Electrical properties of cultured renal tubular cells (OK) grown in confluent monolayers

Schwegler, Johann S.; Heuner, Almut; Silbernagl, Stefan

doi:10.1007/bf00370590

Cited by 20 publications

(7 citation statements)

References 23 publications

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“…Assuming a linear relationship between PD~ and tk+, 100% tk+ would yield a PD m of about -94 mV, and so the intracellular K + concentration can be calculated to be 113 mmol/liter. We previously reported [25] that the K + conductance of OK cells is nearly completely blocked by 3 retool/liter BaC12 in the extracellular fluid. Currentclamp experiments revealed that barium is more effective at negative PD m than at positive membrane potential with respect to the reduction of the overall cell membrane conductance (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…Cell culture and microelectrode measurements have been described elsewhere [25]. In brief, OK cells from passages 98 to 110 were maintained in bicarbonate-buffered Minimal Essential Medium (MEM) at pH 7.4.…”

Section: Methodsmentioning

confidence: 99%

“…The regulation of the intracellular pH of OK cells is mediated by PTH-and ANF-regulated Na+/H + exchange [16,17,22] that can be blocked by amiloride. We recently showed that monolayers of OK cells resemble a leaky epithelium [25] like the proximal tubule. Therefore, a confluent monolayer of OK cells serves as a suitable model for' the study of pHfdependent electrical properties in the proximal renal tubule.…”

Section: Introductionmentioning

confidence: 99%

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pH i -Dependent membrane conductance of proximal tubule cells in culture (OK): Differential effects on K+- and Na+-conductive channels

et al. 1990

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Confluent monolayers of the established opossum kidney cell line were exposed to NH4Cl pulses (20 mmol/liter) during continuous intracellular measurements of pH, membrane potential (PDm) and membrane resistance (R'm) in bicarbonate-free Ringer. The removal of extracellular NH4Cl leads to an intracellular acidification from a control value of 7.33 +/- 0.08 to 6.47 +/- 0.03 (n = 7). This inhibits the absolute K conductance (gK+), reflected by a decrease of K+ transference number from 71 +/- 3% (n = 28) to 26 +/- 6% (n = 5), a 2.6 +/- 0.2-fold rise of R'm, and a depolarization by 24.2 +/- 1.5 mV (n = 52). In contrast, intracellular acidification during a block of gK+ by 3 mmol/liter BaCl2 enhances the total membrane conductance, being shown by R'm decrease to 68 +/- 7% of control and cell membrane depolarization by 9.8 +/- 2.8 mV (n = 17). Conversely, intracellular alkalinization under barium elevates R'm and hyperpolarizes PDm. The replacement of extracellular sodium by choline in the presence of BaCl2 significantly hyperpolarizes PDm and increases R'm, indicating the presence of a sodium conductance. This conductance is not inhibited by 10(-4) mol/liter amiloride (n = 7). Patch-clamp studies at the apical membrane (excised inside-out configuration) revealed two Na(+)-conductive channels with 18.8 +/- 1.4 pS (n = 10) and 146 pS single-channel conductance. Both channels are inwardly rectifying and highly selective towards Cl-. The low-conductive channel is 4.8 times more permeable for Na+ than for K+. Its open probability rises at depolarizing potentials and is dependent on the pH of the membrane inside (higher at pH 6.5 than at pH 7.8).

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Section: Resultsmentioning

confidence: 99%

Section: Methodsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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pH i -Dependent membrane conductance of proximal tubule cells in culture (OK): Differential effects on K+- and Na+-conductive channels

et al. 1990

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“…21 For transcytosis study, OK cells were seeded on Transwell, and grown to complete confluence for 2 days (typical transepithelial resistance approximately 1-5 kV). 72 For uptake study, fluorescein-labeled aKlotho protein with C-terminal 6His tag was added into either apical or basal compartment and incubated for 4 hours. Apical or basal media was then collected for immunoblot for aKlotho with anti-His antibody.…”

Section: Opossum Kidney (Ok) Cellsmentioning

confidence: 99%

Renal Production, Uptake, and Handling of Circulating αKlotho

Shi

Zhang³

et al. 2016

Journal of the American Society of Nephrology

222

254

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ABSTRACTaKlotho is a multifunctional protein highly expressed in the kidney. Soluble aKlotho is released through cleavage of the extracellular domain from membrane aKlotho by secretases to function as an endocrine/paracrine substance. The role of the kidney in circulating aKlotho production and handling is incompletely understood, however. Here, we found higher aKlotho concentration in suprarenal compared with infrarenal inferior vena cava in both rats and humans. In rats, serum aKlotho concentration dropped precipitously after bilateral nephrectomy or upon treatment with inhibitors of aKlotho extracellular domain shedding. Furthermore, the serum half-life of exogenous aKlotho in anephric rats was four-to five-fold longer than that in normal rats, and exogenously injected labeled recombinant aKlotho was detected in the kidney and in urine of rats. Both in vivo (micropuncture) and in vitro (proximal tubule cell line) studies showed that aKlotho traffics from the basal to the apical side of the proximal tubule via transcytosis. Thus, we conclude that the kidney has dual roles in aKlotho homeostasis, producing and releasing aKlotho into the circulation and clearing aKlotho from the blood into the urinary lumen.

