共查询到20条相似文献,搜索用时 11 毫秒
1.
2.
The presence of basolateral Cl− channels in airway epithelium has been reported in several studies, but little is known about their role in the regulation
of anion secretion. The purpose of this study was to characterize regulation of these channels by nitric oxide (NO) in Calu-3
cells. Transepithelial measurements revealed that NO donors activated a basolateral Cl− conductance sensitive to 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid (DIDS) and anthracene-9-carboxylic acid. Apical
membrane permeabilization studies confirmed the basolateral localization of NO-activated Cl− channels. Experiments using 8-bromo cyclic guanosine monophosphate (8Br-cGMP) and selective inhibitors of soluble guanylyl
cyclase and inducible NO synthase (1H-[1, 2, 4] oxadiazolol-[4, 3-a] quinoxalin-1-one [ODQ] and 1400W [N-(3-Aminomethyl)benzyl)acetamidine],
respectively) demonstrated that NO activated Cl− channels via a cGMP-dependent pathway. Anion replacement and 36Cl− flux studies showed that NO affected both Cl− and HCO
3
−
secretion. Two different types of Cl− channels are known to be present in the basolateral membrane of epithelial cells: Zn2+-sensitive ClC-2 and DIDS-sensitive bestrophin channels. S-Nitrosoglutathione (GSNO) activated Cl− conductance in the presence of Zn2+ ions, indicating that ClC-2 channel function was not affected by GSNO. In contrast, DIDS completely inhibited GSNO-activated
Cl− conductance. Bestrophin immunoprecipitation studies showed that under control conditions bestrophin channels were not phosphorylated
but became phosphorylated after GSNO treatment. The presence of bestrophin in airway epithelia was confirmed using immunohistochemistry.
We conclude that basolateral Cl− channels play a major role in the NO-dependent regulation of anion secretion in Calu-3 cells. 相似文献
3.
4.
Winters CJ Zimniak L Mikhailova MV Reeves WB Andreoli TE 《The Journal of membrane biology》2000,177(3):221-230
We have isolated two new and highly homologous cDNAs, mmClC-Ka from mouse outer medulla and mcClC-Ka from mouse cortex. In both cases, mRNA was obtained from the indicated region and subjected to RT-PCR using primers from
the nucleotide sequence of rbClC-Ka, which encodes basolateral Cl− channels (termed rbClC-Ka) in rabbit MTAL. The predicted protein products of mmClC-Ka and mcClC-Ka, mmClC-Ka and mcClC-Ka, respectively, were 85% homologous and had predicted molecular weights of 75 kDa. The predicted protein
sequences for mmClC-Ka and rbClC-Ka had three cytosolic sites—threonine 185, threonine 187 and serine 270—which were absent
in mcClC-Ka. These three moieties represent potential sites for phosphorylation of mmClC-Ka and rbClC-Ka, but not of mcClC-Ka,
and may account for the failure of (ATP + PKA) to increase the open time probability P
o
in basolateral CTAL Cl− channels.
We prepared antisense oligonucleotides specific for nonhomologous regions of these two cDNAs, mmAntisense for mmClC-Ka and mcAntisense for mcClC-Ka. Using anti-rbClC-Ka, a polyclonal antibody to rbClC-Ka, we found that, when transfected into cultured mouse MTAL and CTAL
cells, mmAntisense suppressed the appearance of the 75 kDa band by 50% in vesicles from MTAL but not CTAL cells, while transfection
of MTAL and CTAL cells with mcAntisense suppressed appearance of the 75 kDa band in vesicles from CTAL but not MTAL cells. mmAntisense transfection also prolonged the half-time (T1/2, sec) for 36Cl− efflux in cultured MTAL cells from 82.4 ± 6.8 sec (sem) to 187.8 ± 9.5 sec (n= 5; P= 0.0001) while mcAntisense transfection had no such effect. Conversely, in cultured CTAL cells, mcAntisense transfection prolonged the T1/2 for 36Cl− efflux from 80.9 ± 6.3 sec to 191.8 ± 6.5 sec (n= 5; P= 0.00005), while mmAntisense had no such effect. We conclude that mmClC-Ka and mcClC-Ka may encode the basolateral Cl− channels mediating net Cl− absorption in mouse MTAL and CTAL, respectively.
