Nrp1 and Sema3a expression patterns during trunk NCC migration. (A–E) Longitudinal sections through a 9.5 dpc (25-somite) mouse embryo; only one-half of the section is shown; the neural tube is at the top of each panel; somites are counted backwards from the most recent pair formed. (A–C) Double labeling for the NCC marker p75 (green) and NRP1 (red). Early wave NCCs appeared to migrate alongside intersomitic blood vessels (isv) and expressed p75, but not NRP1 (arrow in A). In addition, some early NCCs migrated along the boundary between anterior and posterior sclerotome (curved arrow). (B) NCCs in the nonsegmented migration staging area (MSA) in the dorsal rostral trunk co-expressed p75 and NRP1 (yellow). (C) A section taken at the same rostral level, but further ventrally through the somites, showed that NCCs in the anterior sclerotome (arrowheads) co-expressed p75 and NRP1 (yellow). (A and C) Mesenchymal cells (wavy arrows) and neural progenitors in the neural tube also expressed NRP1, but at lower levels. (D and E) Triple labeling for Sema3a (blue), p75 (NCCs, green), and endomucin (blood vessels, red). (D) Sema3a was expressed in the posterior sclerotome and the dermomyotome, with highest levels in the anterior and posterior part of the dermomyotome, adjacent to the neighboring intersomitic furrows (clear arrow and arrowhead, respectively). The dimensions of the dermomyotome versus sclerotome and the anterior (a) versus posterior (p) sclerotome are indicated with brackets. (E) In wild-types, most intermediate wave NCCs migrated through the avascular anterior sclerotome (arrowhead), and only a few NCCs migrated alongside intersomitic vessels (arrow) or around the dermomyotome (open arrowhead). (F) Schematic representation of NRP1 and SEMA3A expression during trunk NCC migration in the 9.5 dpc mouse embryo. The firstborn trunk NCCs do not express NRP1 (green) and migrate through the intersomitic furrow or along the anteroposterior boundary within the sclerotome. Most NCCs in the intermediate wave are NRP1-positive (yellow) and migrate through the anterior sclerotome; they settle in the sclerotome or travel ventrally to meet NCCs from the early wave at the dorsal aorta; there, NCCs move rostrocaudally along the dorsal aorta to form the sympathetic anlagen. Abbreviations: da, dorsal aorta; dm, dermomyotome; isf, intersomitic furrow; isv, intersomitic vessel; psv, perisomitic vessel; pvp, perineural vascular plexus; sa, sympathetic anlage; scl, sclerotome. (Scale bars: A–C, 50 μm; D and E, 100 μm.)