Fig. 4. Distribution of sympathetic neuron precursors, spinal nerve fibers, and Sema3A transcripts in E9.25 embryos. (A,B) In situ hybridization analysis on frontal sections of the wild-type (A) and neuropilin 1 homozygous mutant (B) embryos, using an antisense cRNA probe for Mash1. Arrows indicate cells with the Mash1 transcripts. (C,D) Detection of spinal nerve fibers by immunostaining with 2H3 in sections adjacent to A and B. 2H3-positive fibers (arrows) are limited around the neural tube (nt). The staining in blood cells and the internal surface of the digestive tracts is non-specific. (E) In situ hybridization analysis on a frontal section of a wild-type embryo, using an anti-sense cRNA probe for Sema3A. An arrow indicates the notochord. da, dorsal aorta; fl, the forelimb; dm, dermamyotome; nt, neural tube. Scale bar: 100 µm (A-E).