Received: 16 May 2000/Revised: 30 June 2000 相似文献
5.
6.
O. Zegarra-Moran O. Sacco L. Romano G.A. Rossi L.J.V. Galietta 《The Journal of membrane biology》1997,156(3):297-305
The perforated-patch technique was used to study the response of human bronchial cells to extracellular nucleotides. ATP
or UTP (100 μm) elicited a complex response consisting of a large transient membrane current increase followed by a relatively small sustained
level. These two phases were characterized by different current kinetics. Throughout the transient phase (2–3 min) the membrane
current (I
p
) displayed slow activation and deactivation kinetics at depolarizing and hyperpolarizing potentials respectively. At steady-state
(I
s
) the relaxation at hyperpolarizing potential disappeared whereas at positive membrane potentials the current became slightly
deactivating. The I
s
amplitude was dependent on the extracellular Ca2+ concentration, being completely inhibited in Ca2+-free medium. Cell pre-incubation with the membrane-permeable chelating agent BAPTA/AM prevented completely the response to
nucleotides, thus suggesting that both I
p
and I
s
were dependent on intracellular Ca2+. The presence of a hypertonic medium during nucleotide stimulation abolished I
s
leaving I
p
unchanged. On the contrary, niflumic acid, a blocker of Ca2+-activated Cl− channels, prevented completely I
p
without reducing significantly I
s
. 1,9-dideoxyforskolin fully inhibited I
s
but also reduced I
p
. Replacement of extracellular Cl− with aspartate demonstrated that the currents activated by nucleotides were Cl− selective. I
p
resulted five times more Cl− selective than I
s
with respect to aspartate. Taken together, our results indicate that ATP and UTP activate two types of Cl− currents through a Ca2+-dependent mechanism.
Received: 15 August 1996/Revised: 6 December 1996 相似文献
7.
Primary cultures of sea bass gill cells grown on permeable membranes form a confluent, polarized, functional tight epithelium
as characterized by electron microscopy and electrophysiological and ion transport studies. Cultured with normal fetal bovine
serum (FBS) and mounted in an Ussing chamber, the epithelium presents a small short-circuit current (I
sc
: 1.4 ± 0.3 μA/cm2), a transepithelial voltage (V
t
) of 12.7 ± 2.7 mV (serosal positive) and a high transepithelial resistance (R
t
: 12302 ± 2477 Ω× cm2). A higher degree of differentiation and increased ion transport capacities are observed with cells cultured with sea bass
serum: numerous, organized microridges characteristic of respiratory cells are present on the apical cell surface and there
are increased I
sc
(11.9 ± 2.5 μA/cm2) and V
t
(25.9 ± 1.7 mV) and reduced R
t
(4271 ± 568 Ω× cm2) as compared with FBS-treated cells. Apical amiloride addition (up to 100 μm) had no effect on I
sc
. The I
sc
, correlated with an active Cl− secretion measured as the difference between 36Cl− unidirectional fluxes, was partly blocked by serosal ouabain, bumetanide, DIDS or apical DPC or NPPB and stimulated by serosal
dB-cAMP. It is concluded that the chloride secretion is mediated by a Na+/K+/2Cl− cotransport and a Cl−/HCO3
− exchanger both responsible for Cl− entry through the basolateral membrane and by apical cAMP-sensitive Cl− channels. This study gives evidence of a functional, highly differentiated epithelium in cultures composed of fish gill respiratorylike
cells, which could provide a useful preparation for studies on ion transport and their regulation. Furthermore, the chloride
secretion through these cultures of respiratorylike cells makes it necessary to reconsider the previously accepted sea water
model in which the chloride cells are given the unique role of ion transport through fish gills.
Received: 12 July 1996/Revised: 5 November 1996 相似文献
8.
9.
