Cystic fibrosis transmembrane conductance regulator currents in guinea pig pancreatic duct cells: Inhibition by bicarbonate ions

O’Reilly, Catherine M.; Winpenny, John P.; Argent, Barry E.; Gray, Michael A.

doi:10.1016/s0016-5085(00)70372-6

Cited by 63 publications

(73 citation statements)

References 26 publications

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“…We have previously shown using fast whole cell patch clamp recordings (fWCR) that raising extracellular HCO 3 Ϫ inhibits CFTR currents in native guinea pig pancreatic duct cells (26). Our data showed that increasing HCO 3 o Ϫ caused an inhibition of both inward and outward CFTR currents.…”

mentioning

confidence: 54%

“…CFTR inhibition was not caused by changes in either extracellular or intracellular pH or in either the pCO 2 or CO 3 2Ϫ content of the HCO 3 Ϫ -rich solutions. We proposed that this effect of HCO 3 o Ϫ on CFTR was a novel negative feedback mechanism for the control of HCO 3 Ϫ secretion and thus the luminal surface pH in epithelia (26). That external anions can modulate CFTR function is also supported by the recent report of Shcheynikov et al (27).…”

mentioning

confidence: 77%

“…Cells were cultured on glass coverslips for use in patch clamp experiments between 2 and 5 days after plating. Whole cell patch clamp recordings were carried out at room temperature as described previously (26). An EPC-7 amplifier (List Electronic, Darmstadt, Germany) was used to record currents from CHO or BHK cells.…”

Section: Methodsmentioning

confidence: 99%

“…Mean current amplitudes were calculated at E rev Ϯ 60 mV and normalized to cell capacitance (pF) measured using the EPC-7 amplifier. The relative permeability (P x Ϫ/P Cl Ϫ) for each anion (X Ϫ ) compared with Cl Ϫ was determined from changes in E rev (26). Outside out, single channel experiments were conducted on BHK cells at room temperature as described previously (28) at a V m of Ϫ50 mV.…”

Section: Methodsmentioning

confidence: 99%

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Novel Regulation of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Channel Gating by External Chloride

Wright

Gong

Verdon

et al. 2004

Journal of Biological Chemistry

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The cystic fibrosis transmembrane conductance regulator (CFTR) is vital for Cl ؊ and HCO 3 ؊ transport in many epithelia. As the HCO 3 ؊ concentration in epithelial secretions varies and can reach as high as 140 mM, the lumen-facing domains of CFTR are exposed to large reciprocal variations in Cl ؊ and HCO 3 ؊ levels. We have investigated whether changes in the extracellular anionic environment affects the activity of CFTR using the patch clamp technique. In fast whole cell current recordings, the replacement of 100 mM external Cystic fibrosis transmembrane conductance regulator (CFTR) 1 is a cyclic AMP activated epithelial Cl Ϫ channel, the mutation of which causes the potentially fatal inherited disease cystic fibrosis (CF) (1). The CFTR protein is a member of the ABC transporter family and is regulated by phosphorylation (2-4) and by ATP binding and hydrolysis (5-7). Dephosphorylation of CFTR by membrane-bound protein phosphatases is also important in the physiological regulation of channel activity (2,8,9).Although CF is generally considered to result from a defect in Cl Ϫ transport, most of the affected epithelia also transport HCO 3 Ϫ ions. Indeed, recent work suggests that there is a better correlation between defects in CFTR-dependent HCO 3 Ϫ transport than defects in Cl Ϫ transport and the severity of disease (10). HCO 3 Ϫ is an important component of epithelial secretions and, via its buffering role, controls the pH at the epithelial cell surface. It has been reported that CF-affected epithelia, particularly in the gastrointestinal tract, secrete fluid with a more acidic pH than normal epithelia (11,12). Whether this is also true for airway surface liquid is still controversial (13,14), but recent measurements of the pH and HCO 3 Ϫ concentration of fluid secreted by polarized human submucosal gland cells, Calu-3, indicate that these cells are capable of secreting substantial amounts of HCO 3 Ϫ (ϳ80 mM) under appropriate stimulation, a process that would be defective in CF (15). An acidic luminal environment affects the physical properties of mucus (16,17) and promotes bacterial binding to mucins (18,19), both of which may have important implications for CF lung disease (20). As many CF-affected epithelia normally secrete substantial amounts of HCO 3 Ϫ , the luminal HCO 3 Ϫ concentration will vary under different physiological situations. This is particularly the case for the pancreas, a tissue in which CFTR is highly expressed and is severely affected in CF. Although plasma HCO 3 Ϫ is ϳ25 mM, the concentration of HCO 3 Ϫ in pancreatic juice can reach ϳ140 mM in humans, and because pancreatic juice is isotonic with plasma, these high HCO 3 Ϫ concentrations are accompanied by a reciprocal fall in juice Cl Ϫ concentration (21, 22). Thus under physiological conditions, the apical surface of pancreatic duct cells and therefore the extracellular face of CFTR will be exposed to large variations in external Cl Ϫ (Cl o Ϫ ) and HCO 3 Ϫ (HCO 3 o Ϫ ) concentrations. A similar but less extreme situation is also lik...

