scholarly journals Inhibition of ROMK channels by low extracellular K+ and oxidative stress

2013 ◽  
Vol 305 (2) ◽  
pp. F208-F215 ◽  
Author(s):  
Gustavo Frindt ◽  
Hui Li ◽  
Henry Sackin ◽  
Lawrence G. Palmer
Keyword(s):  
Oxidative Stress ◽  
Collecting Duct ◽  
Cell Voltage ◽  
Dependent Manner ◽  
Cortical Collecting Duct ◽  
Pipette Solution ◽  
Whole Cell ◽  
Tert Butyl Hydroperoxide ◽  
Low K ◽  
And Oxidative Stress

We tested the hypothesis that low luminal K+ inhibits the activity of ROMK channels in the rat cortical collecting duct. Whole-cell voltage-clamp measurements of the component of outward K+ current inhibited by the bee toxin Tertiapin-Q ( ISK) showed that reducing the bath concentration ([K+]o) to 1 mM resulted in a decline of current over 2 min compared with that observed at 10 mM [K+]o. However, maintaining tubules in 1 mM [K+]o without establishing whole-cell clamp conditions did not affect ISK. The [K+]o-dependent decline was not prevented by increasing cytoplasmic-side pH or by inhibition of phosphatase activity. It was, however, abolished by the inclusion of 0.5 mM DTT in the pipette solution to prevent oxidation of the intracellular environment. Conversely, treatment of intact tubules with the oxidant H2O2 (100 μM) decreased ISK in a [K+]o-dependent manner. Treatment of the tubules with the phospholipase C inhibitor U73122 prevented the effect of low [K+]o, suggesting the involvement of this enzyme in the process. We examined these effects further using Xenopus oocytes expressing ROMK2 channels. A 50-min exposure to the permeant oxidizing agent tert-butyl hydroperoxide (t-BHP; 500 μM) did not affect outward K+ currents with [K+]o = 10 mM but reduced currents by 50% with [K+]o = 1 mM and by 75% with [K+]o = 0.1 mM. Pretreatment of the oocytes with U73122 prevented the effects of t-BHP. Under conditions of low dietary K intake, K+ secretion by distal nephron segments may be suppressed by a combination of low luminal [K+]o and oxidative stress.

scholarly journals Conservation of Na+ vs. K+ by the rat cortical collecting duct

2011 ◽  
Vol 301 (1) ◽  
pp. F14-F20 ◽  
Author(s):  
Gustavo Frindt ◽  
Véronique Houde ◽  
Lawrence G. Palmer
Keyword(s):  
Collecting Duct ◽  
Immunoblot Analysis ◽  
Cortical Collecting Duct ◽  
Channel Activity ◽  
Distal Nephron ◽  
Whole Cell ◽  
Regulation Of Transport ◽  
Low K ◽  
Na Depletion ◽  
K Depletion

Regulation of transport by principal cells of the distal nephron contributes to maintenance of Na+ and K+ homeostasis. To assess which of these ions is given a higher priority by these cells, we investigated the upregulation of epithelial Na+ channels (ENaC) in the rat cortical collecting duct (CCD) during Na depletion with and without simultaneous K depletion. ENaC activity, assessed as whole cell amiloride-sensitive current in split-open tubules, was 260 ± 40 pA/cell in K-repleted but virtually undetectable (3 ± 1 pA/cell) in K-depleted animals. This difference was confirmed biochemically by the reduced amounts of the cleaved forms of both the α-ENaC and γ-ENaC subunits measured in immunoblots. In contrast, in K-depleted rats, simultaneously reducing Na intake did not affect the activity of ROMK channels, assessed as tertiapin-Q-sensitive whole cell currents, in the CCDs. The lack of Na current in K-depleted animals was the result of reduced levels of aldosterone in plasma, rather than a reduced sensitivity to the hormone. However, rats on a low-Na, low-K diet for 1 wk did not excrete more Na than those on a low-Na, control-K diet for the same period of time. Immunoblot analysis indicated increased levels of the thiazide-sensitive NaCl cotransporter and the apical Na-H exchanger NHE3. This suggests that with reduced K intake, Na balance is maintained despite reduced aldosterone and Na+ channel activity by upregulation of Na+ transport in upstream segments. Under these conditions, Na+ transport by the aldosterone-sensitive distal nephron is reduced, despite the low-Na intake to minimize K+ secretion and urinary K losses.

scholarly journals Dietary K regulates ROMK channels in connecting tubule and cortical collecting duct of rat kidney

