p-Aminohippurate transport in canine tracheal epithelium

1985 ◽  
Vol 59 (5) ◽  
pp. 1585-1589 ◽  
Author(s):  
M. M. Cloutier ◽  
K. M. Lesniak
Keyword(s):  
Short Circuit ◽  
Organic Anion ◽  
Chloride Secretion ◽  
Open Circuit ◽  
Tracheal Epithelium ◽  
Ussing Chambers ◽  
Saturation Kinetics ◽  
Absorptive Flux ◽  
Menten Constant ◽  
Specialized System

The unidirectional fluxes of 20, 100, 500, and 2,000 microM rho-aminohippurate (PAH) were measured under open- and short-circuit conditions in canine tracheal epithelium mounted as flat sheets in Ussing chambers. In tissues pretreated with mucosal indomethacin (10(-6) M) and amiloride (10(-4) M), unidirectional PAH fluxes under short-circuit conditions increased with increasing bath concentrations but there was no significant net PAH transport. After stimulation of chloride secretion by mucosal cyclic adenosine 3′,5′ -cyclic monophosphate (cAMP 10(-3) M), there was a significant increase in the secretory flux of PAH and a significant decrease in the absorptive flux of PAH. This resulted in net PAH secretion that demonstrated saturation kinetics with an apparent Michaelis-Menten constant of 754 microM by Lineweaver-Burk analysis. Intracellular concentrations of PAH were 0.4–1.2 times bath concentrations after pretreatment with indomethacin and amiloride and increased to 2.6–3.3 times bath concentrations after cAMP. Under open-circuit conditions, secretory PAH flux decreased and absorptive flux increased resulting in net PAH absorption. We conclude from these early studies that the canine tracheal epithelium possesses a specialized system for the transport of organic anions in the airways and that this transport system may share many similarities with organic anion transport in the kidney.

Ion transport characteristics of the murine trachea and caecum

1992 ◽  
Vol 82 (6) ◽  
pp. 667-672 ◽  
Author(s):  
S. N. Smith ◽  
E. W. F. W. Alton ◽  
D. M. Geddes
Keyword(s):  
Cystic Fibrosis ◽  
Ion Transport ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Chloride Secretion ◽  
Open Circuit ◽  
Potential Difference ◽  
Cystic Fibrosis Gene ◽  
Tissue Type ◽  
Ussing Chambers

1. The basic defect in cystic fibrosis relates to abnormalities of ion transport in affected tissues, such as the respiratory and gastrointestinal tracts. The identification of the cystic fibrosis gene has enabled studies on the production of a cystic fibrosis transgenic mouse to be undertaken. Knowledge of normal ion transport will be necessary for the validation of any such animal model. We have therefore characterized selected responses of the murine trachea and caecum mounted in ‘mini’ Ussing chambers under open-circuit conditions. 2. Basal values for the trachea were: potential difference, 1.1 mV (sem 0.2; n=18); equivalent short-circuit current, 20.4 μA/cm2 (3.6); conductance, 18.2 mS/cm2 (1.7). Corresponding values for the caecum were: potential difference, 0.7 mV (0.1; n=18); equivalent short-circuit current, 11.0 μA/cm2 (1.6); conductance, 14.5 mS/cm2 (1.4). 3. Amiloride (10 μmol/l) produced a significant (P < 0.001) fall in potential difference of 43.0% (5.7) in the trachea, but had no significant effect in the caecum. 4. Subsequently, one of three protocols was used to assess the capacity of either tissue for chloride secretion. Addition of a combination of forskolin (1 μmol/l) and zardaverine (10 μmol/l) produced rises in the potential difference of 873% (509) in the trachea and 399% (202) in the caecum. Both A23187 (10 μmol/l) and phorbol dibutyrate (10 nmol/l) increased tracheal potential difference by 350% (182) and 147% (47), respectively. Neither had a significant effect in the caecum. 5. Subsequent addition of bumetanide caused a fall in the stimulated potential difference of between 39.8% and 71.7%, depending on secretagogue and tissue type. 6. When a homozygous transgenic cystic fibrosis mouse becomes available, these responses should allow such an animal to be distinguished from normal or heterozygous mice.