Using the whole-cell patch-clamp technique, we examined Cl−-selective currents manifested by strial marginal cells isolated from the inner ear of gerbils. A large Cl−-selective conductance of ∼18 nS/pF was found from nonswollen cells in isotonic buffer containing 150 mm Cl−. Under a quasi-symmetrical Cl− condition, the `instantaneous' current-voltage relation was close to linear, while the current-voltage relation obtained
at the end of command pulses of duration 400 msec showed weak outward rectification. The permeability sequence for anionic
currents was as SCN− > Br−≅ Cl− > F− > NO−
3≅ I− > gluconate−, corresponding to Eisenmann's sequence V. When whole-cell voltage clamped in isotonic bathing solutions, the cells exhibited
volume changes that were accounted for by the Cl− currents driven by the imposed electrochemical potential gradients. The volume change was elicited by lowered extracellular
Cl− concentration, anion substitution and altered holding potentials. The Cl− conductance varied in parallel with cell volume when challenged by bath anisotonicity. The whole-cell Cl− current was only partially blocked by both 5-nitro-2-(3-phenylpropylamino) benzoic acid (NPPB, 0.5 mm) and diphenylamine-2-carboxylic acid (DPC, 1.0 mm), but 4-acetamido-4′-isothiocyanato-stilbene-2,2′-disulfonic acid (SITS, 0.5 mm) was without effect. The properties of the present whole-cell Cl− current resembled those of the single Cl− channel previously found in the basolateral membrane of the marginal cell (Takeuchi et al., Hearing Res.
83:89–100, 1995), suggesting that the volume-correlated Cl− conductance could be ascribed predominantly to the basolateral membrane. This Cl− conductance may function not only in cell volume regulation but also for the transport of Cl− and the setting of membrane potential in marginal cells under physiological conditions.
Received: 15 August 1995/Revised: 3 November 1995 相似文献
10.
Irshad Ali Sheikh Hemanta Koley Manoj K. Chakrabarti Kazi Mirajul Hoque 《The Journal of biological chemistry》2013,288(28):20404-20415
The apical membrane of intestinal epithelia expresses intermediate conductance K+ channel (KCNN4), which provides the driving force for Cl− secretion. However, its role in diarrhea and regulation by Epac1 is unknown. Previously we have established that Epac1 upon binding of cAMP activates a PKA-independent mechanism of Cl− secretion via stimulation of Rap2-phospholipase Cϵ-[Ca2+]i signaling. Here we report that Epac1 regulates surface expression of KCNN4c channel through its downstream Rap1A-RhoA-Rho-associated kinase (ROCK) signaling pathway for sustained Cl− secretion. Depletion of Epac1 protein and apical addition of TRAM-34, a specific KCNN4 inhibitor, significantly abolished cAMP-stimulated Cl− secretion and apical K+ conductance (IK(ap)) in T84WT cells. The current-voltage relationship of basolaterally permeabilized monolayers treated with Epac1 agonist 8-(4-chlorophenylthio)-2′-O- methyladenosine 3′,5′-cyclic monophosphate showed the presence of an inwardly rectifying and TRAM-34-sensitive K+ channel in T84WT cells that was absent in Epac1KDT84 cells. Reconstructed confocal images in Epac1KDT84 cells revealed redistribution of KCNN4c proteins into subapical intracellular compartment, and a biotinylation assay showed ∼83% lower surface expression of KCNN4c proteins compared with T84WT cells. Further investigation revealed that an Epac1 agonist activates Rap1 to facilitate IK(ap). Both RhoA inhibitor (GGTI298) and ROCK inhibitor (H1152) significantly reduced cAMP agonist-stimulated IK(ap), whereas the latter additionally reduced colocalization of KCNN4c with the apical membrane marker wheat germ agglutinin in T84WT cells. In vivo mouse ileal loop experiments showed reduced fluid accumulation by TRAM-34, GGTI298, or H1152 when injected together with cholera toxin into the loop. We conclude that Rap1A-dependent signaling of Epac1 involving RhoA-ROCK is an important regulator of intestinal fluid transport via modulation of apical KCNN4c channels, a finding with potential therapeutic value in diarrheal diseases. 相似文献
11.
