Figure 3.

Analysis of 3D Asteroid Cocultures

(A) Bright-field examples and illustration of asteroids in the absence (left panel) or presence of hAstros with (right panel) or without induction of NGN2.

(B) Progenitor cells, as rosette structures, are not present with NGN2 induction (right panel).

(C) After 35 days, hAstros are dispersed throughout the asteroids.

(D) mGFP reveals the fine-membrane processes emanating from GFAP-positive hAstro branches.

(E) The addition of hAstros significantly increases the number of opposed pre- and post-synaptic markers compared with iNeurons without hAstros; the combination of iNeurons with hAstros increases synaptic density to a further extent. ^∗p < 0.05; ^∗∗p < 0.01.

(F) Costello syndrome-model hAstros (HRAS^G12S) significantly increase synaptic density within asteroids compared with wild-type control (HRAS^WT) and no hAstro addition. Insets are magnified examples of synapses, including perisynaptic astrocyte processes. ^∗∗p < 0.01; ^∗∗∗p < 0.001.

(E and F) Data obtained by random sampling of 3–4 different spheres from each of 3 different independent biological replicates.

Error bars represent SEM. Scale bar, 50 μm.