CFTR-dependent and -independent swelling-activated K+ currents in primary cultures of mouse nephron

2003 ◽  
Vol 284 (4) ◽  
pp. F812-F828 ◽  
Author(s):  
Radia Belfodil ◽  
Hervé Barrière ◽  
Isabelle Rubera ◽  
Michel Tauc ◽  
Chantal Poujeol ◽  
...  
Keyword(s):  
Primary Cultures ◽  
Regulatory Volume Decrease ◽  
Mouse Kidney ◽  
Wild Type ◽  
Collecting Tubule ◽  
K Currents ◽  
Volume Decrease ◽  
Autocrine Mechanism ◽  
Cortical Collecting Tubule

The role of CFTR in the control of K+ currents was studied in mouse kidney. Whole cell clamp was used to identify K+ currents on the basis of pharmacological sensitivities in primary cultures of proximal (PCT) and distal convoluted tubule (DCT) and cortical collecting tubule (CCT) from wild-type (WT) and CFTR knockout (KO) mice. In DCT and CCT cells, forskolin activated a 293B-sensitive K+ current in WT, but not in KO, mice. In these cells, a hypotonic shock induced K+ currents blocked by charybdotoxin in WT, but not in KO, mice. In PCT cells from WT and KO mice, the hypotonicity-induced K+ currents were insensitive to these toxins and were activated at extracellular pH 8.0 and inhibited at pH 6.0, suggesting that the corresponding channel was TASK2. In conclusion, CFTR is implicated in the control of KCNQ1 and Ca2+-sensitive swelling-activated K+ conductances in DCT and CCT, but not in proximal convoluted tubule, cells. In KO mice, impairment of the regulatory volume decrease process in DCT and CCT could be due to the loss of an autocrine mechanism, implicating ATP and adenosine, which controls swelling-activated Cl− and K+channels.

scholarly journals Role of TASK2 Potassium Channels Regarding Volume Regulation in Primary Cultures of Mouse Proximal Tubules

2003 ◽  
Vol 122 (2) ◽  
pp. 177-190 ◽  
Author(s):  
Herve Barriere ◽  
Radia Belfodil ◽  
Isabelle Rubera ◽  
Michel Tauc ◽  
Florian Lesage ◽  
...  
Keyword(s):  
Cell Volume ◽  
Volume Regulation ◽  
Primary Cultures ◽  
Regulatory Volume Decrease ◽  
Cell Volume Regulation ◽  
Proximal Tubules ◽  
Wild Type ◽  
Convoluted Tubules ◽  
K Currents

Several papers reported the role of TASK2 channels in cell volume regulation and regulatory volume decrease (RVD). To check the possibility that the TASK2 channel modulates the RVD process in kidney, we performed primary cultures of proximal convoluted tubules (PCT) and distal convoluted tubules (DCT) from wild-type and TASK2 knockout (KO) mice. In KO mice, the TASK2 coding sequence was in part replaced by the lac-Z gene. This allows for the precise localization of TASK2 in kidney sections using β-galactosidase staining. TASK2 was only localized in PCT cells. K+ currents were analyzed by the whole-cell clamp technique with 125 mM K-gluconate in the pipette and 140 mM Na-gluconate in the bath. In PCT cells from wild-type mice, hypotonicity induced swelling-activated K+ currents insensitive to 1 mM tetraethylammonium, 10 nM charybdotoxin, and 10 μM 293B, but blocked by 500 μM quinidine and 10 μM clofilium. These currents were increased in alkaline pH and decreased in acidic pH. In PCT cells from TASK2 KO, swelling-activated K+ currents were completely impaired. In conclusion, the TASK2 channel is expressed in kidney proximal cells and could be the swelling-activated K+ channel responsible for the cell volume regulation process during osmolyte absorptions in the proximal tubules.

