Na+/Ca2+ exchange regulates Ca2+-dependent duodenal mucosal ion transport and HCO3− secretion in mice

2005 ◽  
Vol 288 (3) ◽  
pp. G457-G465 ◽  
Author(s):  
Hui Dong ◽  
Zachary M. Sellers ◽  
Anders Smith ◽  
Jimmy Y. C. Chow ◽  
Kim E. Barrett
Keyword(s):  
Western Blot ◽  
Ion Transport ◽  
Muscarinic Receptors ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Duodenal Mucosa ◽  
Ussing Chambers ◽  
Intracellular Ca ◽  
Ph Stat ◽  
Stimulation Of

Stimulation of muscarinic receptors in duodenal mucosa raises intracellular Ca2+, which regulates ion transport, including HCO3− secretion. However, the underlying Ca2+ handling mechanisms are poorly understood. The aim of the present study was to determine whether Na+/Ca2+ exchanger (NCX) plays a role in the regulation of duodenal mucosal ion transport and HCO3− secretion by controlling Ca2+ homeostasis. Mouse duodenal mucosa was mounted in Ussing chambers. Net ion transport was assessed as short-circuit current ( Isc), and HCO3− secretion was determined by pH-stat. Expression of NCX in duodenal mucosae was analyzed by Western blot, and cytosolic Ca2+ in duodenocytes was measured by fura 2. Carbachol (100 μM) increased Isc in a biphasic manner: an initial transient peak within 2 min and a later sustained plateau starting at 10 min. Carbachol-induced HCO3− secretion peaked at 10 min. 2-Aminoethoxydiphenylborate (2-APB, 100 μM) or LiCl (30 mM) significantly reduced the initial peak in Isc by 51 or 47%, respectively, and abolished the plateau phase of Isc without affecting HCO3− secretion induced by carbachol. Ryanodine (100 μM), caffeine (10 mM), and nifedipine (10 μM) had no effect on either response to carbachol. In contrast, nickel (5 mM) and KB-R7943 (10–30 μM) significantly inhibited carbachol-induced increases in duodenal mucosal Isc and HCO3− secretion. Western blot analysis showed expression of NCX1 proteins in duodenal mucosae, and functional NCX in duodenocytes was demonstrated in Ca2+ imaging experiments where Na+ depletion elicited Ca2+ entry via the reversed mode of NCX. These results indicate that NCX contributes to the regulation of Ca2+-dependent duodenal mucosal ion transport and HCO3− secretion that results from stimulation of muscarinic receptors.

Regulation of colonic ion transport by GRP. I. GRP stimulates transepithelial K and Na secretion

1996 ◽  
Vol 270 (3) ◽  
pp. C848-C858 ◽  
Author(s):  
T. R. Traynor ◽  
S. M. O'Grady
Keyword(s):  
Ion Transport ◽  
Short Circuit ◽  
Basolateral Membrane ◽  
Short Circuit Current ◽  
Distal Colon ◽  
Transient Increase ◽  
Transport Pathway ◽  
Ussing Chambers ◽  
Intracellular Ca ◽  
Distal Colon Epithelium

Regulation of electrolyte transport across porcine distal colon epithelium by gastrin-releasing peptide (GRP) was examined using mucosal sheets mounted in Ussing chambers. Serosal GRP produced a biphasic response consisting of a transient increase in short-circuit current (ISC) followed by a long-lasting decrease. Indomethacin and tetrodotoxin inhibited the ISC increase without affecting the secondary decrease. Addition of GRP to the mucosal solution produced a decrease in ISC similar to that observed with serosal treatment, but no transient increase in ISC was observed. GRP and bombesin (50% effective concentrations of 26 and 30 nM, respectively) were more effective than neuromedin B in decreasing the ISC, and the GRP receptor antagonist [D-Phe(6)]bombesin(6-13)-O-methyl produced a sixfold dextral shift in the GRP concentration-response relationship. The GRP-stimulated decrease was reduced in the absence of Cl and by serosal bumetanide. Flux measurements showed that GRP increased Rb and Na secretion while having no effect on transepithelial Cl transport. Phosphoinositide turnover was increased by GRP, suggesting that the ion transport changes may be mediated by intracellular Ca concentration. The results of this study demonstrate that GRP stimulates K and Na secretion across the porcine distal colon epithelium and that these processes are dependent, in part, on a bumetanide-sensitive transport pathway located in the basolateral membrane.

