FIG. 3.

Transplantation of mouse-ESC-derived cortical cells into the developing mouse cortex using the TETCaD method. (A) Neural induction of a Sox1-GFP mouse ESC line [22] to a cortical cell fate. The time course of neural induction was 7 (a–e) or 14 days (f–j). DNA (a, f ), Sox1-GFP (b, g and e, j; green), Pax6 (c, h and e, j; red), and Tuj1 (d, i and e, j; blue). Scale bars=30 μm. (B-a) Mouse-ESC-derived cells grafted to the developing mouse cortex (red, CTO positive, arrowheads) using the TETCaD method. (b) Number of grafted ESC-derived cells per brain. Transplantations and cell counting were performed using E12.5, E14.5, and E17.5 mice brains with 20 mM EGTA, and E14.5 without EGTA. The time course of neural induction was 10 days. *P<0.05 and ***P<0.001, t-test. Error bars: SEM. (C) Grafted mouse-ESC-derived cells extend processes in the dorsal cortex. (a–c) A grafted Sox1-GFP-positive cell (arrowhead) with apically (arrow) and basally (double arrowhead) directed processes is recognized with CTO (a) and the expression of Sox1-GFP (b). Scale bar=5 μm. (D) Measurement of the angle of processes relative to the apico-basal axis (dashed line). (a) The angles of processes oriented in the apical (i) and basal (ii) directions were measured as represented. Scale bar=5 μm. (b) Quantified angle of processes extended from grafted cells. The sector is subdivided into bins of 30°, numbered 1, 2, and so on, from the purely apical direction. Neural induction was 7 (blue, n=88) or 14 days (red, n=316). White and yellow dots: the apical surface and the basal pial surface, respectively. Color images available online at www.liebertpub.com/scd