ret Proto-oncogene product is a useful marker of lineage determination in the development of the enteric nervous system in rats

Detailed study of developmental changes in the enteric nervous system is necessary to disclose the pathogenesis of Hirschsprung's and allied disease, some of which have hypoplastic ganglia. Therefore experiments were undertaken to study the fate of neural crest cells that develop in the rat gut during ontogeny. A polyclonal antibody against ret proto-oncogene product (c-Ret protein) and various monoclonal antibodies against neural markers (tyrosine hydroxylase, dopamine beta hydroxylase, microtubule-associated protein 5, microtubule-associated protein 2 and 160-kd neurofilaments) were used to identify neural crest-derived cells in rat embryos (10.5 to 15.5 days' gestation) and adult rats using a double immunostaining method. C-Ret protein was an early marker of lineage determination in the development of the enteric nervous system (11.5-day embryo: E 11.5). C-Ret-positive cells transiently coexpressed tyrosine hydroxylase, which also was observed in the vagal crest-derived precursors of enteric neurons (days E 11.5 to E 13.5). These cells also coexpressed other neural markers in the proximal gut. Expression of neural markers migrated to the distal intestine during development. This study found a discrepancy between the time when these markers appeared in the cranial and when they appeared in the caudal intestine. Tyrosine hydroxylase-positive cells did not appear in the postumbilical gut. The formation of the primitive neural network in the entire myenteric plexus at day E 15.5 was demonstrated by c-Ret protein. Other neural markers were lost or had decreased immunoreactivity throughout the entire intestine of the E 15.5 and adult animals. In conclusion, (1) c- Ret protein is one of the earliest markers of lineage determination in the development of the enteric nervous system, (2) each neural marker is expressed at its own time and differs in spatial developmental lineage, (3) c-Ret protein and other neural markers are transiently expressed by a particular group of neural cells during the embryonic period, (4) there is a subpopulation of cells that has never transiently expressed tyrosine hydroxylase in the postumbilical gut, which may have originated from tissue other than the vagal crest, and (5) the primitive neural network in the myenteric plexus was completed at day E 15.5.