Activation of 5-HT4 receptors facilitates neurogenesis from transplanted neural stem cells in the anastomotic ileum
An orally administered serotonin-4 (5-HT4) receptor agonist, mosapride citrate (MOS), promotes enteric neurogenesis in anastomoses after gut surgery. We performed gut surgery and transplanted 2 × 105 neural stem cells (NSCs) from the embryonic central nervous system after marking them with the cell linker, PKH26. We found that neurons differentiated from transplanted NSCs (PKH [+]) and newborn enteric neurons differentiated from mobilized (host) NSCs (YFP [+]) in the deep granulation tissue of the anastomotic ileum. MOS significantly increased the number of PKH (+) and YFP (+) neurons by 2.5-fold (P < 0.005) (n = 4). The distribution patterns of PKH (+) neurons and YFP (+) neurons were similar along the depth of the anastomosis. A 5-HT4 receptor antagonist, SB-207266, abolished these effects of MOS (n = 4). Our results indicate that neurogenesis from transplanted NSCs is potentiated by activation of 5-HT4 receptors. Thus, a combination of drug administration and cell transplantation could be more beneficial than cell transplantation alone in treating Hirschsprung’s disease and related disorders.To confirm their quality, NSCs (catalog no. F-MUBNF-0101, Cyagen Biosciences) from the hippocampus and SVZ of day-12.5 post-coitus C57BL/6 mouse embryos supplied in a cryovial containing 1 × 106 cells were seeded on a gelatin-coated dish and then cultured as neurospheres in neural stem cell growth medium (catalog no. F-GUXNX-9011, Cyagen Biosciences) for 4 days. Microscopic photographs were taken under an inverted microscope (Nikon Ti-S100, Tokyo, Japan).As the preliminary for cell transplantation, we examined the effects of MOS on NSCs in culture. NSCs formed neurospheres after 4 days of culture in NSC growth medium in controls. BDNF facilitated the formation of enteric neural networks in ES guts, but GDNF did not . We therefore examined the effects of BDNF on NSCs in culture. BDNF (10 ng ml−1) weakly facilitated the outgrowth of projections from neurospheres, and MOS (1 µM) more potently facilitated the outgrowth of projections from neurospheres, but GR (10 µM), a selective 5-HT4 receptor antagonist, abolished MOS-induced effects on neurospheres (Fig. 1). From these results, we judged these NSCs could be used for the following cell transplantation.This research was supported by Grants-in-aid for Scientific Research (KAKENHI) from the Ministry of Education, Culture, Sports, Science, and Technology of Japan (20659210, 23390330, 24650325, 26560280, 15H03057 to M.T. and 23591969 to H.M.). It was partially supported by the Cooperative Study Program of the National Institute for Physiological Sciences.The authors declare no potential conflicts of interest.