Multiple origins and modularity in the spatiotemporal emergence of cerebellar astrocyte heterogeneity
Fig 5
(A) Cumulative proportions of intracluster, cell-to-cell NND. Both double (red) and triple (blue and black) clones show a high degree of clustering when their distributions (large symbols) are compared to random distributions (small symbols; P < 0.001). Symbols represent empirical proportions. Continuous lines are the fitted curves. (B,C) Cluster analysis confirms the presence of subclones in both double and triple clones. The 3D reconstructions of a representative triple (B) and double (C) E12-P30 clone highlight the presence of spaced-out subclones composed of BG and GLAs; (B) also contains WMAs. (B’, C’) Higher magnifications of representative subclones (dashed circles) of the clones in (B) and (C). Gray spheres indicate cells of other clones. Plots in (B”) and (C”) show the number (k) and composition (cell type and number of cells/subclone) of subclones identified in the representative triple (B) and double (C) clones, respectively. Each subclone is arbitrarily associated to a number (subclone n.). (D), Analysis of subclone composition reveals that most (>80%) subclones in both double (red) and triple (blue) clones comprise both BG and GLAs. (E-H), BG:GL ratio distributions in subclones comprising both BG and GLAs. In double clones (E, number of subclones = 55), BG prevails over GLAs (mean ratio = 1.772, equivalent to 0.572 log units, P < 0.001). Subclones in triple clones containing WMAs (F, number of subclones = 49) show no predominance of either BG or GLAs (mean ratio = 0.930, −0.073 log units, P = 0.580), even when WMAs are excluded (G, number of subclones = 53; mean ratio = 0.963, −0.038 log units, P = 0.843), whereas subclones containing only BG and GLAs (H, number of subclones = 64) again display a prevalence of BG over GLAs (mean ratio = 1.310, 0.270 log units, P = 0.017). Number of bins = 10. In each plot, the vertical solid line indicates 1:1 ratio, the vertical dotted line indicates the mean ratio, and the light-colored area indicates the 95% bootstrap confidence interval of the mean. P values are computed with Wilcoxon signed rank test against zero. The numerical data used in panels (A,E,F,G,H) are included in S1 Data. A-P, antero-posterior; BG, Bergmann glia; D-V, dorso-ventral; GLA, granular layer astrocyte; HetC, heterogeneous clone; M-L, medio-lateral; NND, Nearest Neighbor Distance; PCL, Purkinje cell layer; WM, white matter; WMA, white matter astrocyte.