Chloride secretion stimulated by prostaglandin E1 and by forskolin in a canine renal epithelial cell line.

Simmons, Nicholas L.

doi:10.1113/jphysiol.1991.sp018394

Cited by 27 publications

(30 citation statements)

References 48 publications

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“…The biphasic time course of the I sc response to cAMP (Fig. 2) was consistent with previous studies in MDCK cells (43,54) and in A6 cells (4,8,29,44,58) that showed a rapid increase in anion secretion followed by a more gradual increase in Na ϩ absorption. The secretory anion current, represented by NS-I sc , was partially inhibited by 200 M glibenclamide (Figs.…”

Section: Discussionsupporting

confidence: 78%

Inhibition of ENaC by intracellular Cl⁻in an MDCK clone with high ENaC expression

Xie

Schafer

2004

American Journal of Physiology-Renal Physiology

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We examined the effects of intracellular Cl− concentration ([Cl−]i) on the epithelial Na channel (ENaC) in a line of Madin-Darby canine kidney (MDCK) cells (FL-MDCK) with a high rate of Na+ transport produced by stable retroviral transfection with rENaC subunits (Morris RG and Schafer JA. J Gen Physiol 120: 71–85, 2002). Treatment with cAMP (100 μM 8-cpt-cAMP plus 100 μM IBMX) stimulated ENaC-mediated Na+ absorption as well as Cl− secretion via cystic fibrosis transmembrane conductance regulator, which was characterized in α-toxin-permeabilized monolayers to have the anion selectivity sequence NO3− > Br− > Cl− > I−. With the use of FL-MDCK monolayers in which the basolateral membrane was permeabilized by nystatin, the ENaC conductance of the apical membrane [determined from the amiloride-sensitive short-circuit current (AS- Isc) driven by an apical-to-basolateral Na+ concentration gradient] was progressively inhibited by increasing the [Cl−] in the basolateral solution (and hence in the cytosol), but it was insensitive to the [Cl−] in the apical solution. This inhibitory effect of [Cl−]i occurred regardless of the presence or absence of net Cl− transport. However, from fluorometric measurements using the Cl−-sensitive dye 6-methoxy- N-(3-sulfopropyl)-quinolinium in intact FL-MDCK monolayers on permeable supports, cAMP, which activates both Na+ absorption and Cl− secretion, produced a decrease of [Cl−]i from 76 ± 14 to 36 ± 8 mM ( P = 0.03). Thus it might be expected that activation of Cl− secretion by cAMP would lead to stimulation rather than inhibition of ENaC. In the nystatin-treated monolayers, an increase in [Cl−]i from 15 to 145 mM decreased AS- Isc from 24.5 ± 1.0 to 10.2 ± 1.6 μA/cm2. This inhibition of ENaC could be attributed to nearly proportional decreases in the density of ENaC in the apical membrane from 1.91 ± 0.16 to 1.32 ± 0.17 fmol/cm2 and in the intrinsic channel activity (the average current per ENaC subunit) from 13.3 ± 1.2 to 8.2 ± 1.4 μA/fmol.

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Section: Discussionsupporting

confidence: 78%

Inhibition of ENaC by intracellular Cl⁻in an MDCK clone with high ENaC expression

Xie

Schafer

2004

American Journal of Physiology-Renal Physiology

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“…Actually, prostaglandins are able per se to stimulate transepithelial ion transport in MDCK cells (Simmons, 1991). A mechanism through which prostaglandins can be generated is the Ca2+-dependent activation of phospholipase A2.…”

Section: Discussionmentioning

confidence: 99%

Regulation of transepithelial ion transport by two different purinoceptors in the apical membrane of canine kidney (MDCK) cells

Zegarra‐Moran

Romeo

Galietta

1995

British J Pharmacology

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1 The effect of extracellular nucleotides on the transepithelial ion transport of Madin Darby canine kidney cells (MDCK) was investigated. Cells were grown up to confluency on permeable supports and the short circuit current (ISC) was measured with an Ussing chamber-like mini-perfusion system.2 Apical ATP stimulated a biphasic I,C increase consisting of a first rapid and transient peak followed by a broader one. 3 The first peak evoked by ATP was reversibly blocked by basilen blue (BB) in a concentrationdependent fashion, with an EC50 of 7.5 t4M. 4 The P2y receptor agonist, 2-methylthioATP (2-MeSATP) caused a single transient I,, increase that was completely blocked by pretreatment with BB. On the contrary, the P2, agonist, a,p-methylene ATP (a,p-meATP) was almost completely ineffective on UTP essentially induced a monophasic response the time-course of which resembled that of the second peak stimulated by ATP. The agonist potency order was 2-MeSATP>ATP>>UTP, x,P-meATP for the first peak and UTP> ATP>2-MeSATP> oc,-meATP for the second peak.5 Monolayer incubation with the membrane permeable calcium chelator [bis-o-aminophenoxy)-ethane-N,N,NM,N',-tetraacetic acid, tetra(acetoximethyl)-ester] (BAPTA/AM) inhibited the ATP-evoked first peak. 6 The non-hydrolyzable ATP analogue, adenosine-5'-O-(3-thio)-trisphosphate (ATP-'y-S) elicited a biphasic response similar to that of ATP. The P, receptor agonist, 2-chloroadenosine and CGS-21680, were almost unable to induce an I4, increase. These results rule out the involvement of ATP hydrolysis and P, receptor activation as responsible for I4, increase.7 Inhibition of prostaglandins synthesis by indomethacin abolished the second ATP-evoked peak. 8 Chloride replacement with gluconate on both sides of the epithelium completely inhibited the second peak induced by ATP but only reduced the amplitude of the first spike. 9 The results suggest that ATP stimulates I4 increase by two mechanisms. The first one is mediated by a P2y receptor and by intracellular calcium increase. The second induces prostaglandin synthesis probably through a P2U receptor activation.

