scholarly journals Lubiprostone stimulates secretion from tracheal submucosal glands of sheep, pigs, and humans

2009 ◽  
Vol 296 (5) ◽  
pp. L811-L824 ◽  
Author(s):  
N. S. Joo ◽  
J. J. Wine ◽  
A. W. Cuthbert
Keyword(s):  
Short Circuit ◽  
Short Circuit Current ◽  
Gland Secretion ◽  
Single Type ◽  
Surface Epithelium ◽  
Chronic Obstructive ◽  
Apical Surface ◽  
Submucosal Gland ◽  
Anion Secretion ◽  
Secretion Rates

Lubiprostone, a putative ClC-2 chloride channel opener, has been investigated for its effects on airway epithelia (tracheas). Lubiprostone is shown to increase submucosal gland secretion in pigs, sheep, and humans and to increase short-circuit current (SCC) in the surface epithelium of pigs and sheep. Use of appropriate blocking agents and ion-substitution experiments shows anion secretion is the driving force for fluid formation in both glands and surface epithelium. From SCC concentration-response relations, it is shown that for apical lubiprostone Kd = 10.5 nM with a Hill slope of 1.08, suggesting a single type of binding site and, from the speed of the response, close to the apical surface, confirmed the rapid blockade by Cd ions. Responses to lubiprostone were reversible and repeatable, responses being significantly larger with ventral compared with dorsal epithelium. Submucosal gland secretion rates following basolateral lubiprostone were, respectively, 0.2, 0.5, and 0.8 nl gl−1 min−1 in humans, sheep, and pigs. These rates dwarf any contribution surface secretion adds to the accumulation of surface liquid under the influence of lubiprostone. Lubiprostone stimulated gland secretion in two out of four human cystic fibrosis (CF) tissues and in two of three disease controls, chronic obstructive pulmonary disease and idiopathic pulmonary fibrosis (COPD/IPF), but in neither type of tissue was the increase significant. Lubiprostone was able to increase gland secretion rates in normal human tissue in the continuing presence of a high forskolin concentration. Lubiprostone had no spasmogenic activity on trachealis muscle, making it a potential agent for increasing airway secretion that may have therapeutic utility.

scholarly journals SARS-COV-2 induced Diarrhea is inflammatory, Ca2+ Dependent and involves activation of calcium activated Cl channels

2021 ◽  
Author(s):  
Mark Donowitz ◽  
Chung-Ming Tse ◽  
Karol Dokladny ◽  
Manmeet Rawat ◽  
Ivy Horwitz ◽  
...  
Keyword(s):  
Short Circuit ◽  
Ussing Chamber ◽  
Short Circuit Current ◽  
Proximal Colon ◽  
K Channel ◽  
Mrna Levels ◽  
Apical Surface ◽  
Live Virus ◽  
Anion Secretion ◽  
Calcium Buffer

