Neural Tube and Notochord Promote in Vitro Myogenesis in Single Somite Explants

Abstract

We have developed an in vitro assay for examining the neural tube/notochord influence on somite myogenesis. No cells committed to myogenesis (operationally defined as immunopositive staining for myosin heavy chain after a 48-hr culture period) are present in the most caudal somites (I-V) of stage 8-11 chicken embryos, whereas by stage 14-15, the most caudal somites occasionally produce differentiated muscle cells when cultured alone. To test for responsiveness to neural tube, one somite of each pair was cultured in contact with a piece of neural tube; as a control, the contralateral somite was cultured alone to test for prior myogenic commitment of somitic cells. Using this assay, we show that neural tube induces myogenesis in single somite explants, that close somite-neural tube proximity is required, and that the interaction can occur in defined medium. Neural tube induces myogenesis in the most recently formed somites and in segmental plate mesoderm, indicating that somite segmentation is not a prerequisite for neural tube mediated myogenesis in vitro . The assay was also used to determine whether continued exposure to neural tube is required for increasing myogenic cell numbers in somites which already contain cells committed to myogenesis. Since somites containing committed cells exhibit a fourfold increase in the number of myogenic cells when cocultured with neural tube, continued in vitro exposure to neural tube either recruits additional somite cells into the myogenic lineage, and/or it stimulates proliferation of existing myogenic cells. Consistent with this finding, the most rostral neural tube retains its inductive properties as evidenced by the induction of nascent myogenesis in the most caudal somites. Parallel studies of notochord mediated myogenesis indicate that the notochord promotes myogenesis within somites II-VIII, but not within somite I or the segmental plate. This observation suggests either a qualitative or quantitative difference in the mechanism by which the neural tube and notochord communicate with the youngest somite and presomitic mesoderm.