Protein kinase C potentiates cAMP-stimulated mouse duodenal mucosal bicarbonate secretion in vitro

2004 ◽  
Vol 286 (5) ◽  
pp. G814-G821 ◽  
Author(s):  
Bi-Guang Tuo ◽  
Jimmy Y. C. Chow ◽  
Kim E. Barrett ◽  
Jon I. Isenberg
Keyword(s):  
Short Circuit ◽  
Short Circuit Current ◽  
Duodenal Mucosa ◽  
Bicarbonate Secretion ◽  
Dependent Manner ◽  
Concentration Dependent Manner ◽  
Ussing Chambers ◽  
Duodenal Bicarbonate Secretion

PKC has been shown to regulate epithelial Cl- secretion in a variety of models. However, the role of PKC in duodenal mucosal bicarbonate secretion is less clear. We aimed to investigate the role of PKC in regulation of duodenal mucosal bicarbonate secretion. Bicarbonate secretion by murine duodenal mucosa was examined in vitro in Ussing chambers using a pH-stat technique. PKC isoform expression and activity were assessed by Western blotting and in vitro kinase assays, respectively. PMA (an activator of PKC) alone had no effect on duodenal bicarbonate secretion or short-circuit current ( Isc). When PMA and dibutyryl-cAMP (db-cAMP) were added simultaneously, PMA failed to alter db-cAMP-stimulated duodenal bicarbonate secretion or Isc ( P > 0.05). However, a 1-h preincubation with PMA potentiated db-cAMP-stimulated duodenal bicarbonate secretion and Isc in a concentration-dependent manner (from 10-8 to 10-5M) ( P < 0.05). PMA preincubation had no effects on carbachol- or heat-stable toxin-stimulated bicarbonate secretion. Western blot analysis revealed that PKCα, -γ, -ϵ, -θ, -μ, and -ι/λ were expressed in murine duodenal mucosa. Ro 31–8220 (an inhibitor active against PKCϵ, -α, -β, and -γ), but not Gö 6983 (an inhibitor active against PKCα, -γ, -β, and -δ), reversed the potentiating effect of PMA on db-cAMP-stimulated bicarbonate secretion. PMA also time- and concentration-dependently increased the activity of PKCϵ, an effect that was prevented by Ro 31–8220 but not Gö 6983. These results demonstrate that activation of PKC potentiates cAMP-stimulated duodenal bicarbonate secretion, whereas it does not modify basal secretion. The effect of PKC on cAMP-stimulated bicarbonate secretion is mediated by the PKCϵ isoform.

Ion transport across cat and ferret tracheal epithelia

1986 ◽  
Vol 61 (3) ◽  
pp. 1065-1070 ◽  
Author(s):  
R. J. Corrales ◽  
D. L. Coleman ◽  
D. B. Jacoby ◽  
G. D. Leikauf ◽  
H. L. Hahn ◽  
...  
Keyword(s):  
Ion Transport ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Free Medium ◽  
Mucin Secretion ◽  
Ussing Chambers ◽  
Cl Secretion ◽  
Mucosal Side

Sheets of trachea from ferret and cat were mounted in Ussing chambers and continuously short circuited. Under resting conditions, in both the cat and ferret there was little or no Cl secretion, and Na absorption accounted for most of the short-circuit current (Isc). Ouabain (10(-4) M, serosal bath) reduced Isc to zero in 30–60 min. This decline was matched by a decrease in net Na absorption. Amiloride (10(-4) M, luminal bath) caused a significant decrease in Isc and conductance (G) in both species. Bumetanide (10(-4) M, serosal bath) had negligible effects on Isc and G. In both species, isoproterenol increased Isc by stimulating Cl secretion. Methacholine induced equal amounts of Na and Cl secretion, with little change in Isc. In the cat, prostaglandins E2 and F2 alpha and bradykinin increased Isc, responses which were abolished in Cl-free medium. In open-circuited cat tissues, Na flux from the serosal to mucosal side was measured simultaneously with the secretion of nondialyzable 35S. Prostaglandins E1, E2, and F2 alpha, histamine, bradykinin, methacholine and isoproterenol all increased both Na and 35S-mucin secretion.