Effects of amphotericin B on ion and fluid movement across dog tracheal epithelium

1983 ◽  
Vol 55 (4) ◽  
pp. 1257-1261 ◽  
Author(s):  
I. Nathanson ◽  
J. H. Widdicombe ◽  
J. A. Nadel
Keyword(s):  
Amphotericin B ◽  
Short Circuit ◽  
Open Circuit ◽  
Tracheal Epithelium ◽  
Fluid Movement ◽  
Ussing Chambers ◽  
Cl Secretion ◽  
Special Apparatus ◽  
Net Fluxes

Ion fluxes or fluid flow were measured across sheets of dog tracheal epithelium mounted in Ussing chambers or a special apparatus, respectively. Under short-circuit conditions, luminal amphotericin B (3 X 10(-5) M) caused an inhibition of net Cl secretion and an increase in net Na absorption across paired tissues. In paired tissues under resting open-circuit conditions, there was no significant net transepithelial flux of either Cl or Na. Amphotericin B induced significant net fluxes of both Cl and Na toward the serosal side. In separate tissues from the same animals, there was no significant transepithelial fluid movement under resting conditions. Amphotericin B caused a net absorption of fluid. The absorption of salt and fluid in amphotericin B-treated tissues was abolished by ouabain. We conclude that stimulation of active Na transport by amphotericin B leads to fluid absorption. In vivo, the movement of fluid across the dog tracheal epithelium may be dependent on a balance between active Cl secretion and active Na absorption.

Calcium transport in turtle bladder

1987 ◽  
Vol 253 (6) ◽  
pp. R917-R921
Author(s):  
S. Sabatini ◽  
N. A. Kurtzman
Keyword(s):  
Calcium Transport ◽  
Calcium Absorption ◽  
Short Circuit ◽  
Open Circuit ◽  
Calcium Flux ◽  
Bladder Epithelium ◽  
Control Value ◽  
Sodium Calcium ◽  
Turtle Bladder ◽  
Absorptive Flux

Unidirectional 45Ca fluxes were measured in the turtle bladder under open-circuit and short-circuit conditions. In the open-circuited state net calcium flux (JnetCa) was secretory (serosa to mucosa) and was 388.3 +/- 84.5 pmol.mg-1.h-1 (n = 20, P less than 0.001). Ouabain (5 X 10(-4) M) reversed JnetCa to an absorptive flux (serosal minus mucosal flux = -195.8 +/- 41.3 pmol.mg-1.h-1; n = 20, P less than 0.001). Amiloride (1 X 10(-5) M) reduced both fluxes such that JnetCa was not significantly different from zero. Removal of mucosal sodium caused net calcium absorption; removal of serosal sodium caused calcium secretion. When bladders were short circuited, JnetCa decreased to approximately one-third of control value but remained secretory (138.4 +/- 54.3 pmol.mg-1.h-1; n = 9, P less than 0.025). When ouabain was added under short-circuit conditions, JnetCa was similar in magnitude and direction to ouabain under open-circuited conditions (i.e., absorptive). Tissue 45Ca content was approximately equal to 30-fold lower when the isotope was placed in the mucosal bath, suggesting that the apical membrane is the resistance barrier to calcium transport. The results obtained in this study are best explained by postulating a Ca2+-ATPase on the serosa of the turtle bladder epithelium and a sodium-calcium antiporter on the mucosa. In this model, the energy for calcium movement would be supplied, in large part, by the Na+-K+-ATPase. By increasing cell sodium, ouabain would decrease the activity of the mucosal sodium-calcium exchanger (or reverse it), uncovering active calcium transport across the serosa.

Endothelin stimulates chloride secretion across canine tracheal epithelium

1991 ◽  
Vol 261 (2) ◽  
pp. L188-L194 ◽  
Author(s):  
P. I. Plews ◽  
Z. A. Abdel-Malek ◽  
C. A. Doupnik ◽  
G. D. Leikauf
Keyword(s):  
Epithelial Cells ◽  
Short Circuit ◽  
Second Messengers ◽  
Short Circuit Current ◽  
Chloride Secretion ◽  
Tracheal Epithelium ◽  
Membrane Phospholipids ◽  
Cl Secretion ◽  
Tracheal Epithelial Cells ◽  
Tracheal Epithelial

