Recent studies have explored the potential of central nervous system derived neural stem cells (CNS-NSC) to repopulate the enteric nervous system (ENS). However, the exact phenotypic fate of gut transplanted CNS-NSC has not been characterized. The aim of this study was to investigate the effect of the gut microenvironment on phenotypic fate of CNS-NSC in vitro. Using transwell culture, differentiation of mouse embryonic CNS-NSC was studied when co-cultured without direct contact with mouse intestinal longitudinal muscle-myenteric plexus preparations (LM-MP) compared to control non co-cultured cells, in a differentiating medium. Differentiated cells were analyzed by immunocytochemistry and quantitative RT-PCR to assess the expression of specific markers, and by whole cell patch clamp studies for functional characterization of their phenotype. We found that LM-MP co-cultured cells had a significant increase in the numbers of cells that were immune-reactive against the pan neuronal marker β tubulin, neurotransmitters nNOS, ChAT and neuro-peptide VIP and showed an increase in expression of these genes, compared to control cells. Whole cell patch clamp analysis showed that co-culture with LM-MP decreases cell excitability and reduces voltage-gated Na+ currents, but significantly enhances A-current and late afterhyperpolarization (AHP) and increases the expression of AHP generating KCNN1 and 4 channel genes, compared to control cells. In a separate experiment, differentiation of LM-MP co-cultured CNS-NSC produced a significant increase in the numbers of cells that were immune-reactive against the neurotransmitters nNOS, ChAT and the neuro-peptide VIP when compared to CNS-NSC differentiated similarly in presence of neonatal brain tissue. Our results show that the gut microenvironment induces CNS-NSC to produce neurons which share some of the characteristics of classical enteric neurons, further supporting the therapeutic use of these cells for gastrointestinal disorders.
- CNS neural stem cells
- enteric nervous system
- nNOS and KCNN
- Copyright © 2011, American Journal of Physiology- Gastrointestinal and Liver Physiology