Neuromodulation of ion transport in porcine distal colon: NPY reduces secretory actions of leukotrienes

1995 ◽  
Vol 269 (2) ◽  
pp. R426-R431 ◽  
Author(s):  
T. R. Traynor ◽  
D. R. Brown ◽  
S. M. O'Grady
Keyword(s):  
Ion Transport ◽  
Colonic Mucosa ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Distal Colon ◽  
Effective Concentration ◽  
Leukotriene C4 ◽  
Ussing Chambers ◽  
Cl Secretion ◽  
Half Maximal Effective Concentration

Electrical transmural stimulation (ETS) was used to examine the neuroregulation of electrolyte transport in the porcine distal colon. ETS of the colonic mucosa-submucosa mounted in Ussing chambers produced rapid and transient increases in short-circuit current (Isc) that were inhibited 36% by serosal bumetanide, suggesting that a portion of the response may be attributed to Cl secretion. ETS actions were dependent upon stimulus intensity and frequency and were inhibited by tetrodotoxin and omega-conotoxin. Prazosin and pyrilamine had no effect on the mucosal responses to ETS, whereas atropine reduced the responses by 32%. Neuropeptide Y (NPY) also reduced the mucosal responses to ETS up to 60% (half-maximal effective concentration = 17 nM). In addition, the effects of leukotriene C4, previously shown to stimulate Cl secretion via a neuronal pathway, were also inhibited by NPY. These results indicate that cholinergic submucosal neurons play a role in the regulation of epithelial ion transport and that NPY acts as an inhibitory neuromodulator, particularly on leukotriene-sensitive neurons in the porcine distal colon.

Extracellular UTP stimulates electrogenic bicarbonate secretion across CFTR knockout gallbladder epithelium

2000 ◽  
Vol 279 (1) ◽  
pp. G132-G138 ◽  
Author(s):  
Lane L. Clarke ◽  
Matthew C. Harline ◽  
Lara R. Gawenis ◽  
Nancy M. Walker ◽  
John T. Turner ◽  
...  
Keyword(s):  
Cystic Fibrosis ◽  
Short Circuit ◽  
Ussing Chamber ◽  
Short Circuit Current ◽  
Receptor Activation ◽  
Biliary Disease ◽  
Anion Conductance ◽  
Intracellular Ca ◽  
Ph Stat ◽  
Chamber Studies

The loss of cystic fibrosis transmembrane conductance regulator (CFTR)-mediated transepithelial HCO3 − secretion contributes to the pathogenesis of pancreatic and biliary disease in cystic fibrosis (CF) patients. Recent studies have investigated P2Y2 nucleotide receptor agonists, e.g., UTP, as a means to bypass the CFTR defect by stimulating Ca2+-activated Cl− secretion. However, the value of this treatment in facilitating transepithelial HCO3 − secretion is unknown. Gallbladder mucosae from CFTR knockout mice were used to isolate the Ca2+-dependent anion conductance during activation of luminal P2Y2receptors. In Ussing chamber studies, UTP stimulated a transient peak in short-circuit current ( I sc) that declined to a stable plateau phase lasting 30–60 min. The plateau I sc after UTP was Cl− independent, HCO3 − dependent, insensitive to bumetanide, and blocked by luminal DIDS. In pH stat studies, luminal UTP increased both I sc and serosal-to-mucosal HCO3 − flux ( J s→m) during a 30-min period. Substitution of Cl− with gluconate in the luminal bath to inhibit Cl−/HCO3 −exchange did not prevent the increase in J s→mand I sc during UTP. In contrast, luminal DIDS completely inhibited UTP-stimulated increases in J s→m and I sc. We conclude that P2Y2 receptor activation results in a sustained (30–60 min) increase in electrogenic HCO3 − secretion that is mediated via an intracellular Ca2+-dependent anion conductance in CF gallbladder.