ABSTRACTDiarrhea occurs in 2-50% of cases of COVID-19 (∼8% is average across series). The diarrhea does not appear to account for the disease mortality and its contribution to the morbidity has not been defined, even though it is a component of Long Covid or post-infectious aspects of the disease. Even less is known about the pathophysiologic mechanism of the diarrhea. To begin to understand the pathophysiology of COVID-19 diarrhea, we exposed human enteroid monolayers obtained from five healthy subjects and made from duodenum, jejunum, and proximal colon to live SARS-CoV-2 and virus like particles (VLPs) made from exosomes expressing SARS-CoV-2 structural proteins (Spike, Nucleocapsid, Membrane and Envelope). Results: 1) Live virus was exposed apically for 90 min, then washed out and studied 2 and 5 days later. SARS-Cov-2 was taken up by enteroids and live virus was present in lysates and in the apical>>basolateral media of polarized enteroids 48 h after exposure. This is the first demonstration of basolateral appearance of live virus after apical exposure. High vRNA concentration was detected in cell lysates and in the apical and basolateral media up to 5 days after exposure. 2) Two days after viral exposure, cytokine measurements of media showed significantly increased levels of IL-6, IL-8 and MCP-1. 3) Two days after viral exposure, mRNA levels of ACE2, NHE3 and DRA were reduced but there was no change in mRNA of CFTR. NHE3 protein was also decreased. 4) Live viral studies were mimicked by some studies with VLP exposure for 48 h. VLPs with Spike-D614G bound to the enteroid apical surface and was taken up; this resulted in decreased mRNA levels of ACE2, NHE3, DRA and CFTR. 4) VLP effects were determined on active anion secretion measured with the Ussing chamber/voltage clamp technique. S-D614G acutely exposed to apical surface of human ileal enteroids did not alter the short-circuit current (Isc). However, VLPS-D614G exposure to enteroids that were pretreated for ∼24 h with IL-6 plus IL-8 induced a concentration dependent increase in Isc indicating stimulated anion secretion, that was delayed in onset by ∼8 min. The anion secretion was inhibited by apical exposure to a specific calcium activated Cl channel (CaCC) inhibitor (AO1) but not by a specific CFTR inhibitor (BP027); was inhibited by basolateral exposure to the K channel inhibit clortimazole; and was prevented by pretreatment with the calcium buffer BAPTA-AM. 5) The calcium dependence of the VLP-induced increase in Isc was studied in Caco-2/BBe cells stably expressing the genetically encoded Ca2+ sensor GCaMP6s. 24 h pretreatment with IL-6/IL-8 did not alter intracellular Ca2+. However, in IL-6/IL-8 pretreated cells, VLP S-D614G caused appearance of Ca2+waves and an overall increase in intracellular Ca2+ with a delay of ∼10 min after VLP addition. We conclude that the diarrhea of COVID-19 appears to an example of a calcium dependent inflammatory diarrhea that involves both acutely stimulated Ca2+ dependent anion secretion (stimulated Isc) that involves CaCC and likely inhibition of neutral NaCl absorption (decreased NHE3 protein and mRNA and decreased DRA mRNA).

scholarly journals Zinc inhibits cAMP-stimulated Cl secretion via basolateral K-channel blockade in rat ileum

2005 ◽  
Vol 288 (5) ◽  
pp. G956-G963 ◽  
Author(s):  
Kazi Mirajul Hoque ◽  
Vazhaikkurichi M. Rajendran ◽  
Henry J. Binder
Keyword(s):  
Short Circuit ◽  
Basolateral Membrane ◽  
Short Circuit Current ◽  
K Channel ◽  
Isolated Cells ◽  
Rat Ileum ◽  
Anion Secretion ◽  
Cl Secretion ◽  
Ion Absorption ◽  
Isotonic Buffer

Zn, an essential micronutrient and second most abundant trace element in cell and tissues, reduces stool output when administered to children with acute diarrhea. The mechanism by which Zn improves diarrhea is not known but could result from stimulating Na absorption and/or inhibiting anion secretion. The aim of this study was to investigate the direct effect of Zn on intestinal epithelial ion absorption and secretion. Rat ileum was partially stripped of serosal and muscle layers, and the mucosa was mounted in lucite chambers. Potential difference and short-circuit current were measured by conventional current-voltage clamp method.86Rb efflux and uptake were assessed for serosal K channel and Na-K-2Cl cotransport activity, respectively. Efflux experiments were performed in isolated cells preloaded with86Rb in the presence of ouabain and bumetanide, whereas uptake experiments were performed in low-Cl isotonic buffer containing Ba and ouabain. Neither mucosal nor serosal Zn affected glucose-stimulated Na absorption. In contrast, forskolin-induced Cl secretion was markedly reduced by serosal but not mucosal addition of Zn. Zn also substantially reversed the increase in Cl secretion induced by 8-bromoadenosine 3′,5′-cyclic monophosphate (8-BrcAMP) with half-maximal inhibitory concentration of 0.43 mM. In contrast, serosal Zn did not alter Cl secretion stimulated by carbachol, a Ca-dependent agonist. Zn inhibited 8-BrcAMP-stimulated86Rb efflux but not carbachol-stimulated86Rb efflux. Zn had no effect on bumetanide-sensitive86Rb uptake, Na-K-ATPase, or CFTR. We conclude from these studies that Zn inhibits cAMP-induced Cl secretion by blocking basolateral membrane K channels.

