Fig. 5

Effect of Grm3 genetic deletion in vivo and knock-down in vitro on microglial reactivity. a, b Iba1⁺ cells (red)/DAPI (blue) within Cingular white matter of P4 WT and Grm3^−/− mice under basal conditions and after IL-1β injections. Representative photomicrographs at × 10 magnification are shown in a (scale bar = 50 μm). Data (mean ± SEM) are expressed as number of Iba1⁺/mm² in b. Two-way ANOVA followed by the Newman-Keuls multiple comparison; ^$p < 0.05, ^$$p < 0.01 effect of IL-1β + INFγ; ^#p < 0.05, ^##p < 0.01 effect of Grm3^−/−. c, d Iba1⁺ cells within Cingular white matter of P4 WT and Grm3^−/− mice under basal conditions and after IL-1β injections. Representative photomicrographs at × 40 magnification are shown in c (scale bar = 25 μm). The cell area, cell perimeter, and cell circularity were assessed in d. Data (mean ± SEM). Two-way ANOVA followed by the Newman-Keuls multiple comparison; ^$$p < 0.01, ^$$$p < 0.001 effect of IL-1β + INFγ; ^#p < 0.05, ^##p < 0.01, ^###p < 0.001 effect of Grm3^−/−. e, f mRNA expression of pro-inflammatory (e) and anti-inflammatory/immune-regulatory (f) markers in Prefrontal/Cingular cortex isolated from P4 WT and Grm3^−/− mice under basal conditions and after IL-1β injections. Data (mean ± SEM) are relative to the mRNA expression of WT mice. Two-way ANOVA followed by the Newman-Keuls multiple comparison; *p < 0.05, **p < 0.01, ***p < 0.01 effect of IL-1β + INFγ; ^#p < 0.05, ^##p < 0.01, ^###p < 0.001 effect of Grm3^−/−. g Grm3 mRNA expression in primary microglia cultures 48 h after transfection with Grm3 siRNA (25 nM) and with Grm3 siRNA (50 nM). Data (mean ± SEM) are relative to Grm3 expression in the scramble group (SCR). Unpaired t test; ****p < 0.0001. h mGlu3 receptor protein expression in primary microglia cultures 48 h after transfection with Grm3 siRNA (50 nM). Data (mean ± SEM) are relative to mGlu3 receptor protein expression in the scramble group (SCR). Three wells per condition pooled. Unpaired t test; **p < 0.01. i, j mRNA expression of pro-inflammatory (i) and anti-inflammatory/immune-regulatory (j) markers in response to SCR and Grm3 siRNA (50 nM). Data (mean ± SEM) are relative to the mRNA expression for the respective SCR. Unpaired t test; *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001