Extracellular pH alkalinization by Cl−/HCO3− exchanger is crucial for TASK2 activation by hypotonic shock in proximal cell lines from mouse kidney

2007 ◽  
Vol 292 (2) ◽  
pp. F628-F638 ◽  
Author(s):  
S. L'Hoste ◽  
H. Barriere ◽  
R. Belfodil ◽  
I. Rubera ◽  
C. Duranton ◽  
...  
Keyword(s):  
Cell Lines ◽  
Regulatory Volume Decrease ◽  
Inhibitory Effect ◽  
Buffering Capacity ◽  
Mouse Kidney ◽  
Wild Type ◽  
Cytosolic Ph ◽  
Hypotonic Shock ◽  
External Ph ◽  
Volume Decrease

We have previously shown that K+-selective TASK2 channels and swelling-activated Cl− currents are involved in a regulatory volume decrease (RVD; Barriere H, Belfodil R, Rubera I, Tauc M, Lesage F, Poujeol C, Guy N, Barhanin J, Poujeol P. J Gen Physiol 122: 177–190, 2003; Belfodil R, Barriere H, Rubera I, Tauc M, Poujeol C, Bidet M, Poujeol P. Am J Physiol Renal Physiol 284: F812–F828, 2003). The aim of this study was to determine the mechanism responsible for the activation of TASK2 channels during RVD in proximal cell lines from mouse kidney. For this purpose, the patch-clamp whole-cell technique was used to test the effect of pH and the buffering capacity of external bath on Cl− and K+ currents during hypotonic shock. In the presence of a high buffer concentration (30 mM HEPES), the cells did not undergo RVD and did not develop outward K+ currents (TASK2). Interestingly, the hypotonic shock reduced the cytosolic pH (pHi) and increased the external pH (pHe) in wild-type but not in cftr −/− cells. The inhibitory effect of DIDS suggests that the acidification of pHi and the alkalinization of pHe induced by hypotonicity in wild-type cells could be due to an exit of HCO3−. In conclusion, these results indicate that Cl− influx will be the driving force for HCO3− exit through the activation of the Cl−/HCO3− exchanger. This efflux of HCO3− then alkalinizes pHe, which in turn activates TASK2 channels.

Volume regulatory Cl- loss after Na+ pump inhibition in CCT principal cells

1991 ◽  
Vol 260 (2) ◽  
pp. F225-F234 ◽  
Author(s):  
K. Strange
Keyword(s):  
Steady State ◽  
High Selectivity ◽  
Regulatory Volume Decrease ◽  
Disulfonic Acid ◽  
Principal Cells ◽  
Na Pump ◽  
Collecting Tubule ◽  
Volume Decrease ◽  
Rabbit Cortical Collecting Tubule ◽  
Cortical Collecting Tubule

Ouabain caused rabbit cortical collecting tubule (CCT) principal cells to swell 53% and then undergo regulatory volume decrease (RVD) at a rate of 4%/min to a new steady-state volume 10% below control. Reduction of peritubular Cl- concentration transiently depolarized transepithelial potential (Vte) by 36 mV and stimulated the rate of RVD 30-fold. Peritubular application of 0.5 mM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) inhibited RVD 74%. In contrast, luminal Cl- reduction or application of DIDS had no effect on RVD. A 10-fold elevation of perfusate K+ caused volume-regulated cells to swell 23% at a rate of 60%/min. Removal of luminal Cl- had no effect on either the rate or magnitude of K+ swelling. Peritubular or bilateral Cl- removal, however, inhibited the rate of K+ swelling by 96 and 99%, respectively. Substitution of bath Cl- for Br-, SCN-, or I- inhibited the rate of K+ swelling by 40, 38, and 98%, respectively. Surprisingly, NO3- inhibited the rate of K+ swelling by 82%. All Cl- substitutes tested transiently depolarized Vte by 3–49 mV. These results suggest strongly that RVD is mediated by a basolateral Cl- channel with a high selectivity for Cl- over other anions.