2009 ◽  
Vol 296 (2) ◽  
pp. F347-F354 ◽  
Author(s):  
Gustavo Frindt ◽  
Anish Shah ◽  
Johan Edvinsson ◽  
Lawrence G. Palmer
Keyword(s):  
Collecting Duct ◽  
Rat Kidney ◽  
Outward Current ◽  
Equilibrium Potential ◽  
Cortical Collecting Duct ◽  
Pipette Solution ◽  
Connecting Tubule ◽  
Outward Currents ◽  
High K ◽  
Low K

The activity of ROMK channels in rat kidney tubule cells was assessed as tertiapin-Q (TPNQ)-sensitive current under whole cell clamp conditions. With an external K+ concentration of 5 mM and an internal K+ concentration of 140 mM and the membrane potential clamped to 0 mV, TPNQ blocked outward currents in principal cells of the cortical collecting duct (CCD) outer medullary collecting duct and connecting tubule (CNT). The apparent Ki was 5.0 nM, consistent with its interaction with ROMK. The TPNQ-sensitive current reversed at voltages close to the equilibrium potential for K+. The currents were reduced when the pipette solution contained ATP. In the CCD, the average TPNQ-sensitive outward current ( ISK) was 476 ± 48 pA/cell in control animals on a 1% KCl diet. ISK increased to 1,255 ± 140 pA when animals were maintained on a high-K (10% KCl) diet for 7 days and decreased to 314 ± 46 pA after 7 days on a low-K (0.1% KCl) diet. In the CNT, ISK was 360 ± 30 pA on control, 1,160 ± 110 on high-K, and 166 ± 16 pA on low-K diets. The results indicate that ROMK channel activity is highly regulated by dietary K in both the CCD and the CNT.

scholarly journals NF-κB and FosB mediate inflammation and oxidative stress in the blast lung injury of rats exposed to shock waves

2021 ◽  
Author(s):  
Hong Wang ◽  
Wenjuan Zhang ◽  
Jinren Liu ◽  
Junhong Gao ◽  
Le Fang ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Lung Injury ◽  
Shock Waves ◽  
Time Dependent ◽  
Inflammatory Factors ◽  
Control Group ◽  
Dependent Manner ◽  
Total Antioxidant ◽  
Blast Lung Injury ◽  
And Oxidative Stress

Abstract Blast lung injury (BLI) is the major cause of death in explosion-derived shock waves; however, the mechanisms of BLI are not well understood. To identify the time-dependent manner of BLI, a model of lung injury of rats induced by shock waves was established by a fuel air explosive. The model was evaluated by hematoxylin and eosin staining and pathological score. The inflammation and oxidative stress of lung injury were also investigated. The pathological scores of rats’ lung injury at 2 h, 24 h, 3 days, and 7 days post-blast were 9.75±2.96, 13.00±1.85, 8.50±1.51, and 4.00±1.41, respectively, which were significantly increased compared with those in the control group (1.13±0.64; P<0.05). The respiratory frequency and pause were increased significantly, while minute expiratory volume, inspiratory time, and inspiratory peak flow rate were decreased in a time-dependent manner at 2 and 24 h post-blast compared with those in the control group. In addition, the expressions of inflammatory factors such as interleukin (IL)-6, IL-8, FosB, and NF-κB were increased significantly at 2 h and peaked at 24 h, which gradually decreased after 3 days and returned to normal in 2 weeks. The levels of total antioxidant capacity, total superoxide dismutase, and glutathione peroxidase were significantly decreased 24 h after the shock wave blast. Conversely, the malondialdehyde level reached the peak at 24 h. These results indicated that inflammatory and oxidative stress induced by shock waves changed significantly in a time-dependent manner, which may be the important factors and novel therapeutic targets for the treatment of BLI.

Whole cell and unitary amiloride-sensitive sodium currents in M-1 mouse cortical collecting duct cells

1996 ◽  
Vol 270 (4) ◽  
pp. C998-C1010 ◽  
Author(s):  
M. L. Chalfant ◽  
T. G. O'Brien ◽  
M. M. Civan
Keyword(s):  
Collecting Duct ◽  
Cell Permeability ◽  
Na Channel ◽  
Na Channels ◽  
Cortical Collecting Duct ◽  
Ionic Selectivity ◽  
Whole Cell ◽  
Duct Cells ◽  
Excised Patches ◽  
Collecting Duct Cells

