Fig. 2. TuJ1 immunostaining of E4, E7 and E10 control and 3-6 ablated whole-mount gut preparations. (A-D) E4 control gut showing the ENS network in (A) the stomach, where nerve fibres from the vagus nerve are apparent (arrows), and (B) the duodenum. (C) NC-derived cells are not present at the level of the umbilicus. (D) The migration wave-front contains mostly fasciculated axons and no isolated cells. (E-H) 3-6 ablated gut showing numerous vagus nerve fibre projections (arrows) and a reduced ENS network in (E) the stomach and (F) the duodenum. (G) NC-derived cells are not present at the level of the umbilicus and only the NoR is TuJ1+. (H) The migration wave-front has loosely organised TuJ1+ cells and more isolated cells (arrowheads). (I-L) Control E7 gut with characteristic TuJ1+ ENS network in (I) the stomach and (J) the duodenum. (K) At the level of the umbilicus, the NoR projects fibres into the gut wall. (L) TuJ1+ cells have migrated through the cecal buds and extend into the hindgut. NoR fibres project into the gut co-incident with the most advanced cells at the migration wave-front (dashed line). (M-P) 3-6 ablated gut. The TuJ1+ migration wave-front terminates at the caudal extent of the duodenum (N), which represents the anterior extent of the NoR (arrow). The umbilicus level of the intestine (O), the cecal buds and hindgut (P) do not contain NC-derived cells. (Q-T) E10 control gut showing characteristic ENS networks. (U-X) 3-6 ablated gut. There are fewer perivascular extensions (arrow) within the stomach compared with controls. (V) The caudal extent of the migration wave-front in the duodenum is co-incident with the anterior extent of the NoR. Nerve fibres project from the NoR into the gut (V, arrowheads; W,X). NoR, Nerve of Remak; umb, umbilicus; cb, cecal buds; hg, hindgut. Scale bars: 300 µm in A; 100 µm in B.