The voltage-sensitive sodium channel Na v 1.5 (encoded by SCN5A) is expressed in electromechanical organs and is mechanosensitive. This study aimed to determine the mechanosensitive transitions of Na v 1.5 at the molecular level. Na v 1.5 was expressed in HEK 293 cells and mechanosensitivity was studied in cell-attached patches. Patch pressure up to −50 mmHg produced increases in current and large hyperpolarizing shifts of voltage dependence with graded shifts of half-activation and half-inactivation voltages ( V 1/2 ) by ∼0.7 mV mmHg −1 . Voltage dependence shifts affected channel kinetics by a single constant. This suggested that stretch accelerated only one of the activation transitions. Stretch accelerated voltage sensor movement, but not rate constants for gate opening and fast inactivation. Stretch also appeared to stabilize the inactivated states, since recovery from inactivation was slowed with stretch. Unitary conductance and maximum open probability were unaffected by stretch, but peak current was increased due to an increased number of active channels. Stretch effects were partially reversible, but recovery following a single stretch cycle required minutes. These data suggest that mechanical activation of Na v 1.5 results in dose-dependent voltage dependence shifts of activation and inactivation due to mechanical modulation of the voltage sensors.

Interstitial cells of Cajal (ICC) generate the electrical slow wave required for normal gastrointestinal motility. The ionic conductances expressed in human intestinal ICC are unknown. The aim of this study was to determine expression of a Na+ current in human intestinal ICC and to determine the effects of the Na+ current on the slow wave. Visually identified, freshly dissociated, single ICC were verified by the presence of c-kit mRNA by using single-cell RT-PCR. Standard whole cell currents were recorded from patch-clamped ICC held at -100 mV between pulse protocols. A Na+ current was identified in human intestinal ICC. The current activated at -55 mV and peaked at -30 mV. Extracellular N-methyl-d-glucamine abolished and QX-314 (500 microM) blocked the Na+ current, but nifedipine and Ni2+ did not. The Na+ current was activated by shear stress. Single-cell RT-PCR detected mRNA for the Na+ alpha-subunit SCN5A in single human intestinal ICC. Lidocaine (200 microm) and QX-314 (500 microM) decreased slow wave frequency, and stretch increased slow wave frequency. A mechanosensitive Na+ channel current is present in human intestinal ICC and appears to play a role in the control of intestinal motor function.

SignificanceMechanical forces are important for normal gastrointestinal tract function. The enterochromaffin cells in the gastrointestinal epithelium have been proposed, but not previously shown, to be specialized sensors that convert forces into serotonin release, and serotonin released from these cells is important for normal gastrointestinal secretion and motility. The findings in this study show that some enterochromaffin cells are indeed mechanosensitive, and that they use mechanosensitive Piezo2 channels to generate an ionic current that is critical for the intracellular Ca2+ increase, serotonin release, and epithelial fluid secretion.

BACKGROUND & AIMS SCN5A encodes the α-subunit of the voltage-gated sodium channel NaV1.5. Many patients with cardiac arrhythmias caused by mutations in SCN5A also have symptoms of irritable bowel syndrome (IBS). We investigated whether patients with IBS have SCN5A variants that affect the function of Nav1.5. METHODS We performed genotype analysis of SCN5A in 584 persons with IBS and 1380 without (controls). Mutant forms of SCN5A were expressed in HEK-293 cells, and functions were assessed by voltage clamp analysis. A genome-wide association study (GWAS) was analyzed for an association signal for the SCN5A gene, and replicated in 1745 patients in 4 independent cohorts of IBS patients and controls. RESULTS Missense mutations were found in SCN5A in 13/584 patients (2.2%, probands). Diarrhea-predominant IBS (IBS-D) was the most prevalent form of IBS in the overall study population (25%). However, a greater percentage of individuals with SCN5A mutations had constipation-predominant IBS (IBS-C, 31%) than IBS-D (10%, P<.05). Electrophysiologic analysis showed that 10/13 detected mutations disrupted NaV1.5 function (9 reduced and 1 increased function); p.A997T-NaV1.5 had the greatest effect in reducing NaV1.5 function. Incubation of cells that expressed this variant with mexiletine restored their sodium current; administration of mexiletine to 1 carrier of this mutation (who had IBS-C) normalized their bowel habits. In the GWAS and 4 replicated studies, the SCN5A locus was strongly associated with IBS. CONCLUSIONS About 2% of patients with IBS carry mutations in SCN5A. Most of these are loss-of-function mutations that disrupt NaV1.5 channel function. These findings provide a new pathogenic mechanism for IBS and possible treatment options.

The SCN5A-encoded Na(v)1.5 Na(+) channel is expressed in interstitial cells of Cajal and smooth muscle in the circular layer of the human intestine. Patients with mutations in SCN5A are more likely to report gastrointestinal symptoms, especially abdominal pain. Twin and family studies of irritable bowel syndrome (IBS) suggest a genetic basis for IBS, but no genes have been identified to date. Therefore, our aims were to evaluate SCN5A as a candidate gene involved in the pathogenesis of IBS and to determine physiological consequences of identified mutations. Mutational analysis was performed on genomic DNA obtained from 49 subjects diagnosed with IBS who reported at least moderately severe abdominal pain. One patient hosted a loss-of-function missense mutation, G298S, that was not observed in >3,000 reference alleles derived from 1,500 healthy control subjects. Na(+) currents were recorded from the four common human SCN5A transcripts in transfected HEK-293 cells. Comparing Na(v)1.5 with G298S-SCN5A versus wild type in HEK cells, Na(+) current density was significantly less by 49-77%, and channel activation time was delayed in backgrounds that also contained the common H558R polymorphism. Single-channel measurements showed no change in Na(v)1.5 conductance. Mechanosensitivity was reduced in the H558/Q1077del transcript but not in the other three backgrounds. In conclusion, the G298S-SCN5A missense mutation caused a marked reduction of whole cell Na(+) current and loss of function of Na(v)1.5, suggesting SCN5A as a candidate gene in the pathophysiology of IBS.