Extended Data Fig. 8: AMPK activation stimulated autophagy and lipophagy was required in Tsc-deficient cells. Postnatal developmental defects of cerebral cortex in Tsc1^GFAPcKO mice were rescued in 2cKO mice.

a, Mean ± s.e. of the number of GFAP⁺Sox2⁺ NSCs in SVZof Ctrl mice treated with CQ+2DG and Tsc1^GFAPcKO mice treated with vehicle, 2DG, CQ and CQ + 2DG. n = 6 independent experiments. b, Mean ± s.e. of the number of GFAP⁺Nestin⁺BrdU⁺ cell of total BrdU⁺ cells (b) in SVZ of Ctrl mice treated with CQ+2DG and Tsc1^GFAPcKO mice treated with vehicle, 2DG, CQ and CQ + 2DG. n = 6 independent experiments. c, Mean ± s.e. of the number of GFAP⁺Sox2⁺ NSCs in SVZ of Tsc1^GFAPcKO mice treated with vehicle, 2DG, ETO, Rano, TMZ, ETO + 2DG, Rano + 2DG and TMZ + 2DG are shown. n = 5 independent experiments. d, Mean ± s.e. of the number of GFAP⁺Nestin⁺BrdU⁺ cells of total BrdU⁺ cells in SVZ of Ctrl and Tsc1^GFAP cKO mice treated with vehicle, 2DG, ETO, Rano, TMZ, ETO + 2DG, Rano + 2DG and TMZ + 2DG. n = 5 independent experiments. e,f, Mean ± s.e. of the number of DCX⁺ cells (e) and NeuN⁺ cells (f) in SVZ of Ctrl and Tsc1^GFAPcKO mice treated with vehicle, 2DG, ETO, Rano, TMZ, ETO+2DG, Rano+2DG and TMZ+2DG. n = 5 independent experiments. g, Lysates from WT and Tsc2 KO MEFs in normal or glucose-free media were examined by immunoblots with antibodies as indicated. Three independent experiments gave similar results. h, Lysates from WT MEFs, AMPKα1 KO MEFs (2 independent clones), Tsc2 KO MEFs and 2KO MEFs (2 independent clones) were examined by immunoblots with antibodies, as indicated. Three independent experiments gave similar results. i, Lysates were extracted from WT MEFs, AMPKα1 KO MEFs, Tsc2 KO MEFs and 2KO MEFs in glucose-free media with or without BafA1 for 2 h. The levels of LC3-II and vinculin were examined. Three independent experiments gave similar results. j, H&E staining of sagittal sectioned brain indicating hydrocephaly of Tsc1^GFAPcKO lateral ventricle, but not in Ctrl, 2cKO, 2cKI and Fip200^GFAP cKO mice at P21. Arrows indicated SEN-like structures in Tsc1^GFAPcKO brain. Three independent experiments gave similar results. k, Mean ± s.e. of brain weight of Ctrl, Tsc1^GFAPcKO, 2cKO and Fip200^GFAP cKO mice at P7, P14 and P21. n = 3 independent experiments. l, H&E staining of sagittal sectioned brain indicating thicker cortex of Tsc1^GFAPcKO brain, but not in Ctrl, 2cKO, 2cKI and Fip200^GFAP cKO 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. m–o, Mean ± s.e. of cortex thickness of Ctrl, Tsc1^GFAPcKO, 2cKO and Fip200^GFAPcKO mice at two different section positions (m,n) at P7, P14, and P21, and P21 2cKI (o) mice at middle position. n = 4 (m,n) and 5 (o) independent experiments. p–r, Mean ± s.e. of cortex cell size (p) and cortex cell number (q) of Ctrl, Tsc1^GFAPcKO, 2cKO and Fip200^GFAP cKO mice by H&E staining at P21, and cortex TUNEL-positive cell number (r) at P7 and P21. n = 4 (p, q) and 3 (r) 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 (a–f,o–r) or one-way ANOVA (k,m,n,r).