Transport properties of toad kidney epithelia in culture

1981 ◽  
Vol 241 (3) ◽  
pp. C154-C159 ◽  
Author(s):  
F. M. Perkins ◽  
J. S. Handler
Keyword(s):  
Electrical Resistance ◽  
Hormonal Regulation ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Transepithelial Electrical Resistance ◽  
Apical Surface ◽  
Osmotic Water ◽  
Sodium Flux ◽  
Regulation Of Transport ◽  
Osmotic Water Flow

The characteristics of a continuous line of toad kidney epithelial cells (A6) are described. These cells form a monolayer epithelium of high transepithelial electrical resistance (about 5,000 omega . cm2). The cells generate a transepithelial potential difference (apical surface negative) of about 9 mV. The short-circuit current is equivalent to net sodium flux. Net sodium flux is stimulated by aldosterone and by analogues of cAMP. The stimulation is readily reversible. Neither urea permeability nor osmotic water flow is altered by analogues of cAMP. Amiloride eliminates 90% of the short-circuit current. Thus A6 cells form an epithelium with several differentiated properties including hormonal regulation of transport.

Evidence that prostaglandin E2 stimulates chloride secretion in cultured A6 renal epithelial cells

1986 ◽  
Vol 250 (3) ◽  
pp. F511-F515 ◽  
Author(s):  
R. Keeler ◽  
N. L. Wong
Keyword(s):  
Epithelial Cells ◽  
Prostaglandin E2 ◽  
Short Circuit ◽  
Basal Medium ◽  
Short Circuit Current ◽  
Chloride Secretion ◽  
Apical Surface ◽  
Renal Epithelial Cells ◽  
Apical Side ◽  
Net Flux

The effects of prostaglandin E2 (PGE2) on the transport of sodium and chloride were studied in cultured A6 renal epithelial cells. PGE2 on the basolateral but not the apical surface increased transmonolayer short-circuit current (Isc) and conductance. These changes could not be inhibited with amiloride or furosemide in the apical medium. Flux measurements showed that although Isc and net flux of sodium were equal in unstimulated cells, after addition of PGE2 the current increased with no corresponding changes in bidirectional or net flux of sodium. Immersing the cells in sodium-free or chloride-free media inhibited the effects of PGE2. Measurements of the simultaneous fluxes of sodium and chloride showed that after PGE2 was added there was a net flux of chloride from the basal to the apical side (secretion) that was equal to the change in Isc. The effects of PGE2 were inhibited by furosemide in the basal medium. We conclude that PGE2 stimulates a process of chloride secretion in A6 cells.

scholarly journals Apical adenosine regulates basolateral Ca2+-activated potassium channels in human airway Calu-3 epithelial cells

2008 ◽  
Vol 294 (6) ◽  
pp. C1443-C1453 ◽  
Author(s):  
Dong Wang ◽  
Ying Sun ◽  
Wei Zhang ◽  
Pingbo Huang
Keyword(s):  
Epithelial Cells ◽  
Adenosine Receptors ◽  
Short Circuit ◽  
Ussing Chamber ◽  
Specific Inhibitor ◽  
Short Circuit Current ◽  
Airway Epithelial Cells ◽  
Intracellular Camp ◽  
Anion Secretion ◽  
Human Airway

