Division of the cardiac tube into four chambers and two outflow vessels is critical in normal heart development. The septum that converts the original single outflow vessel into an aorta and pulmonary trunk depends on the integrity of the cardiac neural crest cell population that migrates from the caudal hindbrain via the pharyngeal arches into the outflow tract. This cell population subdivides into ectomesenchymal and neural lineages. The ectomesenchymal lineage participates in structural development of the outflow septum while the neural lineage forms the cardiac parasympathetic innervation. The lineage decision is critical because commitment of too many cells to either lineage will affect the other population and result in either congenital cardiac defects or disturbed innervation of the heart. The cellular mechanism for the separation of the neural crest cells into these two lineages is not known. The hypothesis for this proposal is that Mash-1 and the Notch family of genes are involved in the lineage decisions. Many of these genes have recently been targeted for null mutation in mice. The homozygote Mash-1 mutant mouse has no parasympathetic cardiac innervation, indicating an inability of the cardiac neural crest to commit to the neural lineage. In this case, it is not known what happens to the ectomesenchymal lineage. Mice with null mutations in Notch family genes do not survive into the period when cardiac neural crest can be analyzed. We propose to rescue the mutant mouse cardiac neural crest cells by constructing mouse-chick chimeras. The cardiac region of the neural tube will be transplanted into chick embryos lacking the cardiac neural crest. The prospective potency of Mash-1-/- and Notch-/- cells will be determined by analyzing cell fate, proliferation and cell death at three times after implantation of the cells. The role of gene dosage will be determined by mixing cells from wide-type, heterozygote, and homozygote mutant mice prior to implantation into the chick host. Finally, it has been proposed that the development of the coronary vascular tree and Purkinje cells depends on formation of the cardiac ganglia. The development of the conduction system will be assessed in Mash-/- chimeras.
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