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.

NaCl transport across equine proximal colon and the effect of endogenous prostanoids

1990 ◽  
Vol 259 (1) ◽  
pp. G62-G69 ◽  
Author(s):  
L. L. Clarke ◽  
R. A. Argenzio
Keyword(s):  
Ion Transport ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Proximal Colon ◽  
Nacl Transport ◽  
Apparent Lack ◽  
Ussing Chambers ◽  
Cl Secretion

In contrast to in vivo findings, the equine proximal colon fails to demonstrate significant net absorption of Na+ and Cl- under in vitro conditions. The present study was undertaken to determine if endogenous prostanoids are responsible for this apparent lack of ion transport. Proximal colonic tissues from ponies were preincubated in either normal Ringer solution or in Ringer containing 1 microM indomethacin and studied in Ussing chambers containing these solutions. Untreated colonic mucosa demonstrated negligible Na(+)-Cl- absorption in the basal state. In contrast, indomethacin-treated colon significantly absorbed Na+ and Cl-, primarily as the result of an equivalent increase in the mucosal-to-serosal flux of these ions. Preincubation of proximal colon in 0.1 mM ibuprofen-treated Ringer yielded similar results. Treatment of indomethacin colon with 1 mM mucosal amiloride eliminated net Na(+)-Cl- absorption without affecting the short-circuit current (Isc). The Isc in control tissue was significantly greater than in indomethacin-treated tissue and was reduced by 0.1 mM serosal furosemide. Serosal addition of 0.1 microM prostaglandin E2 or 10 mM serosal plus mucosal theophylline to indomethacin-treated tissues abolished net Na(+)-Cl- absorption and increased the Isc to levels indistinguishable from control. In contrast, control tissues were essentially unaffected by these secretagogues. These findings indicated that Na(+)-Cl- absorption in equine proximal colon was electroneutral (possibly involving Na(+)-H+ exchange) and that the tissue was capable of electrogenic Cl- secretion. However, under the in vitro conditions, basal ion transport was dominated by endogenous prostanoids that abolished Na(+)-Cl- absorption and elicited near-maximal electrogenic Cl- secretion.

Effects of tumor promoters on sodium ion transport across frog skin

1985 ◽  
Vol 248 (5) ◽  
pp. C457-C465 ◽  
Author(s):  
M. M. Civan ◽  
D. Rubenstein ◽  
T. Mauro ◽  
T. G. O'Brien
Keyword(s):  
Ion Transport ◽  
Frog Skin ◽  
Phorbol Esters ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Tumor Promoter ◽  
Tumor Promoters ◽  
Transport Effects ◽  
Preliminary Study ◽  
Stimulation Of

Phorbol esters are tumor promoters and mitogens whose effects may be mediated by changes in ion transport across membranes. Clarification of the transport effects of these agents should be facilitated by using a well-characterized model epithelial system whose intracellular and transmural parameters are readily measurable. The current results constitute a preliminary study of the effects of 12-O-tetradecanoylphorbol-13-acetate (TPA), phorbol-12,13-dibutyrate (PDBU), and phorbol on the short-circuit current (Isc) across frog skin. TPA produced two effects: a stimulation of Isc of variable magnitude and a far more constant inhibition of the natriferic action of vasopressin. These effects appear related to the action of TPA as a tumor promoter insofar as PDBU (an active ester) also inhibited the natriferic response to vasopressin, whereas phorbol (inactive as a tumor promoter) had no significant effect. TPA is largely active from the mucosal medium, inhibits the natriferic response to adenosine 3',5'-cyclic monophosphate (cAMP) as well as that to vasopressin, and does not stimulate Isc in the presence of 10(-4) M mucosal amiloride. Inhibition of prostaglandin E1 production by indomethacin had no effect on the actions of TPA. The results indicate that frog skin is a promising model for studying the transport effects of the phorbol esters. The data further suggest that TPA acts on frog skin by activating the physiological amiloride- and cAMP-sensitive channels gating apical Na+ entry from the mucosal medium into the epithelial cells.