In airway epithelial cells, apical adenosine regulates transepithelial anion secretion by activation of apical cystic fibrosis transmembrane conductance regulator (CFTR) via adenosine receptors and cAMP/PKA signaling. However, the potent stimulation of anion secretion by adenosine is not correlated with its modest intracellular cAMP elevation, and these uncorrelated efficacies have led to the speculation that additional signaling pathways may be involved. Here, we showed that mucosal adenosine-induced anion secretion, measured by short-circuit current ( Isc), was inhibited by the PLC-specific inhibitor U-73122 in the human airway submucosal cell line Calu-3. In addition, the Isc was suppressed by BAPTA-AM (a Ca2+ chelator) and 2-aminoethoxydiphenyl borate (2-APB; an inositol 1,4,5-trisphosphate receptor blocker), but not by PKC inhibitors, suggesting the involvement of PKC-independent PLC/Ca2+ signaling. Ussing chamber and patch-clamp studies indicated that the adenosine-induced PLC/Ca2+ signaling stimulated basolateral Ca2+-activated potassium (KCa) channels predominantly via A2B adenosine receptors and contributed substantially to the anion secretion. Thus, our data suggest that apical adenosine activates contralateral K+ channels via PLC/Ca2+ and thereby increases the driving force for transepithelial anion secretion, synergizing with its modulation of ipsilateral CFTR via cAMP/PKA. Furthermore, the dual activation of CFTR and KCa channels by apical adenosine resulted in a mixed secretion of chloride and bicarbonate, which may alter the anion composition in the secretion induced by secretagogues that elicit extracellular ATP/adenosine release. Our findings provide novel mechanistic insights into the regulation of anion section by adenosine, a key player in the airway surface liquid homeostasis and mucociliary clearance.

scholarly journals Regulation of electrolyte transport across cultured endometrial epithelial cells by prolactin

2008 ◽  
Vol 197 (3) ◽  
pp. 575-582 ◽  
Author(s):  
Chatsri Deachapunya ◽  
Sutthasinee Poonyachoti ◽  
Nateetip Krishnamra
Keyword(s):  
Epithelial Cells ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Disulfonic Acid ◽  
Maximum Effect ◽  
Channel Blockers ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Anion Secretion ◽  
Endometrial Epithelial Cells

The effect of prolactin (PRL) on ion transport across the porcine glandular endometrial epithelial cells was studied in primary cell culture using the short-circuit current technique. Addition of 1 μg/ml PRL either to the apical solution or to the basolateral solution produced a peak followed by a sustained increase in Isc, but with a lesser response when PRL was added apically. Basolateral addition of PRL increased the Isc in a concentration-dependent manner with a maximum effect at 1 μg/ml and an effective concentration value of 120 ng/ml. The PRL-stimulated Isc was significantly reduced by pretreatment with an apical addition of 5-nitro-2-(3-phenylpropylamino) benzoic acid (200 μM), diphenylamine-2-carboxylic acid (1 mM) or 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid (200 μM), Cl− channel blockers, but not by amiloride (10 μM), a Na+ channel blocker. In addition, pretreatment with bumetanide (200 μM), a Na+–K+–2Cl− cotransporter inhibitor, in the basolateral solution significantly reduced the PRL-stimulated Isc. Replacement of Cl− or in the bathing solutions also decreased the Isc response to PRL. Pretreatment of the monolayer with AG490 (50 μM), an inhibitor of JAK2 activity significantly inhibited the PRL-induced increase in Isc. Western blot analysis of the porcine endometrial epithelial cells revealed the presence of short isoform of PRL receptor (PRLR-S) that could be regulated by 17β-estradiol. The results of this investigation showed that PRL acutely stimulated anion secretion across the porcine endometrial epithelial cells possibly through PRLR-S present in both apical and basolateral membranes. The PRL response appeared to be mediated by the JAK2-dependent pathway.

scholarly journals Huqi San-Evoked Rat Colonic Anion Secretion through Increasing CFTR Expression

