Abstract

Background: A homozygous mutation of the endothelin B receptor (EDNRB) gene in spotting lethal (sl/sl) rats leads to Hirschsprung's disease (HSCR) with long segmented aganglionosis. However, the effects on the development of the enteric nervous system (ENS) promoted by a heterozygous mutation of the EDNRB gene are not known. The present study aimed to describe and morphometrically assess the phenotypic abnormalities of the ENS in heterozygous (+/sl) EDNRB deficient rats in comparison with homozygous (sl/sl) EDNRB deficient and wild-type (+/+) rats.

Methods: The distal small intestine, caecum, and colon were obtained from sl/sl, +/sl, and +/+ rats. To demonstrate the three dimensional organisation of the ENS, the intestinal wall was microdissected into wholemounts and incubated against the pan-neuronal marker protein gene product 9.5. Assessment of the ENS included morphometric quantification of ganglionic size and density, the number of nerve cells per ganglia, and the diameter of nerve fibre strands within both the myenteric and submucous plexus.

Results: Sl/sl rats were characterised by complete aganglionosis resembling the same histopathological features observed in patients with HSCR. +/sl rats revealed more subtle abnormalities of the ENS: the submucous plexus was characterised by a significantly increased ganglionic size and density, and the presence of hypertrophied nerve fibre strands. Morphometric evaluation of the myenteric plexus did not show statistically significant differences between +/sl and +/+ rats.

Conclusions: In contrast with sl/sl rats, +/sl rats display non-aganglionated malformations of the ENS. Interestingly, these innervational abnormalities resemble the histopathological criteria for intestinal neuronal dysplasia (IND). Although IND has been described in several intestinal motility disorders, the concept of a clearly defined clinical-histopathological entity is still controversially discussed. The present findings support the concept of IND based on clearly defined morphological criteria suggesting a genetic link, and thus may provide a model for human IND. Furthermore, the data underline the critical role of the “gene dose” for the phenotypic effects promoted by the EDNRB/EDN3 system and confirm that the development of the ENS is not an “all or none” phenomenon.