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. Author manuscript; available in PMC: 2015 Apr 16.
Published in final edited form as: Neuron. 2014 Apr 16;82(2):380–397. doi: 10.1016/j.neuron.2014.02.040

Figure 4. Rapid repopulation of the microglia-depleted brain with new cells that differentiate into microglia.

Figure 4

A) To explore microglia homeostasis in the adult brain, 18 month-old wild-type mice were treated with PLX3397 for 28 days to deplete microglia. The inhibitor was withdrawn and groups of mice sacrificed immediately and at 3, 7, 14, and 21 days later (n = 4–5 per group). B–G) IBA1 immunostaining revealed microglia throughout the untreated (control) brains (B) and elimination of microglia in mice treated with PLX3397 (C). New IBA1+ cells appeared throughout the CNS at the 3-day recovery timepoint with very different morphologies to control resident microglia (D). Cell numbers increased by the 7-day recovery timepoint and the morphology of the cells begin to resemble a more ramified state (E). By 14 (F) and 21 (G) days of recovery, the IBA1+ cells resemble ramified microglia and have fully repopulated the entire CNS. H) Quantification of the number of IBA1+ cells in the hippocampal field. I) Analysis of cell body size shows that IBA1+ cells at the 3-day recovery timepoint are much larger than resident microglia. The size of these cells then normalizes over the following recovery timepoints. J) Representative IBA1+ cells from each of the groups, showing the changes in cell morphology and size that occur during repopulation. K) Cells at the 3-day recovery timepoint express a number of unique markers, including CD45, nestin, Ki67, CD34, and c-kit. They also show high immunoreactivity to the IB4 lectin. Notably, IBA1+ cells in control brains and surviving IBA1+ cells in the 0-day recovery group are negative for all these markers. Likewise cells at the 7-, 14-, and 21-day recovery timepoints are also negative for all these markers, highlighting that the initial repopulating cells have a unique morphology and phenotype. Same capital letters above conditions indicates no significant difference (p>0.05) via one-way ANOVA with post-hoc Newman-Keuls Multiple Comparison Test. Error bars indicate SEM. Scale bars represents 20 μM.