The cellular mechanism by which the dermomyotome contributes to the secondwave of myotome development

We have shown that a subset of early postmitotic progenitors that originate s along the medial part of the epithelial somite gives rise to the primary myotome (Kahane, N., Cinnamon, Y. and Kalcheim, C. (1998). Mech. Dev. 74, 5 9-73). Because of its postmitotic nature, further myotome expansion must be achieved by cell addition from extrinsic sources. Here we investigate the mechanism whereby the dermomyotome contributes to this process. Using sever al different methods we found that cell addition occurs from both rostral a nd caudal edges of the dermomyotome, but not directly from its dorsomedial lip (DML). First, labeling of quail embryos with [H-3]thymidine revealed a time-dependent entry of radiolabeled nuclei into the myotome from the entir e rostral and caudal lips of the dermomyotome, but not from the DML. Second , fluorescent vital dyes were injected at specific sites in the dermomyotom e lips and the fate of dye-labeled cells followed by confocal microscopy. C onsistent with the nucleotide labeling experiments, dye-labeled myofibers d irectly emerged from injected epithelial cells from either rostral or cauda l lips. In contrast, injected cells from the DML first translocated along t he medial boundary, reached the rostral or caudal dermomyotome Lips and onl y then elongated into the myotome. These growing myofibers had always one e nd attached to either lip from which they elongated in the opposite directi on. Third, following establishment of the primary myotome, cells along the extreme dermomyotome edges, but not the DML, expressed QmyoD, supporting th e notion that rostral and caudal boundaries generate myofibers. Fourth, abl ation of the DML had only a limited effect on myotomal cell number. Thus, c ells deriving from the extreme dermomyotome lips contribute to uniform myot ome growth in the dorsoventral extent of the myotome. They also account for its expansion in the transverse plane and this is achieved by myoblast add ition in a lateral to medial direction (from the dermal to the sclerotomal sides), restricting the pioneer myofibers to the dermal side of the myotome . Taken together, the data suggest that myotome formation is a multistage p rocess. A first wave of pioneers establishes the primary structure. A secon d wave generated from specific dermomyotome lips contributes to its expansi on. Because dermomyotome lip progenitors are mitotically active within the epithelia of origin but exit the cell cycle upon myotome colonization, they can only provide for limited myotome growth and subsequent waves must take over to ensure further muscle development.