ENTERIC NEUROBLAST-SPECIFIC EXPRESSION OF ENDOTHELIN-B RECEPTORS ALLOWS NORMAL ENS DEVELOPMENT IN THE AGANGLIONOSIS RAT

Abstract

Loss-of-function mutations of the endothelin-B receptor (ETB) cause congenital distal intestinal aganglionosis in mice, rats and humans (Hirschsprung disease). This suggests activation of ETB is critical for normal ENS colonization of the distal gut. The timing and localization of the activation and the downstream events that lead to colonization failure in the absence of activation are unknown. While in situ hybridization experiments in mice fail to identify ETB expression in the hindgut mesenchyme, other studies suggest absence of ETB activation leads to development of a hindgut microenvironment inhibitory to neuroblast colonization. We used a tissue-specific transgenic approach in the rat to investigate the timing and localization of ETB expression sufficient for normal ENS development. The spotting-lethal (sl) rat carries a naturally occurring deletion in the ETB gene resulting in complete absence of functional ETB expression. Homozygous rats exhibit abnormal development of the ENS, similar to that described in Hirschsprung disease. Studies employing a dopamine-β-hydroxylase(DβH)/lacZ transgene indicate enteric neuroblasts are transiently adrenergic during gut colonization and that this colonization is defective in ETB deficient mice. Because in situ hybridization studies in mice show enteric neuroblasts express ETB during gut colonization, we used the DβH promoter to direct ETB transgene expression in sl/sl rats. The DβH promoter limits expression to adrenergic neurons and the adrenals. These transgenic rats exhibit a normal appearing distal ENS, they do not develop megacolon and are otherwise healthy. These findings indicated ETB expression on the developing neuroblasts during the colonization process is sufficient for normal ENS development.