csf1rb is required for HSC-derived microglia development. (A) Scheme of the transgenic lines used to discriminate primitive from definitive microglia in adult zebrafish brains. (B-M) Immunofluorescence on transverse brain sections from Tg(kdrl:Cre; ßactin:Switch-DsRed; mpeg1:EGFP) triple transgenic adult wild-type (B,F,J; n=7), csf1ra−/− (C,G,K; n=6) csf1rb−/− (D,H,L; n=7) and csf1rDM (E,I,M; n=4) fish. GFP (left panels), DsRed (middle panels) and merge of both fluorescence channels (right panels) are shown. (N,O) Quantification of microglia density (GFP+ cells/µm2) (N) and percentage of DsRed+ adult microglia among the whole GFP+ population (O) in each genotype. Each symbol represents a single fish and data are mean±s.e.m. Significances between groups were calculated using the Kruskal–Wallis test with Dunn's post hoc multiple comparisons test [H=19.21 (N); H=19.98 (O)]. *P<0.05, ***P<0.0005. (P,Q) Percentage of GFP+ DsRed+ adult microglia (P) and microglia density (cells/mm3) (Q) at 21, 28, 35 and 50 dpf for each genotype. Each point represents mean±s.e.m. (n=5 individual fish). MG, microglia. Scale bars: 50 µm.
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