Extended Data Fig. 6: Rgl3-specific ligands influence different aspects of mesDA neuron development.
From: Distinct radial glia subtypes regulate midbrain dopaminergic neuron development
a, Heatmap showing selected ligands expressed by Rgl cell types in the developing mouse VM. Ligands are color-coded by activity: green for activation, red for inhibition, and yellow for context-dependent modulation. Pathways are indicated on the left. Color intensity is proportional to the Bayesian estimate of expression, with gray scale indicating values below the significance threshold. b, Representative immunofluorescence images of TH+ neurons on day 35 following treatment with indicated ligands or small molecules. Scale bar, 100 µm. c, Quantification of TH+ neurons (TH+/DAPI+) across conditions. CTRL: 7.91 ± 0.39%; WNT5A: 3.83 ± 0.36% (p = 0.049); IWP2: 1.96 ± 0.05% (p = 0.044); Valsartan: 2.26 ± 0.07% (p = 0.044); AngII: 6.85 ± 0.72%; CGP-42112A: 6.24 ± 0.60%; DKK3: 9.0 ± 0.15%; FGF7: 8.07 ± 0.45%; BMP1: 6.28 ± 0.17%; NBL1: 8.63 ± 0.19%; mean ± s.e.m. d,e, Representative immunofluorescence images used to quantify change in mDA neurogenesis (TH−EdU+/DAPI+ arrowheads) following treatment with selected ligands and small molecules. CTRL: 7.67 ± 1.13%; WNT5A: 6.51 ± 2.83%; IWP2: 1.31 ± 0.54% (p = 0.038); Valsartan: 5.89 ± 2.50%; AngII: 6.06 ± 2.91%; mean ± s.e.m. Scale bar, 100 µm. d,f, Representative immunofluorescence images used to quantify change in proliferation (TH+EdU+/DAPI+ cells) following treatment with selected ligands and small molecules. CTRL: 12.3 ± 0.75%; WNT5A: 11.5 ± 1.12%; IWP2: 3.22 ± 0.18% (p = 0.037); Valsartan: 11.7 ± 1.61%; AngII: 15.2% ± 0.62%; mean ± s.e.m. Scale bar, 100 µm. g,h, Representative immunofluorescence images used to quantify reduced cell death (TUNEL+/DAPI+ cells) following treatment with selected ligands and small molecules. CTRL: 10.8 ± 3.17%; WNT5A: 10.1 ± 5.04%; IWP2: 18.9 ± 5.41%; Valsartan: 23.5 ± 2.67% (p = 0.044); AngII: 13.4 ± 4.00%; mean ± s.e.m. Scale bar, 100 µm. Box plots (c,e,f,h) display interquartile range (IQR), median (indicated by line), and whiskers (1.5× IQR) with individual data points shown as a scatterplot. Asterisks indicate statistical significance (*p < 0.05), determined using two-tailed Student’s t-test with Bonferroni–Holm correction from n = 3 independent differentiations, 2 wells per condition.
