Figure 2. DISC1 regulates progenitor cell proliferation in utero.

(A) DISC1 knockdown cells exhibit cell positioning defects in utero. Control or DISC1 shRNA constructs were electroporated into E13 embryonic mouse brains and the mice were sacrificed at E15. The percentage of GFP cells in each region is shown (n=4, p<0.01). Scale bar=20 µm. CP, cortical plate; IZ, intermediate zone; VZ/SVZ, ventricular zone/subventricular zone.

(B) BrdU incorporation is reduced in DISC1 knockdown brains. Brains of mice electroporated at E13 were pulse labeled with BrdU (100 mg/kg) for 2 hours. Arrows indicate GFP and BrdU double positive cells. The bar graph shows the percentage of BrdU and GFP double positive cells to total GFP positive cells in SVZ (n=4, p<0.0005). Scale bar=20 µm.

(C) Mitotic index of DISC1 silenced cells is reduced in utero. The percentage of GFP positive cells that are also pH3 positive in the VZ is shown (n=4, p<0.01). Scale bar=20 µm. Arrowheads indicate GFP and pH3 double positive cells.

(D) DISC1 knockdown in progenitor cells causes premature cell cycle exit in utero. Control or DISC1 shRNA constructs were electroporated into E13 embryonic brains and BrdU was injected at E15. Mice were sacrificed at E16. The cell cycle exit index is measured as the percentage of the GFP-positive cells that exited the cell cycle (GFP+ BrdU+ Ki67−) divided by total GFP and BrdU double positive (GFP+ BrdU+) cells (n=5, p<0.001). Scale bar=20 µm. White arrows indicate GFP+BrdU+Ki67+ cells. Arrowheads indicate GFP+BrdU+Ki67− cells.

(E) BrdU labeling is increased in DISC1 overexpressing cells in utero. Control or WT-DISC1 plasmids were electroporated into embryonic brains at E14 and BrdU was injected 2 hours before sacrificing at E15. The percentage of GFP positive cells in the SVZ that are also BrdU positive cells is shown (n=3, * p<0.05, ***p<0.005). Scale bar=10 µm.