Summary Gluconate substitution for serosal Cl– reduces the transepithelial short-circuit current (I
sc) and depolarizes shortcircuited frog skins. These effects could result either from inhibition of basolateral K+ conductance, or from two actions to inhibit both apical Na+ permeability (P
Na
ap
) and basolateral pump activity. We have addressed this question by studying whole-and split-thickness frog skins. Intracellular Na+ concentration (C
Na
c
) andP
Na
ap
have been monitored by measuring the currentvoltage relationship for apical Na+ entry. This analysis was conducted by applying trains of voltage pulses, with pulse durations of 16 to 32 msec. Estimates ofP
Na
ap
) and CNa/c were not detectably dependent on pulse duration over the range 16 to 80 msec. Serosal Cl– replacement uniformly depolarized short-circuited tissues. The depolarization was associated with inhibition ofI
sc across each split skin, but only occasionally across the whole-thickness preparations. This difference may reflect the better ionic exchange between the bulk medium and the extracellular fluid in contact with the basolateral membranes, following removal of the underlying dermis in the split-skin preparations.P
Na
ap
was either unchanged or increased, and CNa/c either unchanged or reduced after the anionic replacement. These data are incompatible with the concept that serosal Cl– replacement inhibitsP
Na
ap
and Na, K-pump activity. Gluconate substutition likely reduces cell volume, triggering inhibition of the basolateral K+ channels, consistent with the data and conclusions of S.A. Lewis, A.G. Butt, M.J. Bowler, J.P. Leader and A.D.C Macknight (J. Membrane Biol.
83:119–137, 1985) for toad bladder. The resulting depolarization reduces the electrical force favoring apical Na+ entry. The volume-conductance coupling serves to conserve volume by reducing K+ solute loss. Its molecular basis remains to be identified. 相似文献
12.
M. J. Beilby 《The Journal of membrane biology》1981,62(3):207-218
Summary The changes in the cytoplasmic Cl– concentration, [Cl–]
c
, are monitored at the time of withdrawal (starvation) and subsequent replacement of Cl– in the outside medium. The measurement technique exploits the involvement of Cl– inChara excitation. The transient clamp current due to Cl–,I
Cl, is separated from other excitation transients through Hodgkin-Huxley (HH) equations, which have been adjusted toChara. TheI
Cl amplitude depends on HH parameters, [Cl–]
c
and the maximum membrane conductance to Cl–,
. The results are discussed in terms of these quantities.I
Cl and
were found to fall after 6–10 hr of Cl– starvation, thus supporting the hypothesis that [Cl–
c
decreases in Cl–-free medium. The best HH fit to starved data was obtained with [Cl–
c
=3.5mm. The time-course forI
Cl decline is considerably slower than the time-course of the rise of the starvation-stimulated influx. As cells starved for periods longer than 24 hr are re-exposed to Cl–, it is revealed that while [Cl–]
c
remains low during long starvation,
increases to values greater than those of the normal cells. Such differences among cells starved for various lengths of time have not been detected previously. 相似文献
13.
The rat primary cultured-airway monolayer had been an excellent model for deciphering the ion channel after nystatin permeabilization
of its basolateral or apical membrane (Hwang et al., 1996). After apical membrane permeabilization of rat primary cultured-airway
monolayer, 4,4′-diisothiocyanatostilbene-2,2′-disulfonic acid (DIDS)-sensitive outwardly rectifying depolarization-induced
Cl− (BORDIC) currents were observed across the basolateral membrane in symmetrical NMG-Cl solution in this study. No significant
Cl− current induced by the application of voltage clamping was observed across the apical membrane in symmetrical NMG-Cl solution
after basolateral membrane permeabilization. The halide permeability sequence for BORDIC current was Br−≒ I− > Cl−. BORDIC current was not affected by basolaterally applied bumetanide (0.5 mm). Basolateral DIDS (0.2 mm) but not apical DIDS inhibited CFTR mediated short-circuit current (I
sc
) in an intact monolayer of rat airway epithelia, a T84 human colonal epithelial cell line, and a Calu-3 human airway epithelial
cell line. This is the first report showing that depolarization induced Cl− current is present on the basolateral membrane of airway epithelia.
