Extended Data Fig. 12: Analysis of cortical neurons in the Nfia-cKO mouse line and co-immunoprecipitation with Lhx2 and NPAS3.
From: Inhibitory input directs astrocyte morphogenesis through glial GABABR
a. Antibody staining for BRN2 (Layers II/II), CTIP2 (Layers V), and FOXP2 (Layers VI) layer-specific neuronal markers in the P7 cortex from Nfia-cKO and control; quantification is derived from n = 3 pairs of animals (6 images, GLME model). b. CX-specific transcription factor DEGs increased between P7-P14. c. OB-specific transcription factor DEGs increased between P7-P14. d. Immunoprecipitation of LHX2 and immunoblot of LHX2 and NFIA from the P28 cortex; arrow heads label the proteins of interest. e. Immunoprecipitation of NPAS3 and immunoblot of NPAS3 and SOX9 from P28 cortex; arrow heads label the proteins of interest. f. Quantification of LHX2 expression in virally labelled astrocytes from the P28 cortex of mice where it has been knocked out using guideRNAs in the ROSA-LSL-Cas9-EGFP mouse line; quantification is derived from n = 3 pairs of animals (37, 39 cells; GLME model, ***P = 0.00099). g. Imaging of virally labelled astrocytes from the P28 cortex of Lhx2-cKO mice where Lhx2 has been knocked out using guideRNAs in the ROSA-LSL-Cas9-EGFP mouse line; quantification of morphological complexity via Sholl analysis was derived from n = 3 animals (Lhx2-cKO: 13 mcherry-Cas9-EGFP+ cells, 40 mcherry+Cas9-EGFP+ cells; GLME model with Sidak’s multiple comparisons test, **P = 0.0037; GLME model, **P = 0.006, 0.003). h. RNAscope for Gabbr1 expression in Cas9-EGFP cortical astrocytes lacking Lhx2 and controls at P28; quantitative analysis demonstrating reduction of Gabbr1 expression is derived from n = 3 pairs of animals (51,55 cells; GLME model, *P = 0.015; LME model, **P = 0.0003). Scale bars, 100 μm (a), 20 μm (m). Data represent mean ± s.d. (a, g right), median, minimum value, maximum value and IQR (f, g left, h).