Amiloride-sensitive whole cell currents have been reported in M-1 mouse cortical collecting duct cells (Korbmacher et al., J. Gen. Physiol. 102: 761-793, 1993). We have confirmed that amiloride inhibits the whole cell currents but not necessarily the measured whole cell currents. Anomalous responses were eliminated by removing external Na+ and/or introducing paraepithelial shunts. The amiloride-sensitive whole cell currents displayed Goldman rectification. The ionic selectivity sequence of the amiloride-sensitive conductance was Li+ > Na+ >> K+. Growth of M-1 cells on permeable supports increased the amiloride-sensitive whole cell permeability, compared with cells grown on plastic. Single amiloride-sensitive channels were observed, which conformed to the highly selective low-conductance amiloride-sensitive class [Na(5)] of epithelial Na+ channels. Hypotonic pretreatment markedly slowed run-down of channel activity. The gating of the M-1 Na+ channel in excised patches was complex. Open- and closed-state dwell-time distributions from patches that display one operative channel were best described with two or more exponential terms each. We conclude that 1) study of M-1 whole cell Na+ currents is facilitated by reducing the transepithelial potential to zero, 2) these M-1 currents reflect the operation of Na(5) channels, and 3) the Na+ channels display complex kinetics, involving > or = 2 open and > or = 2 closed states.

scholarly journals Localization of Inward Rectifier Potassium Channel Kir7.1 in the Basolateral Membrane of Distal Nephron and Collecting Duct

2000 ◽  
Vol 11 (11) ◽  
pp. 1987-1994
Author(s):  
KAYOKO OOKATA ◽  
AKIHIRO TOJO ◽  
YOSHIRO SUZUKI ◽  
NOBUHIRO NAKAMURA ◽  
KENJIRO KIMURA ◽  
...  
Keyword(s):  
Potassium Channel ◽  
Collecting Duct ◽  
Basolateral Membrane ◽  
Inward Rectifier ◽  
Kidney Cortex ◽  
Thick Ascending Limb ◽  
Cortical Collecting Duct ◽  
Distal Nephron ◽  
Connecting Tubule ◽  
Low K

Abstract. Inward rectifier potassium channels (Kir) play an important role in the K+ secretion from the kidney. Recently, a new subfamily of Kir, Kir7.1, has been cloned and shown to be present in the kidney as well as in the brain, choroid plexus, thyroid, and intestine. Its cellular and subcellular localization was examined along the renal tubule. Western blot from the kidney cortex showed a single band for Kir7.1 at 52 kD, which was also observed in microdissected segments from the thick ascending limb of Henle, distal convoluted tubule (DCT), connecting tubule, and cortical and medullary collecting ducts. Kir7.1 immunoreactivity was detected predominantly in the DCT, connecting tubule, and cortical collecting duct, with lesser expression in the thick ascending limb of Henle and in the medullary collecting duct. Kir7.1 was detected by electron microscopic immunocytochemistry on the basolateral membrane of the DCT and the principal cells of cortical collecting duct, but neither type A nor type B intercalated cells were stained. The message levels and immunoreactivity were decreased under low-K diet and reversed by low-K diet supplemented with 4% KCl. By the double-labeling immunogold method, both Kir7.1 and Na+, K+-ATPase were independently located on the basolateral membrane. In conclusion, the novel Kir7.1 potassium channel is located predominantly in the basolateral membrane of the distal nephron and collecting duct where it could function together with Na+, K+-ATPase and contribute to cell ion homeostasis and tubular K+ secretion.

Effect of insulin on potassium secretion in rabbit cortical collecting ducts

1992 ◽  
Vol 262 (1) ◽  
pp. F30-F35 ◽  
Author(s):  
H. Furuya ◽  
K. Tabei ◽  
S. Muto ◽  
Y. Asano
Keyword(s):  
Collecting Duct ◽  
Dependent Manner ◽  
Cortical Collecting Duct ◽  
Plasma Insulin Concentration ◽  
Concentration Dependent Manner ◽  
K Secretion ◽  
Potassium Secretion ◽  
Transepithelial Voltage ◽  
K Transport