Received: 7 October 1999/Revised: 24 April 2000 相似文献
14.
John M. Bayliss W. Brian Reeves Thomas E. Andreoli 《The Journal of membrane biology》1990,113(1):49-56
Summary This paper provides the results of studies which characterized conductive36Cl– flux in basolaterally enriched membrane vesicles prepared from rabbit renal outer medulla. Conductive36Cl– uptake was studied under two different experimental conditions. In the first,36Cl– flux was driven by an inside positive voltage created with oppositely directed Cl– and gluconate gradients. In the second, an inwardly direct K+ gradient was used to drive36Cl– uptake. By these two methods, voltage-sensitive36Cl– uptake was shown to comprise about 45 and 65%, respectively, of the initial rates of total36Cl– flux. Separate paired studies demonstrated that the conductive36Cl– uptake was inhibited by the Cl– channel blocker diphenylamine-2-carboxylate (DPC) with an IC50 for DPC of 154 m. The voltagedependent36Cl– uptake had an activation energy of 6.4 kcal/mole. This36Cl– conductance had an anion selectivity sequence of I–>Cl–NO
3
–
gluconate. 相似文献
15.
Summary The present studies examined some of the properties of Cl– channels in renal outer medullary membrane vesicles incorporated into planar lipid bilayers. The predominant channel was anion selective having aP
Cl/P
K ratio of 10 and a unit conductance of 93 pS in symmetric 320mm KCl. In asymmetric KCl solutions, theI-V relations conformed to the Goldman-Hodgkin-Katz equation. Channel activity was voltage-dependent with a gating charge of unity. This voltage dependence of channel activity may account, at least in part, for the striking voltage dependence of the basolateral membrane Cl– conductance of isolated medullary thick ascending limb segments. The Cl– channels incorporated into the planar bilayers were asymmetrical: thetrans surface was sensitive to changes in ionized Ca2+ concentrations and insensitive to reducing KCl concentrations to 10mm, while thecis side was insensitive to changes in ionized Ca2+ concentrations, but was inactivated by reducing KCl concentrations to 50mm. 相似文献
16.
John Cuppoletti Jayati Chakrabarti Kirti Tewari Danuta H. Malinowska 《Cell biochemistry and biophysics》2013,66(1):53-63
In clinical trials, methadone, but not morphine, appeared to prevent beneficial effects of lubiprostone, a ClC-2 Cl? channel activator, on opioid-induced constipation. Effects of methadone and morphine on lubiprostone-stimulated Cl? currents were measured by short circuit current (Isc) across T84 cells. Whole cell patch clamp of human ClC-2 (hClC-2) stably expressed in HEK293 cells and in a high expression cell line (HEK293EBNA) as well as human CFTR (hCFTR) stably expressed in HEK293 cells was used to study methadone and morphine effects on recombinant hClC-2 and hCFTR Cl? currents. Methadone but not morphine inhibited lubiprostone-stimulated Isc in T84 cells with half-maximal inhibition at 100 nM. Naloxone did not affect lubiprostone stimulation or methadone inhibition of Isc. Lubiprostone-stimulated Cl? currents in hClC-2/HEK293 cells, but not forskolin/IBMX-stimulated Cl? currents in hCFTR/HEK293 cells, were inhibited by methadone, but not morphine. HEK293EBNA cells expressing hClC-2 showed time-dependent, voltage-activated, CdCl2-inhibited Cl? currents in the absence (control) and the presence of lubiprostone. Methadone, but not morphine, inhibited control and lubiprostone-stimulated hClC-2 Cl? currents with half-maximal inhibition at 100 and 200–230 nM, respectively. Forskolin/IBMX-stimulated hClC-2 Cl? currents were also inhibited by methadone. Myristoylated protein kinase inhibitor (a specific PKA inhibitor) inhibited forskolin/IBMX- but not lubiprostone-stimulated hClC-2 Cl? currents. Methadone caused greater inhibition of lubiprostone-stimulated currents added before patching (66.1 %) compared with after patching (28.7 %). Methadone caused inhibition of lubiprostone-stimulated Cl? currents in T84 cells and control; lubiprostone- and forskolin/IBMX-stimulated recombinant hClC-2 Cl? currents may be the basis for reduced efficacy of lubiprostone in methadone-treated patients. 相似文献
17.