The endothelins (ET) are a group of isopeptides produced by a number of cells, including canine tracheal epithelial cells. Because these compounds are endogenous peptides that may activate eicosanoid metabolism, we investigated the effects of ET on Cl secretion in canine tracheal epithelium. Endothelin 1 (ET-1) was found to produce a dose-dependent change in short-circuit current (Isc) that increased slowly and reached a maximal value within 10-15 min. When isopeptides of ET were compared, 300 nM ET-1 and ET-2 produced comparable maximal increases in Isc, whereas ET-3 produced smaller changes in Isc (half-maximal concentrations of 2.2, 7.2, and 10.4 nM, respectively). Ionic substitution of Cl with nontransported anions, iodide and gluconate, reduced ET-1-induced changes in Isc. Furthermore, the response was inhibited by the NaCl cotransport inhibitor, furosemide. In paired tissues, ET-1 significantly increased mucosal net 36Cl flux without significant effect on 22Na flux. The increase in Isc induced by ET was diminished by pretreatment with indomethacin. The second messengers mediating the increase in Isc were investigated in cultured canine tracheal epithelial cells. ET-1 stimulated the release of [3H]arachidonate from membrane phospholipids, increased intracellular Ca2+ (occasionally producing oscillations), and increased adenosine 3',5'-cyclic monophosphate accumulation. The latter was diminished by indomethacin. Thus ET is a potent agonist of Cl secretion (with the isopeptides having the following potency: ET-1 greater than or equal to ET-2 greater than ET-3) and acts, in part, through a cyclooxygenase-dependent mechanism.

Mechanisms of sodium and chloride transport across equine tracheal epithelium

1990 ◽  
Vol 259 (6) ◽  
pp. L459-L467 ◽  
Author(s):  
G. J. Tessier ◽  
T. R. Traynor ◽  
M. S. Kannan ◽  
S. M. O3'Grady
Keyword(s):  
Chloride Transport ◽  
Short Circuit ◽  
Basolateral Membrane ◽  
Short Circuit Current ◽  
Ringer Solution ◽  
Tracheal Epithelium ◽  
Ussing Chambers ◽  
Cl Secretion ◽  
Cotransport System ◽  
Ethanesulfonic Acid

Equine tracheal epithelium, stripped of serosal muscle, mounted in Ussing chambers, and bathed in plasmalike Ringer solution generates a serosa-positive transepithelial potential of 10–22 mV and a short-circuit current (Isc) of 70–200 microA/cm2. Mucosal amiloride (10 microM) causes a 40–60% decrease in Isc and inhibits the net transepithelial Na flux by 95%. Substitution of Cl with gluconate resulted in a 30% decrease in basal Isc. Bicarbonate substitution with 20 mM N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid decreased the Isc by 21%. The Cl-dependent Isc was inhibited by serosal addition of 1 mM amiloride. Bicarbonate replacement or serosal amiloride (1 mM) inhibits the net Cl flux by 72 and 69%, respectively. Bicarbonate replacement significantly reduces the effects of serosal amiloride (1 mM) on Isc, indicating its effect is HCO3 dependent. Addition of 8-bromoadenosine 3',5'-cyclic monophosphate (8-BrcAMP; 100 microM) causes a 40% increase in Isc. This effect is inhibited by subsequent addition of 10 microM serosal bumetanide. Bumetanide (10 microM) reduces net Cl secretion following stimulation with 8-BrcAMP (100 microM). Serosal addition of BaCl2 (1 mM) causes a reduction in Isc equal to that following Cl replacement in the presence or absence of 100 microM cAMP. These results suggest that 1) Na absorption depends on amiloride-inhibitable Na channels in the apical membrane, 2) Cl influx across the basolateral membrane occurs by both a Na-H/Cl-HCO3 parallel exchange mechanism under basal conditions and by a bumetanide-sensitive Na-(K?)-Cl cotransport system under cAMP-stimulated conditions, and 3) basal and cAMP-stimulated Cl secretion depends on Ba-sensitive K channels in the basolateral membrane.

