PhD

dissertationThe question of which embryonic structures arise from particular regions of the early embryo is central to a better understanding of normal development. Marking undifferentiated cells with a dye that can later be recognized is one way of examining this question. Such fate maps have traditionally been generated by the use of vital stains as cellular markers; however, the fading and dilution of the dye render this technique unsatisfactory. The availability of nonfading heritable lineage tracers has made it possible to reexamine prospective cell fates more accurately and reliably. In the present work, horseradish peroxidase was used to label whole Xenopus embryos, from which grafts were later taken and transplanted orthotopically to unlabeled embryos to produce fate maps of the different germ layers from stage 10 to stage 23, and of the subdivisions of nervous system from stage 14 to stage 23. A second central question in embryology concerns the mechanism by which some cells follow one developmental pathway whereas their sisters may follow another. To understand this problem one needs first to determine over what developmental period the cell's fate becomes restricted. In the second series of experiments, horseradish peroxidase and lissamine-rhodamine-dextran-lysine labeled single cells and grafts were transplanted orthotopically, heterotopically, isochronically or heterochronically at the 512-cell stage (mid-blastula), stage 10 and stage 14 to examine the state of cell determination. At the 512-cell stage, single labeled cells were taken from one region and transplanted stage, single labeled cells were taken from one region and transplanted to the same or a different region of an unlabeled 512-cell stage embryo. These experiments were repeated with small clusters of cells from stage 10 embryos, and the results showed that the labeled cells regulated and developed according to their new environment. This was not the case when cells were transplanted at stage 14. These results indicate that embryonic cells from certain regions at the 512-cell stage and stage 10 in Xenopus are not determined irreversibly, whereas cells from the neural plate and neural fold at stage 14 appear to have become irreversibly determined. The results of heterochronic reciprocal transplantations between stage 10 and stage 14 further supported these observations