Figure 2: Transient exposure of dorsalizing factors can induce patterned dorsomedial telencephalic tissues.

(a) Schematic of mouse medial pallium and neighbouring tissues at E12.5–13.5. (b) Schematic of condition to induce medial pallium tissues. (c) Bright-field view of aggregates cultured in condition 3 on day 36. (d,e) By shortening the period (day 18–21), the aggregates expressed Foxg1::Venus with patterning into Foxg1::Venus⁺ (arrow) and Venus⁻ (arrowhead) NE domains. (f) Histogram of percentage of Foxg1::Venus⁻ protrusion and Foxg1::Venus⁺ main body induction. Patterning of Foxg1::Venus⁺/Venus⁻ NE domains were induced in around 70–80% of aggregates (bars in graph, s.e.m.). (g–l) The Venus⁻ NE domains of aggregates (g) showed TTR⁺/Lmx1a⁺ in distal parts (h,i) and Lmx1a⁺/TTR⁻ in proximal parts (j–l, box in h). (m–o) In hESC-derived NE, Foxg1::Venus⁺ main bodies expressed Lef1 (m) and Lhx2 (n). (o) On the basal side, the thick cell layer of the aggregate expressed Tbr1 and Lhx5. (p) Foxg1::Venus and Lef1 co-expressed in continuous NE (p,q, box in h), suggesting medial pallium-like NE generation. (r) Schematic of hESC-derived dorsomedial telencephalic tissues. Choroid plexus-, hem- and medial pallium-like regions were continuously generated as seen in vivo. (s) Schematic of method to examine Wnts/BMPs expression in Venus⁻ protrusions. The aggregates were cut into Venus⁻ protrusions and Venus⁺ main bodies at day 35–40, and wnts/bmps expression were examined using RT–qPCR. (t) qPCR for genes expressed in Venus⁻ protrusion versus Venus⁺ main body (**P<0.01; ***P<0.001, n=3, unpaired t-test). foxg1 was significantly attenuated in Venus⁻ protrusions, and a significant increase in lmx1a/ttr mRNA expression in Venus⁻ protrusions was confirmed. wnt 2b/3a/5a and bmp 4/6 significantly increased in Venus⁻ protrusions. (u) Schematic summary of conditions examined in Figs 1 and 2. Scale bars, 500 μm (c–e); 200 μm (g–p). Bars in graph, s.e.m. Nuclear counter staining (blue), 4,6-diamidino-2-phenylindole (DAPI).