Amiloride-sensitive sodium channels in rabbit cortical collecting tubule primary cultures

1991 ◽  
Vol 261 (6) ◽  
pp. F933-F944 ◽  
Author(s):  
B. N. Ling ◽  
C. F. Hinton ◽  
D. C. Eaton
Keyword(s):  
High Selectivity ◽  
Primary Cultures ◽  
Principal Cell ◽  
Na Channel ◽  
Open Probability ◽  
Inward Current ◽  
Collecting Tubule ◽  
Apical Membranes ◽  
Rabbit Cortical Collecting Tubule ◽  
Cortical Collecting Tubule

Patch-clamp methodology was applied to principal cell apical membranes of rabbit cortical collecting tubule (CCT) primary cultures grown on collagen supports in the presence of aldosterone (1.5 microM). The most frequently observed channel had a unit conductance of 3-5 pS, nonlinear current-voltage (I-V) relationship, Na permeability (PNa)-to-K permeability (PK) ratio greater than 19:1, and inward current at all applied potentials (Vapp) less than +80 mV (n = 41). Less frequently, an 8- to 10-pS channel with a linear I-V curve, PNa/PK less than 5:1, and inward current at Vapp less than +40 mV was also observed (n = 7). Luminal amiloride (0.75 microM) decreased the open probability (Po) for both of these channels. Mean open time for the high-selectivity Na+ channel was 2.1 +/- 0.5 s and for the low-selectivity Na+ channel was 50 +/- 12 ms. In primary cultures grown without aldosterone the high-selectivity Na+ channel was rarely observed (1 of 32 patches). Lastly, a 26- to 35-pS channel, nonselective for Na+ over K+, was not activated by cytoplasmic Ca2+ or voltage nor inhibited by amiloride (n = 17). We conclude that under specific growth conditions, namely permeable transporting supports and chronic mineralocorticoid hormone exposure, principal cell apical membranes of rabbit CCT primary cultures contain 1) both high-selectivity and low-selectivity, amiloride-inhibitable Na+ channels and 2) amiloride-insensitive, nonselective cation channels.

Evidence of corticosteroid action along the nephron

1984 ◽  
Vol 246 (2) ◽  
pp. F111-F123 ◽  
Author(s):  
D. Marver
Keyword(s):  
Collecting Duct ◽  
Type I ◽  
Type Ii ◽  
Cortical Collecting Duct ◽  
Distal Nephron ◽  
Type Iii ◽  
Collecting Ducts ◽  
Collecting Tubule ◽  
Cortical Collecting Tubule

The kidney contains three classes of corticosteroid-binding proteins receptors. They include a mineralocorticoid-specific (Type I), a glucocorticoid-specific (Type II), and a corticosterone-specific (Type III) site. The Type I and Type III sites roughly parallel each other along the nephron, with maximal binding occurring in the late distal convoluted or connecting segment and the cortical and medullary collecting ducts. Type II sites occur throughout the nephron, with maximal concentrations appearing in the proximal tubule and the late distal convoluted-cortical collecting duct region. The function of the Type I sites in the connecting segment is unclear since chronic mineralocorticoid therapy does not influence the potential difference in this segment as it does in the cortical collecting tubule. Furthermore, the specific role of Type II versus Type III sites in the distal nephron is unknown. Finally, the possible influence of sodium on both latent and steroid-induced renal cortical and medullary Na-K-ATPase is discussed.

