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Fig. 4. Commissural axons are misguided following misexpression of Math1::caBMPRIB-IRES-fGFP in stage 15 chick spinal cords. (A-D) Electroporation of either Math1::fGFP (A,B) or Math1::caBMPRIA-IRES-fGFP (C,D) constructs results in GFP (blue) in all commissural neural processes, including the trailing processes (open arrowhead, B,D) and Axonin1⁺ axons (green) that cross the FP (arrowhead, B,D). Antibodies against Axonin1 also transiently label MNs (m). (E,F) By contrast, electroporation with a Math1::caBMPRIB-IRES-fGFP construct results in commissural axons being mispolarized towards the lumen (arrow, F) and stalling (arrowhead, F). (G,H) The GFP⁺ axons in control fillet preparations (G; dotted yellow lines in G and H indicate position of the RP and FP) project robustly to the FP, whereas GFP⁺ axons in the Math1::caBMPRIB-IRES-fGFP fillets (H) do not enter the ventral spinal cord (arrowhead). (I,J) The extent of the commissural axon outgrowth was quantified in stage 22/23 embryos by determining the number of GFP⁺ commissural axons that crossed lines drawn (J) in the mid-dorsal (MD), intermediate (INT) and mid-ventral (MV) spinal cord, and the FP. Of the control commissural neurons extending axons to the MD line, over 55% of these axons subsequently project to the MV line (n=158 sections from 10 embryos). By contrast, less than 23% of the Math1::caBMPRIB-IRES-fGFP commissural axons that extend to the MD line subsequently reach the MV line (n=145 sections, 15 embryos), a figure significantly different from control (P<2.4x10^-31). Scale bar in F: 100 µm for A-F.

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