Role of CFTR in chloride secretion across human tracheal epithelium

1995 ◽  
Vol 269 (5) ◽  
pp. L561-L566 ◽  
Author(s):  
B. Q. Shen ◽  
R. J. Mrsny ◽  
W. E. Finkbeiner ◽  
J. H. Widdicombe
Keyword(s):  
Apical Membrane ◽  
Short Circuit ◽  
Primary Cultures ◽  
Short Circuit Current ◽  
Chloride Secretion ◽  
Culture Conditions ◽  
Tracheal Epithelium ◽  
Disulfonic Acid ◽  
Acetoxymethyl Ester ◽  
Cl Channels

We have tested two hypotheses: 1) the cystic fibrosis transmembrane conductance regulator (CFTR) represents the predominant Cl conductance in the apical membrane of human tracheal epithelium, and 2) CFTR in this tissue is close to maximally activated under baseline conditions. In support of the first hypothesis, we found 1) when the level of differentiation of cultures was varied by varying the culture conditions, there was a significant positive correlation between the levels of CFTR and the magnitude of mediator-induced Cl secretion. 2) Amiloride-insensitive baseline short-circuit current (Isc) and mediator-induced increases in Isc were inhibited by diphenylamine-2-carboxylic acid (DPAC) but not by 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS), a pharmacology consistent with passage of apical membrane Cl current through CFTR; Ca-activated Cl channels are inhibited by DIDS but not by DPAC. 3) Raising temperature from 22 degrees to 37 degrees C increased 125I efflux, and this increase was inhibited by DPAC and blockers of protein kinase A, but not by DIDS or 1,2-bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid-acetoxymethyl ester. In support of the second hypothesis, we have earlier shown [M. Yamaya, W.E. Finkbeiner, S.Y. Chun, and J.H. Widdicombe. Am. J. Physiol. 262 (Lung Cell. Mol. Physiol. 6): L713-L724, 1992] that adenosine 3',5'-cyclic monophosphate (cAMP)-elevating agents are essentially without effect on Isc across primary cultures of human tracheal epithelium. Here, we further show that these agents are also usually without effect on 125I efflux; the mean increase in efflux in response to elevating cAMP was approximately 20% that of raising temperature from 22 degrees to 37 degrees C.

Tin protoporphyrin induces intestinal chloride secretion by inducing light oxidation processes

2007 ◽  
Vol 292 (5) ◽  
pp. C1906-C1914 ◽  
Author(s):  
Aliye Uc ◽  
Krzysztof J. Reszka ◽  
Garry R. Buettner ◽  
John B. Stokes
Keyword(s):  
Short Circuit ◽  
Specific Inhibitor ◽  
Chloride Secretion ◽  
Epithelial Cell Line ◽  
Intestinal Epithelial ◽  
Paramagnetic Resonance ◽  
Ussing Chambers ◽  
Oxidative Processes ◽  
Tin Protoporphyrin ◽  
Intestinal Chloride Secretion

Heme induces Cl− secretion in intestinal epithelial cells, most likely via carbon monoxide (CO) generation. The major source of endogenous CO comes from the degradation of heme via heme oxygenase (HO). We hypothesized that an inhibitor of HO activity, tin protoporphyrin (SnPP), may inhibit the stimulatory effect of heme on Cl− secretion. To test this hypothesis, we treated an intestinal epithelial cell line (Caco-2 cells) with SnPP. In contrast to our expectations, Caco-2 cells treated with SnPP had an increase in their short-circuit currents ( Isc) in Ussing chambers. This effect was observed only when the system was exposed to ambient light. SnPP-induced Isc was caused by Cl− secretion because it was inhibited in Cl−-free medium, with ouabain or 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB). The Cl− secretion was not via activation of the CFTR, because a specific inhibitor had no effect. Likewise, inhibitors of adenylate cyclase and guanylate cyclase had no effect on the enhanced Isc. SnPP-induced Isc was inhibited by the antioxidant vitamins, α-tocopherol and ascorbic acid. Electron paramagnetic resonance experiments confirmed that oxidative reactions were initiated with light in cells loaded with SnPP. These data suggest that SnPP-induced effects may not be entirely due to the inhibition of HO activity but rather to light-induced oxidative processes. These novel effects of SnPP-photosensitized oxidation may also lead to a new understanding of how intestinal Cl− secretion can be regulated by the redox environment of the cell.