Insulin is known to play an important role in the regulation of extrarenal K homeostasis. Previous clearance studies have shown that insulin decreases urinary K excretion, but the responsible nephron segments have not been identified. In this microperfusion study, in vitro, the effect of insulin on K transport in the cortical collecting duct (CCD), which is thought to be an important segment for regulation of the final urinary K excretion, was investigated. Basolateral insulin (10(-6) M) significantly inhibited net K secretion by 20% (mean JK = -26.2 +/- 4.2 peq.mm-1.min-1 for controls compared with -21.1 +/- 3.4 with insulin, P less than 0.001) and depolarized the transepithelial voltage (VT, from -14.6 +/- 3.5 to -10.8 +/- 3.5 mV, P less than 0.005), recovery did not occur over 60 min. Insulin (10(-11)-10(-5) M) depressed K secretion and depolarized the VT in a concentration-dependent manner. The half-maximal concentration was 5 x 10(-10) M, which is within the physiological range of plasma insulin concentration. In tubules of deoxycorticosterone acetate-treated rabbits, insulin also produced a significant fall in K secretion (from -43.4 +/- 7.5 to -36.1 +/- 5.7 peq.mm-1.min-1, P less than 0.05). Although luminal Ba (2 mM) decreased K secretion (from -14.4 +/- 2.9 to -7.0 +/- 1.7 peq.mm-1.min-1), basolateral insulin (10(-6) M) inhibited K secretion further (to -4.7 +/- 1.3 peq.mm-1.min-1, P less than 0.01).(ABSTRACT TRUNCATED AT 250 WORDS)

Acute reduction in whole cell conductance in anoxic turtle brain

1999 ◽  
Vol 277 (3) ◽  
pp. R887-R893 ◽  
Author(s):  
H. S. Ghai ◽  
L. T. Buck
Keyword(s):  
Brain Slices ◽  
Cell Voltage ◽  
Dependent Manner ◽  
Recording Electrode ◽  
Whole Cell ◽  
Artificial Cerebrospinal Fluid ◽  
Intracellular Ca ◽  
Specific Antagonist ◽  
Acute Changes ◽  
Dose Dependent

We tested the effect of anoxia, a “mimic” turtle artificial cerebrospinal fluid (aCSF) consisting of high Ca2+ and Mg2+ concentrations and low pH and adenosine perfusions, on whole cell conductance ( G w) in turtle brain slices using a whole cell voltage-clamp technique. With EGTA in the recording electrode, anoxic or adenosine perfusions did not change G w significantly (values range between 2.15 ± 0.24 and 3.24 ± 0.56 nS). However, perfusion with normoxic or anoxic mimic aCSF significantly decreased G w. High [Ca2+] (4.0 or 7.8 mM) perfusions alone could reproduce the changes in G w found with the mimic perfusions. With the removal of EGTA from the recording electrode, G wdecreased significantly during both anoxic and adenosine perfusions. The A1-receptor agonist N 6-cyclopentyladenosine reduced G w in a dose-dependent manner, whereas the A1-receptor specific antagonist 8-cyclopentyl-1,3-dipropylxanthine blocked both the adenosine- and anoxic-mediated changes in G w. These data suggest a mechanism involving A1-receptor-mediated changes in intracellular [Ca2+] that result in acute changes in G w with the onset of anoxia.

Conductive properties of a rabbit cortical collecting duct cell line: regulation by isoproterenol

1992 ◽  
Vol 262 (4) ◽  
pp. F578-F582 ◽  
Author(s):  
P. Dietl ◽  
N. Kizer ◽  
B. A. Stanton
Keyword(s):  
Protein Kinase ◽  
Protein Kinase A ◽  
Cell Line ◽  
Collecting Duct ◽  
Cortical Collecting Duct ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Conductive Properties ◽  
Cl Conductance ◽  
Kinase A

The present study was carried out to characterize the membrane conductive properties of RCCT-28A cells, a continuous cell line derived from rabbit cortical collecting duct (CCD). RCCT-28A cells have many phenotypic properties of acid-secreting intercalated cells (A-IC). Using the whole cell patch-clamp technique, we found that the cells are conductive to Cl-, but not to Na+ or K+. The beta-adrenergic agonists isoproterenol (2 x 10(-6) M) and adenosine 3',5'-cyclic monophosphate (cAMP, 10(-4) M) increased the whole cell Cl- conductance. Protein kinase A (150 nM) in the patch pipette (i.e., intracellular solution) also increased whole cell Cl- conductance. Because isoproterenol increases cAMP levels in these cells, we conclude that isoproterenol stimulates the Cl- conductance by increasing cell cAMP, which in turn activates protein kinase A. In contrast, vasopressin does not increase cAMP in these cells and did not increase the Cl- conductance. In conclusion, these experiments show that RCCT-28A cells, like A-IC, are conductive only to Cl-. Thus RCCT-28A cells are a good model with which to study Cl- channels in the collecting duct.

scholarly journals β-Lapachone Induces Acute Oxidative Stress in Rat Primary Astrocyte Cultures that is Terminated by the NQO1-Inhibitor Dicoumarol

