Swelling-activated potassium currents of Ehrlich ascites tumour cells

Riquelme, Gloria; Sepúlveda, Francisco V.; Jørgensen, F.; Pedersen, Susanne; Hoffmann, Else K.

doi:10.1016/s0005-2736(98)00006-6

Cited by 33 publications

(52 citation statements)

References 14 publications

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“…Figure 4B shows the quantification of pixel intensities in the phalloidin-labeled cells, under isotonic and hypotonic conditions, in control cells and after cholesterol depletion and enrichment, respectively. As shown, in control cells both cytoplasmic (i.e., predominantly stress-fiber associated) and cortical F-actin levels were reduced by ϳ50% after 2 min of hypotonic swelling, consistent with previous findings in EAT cells (53). Under isotonic conditions, cholesterol depletion tended to increase cytoplasmic F-actin, although not quite significantly (P ϭ 0.11), whereas cholesterol enrichment significantly decreased the cortical, but not the cytoplasmic, F-actin level.…”

Section: Characterization Of Swelling-activated CLsupporting

confidence: 92%

“…Instead, as discussed in Ref. 53, this deviation from the theoretical value is likely to reflect a dilution of the intracellular solution, arising because the osmotic water inflow into Ehrlich cells is faster than the solution exchange via the patch pipette. From the Nernst equation, it may be calculated that the discrepancy between the calculated and experimental V rev corresponds to a 38% dilution of the intracellular solution in the 190 mosmol/l bath solution.…”

Section: Characterization Of Swelling-activated CLmentioning

confidence: 95%

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Cholesterol modulates the volume-regulated anion current in Ehrlich-Lettre ascites cells via effects on Rho and F-actin

Klausen

Hougaard²,

Hoffmann³

et al. 2006

American Journal of Physiology-Cell Physiology

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The mechanisms controlling the volume-regulated anion current (VRAC) are incompletely elucidated. Here, we investigate the modulation of VRAC by cellular cholesterol and the potential involvement of F-actin, Rho, Rho kinase, and phosphatidylinositol-(4,5)-bisphosphate [PtdIns(4,5)P2] in this process. In Ehrlich-Lettre ascites (ELA) cells, a current with biophysical and pharmacological properties characteristic of VRAC was activated by hypotonic swelling. A 44% increase in cellular cholesterol content had no detectable effects on F-actin organization or VRAC activity. A 47% reduction in cellular cholesterol content increased cortical and stress fiber-associated F-actin content in swollen cells. Cholesterol depletion increased VRAC activation rate and maximal current after a modest (15%), but not after a severe (36%) reduction in extracellular osmolarity. The cholesterol depletion-induced increase in maximal VRAC current was prevented by F-actin disruption using latrunculin B (LB), while the current activation rate was unaffected by LB, but dependent on Rho kinase. Rho activity was decreased by ∼20% in modestly, and ∼50% in severely swollen cells. In modestly swollen cells, this reduction was prevented by cholesterol depletion, which also increased isotonic Rho activity. Thrombin, which stimulates Rho and causes actin polymerization, potentiated VRAC in modestly swollen cells. VRAC activity was unaffected by inclusion of a water-soluble PtdIns(4,5)P2 analogue or a PtdIns(4,5)P2-blocking antibody in the pipette, or neomycin treatment to sequester PtdIns(4,5)P2. It is suggested that in ELA cells, F-actin and Rho-Rho kinase modulate VRAC magnitude and activation rate, respectively, and that cholesterol depletion potentiates VRAC at least in part by preventing the hypotonicity-induced decrease in Rho activity and eliciting actin polymerization.

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Section: Characterization Of Swelling-activated CLsupporting

confidence: 92%

Section: Characterization Of Swelling-activated CLmentioning

confidence: 95%

Cholesterol modulates the volume-regulated anion current in Ehrlich-Lettre ascites cells via effects on Rho and F-actin

Klausen

Hougaard²,

Hoffmann³

et al. 2006

American Journal of Physiology-Cell Physiology

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“…We now show that CFTR is needed for an effective cell volume regulation in human airways exposed to hypotonic conditions. Regulatory volume decrease in both murine (14) and human tracheal cells (this study) depends on the coordinated activity of Cl -dependent IK channels (43). The fact that the ⌬F508-CF cells (CFT1 and CFT1-LC3), unlike the rescued CFT1-LCFSN (CFTR) cells, showed IK activity but no swelling-induced activation pointed to a defect in the regulation of the channel protein, rather than alterations in its expression.…”

Section: Discussionmentioning

confidence: 64%

Defective regulatory volume decrease in human cystic fibrosis tracheal cells because of altered regulation of intermediate conductance Ca ²⁺ -dependent potassium channels

Vázquez

Nobles

Valverde

2001

Proc. Natl. Acad. Sci. U.S.A.

