Figure 2

The explant culture system recapitulates the initiation of Hoxb4 expression in the neural tube and reveals a stage-dependent requirement of somites. (A-F) As schematized in the left of the panel, explants of neural tube were taken with flanking somites at the level of somites 1 to 5, from embryos at 2 (A, D), 6 (B, E) or 8 (C, F) somite stages, and either fixed immediately (A-C) or cultured for 24 hours (D-F). Immediately fixed explants show no or faint expression of Hoxb4 depending on the stage of explanting, reflecting the normal expression (A-C, n = 3/3 at each stage). After 24 hours of incubation, the explants show strong expression of Hoxb4 throughout the length of the neural tube (D-F, n = 6/6 at each stage). Indicated above each panel are the somite stages (S) at which explants were dissected. (G-R) Neural tube explants were taken either with (G-L) or without (M-R) flanking somites, at stages between 3 and 8 somites (3S to 8S) and cultured for 24 hours. All explants that included somites strongly express Hoxb4 homogenously in the neural tube (G-L, n = 6/6 at each stage). When somites are removed, Hoxb4 expression is absent in explants from 3 to 5 somite stages (M-O, n = 6/6 at each stage). In explants taken at the 6 somite stage, weak expression is observed (P, n = 8/8). It is at the 7 and 8 somite stages that the explants express Hoxb4 strongly (Q, R, n = 6/6 at each stage). Scale bar; 200 μm.