The expression patterns of sema3a1 and nrp1a correlate with extension of CaP axons along the common pathway. In situ hybridization (purple) for sema3a1 (A,B) or nrp1a (C-E) with immunostaining (brown) by mAb Znp1 that labels primary motor axons in 26-somite stage (22 hpf) embryos. Unless otherwise noted, embryos are oriented with rostral leftwards and dorsal upwards. (A) A transverse section of the trunk with dorsal upwards showing that sema3a1 is expressed in the dorsal and ventral regions of the myotome and less so in the horizontal myoseptal region (brackets). Asterisks indicate CaP motoneurons whose axons extend along the medial surface of the myotome. Sm, somite; Nc, notochord; Sc, spinal cord. Broken line indicates the level of the horizontal section shown in B. (B) A horizontal section with rostral leftwards, showing that the myotome cells immediately adjacent to the notochord, which CaP axons (arrowheads) extend upon, express little to no sema3a1 (black arrow). However, the more lateral cells express higher levels of sema3a1 (white arrow). Broken lines indicate somite borders. (C) Lateral view of the trunk showing that nrp1a is expressed segmentally in ventral spinal neurons that, based upon their axon trajectory, correspond to CaP neurons. Presumptive VaP (variable primary) neurons that arise in about half of the hemisegments as equivalent pair of CaP, but later die, may also express nrp1a. The expression of nrp1a declines in more anterior and developed CaPs. (D) nrp1a is expressed by CaP motoneurons (asterisk) while they are extending along the common pathway. (E) nrp1a expression is much reduced in CaP neurons (asterisk) with axons (mAb Znp1 immunostained in brown) extending onto the specific ventral pathway. Arrowheads in D,E indicates the position of the horizontal myoseptum. Scale bars: 20 μm.
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