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. Author manuscript; available in PMC: 2019 Dec 17.
Published in final edited form as: Nat Neurosci. 2019 Jun 17;22(7):1075–1088. doi: 10.1038/s41593-019-0419-y

Figure 5. CX3CL1 is necessary for TC input engulfment and elimination after sensory lesioning.

Figure 5.

a-b, VGluT2 immunolabeled TC inputs within tangential sections of control and deprived barrel cortices in Cx3cl1+/+ (a) and Cx3cl1−/− mice (b). Scale bar, 150 μm c, Quantification of fluorescence intensity of VGluT2-positive TC input 6 d after deprivation shows a significant decrease in VGluT2 fluorescence intensity in Cx3cl1+/+ mice 6 d post-deprivation, which is blocked in Cx3cl1−/− littermates. Data normalized to the control, non-deprived hemisphere within each animal. (Two-Way ANOVA with Sidak’s post hoc, n = 4 animals per genotype, P <0.0001, t = 28.3, df = 12). d-f, Quantification of high magnification images of synaptic components in the barrel centers 6 d after whisker removal reveals a significant decrease in structural synapses (d, VGluT2 colocalized with Homer; Two-Way ANOVA with Sidak’s post hoc, n = 4 animals per genotype, Cx3cl1+/+ control vs deprived, P <0.0001, t =11.66, df = 12) and VGluT2-positive presynaptic terminals (e; Two-Way ANOVA with Sidak’s post hoc, n = 4 animals per genotype, Cx3cl1+/+ control vs deprived, P <0.0001, t = 7.418, df = 12) in Cx3cl1+/+ mice but no significant change in Cx3cl1−/− littermates (Colocalized Area, P = 0.0642, t = 2.415, df = 12; VGlut2 Area, P = 0.1071, t =2.125, df = 12). There was no significant change in density of homer immunoreactivity in Cx3cl1+/+ or Cx3cl1−/− mice following whisker deprivation (f; Two-Way ANOVA with Sidak’s post hoc, n = 4 animals per genotype, no significance). Data normalized to the control, non-deprived hemisphere within each animal. g-h, Representative microglia from the deprived barrel cortex of Cx3cl1+/+ (g) and Cx3cl1−/− (h) mice. Left panel displays raw fluorescent image with microglia (Anti-Iba1, green) VGluT2 inputs (red) and lysosomes (Anti-CD68, blue) labeled. Right panels shows 3D-surface rendering of these cells. Engulfed VGluT2 (red) immunoreactive TC inputs within microglia are visualized in Cx3cl1+/+ microglia (g, arrows)) but not Cx3cl1−/− microglia (h). Scale bars, 5 μm. i, Quantification of VGlut2 engulfment 24 h after whisker removal reveals that Cx3cl1−/− microglia fail to engulf TC inputs following sensory deprivation. Data normalized to engulfment in the control hemisphere within each animal. (Two-Way ANOVA with Sidak’s post hoc, n = 4 littermates per genotype, Cx3cl1+/+ control vs deprived P = 0.0111, t = 3.369, df = 12; Cx3cl1−/− control vs deprived P = 0.5963, t = 0.9422, df = 12). All data presented as mean ± SEM.