Flupirtine enhances NHE-3-mediated Na+ absorption in rat colon via an ENS-dependent mechanism

Abstract

Recent studies in our lab have shown that the K_V7 channel activator, flupirtine, inhibits colonic epithelial Cl^- secretion through effects on submucosal neurons of the enteric nervous system (ENS). We hypothesized that flupirtine would also stimulate Na⁺ absorption as a result of reduced secretory ENS input to the epithelium. To test this hypothesis, unidirectional ²²Na⁺ fluxes were measured under voltage-clamped conditions. Pharmacological approaches using an Ussing-style recording chamber combined with immunofluorescence microscopy techniques were used to determine the effect of flupirtine on active Na⁺ transport in the rat colon. Flupirtine stimulated electroneutral Na⁺ absorption in partially seromuscular-stripped colonic tissues, while simultaneously inhibiting short-circuit current (I_SC; i.e., Cl^- secretion). Both of these effects were attenuated by pretreatment with the ENS inhibitor, tetrodotoxin. The Na⁺/H⁺ exchanger isoform 3 (NHE-3)-selective inhibitor, S3226, significantly inhibited flupirtine-stimulated Na⁺ absorption, whereas the NHE-2-selective inhibitor HOE-694 did not. NHE-3 localization near the apical membranes of surface epithelial cells was also more apparent in flupirtine-treated colon versus control. Flupirtine did not alter epithelial Na⁺ channel (ENaC)-mediated Na⁺ absorption in distal colonic tissues obtained from hyperaldosteronaemic rats and had no effect in the normal ileum but did stimulate Na⁺ absorption in the proximal colon. Finally, the parallel effects of flupirtine on I_SC (Cl^- secretion) and Na⁺ absorption were significantly correlated with each other. Together, these data indicate that flupirtine stimulates NHE-3-dependent Na⁺ absorption, likely as a result of reduced stimulatory input to the colonic epithelium by submucosal ENS neurons.NEW & NOTEWORTHY We present a novel mechanism regarding regulation of epithelial ion transport by enteric neurons. Activation of neuronal K_V7 K⁺ channels markedly stimulates Na⁺ absorption and inhibits Cl^- secretion across the colonic epithelium. This may be useful in developing new treatments for diarrheal disorders, such as irritable bowel syndrome with diarrhea (IBS-D).

Keywords: IBS-D; KCNQ; Ussing chamber; enteric nervous system.