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“…Over the past years the opossum kidney (OK) cell line [1] has become a widely used model for the investigation of basic proper ties of proximal tubular transport systems such as ion channels [2][3][4], Na+/H+ ex changer [5][6][7][8], Na^-phosphate colransporter [9][10][11] and Na*-coupled amino acid and hexose transporter [12,13].…”

Section: Introductionmentioning

confidence: 99%

K<sup>+</sup>-, Cl<sup>–</sup>- and Nonselective Channels in Opossum Kidney Cells

Hollunder-Reese

Mohrmann

Greger

1991

Cell Physiol Biochem

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The opossum kidney (OK) cell line serves as a model for proximal tubular cells in many respects. The aim of the present study was to characterize the ion channels found in this cell line and to examine their regulation. OK cells of passages 85–92 were grown on glass coverslips in Dulbecco’s modified Eagle’s medium. Cells were used in a subconfiuent state. Standard patch-clamp experiments were carried out predominantly in the inside/out configuration. Four types of ion channels were found: large-conductance K⁺ channels (bK⁺), intermediate-conductance K⁺ channels (iK⁺), Cl^– channels and nonselective (NS) cation channels. The bK⁺ channel was Ca²⁺-dependent. It had a mean conductance of 174 ± 45 pS (n = 21) at a clamp voltage (V_c) = 0 mV (pipette KCl/bath NaCl). The channel was inhibited by Ba²⁺ (10^–3 mol/l, n = 7) in a voltage-dependent manner; the half-maximal inhibition (IC₅₀) was ≈ 10^–4 mol/l at V_c = 0 mV. Quinidine (10^–5 mol/l, n ≈ 4) as well as verapamil (≧ 10^–5 mol/l, n = 4) and diltiazem (5 × 10^-4 mol/l, n = 4) induced fast flickering and a reduction in mean open probability. Tetraethylammonium+ (10^–2 mol/l) blocked from the outside of the membrane only. The less frequently observed iK⁺ channel had a mean conductance of 62 ± 24 pS (n = 10). The channel showed no Ca²⁺ dependence but inactivation by Mg²⁺ at 10^–3 mol/l. The channel could be blocked reversibly by diltiazem (5 × 10^–4 mol/l, n = 1) and verapamil (10^–4 mol/l, n = 1). The Cl^– channel was outwardly rectifying and had an outward conductance of 51 ± 29 pS (g+) and an inward conductance of 36 ± 17pS (g; 5-Nitro-2-(3-phenylpropylamino)-benzoate at 10^–5 mol/l (n = 3) induced a reversible flicker block. The NS channel had a mean conductance of 27 ± 6 pS (n = 5) and did not rectify.

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Electrical properties of cultured renal tubular cells (OK) grown in confluent monolayers

Abstract: 1. The membrane potential of OK cells is largely determined by a pH-sensitive, barium-blockable K+-conductance. 2. Amiloride-blockable Na+/H+-exchange is reflected by membrane potential changes via this K+-conductance. 3. Monolayers of OK cells are electrically leaky.

Cited by 20 publications

References 23 publications

pH i -Dependent membrane conductance of proximal tubule cells in culture (OK): Differential effects on K+- and Na+-conductive channels

pH i -Dependent membrane conductance of proximal tubule cells in culture (OK): Differential effects on K+- and Na+-conductive channels

Renal Production, Uptake, and Handling of Circulating αKlotho

K<sup>+</sup>-, Cl<sup>–</sup>- and Nonselective Channels in Opossum Kidney Cells

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Electrical properties of cultured renal tubular cells (OK) grown in confluent monolayers

Abstract: 1. The membrane potential of OK cells is largely determined by a pH-sensitive, barium-blockable K+-conductance. 2. Amiloride-blockable Na+/H+-exchange is reflected by membrane potential changes via this K+-conductance. 3. Monolayers of OK cells are electrically leaky.

Cited by 20 publications

References 23 publications

pH i -Dependent membrane conductance of proximal tubule cells in culture (OK): Differential effects on K+- and Na+-conductive channels

pH i -Dependent membrane conductance of proximal tubule cells in culture (OK): Differential effects on K+- and Na+-conductive channels

Renal Production, Uptake, and Handling of Circulating αKlotho

K<sup>+</sup>-, Cl<sup>&ndash;</sup>- and Nonselective Channels in Opossum Kidney Cells

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Product

Resources

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K<sup>+</sup>-, Cl<sup>–</sup>- and Nonselective Channels in Opossum Kidney Cells