2015 ◽  
Vol 2015 ◽  
pp. 1-12 ◽  
Author(s):  
Xiaowei Xue ◽  
Zhengming Shi ◽  
Wen Wang ◽  
Xiaotong Yu ◽  
Ping Feng ◽  
...  
Keyword(s):  
Protein Content ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Distal Colon ◽  
Transmembrane Conductance Regulator ◽  
Transmembrane Conductance ◽  
Anion Secretion ◽  
Mrna Transcripts ◽  
And Function ◽  
Cystic Fibrosis Transmembrane

Huqi San (HQS) is a Chinese herbal preparation of eight medicinal herbs that promote diuresis, detoxification, blood circulation, and cholestasis. Defects in transporter expression and function can cause cholestasis and jaundice. However, the mechanism of the cholestasis underlying HQS effects, especially on the gastrointestinal tract ion secretion, has not been elucidated. Real-time RT-PCR and Western blotting were used to study the expression and localization of cystic fibrosis transmembrane conductance regulator (CFTR) andα-ENaC in rat alimentary tract, and then the effect of HQS on the ion transport in rat distal colon mucosa was investigated using the short-circuit current (ISC) technique. The results showed that pretreatment with HQS significantly enhanced mRNA transcripts and protein content of CFTR in liver and distal colon but notα-ENaC in alimentary organs. HQS increasesISCand decreases the transepithelial resistance. Pretreatment with epithelial Na+channel blocker did not affect theISCresponses elicited by HQS, but removal of extracellular Cl−or pretreatment with Cl−channel or Na+-K+-2Cl−cotransporter blocker inhibited HQS-elicitedISCresponses. These findings demonstrated that HQS, RA, and RP can stimulate Cl−secretion in the distal colon by increasing the mRNA transcripts and protein content of CFTR in liver and distal colon.

scholarly journals THE INHIBITORY EFFECT OF STROPHANTHIDIN ON SECRETION BY THE ISOLATED GASTRIC MUCOSA

1959 ◽  
Vol 42 (6) ◽  
pp. 1233-1239 ◽  
Author(s):  
I. L. Cooperstein
Keyword(s):  
Gastric Mucosa ◽  
Chloride Transport ◽  
Short Circuit ◽  
Membrane Conductance ◽  
Short Circuit Current ◽  
Inhibitory Effect ◽  
Transport Systems ◽  
Anion Secretion ◽  
Diffusion Component ◽  
Sodium And Potassium

The unidirectional fluxes of Na+ and Cl- were measured across the isolated gastric mucosa of the bullfrog (R. catesbiana). The addition of strophanthidin, a cardiac aglycone, resulted in marked reductions of the spontaneous potential and short-circuit current. Associated with these changes, the isolated gastric mucosa ceased secreting chloride and hydrogen ion. Although the active component of chloride transfer was inhibited, the exchange diffusion component seemed to increase. No significant changes in membrane conductance or sodium flux were noted. Possible mechanisms of strophanthidin inhibition were discussed in view of its effect on chloride transport across the gastric mucosa and on sodium and potassium transfer in other tissues. It was concluded that the cardiac glycosides may not be specific inhibitors of sodium and potassium transport. This non-specific inhibition suggests that active chloride transport is affected by strophanthidin directly and/or anion secretion is dependent upon normal functioning of cation transport systems in the tissue.