Difference in the Na affinity of Na(+)-K(+)-ATPase along the rabbit nephron: modulation by K

1990 ◽  
Vol 259 (2) ◽  
pp. F246-F250 ◽  
Author(s):  
C. Barlet-Bas ◽  
L. Cheval ◽  
C. Khadouri ◽  
S. Marsy ◽  
A. Doucet
Keyword(s):  
Atpase Activity ◽  
Thick Ascending Limb ◽  
Kidney Cells ◽  
Nephron Segments ◽  
Adaptive Property ◽  
Collecting Tubule ◽  
Pump Activity ◽  
K Concentration ◽  
Cortical Collecting Tubule

The sensitivity of Na(+)-K(+)-ATPase to Na was determined in single segments of rabbit nephron isolated by microdissection. In the cortical collecting tubule (CCT), Na(+)-K(+)-ATPase was threefold more sensitive to Na (apparent K0.5 approximately 3 mM) than in proximal convoluted tubule and cortical thick ascending limb (apparent K0.5 approximately 10 mM). Furthermore, increasing K concentration from 5 to greater than 100 mM markedly reduced the affinity of the pump for Na in all three nephron segments. In fact, the main shift in Na affinity occurred when K changed from 100 to 120 mM; in the CCT, increasing K concentration from 100 to 120 mM while maintaining Na concentration at 10 mM reduced Na(+)-K(+)-ATPase activity by greater than 35%. These findings confirm that, in kidney cells as in other cells, intracellular Na limits the rate of Na(+)-K(+)-ATPase. Thus any alteration of intracellular Na concentration modifies the pump activity in a way that contributes to the restoration of intracellular Na homeostasis. This adaptive property is particularly efficient in the collecting tubule in which the apparent K0.5 of the pump for Na is close to normal intracellular Na concentration. Furthermore, changes in intracellular K concentration, which usually accompany those of Na so as to maintain the total cation concentration constant, potentiate the regulatory role of Na through modifications of its affinity for the pump.

RVD in principal and intercalated cells of rabbit cortical collecting tubule

1988 ◽  
Vol 255 (5) ◽  
pp. C612-C621 ◽  
Author(s):  
K. Strange
Keyword(s):  
Mammalian Cells ◽  
Control Volume ◽  
Regulatory Volume Decrease ◽  
Isotonic Saline ◽  
Cell Types ◽  
Intercalated Cells ◽  
Intercalated Cell ◽  
Collecting Tubule ◽  
Principal And Intercalated Cells ◽  
Cortical Collecting Tubule

Cells of the rabbit renal cortical collecting tubule possess significant regulatory volume decrease (RVD) capabilities. After a 100-mosmol/kg reduction in peritubular osmolality, principal and intercalated cells swell 40-45 and 30-35%, respectively, and immediately activate RVD mechanisms. Both cell types downregulate their volume to within 5-6% of control volume at initial rates of 3-6%/min. Return to isotonic saline causes both cell types to shrink (isotonic shrinkage) 25-35% below control volume due to the loss of osmotically active intracellular solutes during RVD. In most mammalian cells studied to date, RVD is mediated largely by passive KCl efflux via KCl cotransport, parallel K+ and Cl- channels, or parallel K+-H+ and Cl- -HCO3- exchange mechanisms. Peritubular application of 0.1 mM ouabain (0 Na+ lumen), bilateral CO2-HCO3- removal, or bilateral application of 0.02 mM bumetanide, 2.0 mM Ba2+, 2.0 mM anthracene-9-carboxylic acid, or 0.5 mM SITS had no significant effect on rates or magnitudes of RVD and isotonic shrinkage in either cell type. Bilateral elevation of K+ from 5 to 52.5 mM reverses or reduces the electrochemical gradient for K+ movement, causing accumulation of this ion in the cytoplasm, but had no effect on the rates or magnitude of principal and intercalated cell RVD. Principal and intercalated cells from K+- or Cl- -depleted tubules (1 h bilateral perfusion with K+- or Cl- -free saline at 37 degrees C) showed normal rates and magnitudes of RVD in K+- or Cl- -free hypotonic saline. Taken together, these results argue against a significant role of passive KCl efflux pathways in mediating principal and intercalated cell RVD.