Previous studies have shown that the whole-cell current-voltage (I-V) relation of unstimulated sheep parotid cells is dominated by two K+ conductances, one outwardly and the other inwardly rectifying. We now show that once these K+ conductances are blocked by replacement of pipette K+ with Na+ and by the addition of 5 mmol/liter CsCl to the bath, there remains an outwardly rectifying conductance with a reversal potential of 0 mV. Replacement of 120 mmol/liter NaCl in the pipette solution with an equimolar amount of Na-glutamate shifted the reversal potential of this residual current to -55 mV, indicating that the conductance was Cl? selective. The Cl? current was activated by increasing the free Ca2+ in the pipette solution from 10 to 100 nmol/liter. When the Ca2+ concentration in the pipette solution was 10 nmol/liter, the relaxations observed in response to membrane depolarization could be fitted with a single exponential, whose time constant increased from 81 to 183 ms as the pipette potential was increased from -30 to +60 mV. Relaxation analysis showed that the current was activated by membrane depolarization. Reversal potential measurements in experiments in which external Cl? was replaced with various anions, gave the following relative permeabilities: SCN- (1.80) > I- (1.09) > CI- (1) > NO 3 - (0.92) > Br- (0.75). The relative conductances were: SCN- (2.18) > I- (1.07) > Cl? (1.00) > Br- (0.91) > NO 3 - (0.50). The Cl? current was blocked by NPPB (ID50 ≈ 10 μm), DIDS (10 or 30 μmol/liter) and furosemide (100 μmol/liter). 相似文献
18.
Christina M. Szalinski Christopher J. Guerriero Wily G. Ruiz Brianne E. Docter Youssef Rbaibi Núria M. Pastor-Soler Gerard Apodaca Manojkumar A. Puthenveedu Ora A. Weisz 《PloS one》2013,8(1)
Localized synthesis of phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P2] at clathrin coated pits (CCPs) is crucial for the recruitment of adaptors and other components of the internalization machinery, as well as for regulating actin dynamics during endocytosis. PtdIns(4,5)P2 is synthesized from phosphatidylinositol 4-phosphate by any of three phosphatidylinositol 5-kinase type I (PIP5KI) isoforms (α, β or γ). PIP5KIβ localizes almost exclusively to the apical surface in polarized mouse cortical collecting duct cells, whereas the other isoforms have a less polarized membrane distribution. We therefore investigated the role of PIP5KI isoforms in endocytosis at the apical and basolateral domains. Endocytosis at the apical surface is known to occur more slowly than at the basolateral surface. Apical endocytosis was selectively stimulated by overexpression of PIP5KIβ whereas the other isoforms had no effect on either apical or basolateral internalization. We found no difference in the affinity for PtdIns(4,5)P2-containing liposomes of the PtdIns(4,5)P2 binding domains of epsin and Dab2, consistent with a generic effect of elevated PtdIns(4,5)P2 on apical endocytosis. Additionally, using apical total internal reflection fluorescence imaging and electron microscopy we found that cells overexpressing PIP5KIβ have fewer apical CCPs but more internalized coated structures than control cells, consistent with enhanced maturation of apical CCPs. Together, our results suggest that synthesis of PtdIns(4,5)P2 mediated by PIP5KIβ is rate limiting for apical but not basolateral endocytosis in polarized kidney cells. PtdIns(4,5)P2 may be required to overcome specific structural constraints that limit the efficiency of apical endocytosis. 相似文献
19.
