The broad objective of this research is to determine how cells generate morphogenetic movements. The specific objective is to learn how deep mesodermal cells produce the autonomous narrowing (convergence) and lengthening (extension) of the dorsal axial tissues of the gastrula and neurula of the anuran amphibian, Xenopus laevis. Previous work showed that convergence and extension is accomplished by active rearrangement of several layers of cells to form fewer layers of greater area (radial intercalation) and several rows of cells to form fewer rows of greater length and lesser width (mediolateral intercalation). The proposed research will show how the direction of intercalation is determined and how the motile activities of individual cells contribute to the process. Regions of the embryo are explanted into a culture system of Dr. Keller's design that allows direct recording and manipulation of cell behavior. Paths of cell movement, protrusive activity and contact behavior in normal and microsurgically altered explants will be recorded with an optical disc recorder after digital video image processing. Two types of microscopy will be used: normal epi-illumination and low-light level fluorescence microscopy of cells labeled with DiI, a fluorescent lipid. These methods resolve the finest details of motility, which will be analyzed quantitatively and correlated with immunocytochemical analysis of the cytoskeleton of fixed explants and with cell structure seen by electron microscopy. Cell fates will be determined by tracing with fluorescein dextran amine. These data will show how the paths of cell movement are controlled and how protrusive activity of individual cells results in their intercalation. Convergence and extension by cell intercalation is an early, essential step in development of the dorsal axial tissues, the head, the vertebral column and central nervous system of all vertebrates examined thus far, and thus is a fundamental process in development. Understanding how cells intercalate will be a major contribution to our knowledge about how cells function in morphogenesis. %%% This work will provide a detailed description of the movements of cells deep in the early developing embryo that lead to the tissue shape changes. These ultimately cause the early foldings of the embryonic tissues to shape form the adult animal.
