Figure 2.

Microglia are required for tissue integrity at the embryonic CSA and CSB

(A) Hoechst staining of hemibrain coronal sections from E14.5 or E15.5 embryos, showing CSA integrity in controls (open arrowhead) and cavitary lesions in the absence of microglia (solid arrowheads) (n_controls = 27, n_Pu1KO = 6, n_AFS = 4, n_PLX3397 = 8, n_PLX3397-E12 = 11, n_CSF1RFire = 7); and from E18.5 embryos, showing the localization of CSA cavitary lesions, bordering the neocortex (Ncx), striatum (Str), and amygdala (Am) (n_controls = 35, n_Pu1KO = 5, n_AFS = 7, n_PLX3397 = 13, n_PLX3397-E12 = 11, n_CSF1RFire = 6).

(B) L1-immunolabeling stains the corpus callosum (CC) and fornix (Fx), highlighting CSB integrity in controls (open arrowheads) and cavitary lesions (solid arrowheads) in various models disrupting microglial colonization (n_controls = 21, n_Pu1KO = 4, n_AFS = 5, n_PLX3397 = 9, n_PLX3397-E12 = 11, n_CSF1RFire = 5).

(C) Cavity reconstruction (yellow) after whole hemibrain clearing using iDISCO, highlighting the absence of cavities in controls and stereotypically located cavities in two depletion models (n_controls = 4, n_AFS = 3, n_PLX3397 = 4) and enabling 3D quantification of lesion volumes (Imaris Software).

(D) Whole-head MRI scans of E15.5 or P0 mice, showing low-intensity signals confirming the formation of lesions at the CSA of PLX3397-treated embryos (yellow solid arrowheads) in contrast to controls (yellow open arrowheads) (n_controls-E15 = 4, n_PLX3397-E15 = 3, n_controls-P0 = 3, n_PLX3397-P0 = 3).

(E) Whole-head MRI scans of E15.5 or E18.5 mice, showing low-intensity signals confirming the formation of lesions at the CSA of Csf1r^{ΔFIRE/ΔFIRE} embryos (yellow solid arrowheads) in contrast to controls (yellow open arrowheads; n_controls-E15 = 3, n_{CSF1RFire-E15} = 4, n_controls-E18 = 4, n_{CSF1RFire-E18} = 4).

(F) Whole-head MRI scans of E18.5 mice, showing low-intensity signals confirming the formation of lesions at the CSB of Csf1r^{ΔFIRE/ΔFIRE} embryos (yellow solid arrowheads) in contrast to controls (yellow open arrowheads) (n_controls-E18 = 4, n_{CSF1RFire-E18} = 4).

Graphs show means ± SEM. Mann-Whitney U tests were performed for statistical comparison, ^∗∗p < 0.01. Scale bars: 200 μm in (A) and (B); 800 μm in (C); and 1 mm (D)–(F).

Am, amygdala; CC, corpus callosum; CSA, cortico-striato-amygdalar boundary; CSB, cortico-septal boundary; Fx, fornix; Ncx, neocortex; OB, olfactory bulb; Se, Septum; Str, striatum.

See also Figure S2 and Video S1. Lateral rotation of an AFS-treated E16.5 hemibrain cleared by iDISCO, related to Figure 2, Video S2. Lateral rotation of a PLX3397-treated E16.5 hemibrain cleared by iDISCO, related to Figure 2, Video S3. Rostrocaudal progression through an E16.5 control hemibrain cleared by iDISCO, related to Figure 2, Video S4. Rostrocaudal progression through a PLX3397-treated E16.5 hemibrain cleared by iDISCO, related to Figure 2.