Cremaschi D Porta C Bottà G Bazzini C Baroni MD Garavaglia M 《The Journal of membrane biology》2000,176(1):53-65
Cl− apically enters the epithelium of rabbit gallbladder by a Na+-Cl− symport, sensitive to hydrochlorothiazide (HCTZ). Since HCTZ also activates an apical SITS-sensitive Cl− conductance (G
Cl
), the symport inhibition might be merely due to a short circuit of the symport by G
Cl
rather than to a direct action of HCTZ on the symporter. To examine whether the symport is directly inhibited by HCTZ and
whether the symporter belongs to the family of thiazide-sensitive cotransporters (TSC), radiochemical measurements of the
apical Cl− uptake, electrophysiological determinations of intracellular Cl− and Na+ activities (a
i,Cl
and a
i,Na
) with selective theta microelectrodes and molecular biology methods were used. The 36Cl− uptake proved to be a measurement of the apical unidirectional Cl− influx (J
mc
) and of the symport only (without backflux components), with measuring times of 45 sec under all experiment conditions; its
inhibition by HCTZ was unaffected by G
Cl
activation or abolition. After HCTZ treatment the decrease in a
i,Cl
(measured as the initial rate or in 3 min) was larger than the decrease in a
i,Na
. The difference was reduced to one third in a group of epithelia in which the elicited G
Cl
was reduced to one third; moreover it was abolished in any case when G
Cl
was abolished with 10−4
m SITS. The SITS-insensitive rate of a
i,Cl
decrease was equal to that of the a
i,Na
decrease in any case. Thus the a
i,Cl
decrease displays a component dependent on G
Cl
activation and a second component dependent on symport inhibition. Using the RT-PCR technique a cDNA fragment was obtained
that was 99% identical to the corresponding region of the rabbit renal TSC isoform. The results indicate that in rabbit gallbladder
epithelium HCTZ displays a dual action, namely G
Cl
activation and Na+-Cl− symport inhibition. This Na+-Cl− symporter is the first TSC found to be functionally expressed in a nonrenal or nonrenal-like epithelium.
Received: 29 July 1999/Revised: 23 March 2000 相似文献
20.
Ning Yang Zhen Lei Xiaoyu Li Junhan Zhao Tianjian Liu Nannan Ning Ailin Xiao Linlin Xu Jingxin Li 《PloS one》2014,9(1)
Chloroquine (CQ), a bitter tasting drug widely used in treatment of malaria, is associated gastrointestinal side effects including nausea or diarrhea. In the present study, we investigated the effect of CQ on electrolyte transport in rat ileum using the Ussing chamber technique. The results showed that CQ evoked an increase in short circuit current (ISC) in rat ileum at lower concentration (≤5×10−4 M ) but induced a decrease at higher concentrations (≥10−3 M). These responses were not affected by tetrodotoxin (TTX). Other bitter compounds, such as denatoniumbenzoate and quinine, exhibited similar effects. CQ-evoked increase in ISC was partly reduced by amiloride(10−4 M), a blocker of epithelial Na+ channels. Furosemide (10−4 M), an inhibitor of Na+-K+ -2Cl− co-transporter, also inhibited the increased ISC response to CQ, whereas another Cl− channel inhibitor, CFTR(inh)-172(10−5M), had no effect. Intriguingly, CQ-evoked increases were almost completely abolished by niflumic acid (10−4M), a relatively specific Ca2+-activated Cl− channel (CaCC) inhibitor. Furthermore, other CaCC inhibitors, such as DIDS and NPPB, also exhibited similar effects. CQ-induced increases in ISC were also abolished by thapsigargin(10−6M), a Ca2+ pump inhibitor and in the absence of either Cl− or Ca2+ from bathing solutions. Further studies demonstrated that T2R and CaCC-TMEM16A were colocalized in small intestinal epithelial cells and the T2R agonist CQ evoked an increase of intracelluar Ca2+ in small intestinal epithelial cells. Taken together, these results demonstrate that CQ induces Cl− secretion in rat ileum through CaCC at low concentrations, suggesting a novel explanation for CQ-associated gastrointestinal side-effects during the treatment of malaria. 相似文献