Fig. 3. Regionalization of the myogenic core of BA1 by CPM and SpM cells. (A,A') Injection of DiI to the CPM (A) of a St. 8 chick embryo. After 36 hours incubation, the labeled cells streamed toward BA1. Notably, some of the CPM cells were seen in the distal outflow tract (A'). (A'',A''') Injection of DiI into the AHF/SpM (A'') of a St. 8 embryo. After 36 hours incubation, labeled cells populated the outflow tract; some of the cells were seen in the distal portion of BA1 (A'''). (B,B') Double injection of DiI (CPM) and DiO (AHF/SpM) into a St. 8 embryo (B) as shown in the transverse section (B') (n=22). (C-C'') After 18 hours incubation (C), at St. 11, the DiI was found in the CPM (C') anterior to the location of the DiO in the outflow tract (C''). (D-D'') After 24 hours (St. 12, D) the DiI was found at the most-proximal area of the developing BA (D'), as well as in the outflow tract (yellow staining in D''). The DiO was located in the distal part of the forming BA (D') and in the cardiac outflow tract (D''). (E-E'') After 48 hours (St. 16, E) the DiI was found in the most-proximal area of the mesoderm core, whereas the DiO was located in the distal part of the BA1 core (E'). BA1 viewed at higher resolution (E'') demonstrates that the dye is restricted to the mesodermal core, and doesn't appear in the neural crest cells (nc) surrounding the core. (F,G) DiO was injected into the AHF/SpM of St. 8 embryos that were subsequently immunostained (red) for the AHF markers Isl1 (F) and Nkx2.5 (G). Co-localization of the DiO and the two AHF markers is shown in the distal part of the arch. Note that the DiO only labeled mesoderm cells (Isl1⁺ and Nkx2.5⁺), and not neural crest cells. oft, outflow tract; nt, neural tube; BA1, branchial arch 1; mes, mesoderm; hrt, heart; ph, pharynx; ecto, ectoderm; CPM, cranial paraxial mesoderm; AHF/SpM, anterior heart field/splanchnic mesoderm; ov, otic vesicle.