2020 ◽  
Vol 45 (10) ◽  
pp. 2442-2455
Author(s):  
Johann Steinmeier ◽  
Sophie Kube ◽  
Gabriele Karger ◽  
Eric Ehrke ◽  
Ralf Dringen
Keyword(s):  
Oxidative Stress ◽  
Dependent Manner ◽  
Oxygen Species ◽  
Glutathione Disulfide ◽  
Cellular Accumulation ◽  
Primary Astrocyte ◽  
Cultured Astrocytes ◽  
Rat Astrocytes ◽  
Quinone Acceptor ◽  
And Oxidative Stress

Abstract β-lapachone (β-lap) is reduced in tumor cells by the enzyme NAD(P)H: quinone acceptor oxidoreductase 1 (NQO1) to a labile hydroquinone which spontaneously reoxidises to β-lap, thereby generating reactive oxygen species (ROS) and oxidative stress. To test for the consequences of an acute exposure of brain cells to β-lap, cultured primary rat astrocytes were incubated with β-lap for up to 4 h. The presence of β-lap in concentrations of up to 10 µM had no detectable adverse consequences, while higher concentrations of β-lap compromised the cell viability and the metabolism of astrocytes in a concentration- and time-dependent manner with half-maximal effects observed for around 15 µM β-lap after a 4 h incubation. Exposure of astrocytes to β-lap caused already within 5 min a severe increase in the cellular production of ROS as well as a rapid oxidation of glutathione (GSH) to glutathione disulfide (GSSG). The transient cellular accumulation of GSSG was followed by GSSG export. The β-lap-induced ROS production and GSSG accumulation were completely prevented in the presence of the NQO1 inhibitor dicoumarol. In addition, application of dicoumarol to β-lap-exposed astrocytes caused rapid regeneration of the normal high cellular GSH to GSSG ratio. These results demonstrate that application of β-lap to cultured astrocytes causes acute oxidative stress that depends on the activity of NQO1. The sequential application of β-lap and dicoumarol to rapidly induce and terminate oxidative stress, respectively, is a suitable experimental paradigm to study consequences of a defined period of acute oxidative stress in NQO1-expressing cells.

Oxytocin affects apical sodium conductance in rabbit cortical collecting duct

1993 ◽  
Vol 265 (4) ◽  
pp. F487-F503 ◽  
Author(s):  
T. Inoue ◽  
M. Naruse ◽  
M. Nakayama ◽  
K. Kurokawa ◽  
T. Sato
Keyword(s):  
Receptor Antagonist ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Physiological Role ◽  
Free Calcium ◽  
Second Phase ◽  
Dependent Manner ◽  
Cortical Collecting Duct ◽  
Acetoxymethyl Ester ◽  
Dose Dependent

The physiological role of oxytocin (OT) in the kidney is still unclear, although autoradiographic data have shown the existence of OT receptors in the rat kidney. We examined the effect of OT in the microperfused rabbit cortical collecting duct (CCD) by using conventional cable analysis and microscope photometry. On addition of 10(-9) M OT to the bath, the lumen-negative transepithelial voltage (VT) transiently increased and the transepithelial resistance (RT) and the fractional resistance of the apical membrane (FRA) (1st phase) both decreased. After this initial change, the lumen-negative VT gradually decreased below its baseline level and RT and FRA (second phase) both increased. These electrical changes were dose dependent and were prevented by the addition of 10(-5) M amiloride to the lumen. Although responses to OT were not prevented by 10(-9) M arginine vasopressin (AVP) or 10(-6) M of a V1-receptor antagonist (OPC-21268) or V2-receptor antagonist (OPC-31260), they were inhibited by the addition of the specific OT antagonist des-Gly-NH2-[d(CH2)3,Tyr(Me),Thr]OVT. Additional studies of intracellular free calcium ([Ca2+]i) revealed that 10(-8)-10(-6) M OT caused an increase in [Ca2+]i in CCD in a dose-dependent manner. Also, pretreatment with 2 x 10(-8) M bis-(aminophenoxy)ethane-tetraacetic acid-acetoxymethyl ester, an intracellular Ca2+ chelator, abolished the electrical and [Ca2+]i responses to OT. Pretreatment with 5 x 10(-4) M 8-(4-chlorophenylthio)-adenosine 3',5'-cyclic monophosphate (CPT-cAMP) partially prevented the electrical responses to OT, thus reducing the decrease in lumen-negative VT below its basal level and the increase in RT after the 1st phase. These data show that OT affects the apical Na+ conductance of collecting duct cells through OT receptors distinct from the AVP receptors and that the effect of OT may, at least in part, be brought about by a mechanism(s) dependent on the increase in [Ca2+]i and cAMP production.

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