Figure 3.

Perturbation of vascular and neuronal development in E15 Vegf^ΔTie2-Cre telencephalon. A–F, Reduced PECAM-1⁺ endothelial cells (B, yellow asterisk), laminin⁺ vessels (D, red arrows), and isolectin B4⁺ vessels (F, yellow arrows) in E15 Vegf^ΔTie2-Cre pallium (B, D, F) compared with Vegf^fl/fl pallium (A, C, E). C, E, Orange arrows indicate well-formed vascular network in Vegf^fl/fl pallium. G, H, Ki67 labeling marks the proliferating progenitor pool in Vegf^fl/fl telencephalon (G), whereas Ki67-positive progenitors spanned the entire pallium of Vegf^ΔTie2-Cre embryos (H). Boxed regions were magnified; white asterisks indicate Ki67-negative CP (G), and yellow asterisks indicate abnormal proliferation (H). I, J, MAP2 immunoreactivity reveals normal neuronal differentiation zone of E15 Vegf^fl/fl pallium (I, white asterisks), which is reduced in Vegf^ΔTie2-Cre pallium (J, yellow asterisks). A band of MAP2⁺ cells were abnormally positioned in the Vegf^ΔTie2-Cre IZ-SVZ (J, yellow arrows). K, High-magnification image of MAP2⁺ neurons in the Vegf^ΔTie2-Cre SVZ. L, M, Tbr1 immunoreactivity in E15 Vegf^fl/fl (L) and Vegf^ΔTie2-Cre (M) pallium. Boxed regions are magnified. The uniform arrangement of CP cells in Vegf^fl/fl pallium (L, white asterisk) was altered in Vegf^ΔTie2-Cre pallium with abnormally thicker (M, yellow arrow) and thinner (M, yellow asterisk) cell arrangement in the CP. N, O, RC2 immunostaining reveals continuous radial glial fibers extending from VZ to pial surface in Vegf^fl/fl pallium (N) and shortened radial glial processes in Vegf^ΔTie2-Cre pallium (O, red asterisks). P, Q, GAD65/67 immunoreactivity shows decreased stream of GABA neurons in Vegf^ΔTie2-Cre pallium extending to the medial edge (Q, yellow asterisks) compared with Vegf^fl/fl pallium (P, white asterisks). R–U, Calbindin⁺ and calretinin⁺ neurons were decreased (S, U, yellow asterisks) in E15 Vegf^ΔTie2-Cre pallium compared with Vegf^fl/fl pallium (R, T, white asterisks). Calretinin immunoreactivity was decreased in the hippocampal primodium of Vegf^ΔTie2-Cre embryos (U, yellow arrow). V, W, Uniform calretinin labeling in MZ and SP of Vegf^fl/fl embryos (V) and irregular calretinin labeling outside MZ in Vegf^ΔTie2-Cre telencephalon (W). W, The boxed region was magnified, and yellow arrows point to calretinin⁺ cells outside MZ. X, Y, CHL1 immunostaining reveals axonal tracts in Vegf^fl/fl (X) and Vegf^ΔTie2-Cre (Y) pallium. Y, Yellow arrow points to displaced axonal tract. DAPI (blue) was used to label nuclei (A, B, G–M, P–Y). Scale bars: A (applies also to B, E–J, L, M, P–U, X, Y), 100 μm; C (applies also to D), 75 μm; N (applies to O, V, W), 50 μm; magnified inset in G (applies to magnified insets in H, L, M), 50 μm; W (magnified inset), 25 μm. n = 8.