A guide to Ussing chamber studies of mouse intestine

Clarke, Lane L.

doi:10.1152/ajpgi.90649.2008

Cited by 328 publications

(308 citation statements)

References 54 publications

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“…To know if the above functional modifications observed in response to HFdiet resulted in rupture of the physical integrity of the epithelial barrier, we examined the ionic conductance of SI segments in HF-and SD-fed mice. Indeed, in the intestine, the paracellular pathway accounts for at least 90% of the total transepithelial ionic conductance as measured in Ussing chambers (53). In the duodenum and jejunum, the measured ionic conductance remained unchanged between HF-fed and SD-fed mice (Fig.…”

Section: Hf Diet Alters the Functional Integrity Of The Ileum Via Cftmentioning

confidence: 85%

“…Forskolin, an activator of adenylate cyclase, activates CFTR, and deletion of the Cftr gene abolishes its effect on epithelial Cl − and HCO 3 − (51, 52). We examined electrogenic activities reflecting the secretion of Cl − and HCO 3 − on the luminal side of the ileal mucosa in HF-fed mice and in SD-fed mice using Ussing chambers (53). Forskolin-induced Cl − and HCO 3 − secretion (Isc ratio) was 37% lower in the distal ileum of HF-fed mice than in SD-fed mice (Fig.…”

Section: Hf Diet Alters the Functional Integrity Of The Ileum Via Cftmentioning

confidence: 99%

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High-fat diet modifies the PPAR-γ pathway leading to disruption of microbial and physiological ecosystem in murine small intestine

Tomas

Mulet

Saffarian

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

203

146

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Diet is among the most important factors contributing to intestinal homeostasis, and basic functions performed by the small intestine need to be tightly preserved to maintain health. Little is known about the direct impact of high-fat (HF) diet on small-intestinal mucosal defenses and spatial distribution of the microbiota during the early phase of its administration. We observed that only 30 d after HF diet initiation, the intervillous zone of the ileum—which is usually described as free of bacteria—became occupied by a dense microbiota. In addition to affecting its spatial distribution, HF diet also drastically affected microbiota composition with a profile characterized by the expansion of Firmicutes (appearance of Erysipelotrichi), Proteobacteria (Desulfovibrionales) and Verrucomicrobia, and decrease of Bacteroidetes (family S24-7) and Candidatus arthromitus. A decrease in antimicrobial peptide expression was predominantly observed in the ileum where bacterial density appeared highest. In addition, HF diet increased intestinal permeability and decreased cystic fibrosis transmembrane conductance regulator (Cftr) and the Na-K-2Cl cotransporter 1 (Nkcc1) gene and protein expressions, leading to a decrease in ileal secretion of chloride, likely responsible for massive alteration in mucus phenotype. This complex phenotype triggered by HF diet at the interface between the microbiota and the mucosal surface was reversed when the diet was switched back to standard composition or when mice were treated for 1 wk with rosiglitazone, a specific agonist of peroxisome proliferator-activated receptor-γ (PPAR-γ). Moreover, weaker expression of antimicrobial peptide-encoding genes and intervillous bacterial colonization were observed in Ppar-γ–deficient mice, highlighting the major role of lipids in modulation of mucosal immune defenses.

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Section: Hf Diet Alters the Functional Integrity Of The Ileum Via Cftmentioning

confidence: 85%

Section: Hf Diet Alters the Functional Integrity Of The Ileum Via Cftmentioning

confidence: 99%

High-fat diet modifies the PPAR-γ pathway leading to disruption of microbial and physiological ecosystem in murine small intestine

Tomas

Mulet

Saffarian

et al. 2016

Proc. Natl. Acad. Sci. U.S.A.

203

146

View full text Add to dashboard Cite

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“…Animal experiments and protocols used to demonstrate the Ussing chamber method and ileal loop experiments were approved by the Institutional Animal Ethics Committee (NICED/CPCSEA/AW/(225)/IAEC-MH/3). Following euthanasia by CO 2 inhalation, the mouse intestine was removed by sharp dissection but not pulling the intestine from its mesenteric attachment to avoid damaging the epithelium, as described elsewhere (47). The intestinal section was opened longitudinally along the mesenteric border so that the antimesenteric mucosa will be situated in the aperture of the chamber.…”

Section: Mutagenesis and Construction Of Ano6 In Pmxs-ires-neomentioning

confidence: 99%

Anoctamin 6 Contributes to Cl− Secretion in Accessory Cholera Enterotoxin (Ace)-stimulated Diarrhea

