Fig. 4. Chemotaxis in MGE-derived cells requires leading process branching. (A) Schematic diagram of experimental design. (B-D) Migration of MGE-derived cells in response to mock-transfected (B) or Nrg1-transfected (C,D) COS cells aggregates cultured in matrigel matrices for 36 hours in the presence of vehicle solution (B,C) or the ROCK inhibitor Y27632 (30 µM) (D). COS cells were also transfected with dsRed to aid their visualization. Broken lines indicate the limits of the explants before culture. (B'-D') Confocal images of cells migrating through Sector 1, as defined in the schematic shown in A, in control (B'), Nrg1 (C') and in Nrg1+Y27632 (30 µM) (D')-treated explants. Solid and open arrowheads indicate branched and non-branched interneurons, respectively. (E) Schematic view of the method used to quantify the orientation of cells ( angle). (F) Quantification of angle in Sector 1. Bars show mean±s.e.m. 37.02±2.14° (control, n=167 cells from three independent experiments), 49.65±1.29° (Nrg1, n=520 cells from three independent experiments) and 38.10±1.41° (Nrg1 +Y27631, n=381 cells from three independent experiments). t-test, ^***P<0.001. (G) Quantification of percentage of neurons located in Sector 1 with at least two leading process branches. Bars show mean±s.e.m. 54.85±7.41% (Ctrl, n=161 cells from three independent experiments), 72.21±4.49% (Nrg1, n=528 cells from three independent experiments) and 55.73±5.34% (Nrg1+Y27631, n=317 cells from three independent experiments). t-test, ^*P<0.05. Scale bars: 100 µm in B,C,D; 40 µm B',C',D'.