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“…C Mean±SEM (n=5 pairs) for the effect of 0.1 mM furosemide on the initial rate of acidification of the cell cytosol following basal exposure to 5 mM NH 4 Cl tion of 10 -5 M forskolin stimulated inward I sc by 5.2±0.8 µA cm -2 (n=12) which is due to transepithelial anion secretion [3] Application of 0.2 mM furosemide during the maintained phase to the basal solutions resulted in only small reductions of the secretory current to 4.6±0.8 µA cm -2 (n=12) after 5 min incubation (83.5±5.3%, n=12, of the maintained response is insensitive to furosemide). Thus, in contrast to other secretory epithelia such as intestinal or MDCK epithelia [8,26,27,28,31], NKCC1 is not the major determinant of cAMP-stimulated anion secretion at the basolateral membrane of mIMCD-K2 cells.…”

Section: Transepithelial Anion Secretionmentioning

confidence: 98%

Expression and role of sodium, potassium, chloride cotransport (NKCC1) in mouse inner medullary collecting duct (mIMCD-K2) epithelial cells

Glanville

Kingscote

Thwaites

et al. 2001

Pflügers Arch - Eur J Physiol

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Loop-diuretic-sensitive 86Rb+(K+) transmembrane fluxes were determined in cells of a mouse inner medullary collecting duct cell line (mIMCD-K2). The furosemide-sensitive (0.1 mM) influx was a substantial fraction of the total influx (0.39+/-0.04 or 0.42+/-0.03, n=5 in the presence or absence of ouabain, respectively). Furosemide also reduced 86Rb+(K+) efflux by a similar fraction (0.46). RT-PCR analysis revealed expression of mRNA for the Na+-K+-2Cl- cortransporter-1 (NKCC1), but not NKCC2. Loop-diuretic-sensitive 86Rb+(K+) influx was confined to the basolateral membrane, confirming its localisation there. The physiological properties of NKCC1 expressed in mIMCD-K2 cells, including the dependence upon medium Na+, K+ and Cl- and the relative sensitivity to loop diuretics as assessed by the concentration required for half-maximal inhibition (IC50) (bumetanide 3.3+/-1.4x10-7 M>piretanide 2.5+/-0.15x10-6 M>furosemide 2.3+/-1.2x10-5 M) were typical for NKCC1. Possible functions of NKCC1 were tested; furosemide did not inhibit the majority of forskolin-stimulated secretory short-circuit current (Isc) (83.5+/-5.3% of the maintained response at 5 min). Secondly, total 86Rb+(K+) influx was stimulated markedly when external osmolarity was increased to 600 mosmol/l by mannitol due to an increase via NKCC1 from 55+/-11 to 191+/-2 nmol/106 cells per 15 min, (both n=4, P<0.01). In contrast, 10-5 M forskolin did not stimulate total 86Rb+(K+) influx. Finally, the ability of both K+ and NH4+ to compete for ouabain-insensitive 86Rb+(K+) influx via NKCC1 was confirmed with similar concentrations for half-maximal influx reduction (K0.5). Apical exposure to NH4+ elicited rapid cytosolic alkalinisation in 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein (BCECF)-loaded epithelial layers, consistent with selective permeability of the apical membrane to NH3. Conversely, NH4+ (5 mM) at the basal cell surface resulted in progressive acidification, the initial rate being reduced by 43% by furosemide. We conclude that NKCC1 participates in selective uptake of NH4+ at the basal surface, and that IMCD may function in direct NH4+ deposition to urine.

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Chloride secretion stimulated by prostaglandin E1 and by forskolin in a canine renal epithelial cell line.

Cited by 27 publications

References 48 publications

Inhibition of ENaC by intracellular Cl⁻in an MDCK clone with high ENaC expression

Inhibition of ENaC by intracellular Cl⁻in an MDCK clone with high ENaC expression

Regulation of transepithelial ion transport by two different purinoceptors in the apical membrane of canine kidney (MDCK) cells

Expression and role of sodium, potassium, chloride cotransport (NKCC1) in mouse inner medullary collecting duct (mIMCD-K2) epithelial cells

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Chloride secretion stimulated by prostaglandin E1 and by forskolin in a canine renal epithelial cell line.

Cited by 27 publications

References 48 publications

Inhibition of ENaC by intracellular Cl−in an MDCK clone with high ENaC expression

Inhibition of ENaC by intracellular Cl−in an MDCK clone with high ENaC expression

Regulation of transepithelial ion transport by two different purinoceptors in the apical membrane of canine kidney (MDCK) cells

Expression and role of sodium, potassium, chloride cotransport (NKCC1) in mouse inner medullary collecting duct (mIMCD-K2) epithelial cells

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Inhibition of ENaC by intracellular Cl⁻in an MDCK clone with high ENaC expression

Inhibition of ENaC by intracellular Cl⁻in an MDCK clone with high ENaC expression