Background-Cellular changes associated with diabetic and idiopathic gastroparesis are not well described.Aim-Describe histologic abnormalities in gastroparesis and compare findings in idiopathic versus diabetic gastroparesis.Methods-Full thickness gastric body biopsies were obtained from 40 gastroparetics (20 diabetic) and matched controls. Sections were stained for H&E and trichrome, and immunolabeled with antibodies against PGP 9.5, nNOS, VIP, substance P and tyrosine hydroxylase to quantify nerves, S100β for glia, Kit for interstitial cells of Cajal (ICC), CD45 and CD68, for immune cells and smoothelin for smooth muscle cells. Tissue was also examined by transmission electron microscopy (TEM).Results-Histological abnormalities were found in 83% of patients. Most common defects were loss of ICC with remaining ICC showing injury, an abnormal immune infiltrate containing macrophages, and decreased nerve fibers. On light microscopy, no significant differences were found between diabetic and idiopathic gastroparesis with the exception of nNOS expression which was decreased in more idiopathic gastroparetics (40%) compared to diabetic (20%) patients by visual grading. On electron microscopy, a markedly increased connective tissue stroma was present in both disorders.Conclusion-This study suggests that on full thickness biopsies, cellular abnormalities are found in the majority of patients with gastroparesis. Most common findings were loss of Kit expression suggesting loss of ICC and an increase in CD45 and CD68 immunoreactivity. These findings suggest that examination of tissue can lead to valuable insights into the pathophysiology of these disorders and offers hope that new therapeutic targets can be found.

Background & Aims Chronic unexplained nausea and vomiting (CUNV) is a debilitating disease of unknown cause. Symptoms of CUNV substantially overlap with those of gastroparesis, so the diseases therefore may share pathophysiologic features. We investigated this hypothesis by quantifying densities of interstitial cells of Cajal (ICCs) and mapping slow wave abnormalities in patients with CUNV vs controls. Methods Clinical data and gastric biopsy specimens were collected from 9 consecutive patients with at least 6 months of continuous symptoms of CUNV, but normal gastric emptying, treated at the University of Mississippi Medical Center, and from 9 controls (individuals undergoing bariatric surgery but free of gastrointestinal disease or diabetes). ICCs were counted and ultrastructural analyses were performed on tissue samples. Slow-wave propagation profiles were defined by high-resolution electrical mapping (256 electrodes; 36 cm2). Results from patients with CUNV were compared to those of controls as well as patients with gastroparesis who were previously studied by identical methods. Results Patients with CUNV had fewer ICCs than controls (mean 3.5 vs 5.6 bodies/field; P<.05), with mild ultrastructural abnormalities in the remaining ICCs. Slow-wave dysrhythmias were identified in all 9 subjects with CUNV vs only 1/9 controls. Dysrhythmias included abnormalities of initiation (stable ectopic pacemakers, unstable focal activities) and conduction (retrograde propagation, wave front collisions, conduction blocks, and re-entry), operating across bradygastric, normal (range 2.4−3.7 cycles/min), and tachygastric frequencies; dysrhythmias showed velocity anisotropy (mean 3.3 mm/s longitudinal vs 7.6 mm/s circumferential, P<.01). ICCs were less depleted in patients with CUNV than those with gastroparesis (mean 3.5 vs 2.3 bodies/field; P<.05), but slow-wave dysrhythmias were similar between groups. Conclusions This study defined cellular and bioelectrical abnormalities in patients with CUNV, including the identification of slow-wave re-entry. Pathophysiologic features of CUNV were observed to be similar to those of gastroparesis, indicating that they could be spectra of the same disorder. These findings offer new insights into the pathogenesis of CUNV and may help to inform future treatments.

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.

The effect of age on the anatomy and function of the human colon is incompletely understood. The prevalence of disorders in adults such as constipation increase with age but it is unclear if this is due to confounding factors or age-related structural defects. The aim of this study was to determine number and subtypes of enteric neurons and neuronal volumes in the human colon of different ages. Normal colon (descending and sigmoid) from 16 patients (9 male) was studied; ages 33–99. Antibodies to HuC/D, ChAT, nNOS, and PGP9.5 were used. Effect of age was determined by testing for linear trends using regression analysis. In the myenteric plexus, number of Hu-positive neurons declined with age (slope = −1.3 neurons/mm/10yrs, p =0.03). The number of ChAT-positive neurons also declined with age (slope = −1.1 neurons/mm/10yrs of age, p=0.02). The number of nNOS-positive neurons did not decline with age. As a result, the ratio of nNOS to Hu increased (slope= 0.03 per 10yrs of age, p=0.01). In the submucosal plexus, the number of neurons did not decline with age (slope = − 0.3 neurons/mm/10 yrs, p =0.09). Volume of nerve fibers in the circular muscle and volume of neuronal structures in the myenteric plexus did not change with age. In conclusion, the number of neurons in the human colon declines with age with sparing of nNOS- positive neurons. This change was not accompanied by changes in total volume of neuronal structures suggesting compensatory changes in the remaining neurons.

Background-Cellular changes associated with diabetic (DG) and idiopathic gastroparesis (IG) have recently been described from patients enrolled in the Gastroparesis Clinical Research Consortium. The association of these cellular changes with gastroparesis symptoms and gastric emptying is unknown.Aim-Relate cellular changes to symptoms and gastric emptying in patients with gastroparesis.Methods-Earlier, using full thickness gastric body biopsies from 20 DG, 20 IG and 20 matched controls, we found decreased interstitial cells of Cajal (ICC) and enteric nerves and an increase in immune cells in both DG and IG. Here, demographic, symptoms (gastroparesis cardinal symptom index score), and gastric emptying were related to cellular alterations using Pearson's correlation coefficients.Results-ICC counts inversely correlated with 4 hours gastric retention in DG but not in IG (r= −0.6, p=0.008, DG, r=0.2, p=0.4, IG). There was also a significant correlation between loss of ICC and enteric nerves in DG but not in IG (r=0.5, p=0.03 for DG, r=0.3, p=0.16, IG). IG with a myenteric immune infiltrate scored higher on the average GCSI (3.6±0.7 vs 2.7±0.9, p=0.05) and nausea score (3.8±0.9 vs 2.6±1.0, p=0.02) as compared to those without an infiltrate. Conclusions-In

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 gaseous molecule hydrogen sulfide (H 2 S) has been proposed as an endogenous signal molecule and neuromodulator in mammals. Using a newly developed method, we report here for the first time the ability of intact and living brain and colonic tissue in the mouse to generate and release H 2 S. This production occurs through the activity of two enzymes, cystathionine-c-lyase and cystathionine-b-synthase. The quantitative expression of messenger RNA and protein localization for both enzymes are described in the liver, brain, and colon. Expression levels of the enzymes vary between tissues and are differentially distributed. The observation that, tissues that respond to exogenously applied H 2 S can endogenously generate the gas, strongly supports its role as an endogenous signal molecule.

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.