The enteric nervous system (ENS) is the most complex and largest portion of the peripheral nervous system. The ENS is comprised of a complex series of interconnected neurons and glia that control intestinal motility and regulate blood flow within the gut wall. During development, ENS arises from vagal neural crest cells that emigrate from the neural tube at caudal hindbrain levels. Upon reaching the foregut, the vagal-derived enteric neural crest cells migrate caudally along the developing gut and differentiate into enteric neurons and glial cell types. Abnormalities in migration to or along the gut or along resul in birth anomalies like Hirschsprung's Disease, in which the distal portion of the intestine is devoid of enteric neurons or glia, resulting in megacolon and chronic constipation. While much research attention has focused on the functional analysis of enteric neural crest cell migration in the gut during later phases of ENS development, the molecular mechanisms that regulate their migration and the designation of early vagal and enteric neural crest cell identity is poorly understood. The experiments in this proposal will utilize zebrafish and chicken embryos in order to enhance our understanding of the mechanisms that regulate early phases of vagal neural crest and ENS development. Both chicken and zebrafish embryos develop externally, allowing for easy experimental manipulation and observation. Furthermore, zebrafish embryos are transparent during development allowing for high-resolution analysis of neural crest cell migration. The specific aims are to 1. Investigate the role of Meis3 in development of the zebrafish enteric neural crest, and 2. Elucidate the role and regulation of Hoxb5 in vagal and enteric neural crest development. The results of these experiments will significantly improve our basic understanding of enteric neural crest cell migration and fate specification, thus providing a solid foundation on which to base upstream translational research endeavors.