Mucosal histamine inhibits Na absorption and stimulates Cl secretion across equine tracheal epithelium

1991 ◽  
Vol 261 (6) ◽  
pp. L456-L461 ◽  
Author(s):  
G. J. Tessier ◽  
T. R. Traynor ◽  
M. S. Kannan ◽  
S. M. O'Grady
Keyword(s):  
Apical Membrane ◽  
Short Circuit ◽  
Membrane Surface ◽  
Short Circuit Current ◽  
Ringer Solution ◽  
Tracheal Epithelium ◽  
Ussing Chambers ◽  
Cl Secretion ◽  
Subsequent Addition ◽  
Over Time

When the equine tracheal epithelium is mounted in Ussing chambers and bathed in plasma-like Ringer solution, the tissue generates a lumen-negative transepithelial potential (PD) of 22 mV and a short-circuit current (Isc) of 70-200 microA/cm2. Mucosal addition of 10 microM histamine produces a transient increase in the Isc followed by a return to baseline or below. Mucosal addition of 2 microM diphenhydramine inhibits the Isc response to mucosal histamine, whereas 100 microM mucosal cimetidine produces no effect. The average initial increases in Isc over time for mucosal vs. serosal histamine addition are significantly different (17.32 +/- 2.8 and 3.76 +/- 0.69 microA/min, respectively). Pretreatment with mucosal amiloride significantly prolongs the effect of mucosal histamine on Isc over a 20-min period from 4.73 +/- 0.33 to 15.48 +/- 3.16 microA. When Cl is replaced by gluconate, mucosal histamine addition results in a gradual decrease in Isc and significantly reduces the effect of mucosal amiloride on Isc from 80.8% to 54.9%. Mucosal histamine inhibits the net transepithelial Na flux by 42% and stimulates the secretion of Cl by 106%. Subsequent addition of serosal bumetanide decreases net Cl secretion by 70% These results suggest that histamine stimulates bumetanide-sensitive Cl secretion and inhibits amiloride-sensitive Na absorption; these effects are mediated by H1 receptors at the apical membrane surface

Prostaglandin-induced recovery of barrier function in porcine ileum is triggered by chloride secretion

1999 ◽  
Vol 276 (1) ◽  
pp. G28-G36 ◽  
Author(s):  
Anthony T. Blikslager ◽  
Malcolm C. Roberts ◽  
Robert A. Argenzio
Keyword(s):  
Short Circuit ◽  
Second Messengers ◽  
Short Circuit Current ◽  
Chloride Secretion ◽  
Ringer Solution ◽  
Signal Recovery ◽  
Osmotic Gradient ◽  
Current Increase ◽  
Ussing Chambers

We have previously shown that PGI2 and PGE2 have a synergistic role in restoring electrical transepithelial resistance ( R) in ischemia-injured porcine ileum via the second messengers Ca2+ and cAMP. Because Ca2+ and cAMP stimulate Cl− secretion, we assessed the role of PG-induced Cl−secretion in recovery of R. Mucosa from porcine ileum subjected to ischemia for 45 min was mounted in Ussing chambers and bathed in indomethacin and Ringer solution. Addition of PGs stimulated a twofold increase in R, which was preceded by elevations in short-circuit current (increase of 25 μA/cm2). The PG-induced effect on R was partially inhibited with bumetanide, an inhibitor of Cl− secretion. The remaining elevations in R were similar in magnitude to those induced in ischemic tissues by amiloride, an inhibitor of Na+ absorption. Treatment with 10−4 M 8-bromo-cGMP or 300 mosM mucosal urea resulted in elevations in R similar to those attained with PG treatment. PGs signal recovery of Rvia induction of Cl−secretion and inhibition of Na+absorption, possibly by establishing a transmucosal osmotic gradient.

Changes in intracellular sodium with chloride secretion in dog tracheal epithelium

1986 ◽  
Vol 250 (4) ◽  
pp. C646-C650 ◽  
Author(s):  
S. R. Shorofsky ◽  
M. Field ◽  
H. A. Fozzard
Keyword(s):  
Steady State ◽  
Kinetic Models ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Chloride Secretion ◽  
Tracheal Epithelium ◽  
Cl Secretion ◽  
Intracellular Na Activity ◽  
Sufficient Energy ◽  
Stimulation Of

Na-selective microelectrodes were employed to investigate the mechanism of Cl secretion by canine tracheal epithelium. In control tissues with a mean short-circuit current (Isc) of 30.1 microA/cm2, the intracellular Na activity (aiNa) was 10.7 mM. Following steady-state stimulation of Cl secretion with epinephrine (Isc = 126.4 microA/cm2), aiNa was 21.3 mM. These data indicate that there is sufficient energy in the Na gradient to drive Cl secretion by this tissue. When analyzed with simple kinetic models for the Na-K pump, they also suggest that the basolateral entry step involves the Na-K-2Cl cotransporter.

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