Adenosine 3',5'-cyclic monophosphate stimulates chloride secretion in A6 epithelia

1986 ◽  
Vol 251 (5) ◽  
pp. C810-C814 ◽  
Author(s):  
M. Yanase ◽  
J. S. Handler
Keyword(s):  
Short Circuit ◽  
Short Circuit Current ◽  
Chloride Secretion ◽  
Apical Surface ◽  
Cyclic Monophosphate ◽  
A6 Epithelia ◽  
In Series ◽  
Cl Channels ◽  
Solution Bathing ◽  
Stimulation Of

Basal and aldosterone-stimulated short-circuit current (Isc) of A6 epithelia are known to be equivalent to net apical to basal Na flux and are completely inhibited by 0.05 mM amiloride added to the solution bathing the apical surface of the epithelium. In the absence of amiloride, the Isc stimulated by adenosine 3',5'-cyclic monophosphate (cAMP) is also equivalent to net apical to basal Na flux. However, amiloride does not completely inhibit the cAMP-stimulated Isc. In this study, the cAMP-stimulated, amiloride-insensitive Isc was characterized, using vasopressin or forskolin to raise cell cAMP. After basal Isc is inhibited by amiloride, forskolin stimulates Isc, conductance, and bidirectional 36Cl flux. Stimulation of Isc depends on the presence of both Na and Cl; stimulation of conductance depends on the presence of Cl. 36Cl flux studies showed that the cAMP-stimulated, amiloride-insensitive Isc is equivalent to net Cl flux. It is inhibited by ouabain and by furosemide or bumetanide added to the solution bathing the basal surface of the epithelium. In view of the effect of cAMP in some other epithelia, we suggest that cAMP activates apical membrane Cl channels that are in series with a Na-K-Cl cotransporter in the basolateral plasma membrane.

NH2-terminal modification of a channel-forming peptide increases capacity for epithelial anion secretion

2001 ◽  
Vol 280 (3) ◽  
pp. C451-C458 ◽  
Author(s):  
James R. Broughman ◽  
Kathy E. Mitchell ◽  
Roger L. Sedlacek ◽  
Takeo Iwamoto ◽  
John M. Tomich ◽  
...  
Keyword(s):  
Therapeutic Potential ◽  
Short Circuit ◽  
Ussing Chamber ◽  
Glycine Receptor ◽  
Short Circuit Current ◽  
Disulfonic Acid ◽  
Α Subunit ◽  
Anion Secretion ◽  
Anion Conductance ◽  
Selective Channel

A synthetic, channel-forming peptide, derived from the α-subunit of the glycine receptor (M2GlyR), has been synthesized and modified by adding four lysine residues to the NH2 terminus (N-K4-M2GlyR). In Ussing chamber experiments, apical N-K4-M2GlyR (250 μM) increased transepithelial short-circuit current ( I sc) by 7.7 ± 1.7 and 10.6 ± 0.9 μA/cm2 in Madin-Darby canine kidney and T84 cell monolayers, respectively; these values are significantly greater than those previously reported for the same peptide modified by adding the lysines at the COOH terminus (Wallace DP, Tomich JM, Iwamoto T, Henderson K, Grantham JJ, and Sullivan LP. Am J Physiol Cell Physiol 272: C1672–C1679, 1997). N-K4-M2GlyR caused a concentration-dependent increase in I sc ( k [1/2] = 190 μM) that was potentiated two- to threefold by 1-ethyl-2-benzimidazolinone. N-K4-M2GlyR-mediated increases in I sc were insensitive to changes in apical cation species. Pharmacological inhibitors of endogenous Cl− conductances [glibenclamide, diphenylamine-2-dicarboxylic acid, 5-nitro-2-(3-phenylpropylamino)benzoic acid, 4,4′-dinitrostilben-2,2′-disulfonic acid, indanyloxyacetic acid, and niflumic acid] had little effect on N-K4-M2GlyR-mediated I sc. Whole cell membrane patch voltage-clamp studies revealed an N-K4-M2GlyR-induced anion conductance that exhibited modest outward rectification and modest time- and voltage-dependent activation. Planar lipid bilayer studies yielded results indicating that N-K4-M2GlyR forms a 50-pS anion conductance with a k [1/2] for Cl−of 290 meq. These results indicate that N-K4-M2GlyR forms an anion-selective channel in epithelial monolayers and shows therapeutic potential for the treatment of hyposecretory disorders such as cystic fibrosis.

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