Effect of butyrate on membrane potential, ionic currents and intracellular Ca²⁺ concentration in cultured rat myenteric neurones

Abstract

Myenteric neurones from 1-10-day-old rats were isolated from the small and large intestine by enzymatic digestion with collagenase. Single cells were collected and kept in culture for up to 1 week. After 1-5 days in culture, membrane potential and ionic currents were measured with the whole-cell patch-clamp technique. The intracellular Ca2+ concentration was measured with the fura-2 method. The short-chain fatty acid butyrate (50 mmol L-1) induced a reversible hyperpolarization of the myenteric neurones by about 10 mV. This hyperpolarization was concomitant with an inhibition of a TTX-sensitive Na+ current. The hyperpolarization could be suppressed by intracellular application of Cs+, a nonselective K+ channel blocker. Fura-2 experiments revealed that butyrate induced an increase of the intracellular Ca2+ concentration. The butyrate response was suppressed by thapsigargin, indicating that butyrate stimulates the release of intracellular Ca2+. This release is responsible for the voltage response, because intracellular chelation of Ca2+ inhibited the butyrate induced hyperpolarization. Consequently, butyrate acts on enteric neurones by releasing Ca2+ from intracellular stores with the consequence of the activation of K+ channels, followed by a hyperpolarization.

P2X2receptors are essential for [Ca2+]iincreases in response to ATP in cultured rat myenteric neurons

“…Therefore, these immunocytochemical data suggest that the major classes of myenteric neurons are motor and interneurons and a small population of them are AH-type sensory neurons. Similar to the present results, it has been reported that there are few AH-type neurons in cultured rat myenteric neurons by patch-clamp study (29).…”

The peristaltic reflex induced by short-chain fatty acids is mediated by sequential release of 5-HT and neuronal CGRP but not BDNF

“…In general, the excitatory effects of SCFAs have been thought to be dependent on enteric neural reflexes that lead to activation of enteric cholinergic motor neurons (11,21,24,29,33), although direct effects on smooth muscle cells (28) and enteric neurons (2,17,18) as well as effects mediated by local release of prostaglandin have also been demonstrated (29). There is also general agreement that the SCFA-induced reflex is dependent on the presence of an intact mucosa (2,11,21,24,29,39).…”

“…enteric nervous system; neurotrophins; neuropeptides; colon; gastrointestinal motility; calcitonin gene-related peptide; brain-derived neurotrophic factor; serotonin THE SHORT-CHAIN FATTY ACIDS (SCFA), acetate, propionate, and butyrate, produced by a bacterial fermentation of dietary fiber in the colon, are an important source of energy for colonic epithelial cells (4,8,9,34). These anions regulate various colonic functions, including colonic electrolyte transport and motor activity and the growth, proliferation, and differentiation of colonic mucosal cells (4,6,7,9,18,24,35,36,39). Their concentration in the colon of most mammals, including humans, varies with the nature and availability of the substrate, the type and amount of microflora present in the colon, the pH, the rate of transit of luminal contents, and the site of measurement within the colon (1, 4, 10, 37, 38).…”

Effects of H2O2 at rat myenteric neurones in culture

“…The inward current at cultured rat myenteric neurones from newborn rats has been demonstrated to be mediated by a tetrodotoxin-sensitive inflow of Na + (Haschke et al 2002). Tetrodotoxin (TTX) is a neurotoxin, which inhibits voltage-dependent Na + currents in many neurones (Catterall 1980), although tetrodotoxin-resistant Na + channels are known, too (see e.g.…”

“…Changes in the intracellular Ca 2+ concentration were measured using the Ca 2+-sensitive fluorescent dye fura-2 as described previously (Haschke et al 2002). The neurones grown on coverslips were loaded for 60 min with 2.5 μmol/l fura-2 acetoxymethylester (fura-2AM) in the presence of 0.05 g/l pluronic acid.…”