scholarly journals CD4+T cells mediate superantigen-induced abnormalities in murine jejunal ion transport

1998 ◽  
Vol 275 (1) ◽  
pp. G29-G38 ◽  
Author(s):  
Derek M. McKay ◽  
Michelle A. Benjamin ◽  
Jun Lu
Keyword(s):  
T Cells ◽  
Ion Transport ◽  
Short Circuit ◽  
Interleukin 2 ◽  
Elevated Serum ◽  
Short Circuit Current ◽  
Serum Levels ◽  
Scid Mice ◽  
Ussing Chambers ◽  
Ifn Γ

The immunomodulatory properties of bacterial superantigens (SAgs) have been defined, yet comparatively little is known of how SAgs may affect enteric physiology. Staphylococcus aureus enterotoxin B (SEB) was used to examine the ability of SAgs to alter epithelial ion transport. BALB/c mice, severe combined immunodeficient (SCID, lack T cells) mice, or SCID mice reconstituted with lymphocytes or CD4+T cells received SEB intraperitoneally, and jejunal segments were examined in Ussing chambers; controls received saline only. Baseline short-circuit current ( Isc, indicates net ion transport) and Iscresponses evoked by electrical nerve stimulation, histamine, carbachol, or forskolin were recorded. Serum levels of interleukin-2 (IL-2) and interferon-γ (IFN-γ) were measured. SEB-treated BALB/c mice showed elevated serum IL-2 and IFN-γ levels, and jejunal segments displayed a time- and dose-dependent increase in baseline Isccompared with controls. Conversely, evoked ion secretion was selectively reduced in jejunum from SEB-treated mice. Elevated cytokine levels and changes in jejunal Iscwere not observed in SEB-treated SCID mice. In contrast, SCID mice reconstituted with T cells were responsive to SEB challenge as shown by increased cytokine production and altered jejunal Iscresponses that were similar to those observed in jejunum from SEB-treated BALB/c mice. We conclude that exposure to a model bacterial SAg causes distinct changes in epithelial physiology and that these events can be mediated by CD4+T cells.

Electrolyte transport in rabbit cecum. I. Effect of RDEC-1 infection

1989 ◽  
Vol 256 (4) ◽  
pp. G721-G726 ◽  
Author(s):  
Y. H. Tai ◽  
T. P. Gage ◽  
C. McQueen ◽  
S. B. Formal ◽  
E. C. Boedeker
Keyword(s):  
Ion Transport ◽  
Short Circuit ◽  
Short Circuit Current ◽  
Fluid Secretion ◽  
Coli Strain ◽  
High Rate ◽  
Ussing Chambers ◽  
Induced Changes ◽  
Tight Junction Disruption ◽  
Net Fluxes

To investigate the characteristics of intestinal ion and fluid secretion induced by the adherent, effacing enteropathogenic Escherichia coli strain RDEC-1, we infected weanling rabbits with 10(7)-10(8) RDEC-1 organisms and then studied cecal ion transport under short-circuit conditions in Ussing chambers. Results in tissues with confluent adherent organisms were compared with those in uninfected ceca and in ceca stimulated with dibutyryl adenosine 3',5'-cyclic monophosphate (DBcAMP). The short-circuited cecum normally absorbed Na and Cl, secreted bicarbonate (as represented by the residual ion flux), and displayed a high rate of nondiffusional Na and Cl transport. RDEC-1 infection did not alter the short-circuit current (Isc), but it increased the conductance (Gt), decreased the potential difference (PD), abolished net Na absorption, and reversed Cl absorption to secretion. The changes in Na and Cl net fluxes may be explained by inhibition of a Na-Cl linked absorptive process. In contrast, DBcAMP significantly increased the Isc, PD, and Gt, decreased net Na flux, and abolished net Cl absorption by stimulating electrogenic Cl secretion. These results suggest that RDEC-1-induced changes in cecal ion transport are not mediated by cAMP. The reduction in Na-Cl linked absorption is consistent with anatomic changes in the apical surfaces of absorptive epithelial characteristic of effacing enteroadherence, whereas the increased conductance is consistent with tight junction disruption seen with RDEC-1 infection.

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