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confidence: 54%

mentioning

confidence: 77%

Section: Methodsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Novel Regulation of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Channel Gating by External Chloride

Wright

Gong

Verdon

et al. 2004

Journal of Biological Chemistry

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“…A similar model to explain HCO 3 -secretion by the pancreatic duct of non-rodent eutherian mammals has been proposed (Park et al, 2010;Steward et al, 2005). CFTR from eutherian mammals also has a relatively low P HCO3 (Gray et al, 1993;Gray et al, 1990;Linsdell et al, 1997;O'Reilly et al, 2000;Poulsen et al, 1994). However, in the pancreatic duct cells it has been suggested that HCO 3 -secretion occurs via CFTR because there is no mechanism on the basolateral membrane to accumulate Cl -, whereas there is a mechanism for HCO 3 -accumulation (Fernández-Salazar et al, 2004).…”

Section: Discussionmentioning

confidence: 89%

The distribution and expression of CFTR restricts electrogenic anion secretion to the ileum of the brushtail possum, Trichosurus vulpecula

Gill

Bartolo

Demmers

et al. 2011

Journal of Experimental Biology

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SUMMARYIn eutherian mammals, fluid secretion is essential for intestinal function. This is driven by electrogenic Cl -secretion, which involves a NaK2Cl cotransporter (NKCC1) in the enterocyte basolateral membrane and the cystic fibrosis transmembrane conductance regulator (CFTR) in the apical membrane. However, in the possum ileum, NKCC1 expression is low and secretagogues stimulate electrogenic HCO 3 -secretion driven by a basolateral NaHCO 3 cotransporter (pNBCe1). Here we investigated whether electrogenic anion secretion occurs in possum duodenum and jejunum and determined the role of CFTR in possum intestinal anion secretion. Prostaglandin E 2 (PGE 2 ) and forskolin stimulated a large increase in ileal short-circuit current (I sc ), consistent with electrogenic HCO 3 -secretion, but had little effect on the duodenal and jejunal I sc . Furthermore, 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) and N-(2-naphthalenyl)-[(3,5-dibromo-2,4-dihydroxyphenyl)methylene]glycine hydrazide (GlyH101) inhibited cloned possum CFTR in cultured cells and the PGE 2 -stimulated ileal I sc , implicating CFTR in ileal HCO 3 -secretion. Consistent with this, CFTR is expressed in the apical membrane of ileal crypt and lower villous cells, which also express pNBCe1 in the basolateral membrane. In contrast, duodenal and jejunal CFTR expression is low relative to the ileum. Jejunal pNBCe1 expression is also low, whereas duodenal and ileal pNBCe1 expression are comparable. All regions have low NKCC1 expression. These results indicate that cAMP-dependent electrogenic Cl -secretion does not occur in the possum small intestine because of the absence of CFTR and NKCC1. Furthermore, CFTR functions as the apical anion conductance associated with HCO 3 -secretion and its distribution limits electrogenic HCO 3 -secretion to the ileum.

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How to Measure CFTR-Dependent Bicarbonate Transport: From Single Channels to the Intact Epithelium

Hug

Clarke

Gray

2011

Methods in Molecular Biology

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Bicarbonate serves many functions in our body. It is the predominant buffer maintaining a physiological pH in the blood and within our cells. It is also essential for proper digestion of nutrients and solubilization of complex protein mixtures, such as digestive enzymes and mucins, in epithelial secretions. Transepithelial HCO3- transport also drives net fluid secretion in many epithelial tissues including those in the gastrointestinal and reproductive tracts as well as the airways. Indeed, defective bicarbonate secretion is a hallmark of the pathophysiology in the pancreas of most patients suffering from cystic fibrosis. Some, but not all, disease-causing mutations in the CF gene lead to impaired bicarbonate transport when expressed in heterologous systems. Recently developed pharmacological modulators of mutant CFTR have demonstrated an ability to activate chloride transport but little is known about whether they also increase the secretion of bicarbonate. It is therefore essential to assay bicarbonate transport when studying the effect of small molecules on CFTR function. However, due to the chaotropic nature of the ion, the measurement of the absolute bicarbonate concentration and its permeability through CFTR is far from trivial. In this chapter we will review some of the techniques available to measure bicarbonate transport through single ion channels, individual cells, and intact epithelial layers.

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Cystic fibrosis transmembrane conductance regulator currents in guinea pig pancreatic duct cells: Inhibition by bicarbonate ions

Cited by 63 publications

References 26 publications

Novel Regulation of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Channel Gating by External Chloride

Novel Regulation of Cystic Fibrosis Transmembrane Conductance Regulator (CFTR) Channel Gating by External Chloride

The distribution and expression of CFTR restricts electrogenic anion secretion to the ileum of the brushtail possum, Trichosurus vulpecula

How to Measure CFTR-Dependent Bicarbonate Transport: From Single Channels to the Intact Epithelium

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