Aoun

Hayashi

Sheikh

et al. 2016

Journal of Biological Chemistry

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Edited by F. Anne StephensonAccessory cholera enterotoxin (Ace) of Vibrio cholerae has been shown to contribute to diarrhea. However, the signaling mechanism and specific type of Cl ؊ channel activated by Ace are still unknown. We have shown here that the recombinant Ace protein induced I Cl of apical plasma membrane, which was inhibited by classical CaCC blockers. Surprisingly, an Ace-elicited rise of current was neither affected by ANO1 (TMEM16A)-specific inhibitor T16A (inh) -AO1(TAO1) nor by the cystic fibrosis transmembrane conductance regulator (CFTR) blocker, CFTR inh-172. Ace stimulated whole-cell current in Caco-2 cells. However, the apical I Cl was attenuated by knockdown of ANO6 (TMEM16F). This impaired phenotype was restored by

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“…Samples were placed in scintillation vials with 5 ml of scintillation cocktail (Perkin Elmer) and read on a Multi-Purpose Scintillation Counter (model LS6500; Beckman Coulter, Mississauga, ON, Canada). Flux was calculated as described previously (Clarke, 2009;Wilson et al, 2012) and is as follows:…”

Section: Methodsmentioning

confidence: 99%

Acute exposure to ergot alkaloids from endophyte-infected tall fescue does not alter absorptive or barrier function of the isolated bovine ruminal epithelium

Foote

Penner

Walpole

et al. 2014

animal

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Ergot alkaloids in endophyte-infected (Neotyphodium coenophialum) tall fescue (Lolium arundinaceum) have been shown to cause a reduction in blood flow to the rumen epithelium as well as a decrease in volatile fatty acids (VFA) absorption from the washed rumen of steers. Previous data also indicates that incubating an extract of endophyte-infected tall fescue seed causes an increase in the amount of VFA absorbed per unit of blood flow, which could result from an alteration in the absorptive or barrier function of the rumen epithelium. An experiment was conducted to determine the acute effects of an endophyte-infected tall fescue seed extract (EXT) on total, passive or facilitated acetate and butyrate flux across the isolated bovine rumen as well as the barrier function measured by inulin flux and tissue conductance (G t ). Flux of ergovaline across the rumen epithelium was also evaluated. Rumen tissue from the caudal dorsal sac of Holstein steers ( n = 6), fed a common diet, was collected and isolated shortly after slaughter and mounted between two halves of Ussing chambers. In vitro treatments included vehicle control (80% methanol, 0.5% of total volume), Low EXT (50 ng ergovaline/ml) and High EXT (250 ng ergovaline/ml). Results indicate that there is no effect of acute exposure to ergot alkaloids on total, passive or facilitated flux of acetate or butyrate across the isolate bovine rumen epithelium ( P > 0.51). Inulin flux ( P = 0.16) and G t ( P > 0.17) were not affected by EXT treatment, indicating no alteration in barrier function due to acute ergot alkaloid exposure. Ergovaline was detected in the serosal buffer of the High EXT treatment indicating that the flux rate is~0.25 to 0.44 ng/cm 2 per hour. Data indicate that specific pathways for VFA absorption and barrier function of the rumen epithelium are not affected by acute exposure to ergot alkaloids from tall fescue at the concentrations tested. Ergovaline has the potential to be absorbed from the rumen of cattle that could contribute to reduced blood flow and motility and lead to reduced growth rates of cattle.Keywords: bovine, ergovaline absorption, fescue toxicosis, volatile fatty acid absorption ImplicationsResults from this experiment show that ergot alkaloids present in endophyte-infected tall fescue do not have an impact on nutrient absorption or barrier function of the isolated rumen epithelium of cattle following acute exposure. This indicates that previously observed reductions in nutrient absorption in live animals exposed to ergot alkaloids requires chronic exposure or is driven by the reduced blood flow. Results also show that ergovaline, one of the main causative agents of fescue toxicosis, is absorbed across the rumen epithelium, although it is at a fairly slow rate ∼1% of a daily dose.

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A guide to Ussing chamber studies of mouse intestine

Abstract: Clarke LL. A guide to Ussing chamber studies of mouse intestine.

Cited by 328 publications

References 54 publications

High-fat diet modifies the PPAR-γ pathway leading to disruption of microbial and physiological ecosystem in murine small intestine

High-fat diet modifies the PPAR-γ pathway leading to disruption of microbial and physiological ecosystem in murine small intestine

Anoctamin 6 Contributes to Cl− Secretion in Accessory Cholera Enterotoxin (Ace)-stimulated Diarrhea

Acute exposure to ergot alkaloids from endophyte-infected tall fescue does not alter absorptive or barrier function of the isolated bovine ruminal epithelium

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