Figure 4Micrograph of coronal sections of E17 mouse cerebral cortices electroporated at E14 with GFP-miR-neg vector (CTRL) and GFP-miRNA-Cstb (KD), analyzed 3 dpe and immunostained with GFP. Ventricle (V) and cortical plate (CP) are indicated. The dashed lines represent the apical surface of the ventricles. Distribution of electroporated GFP+ cells in the mouse cortex. The cortex was subdivided in 5 equal Bins -Bin1 corresponded to apical side and Bin5 to pial side of the cortex and GFP+ cells calculated in each Bin. Nuclei (blue) are stained with DAPI. Micrograph of coronal sections of E17 mouse cerebral cortices electroporated at E14 with GFP-empty vector control and GFP-miRNA-Cstb, analyzed 3 dpe and immunostained with GFP and Ki67. Ventricle (V) is indicated. The dashed lines represent the apical surface of the ventricles. Quantification of the total number of proliferating Ki67+ cells/area (µm2) of ventricular structures transfected with GFP-miR-neg vector or GFP-miRNA- Cstb in (C). Nuclei (blue) are stained with DAPI. Micrograph sections of d40 hCOs electroporated with GFP-CSTB expressing vector or GFP-R68X mutant and analyzed 5 dpe. Sections were immunostained for GFP and Ki67. The dashed lines represent the apical surface of the cavities/ventricles (V-like). The cortical plate like side is indicated (CP-like). Quantification of the total number of proliferating KI67+ cells/area(µm2)(F) and Ki67+GFP+ cells/area(µm2)(G) of ventricles like structures transfected with GFP-empty vector control, GFP-CSTB or GFP-R68X mutant in (E). Micrograph of coronal sections of E16 mouse cerebral cortices electroporated at E14 with GFP-empty vector control, GFP-Cstb or GFP-R68X, analyzed 2 dpe and immunostained with GFP and Phospho-Histone H3 (PH3). Ventricle (V) and cortical plate (CP) are indicated. The dashed lines represent the apical surface of the ventricles. Distribution of electroporated GFP+ cells in the mouse cortex. The cortex was subdivided in 5 equal Bins -Bin1 corresponded to apical side and Bin5 to pial side of the cortex and GFP+ cells calculated in each Bin. Distribution of mitotic progenitor PH3+ cells in the mouse cortex. The cortex was subdivided in 5 equal Bins -Bin1 corresponded to apical side and Bin5 to pial side of the cortex and PH3+ cells calculated in each Bin. K.Quantification of the total number of proliferating PH3+ cells/area(µm2) of ventricular structures transfected with GFP-empty vector control, GFP-Cstb or GFP-R68X mutant in (H). Micrograph of coronal sections of E16 mouse cerebral cortices electroporated at E14 with GFP-empty vector control, GFP-Cstb or GFP-R68X mutant, analyzed 2 dpe and immunostained with GFP and Tbr2. Ventricle (V) is indicated. The dashed lines represent the apical surface of the ventricles. Quantification of the total number of Tbr2+ cells/ cortical area (µm2) in (L). Data shown as Z-scores relative to the mean of GFP control vector. Micrograph of coronal sections of E17 mouse cerebral cortices electroporated at E14 co-electroporated with mCherry expressing vector and HA-Cstb or R68X, and analyzed 3 dpe. Immunostaining with RFP to identify electroporated cells and GFP to identify migrating interneurons in the GAD65-GFP transgenic mouse line. Ventricle (V) is indicated. The dashed lines represent the apical surface of the ventricles. Quantification of the total number of GAD65-GFP interneurons/ cortical area (µm2) in (N). Data shown as Z-scores relative to the mean of GFP control vector. Nuclei (blue) are stained with DAPI.