DEV02312 Supplementary Material

Files in this Data Supplement:

• Supplemental Figure 1 -

  Fig. S1. The extent of disorganization of the dorsal funiculus becomes more severe at later stages. Transverse sections stained by Nissl staining (A,B), neurofilament (C,D) and TrkA (E,F) immunohistochemistry in the E14.5 wild-type (A,C,E) and netrin 1 mutant (B,D,F) dorsal horn. (A,B) In wild-type mice, many small neurons are uniformly distributed in the superficial layer of the dorsal horn. In the netrin 1 mutant dorsal horn, the distribution of dorsal cells is severely disrupted, and some neurophil islands are ectopically observed (arrowheads in B). (C,D) In netrin 1 mutant mice, there are many neurofilament-positive misplaced axon bundles throughout the dorsal horn (arrows in D), whereas the dorsal funiculus in wild-type mice is correctly formed in the dorsal spinal cord (arrowhead in C). (E,F) In both animals, TrkA-positive fibers are present in the superficial layer of the dorsal horn in spite of the disorganization of the dorsal funiculus in netrin 1 mutant mice (arrowheads). Scale bar: 100 mm.

• Supplemental Figure 2 -

  Fig. S2. Early-born neurons express dorsally derived netrin 1. (A,B) Immunostaining for BrdU after ISH for lacZ in the E12.5 dorsolateral spinal cord. BrdU was injected at E10.5. (C,D) X-gal staining at E14.5. (A,C) Netrin 1 heterozygous mice. (B,D) Netrin 1 mutant mice. (A,B) LacZ-expressing cells in the E12.5 dorsolateral spinal cord are colabeled with BrdU in both animals (arrowheads). ML, mantle layer; MZ, marginal zone. (C,D) Many X-gal-positive cells are abnormally distributed in the dorsolateral spinal cord of netrin 1 mutant mice (arrowhead in D). Scale bar: 25 mm in A,B; 100 mm in C,D.

• Supplemental Figure 3 -

  Fig. S3. Dcc mutant mice show disorganization of the dorsal funiculus at E13.5. Double staining with the Nissl method and neurofilament immunohistochemistry (A,B) and TrkA immunostaining (C,D) in the E13.5 spinal cord. (A,C) Wild-type mice. (B,D) Dcc mutant mice. At E13.5, the dorsal funiculus of Dcc mutants is obviously disorganized, and many fibers, including TrkA-positive fibers, enter the gray matter from the lateral portion of the dorsal funiculus (arrows in B,D) as well as the dorsomedial region, as observed in wild-type mice (arrowheads in A,B). Scale bar: 100 mm.