Extended Data Fig. 9: Postnatal developmental defects of cerebral cortex in Tsc1^GFAPcKO mice were rescued in 2cKO mice and by rapamycin treatment.

a, Immunofluorescence of pS6RP, NeuN and DAPI in cortex of Ctrl, Tsc1^GFAPcKO, 2cKO and Fip200^GFAP cKO mice at P21. Inset shows details of pS6RP and NeuN in boxed area. Four independent experiments gave similar results. b,c, Mean ± s.e. of the percentage of pS6RP⁺NeuN⁺ of NeuN⁺ cells (b) and the cell size of NeuN⁺ cells (c) in the cortex of Ctrl, Tsc1^GFAP cKO, 2cKO, and Fip200^GFAP cKO mice at P21. n = 4 independent experiments, at least 300 cells counted. d, Immunofluorescence of GFAP and DAPI in SVZ of Ctrl, Tsc1^GFAPcKO, 2cKO and Fip200^GFAPcKO mice at P21. Four independent experiments gave similar results. e, Mean ± s.e. of the relative percentage of GFAP⁺ cells in the cortex of Ctrl, Tsc1^GFAPcKO and 2cKO mice at P21. n = 4 independent experiments, at least 300 cells counted. f, Immunofluorescence of pS6RP, NeuN and DAPI in the cortex of rapamycin treated Ctrl and Tsc1^GFAP mice at P21. Inset shows details of pS6RP and NeuN in boxed area. The dashed lines indicated the boundaries of cortex and CC. Four independent experiments gave similar results. g, Mean ± s.e. of the percentage of pS6RP⁺NeuN⁺ of NeuN⁺ cells in the cortex of rapamycin treated Ctrl andTsc1^GFAPcKO mice at P21. n = 4 independent experiments, at least 300 cells counted. h, Mean ± s.e. of the brain weight of rapamycin treated Ctrl andTsc1^GFAPcKO mice at P21. n = 3 independent experiments. i, H&E staining of sagittal sectioned brain indicating cortex of Ctrl, Tsc1^GFAPcKO, 2cKO, 2cKI and Fip200^GFAPcKO mice at P21. The sagittal sections for middle of the brain (middle position) and 1 mm lateral to the middle position were stained. The dashed lines indicated the boundaries of cortex and CC. Four independent experiments gave similar results. j, Mean ± s.e. of cortex thickness of Ctrl, Tsc1^GFAPcKO, 2cKO and Fip200^GFAP cKO mice at two different section positions at P21. n = 3 (middle) or 4 (–1 mm) independent experiments. CC, corpus callosum; hippo, hippocampus; I-VI indicates layers of cerebral cortex. Scale bar, 50 μm. Data were analysed by one-way ANOVA with Tukey’s post-hoc test (b,c,e,s,h,j).