Role of neuregulin 1 in regulating the development of enteric nervous system in zebrafish

Abstract

Objective To explore the role of Neuregulin 1(NRG1) in the development of intestinal nervous system. Methods Zebrafish NRG1 gene was cloned, NRG1 homology and phylogenetic tree were analyzed in multiple species. In situ hybridization was employed for detecting the expression of NRG1 in adult zebrafish intestine and embryos. Zebrafish NRG1 knockdown model was established by a microinjection of morpholino antisense oligonucleotides. Phenotypes, differentiation and migration of intestinal neuron were shown by in situ hybridization and immunofluorescence. Results Evolutionarily conserved, NRG1 had a high homology between zebrafish and human counterparts. It was expressed in mucosal layer but not muscular layer or serous membrane of adult zebrafish intestine. NRG1 was expressed in larval gut since early embryonic period. The expression spectrum matched enteric nervous development phase. In NRG1 knockdown model, embryonic hatching was delayed. And there were head deformity and short and distorted body. Most importantly, intestinal neuronal differentiation and migration process were inhibited by a down-regulation of NRG1. Survival rate was lower in embryos injected by NRG1-MO than control group[29.0%(196/677) vs 87.5%(446/510)]. And the phenotypic abnormality rate was higher than control group[21.5%(40/186)vs 5.5%(23/415)]. Differentiated intestinal neurons in NRG1 knock-down embryos were less than control group[(3.33±1.53) vs(43.00±13.23)]. And the anterio-posterior migration of enteric neurons was blocked in NRG1 knock-down embryos. Conclusions An aberrant expression of NRG1 may play an important role in the pathology of HD. The intestinal nervous development in zebrafish is imitated by interfering the NRG1 expression. And a suitable model has been established for studying the mechanism of HD. Key words: Enteric nervous system; Zebrafish; Hirschsprung disease