Regulation of renal Na+-K+-ATPase in the rat: role of increased potassium transport

1986 ◽  
Vol 251 (2) ◽  
pp. F199-F207
Author(s):  
S. K. Mujais ◽  
M. A. Chekal ◽  
J. P. Hayslett ◽  
A. I. Katz
Keyword(s):  
Enzyme Activity ◽  
Atpase Activity ◽  
Potassium Transport ◽  
High Potassium ◽  
Dietary Potassium ◽  
Collecting Tubule ◽  
Physiological Dose ◽  
Potassium Loading ◽  
Cortical Collecting Tubule

The purpose of this study was to characterize the alterations in collecting tubule Na+-K+-ATPase activity produced by sustained increments in dietary potassium in the rat and to evaluate the role of aldosterone in their generation. In adrenal-intact animals, feeding a high-potassium diet (10-fold that of control) or administration of a high physiological dose of aldosterone (5 micrograms X 100 g-1 X day-1), which simulates the delivery rate of this hormone during potassium loading (both for 7 days), caused marked increments in Na+-K+-ATPase activity in the cortical collecting tubule (CCT) but had no effect on the enzyme in the inner stripe of the medullary collecting tubule (MCT). A significant increase in enzyme activity was also observed after smaller dietary potassium increments (2.5 and 5 times the control) and after 4 (but not 2) days of dietary potassium load. In adrenalectomized rats provided with physiological replacement doses of corticosterone and aldosterone (0.8 micrograms X 100 g-1 X day-1), Na+-K+-ATPase activity in both CCT and MCT was similar to that of adrenal-intact controls but remained unchanged after 7 days on the potassium-enriched (10-fold) diet. In contrast, adrenalectomized animals receiving the high physiological dose of aldosterone displayed an increase in Na+-K+-ATPase activity of CCT comparable with that of adrenal-intact animals, whereas the enzyme activity in the MCT was unaffected. In conclusion, 1) following chronic potassium loading Na+-K+-ATPase activity increases significantly in the CCT with no change in its activity in the inner stripe of the MCT.(ABSTRACT TRUNCATED AT 250 WORDS)

Volume-activated K+ and Cl- pathways of dissociated epithelial cells (MDCK): role of Ca2+

1990 ◽  
Vol 258 (5) ◽  
pp. C827-C834 ◽  
Author(s):  
A. Rothstein ◽  
E. Mack
Keyword(s):  
Mdck Cells ◽  
Regulatory Volume Decrease ◽  
Disulfonic Acid ◽  
Ionophore A23187 ◽  
Madin Darby Canine Kidney ◽  
Osmotic Swelling ◽  
Volume Activation ◽  
Darby Canine Kidney ◽  
Volume Decrease

Osmotic swelling of dissociated Madin-Darby canine kidney (MDCK) cells in NaCl medium is followed by shrinking (regulatory volume decrease, or RVD) or in KCl medium by secondary swelling. The cation ionophore gramicidin has little effect on volumes of isotonic cells but accelerates volume-activated changes in either medium. Immediately after hypotonic exposure, the membrane becomes transiently hyperpolarized followed by depolarization. The depolarization phase is diminished by the anion transport inhibitor 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). Swelling is also associated with an almost immediate increase in Ca2+ influx and elevation of cytoplasmic Ca2+ ([Ca2+]i) preceding RVD. In Ca2(+)-free medium, [Ca2+]i rapidly declines to a low level. Osmotic swelling, under these circumstances, is associated with a small transient increase in [Ca2+]i, but RVD or secondary swelling (in KCl) are minimal. Under these conditions, addition of gramicidin or the Ca2(+)-ionophore A23187 induces significant volume changes, although not as large as those found in the presence of Ca2+. Quinine inhibits RVD in the absence of gramicidin, but not in its presence; oligomycin C, DIDS, and trifluoperazine, on the other hand, inhibit in the presence of the ionophore. These findings suggest that in MDCK cells RVD involves activation of distinct conductive K+ and Cl- pathways which allow escape of KCl and osmotically obligated water and that activation of both pathways is associated with elevated [Ca2+]i derived largely from volume activation of a Ca2(+)-influx pathway.

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