Abstract
Extending well beyond the original use of propagating neural precursors from the central nervous system and dorsal root ganglia, neurosphere medium (NSM) and self-renewal medium (SRM) are two distinct formulas with widespread popularity in enteric neural stem cell (ENSC) applications. However, it remains unknown what growth factors or nutrients are crucial to ENSC development, let alone whether the discrepancy in their components may affect the outcomes of ENSC culture. Dispersed enterocytes from murine fetal gut were nurtured in NSM, SRM or their modifications by selective component elimination or addition to assess their effects on ENSC development. NSM generated neuriteless neurospheres, whereas SRM, even deprived of chicken embryo extract, might wire ganglia together to assemble neural networks. The distinct outcomes came from epidermal growth factor, which inhibited enteric neuronal wiring in NSM. In contrast, basic fibroblast growth factor promoted enteric neurogenesis, gangliogenesis, and neuronal wiring. Moreover, vitamin A derivatives might facilitate neuronal maturation evidenced by p75 downregulation during ENSC differentiation toward enteric neurons to promote gangliogenesis and network assembly. Our results might help to better manipulate ENSC propagation and differentiation in vitro, and open a new avenue for the study of enteric neuronal neuritogenesis and synaptogenesis.