way ANOVA followed by Dunnett’s test). Error bars represent the mean ± SEM.

Back to article page