Fig. 2: Macrophages/microglia are required for angiogenesis in the ventricular zone of the GEs.

a. Live imaging of Cx3cr1^+/GFP macrophages/microglia with nascent vasculature in the GEs. (i) Schematic diagrams of imaging setup for E12.5 LGE in Cx3cr1^+/GFP mice. The embryo remains attached to placenta while bathed in artificial cerebrospinal fluid (aCSF). (ii) Max projection of an in vivo two-photon image with Cx3cr1^+/GFP cells in LGE associated with blood vessels illuminated with Texas Red dextran. (iii) Still frames of timelapse images of three highlighted Cx3cr1^+/GFP cells, with no. 1 extending the process into the blood vessel and taking up dextran (white arrow), no. 2 rolling within the blood vessel before releasing into the circulation and no. 3 moving along the surface of a blood vessel. (iv) Extravasation of a Cx3cr1^+/GFP cell between the lumen and abluminal side. Orthogonal views show the macrophage against one vessel wall on the luminal side at time 0 min and against the vessel wall on the abluminal side at 20:26 min. b, IEM using IBA1 antibody shows IBA1⁺ macrophages and microglia directly attached to the endothelial cells in MGE of E12.5 mouse brain. This was performed in three biological replicates. c,e,g, Loss of CSF1R leads to complete ablation of IBA1⁺ cells in GE (c), VZ/SVZ of the pallium (e) and the cortical plate (CP) (g). The red boxes in the schematic diagrams show the regions captured in confocal images. d,f,h, Quantification of densities of IBA⁺ cells and IB4⁺ blood vessels in the VZ of GE (d), the SVZ/VZ of pallium (f) and the CP (h). The dashed lines indicate the regions in which blood vessel quantifications are performed. Statistics in d, f and h use a two-tailed, unpaired Student’s t-test, and the data represent the mean ± standard error of the mean. n.s., not significant. n indicates the number of independent biological samples used for quantification.