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The cystic fibrosis transmembrane conductance regulator (CFTR) protein has the ability to function as both a chloride channel and a channel regulator. The loss of these functions explains many of the manifestations of the cystic fibrosis disease (CF), including lung and pancreatic failure, meconium ileus, and male infertility. CFTR has previously been implicated in the cell regulatory volume decrease (RVD) response after hypotonic shocks in murine small intestine crypts, an effect associated to the dysfunction of an unknown swelling-activated potassium conductance. In the present study, we investigated the RVD response in human tracheal CF epithelium and the nature of the volume-sensitive potassium channel affected. Neither the human tracheal cell line CFT1, expressing the mutant CFTR-⌬F508 gene, nor the isogenic vector control line CFT1-LC3, engineered to express the ␤gal gene, showed RVD. On the other hand, the cell line CFT1-LCFSN, engineered to express the wild-type CFTR gene, presented a full RVD. Patch-clamp studies of swelling-activated potassium currents in the three cell lines revealed that all of them possess a potassium current with the biophysical and pharmacological fingerprints of the intermediate conductance Ca 2؉ -dependent potassium channel (IK, also known as KCNN4). However, only CFT1-LCFSN cells showed an increase in IK currents in response to hypotonic challenges. Although the identification of the molecular mechanism relating CFTR to the hIK channel remains to be solved, these data offer new evidence on the complex integration of CFTR in the cells where it is expressed. CFTR ͉ ⌬F508 ͉ chloride secretion ͉ airways ͉ hIKK

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“…Various K ϩ channels have been shown to be implicated in RVD depending on cell type; i.e., 1) Ca 2ϩ -activated small conductance K ϩ (SK) channels (24,42), intermediate conductance K ϩ (IK) channels (3,7,24,25,46), and Big conductance K ϩ (BK) channels (10,23); 2) stretch-activated K ϩ channels; 3) voltage-dependent channels; 4) the MinK channel; and 5) the two-pore domain K ϩ channel (4,(31)(32)(33)(34)(35)37). The volume-sensitive ion channels in EATC are the volume-regulated anion channel (VRAC) (38) and the TWIK-related acid-sensitive K ϩ channel-2 (TASK-2) (22,33,41), which belongs to the family of two-pore domain channels (K 2p ). TASK-2 is sensitive to clofilium (35), independent of intracellular Ca 2ϩ (35,41), and highly sensitive to external pH (21).…”

mentioning

confidence: 99%

“…The volume-sensitive ion channels in EATC are the volume-regulated anion channel (VRAC) (38) and the TWIK-related acid-sensitive K ϩ channel-2 (TASK-2) (22,33,41), which belongs to the family of two-pore domain channels (K 2p ). TASK-2 is sensitive to clofilium (35), independent of intracellular Ca 2ϩ (35,41), and highly sensitive to external pH (21). Besides being the swelling-activated K ϩ channel in EATC, TASK-2 has also been found to be involved in volume regulation in kidney cells (4) and in murine spermatozoa (1).…”

mentioning

confidence: 99%

Activation of the TASK-2 channel after cell swelling is dependent on tyrosine phosphorylation

Kirkegaard

Lambert

Gammeltoft

et al. 2010

American Journal of Physiology-Cell Physiology

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Kirkegaard SS, Lambert IH, Gammeltoft S, Hoffmann EK. Activation of the TASK-2 channel after cell swelling is dependent on tyrosine phosphorylation. Am J Physiol Cell Physiol 299: C844 -C853, 2010. First published July 14, 2010; doi:10.1152/ajpcell.00024.2010.-The swelling-activated K ϩ currents (I K,vol ) in Ehrlich ascites tumor cells (EATC) has been reported to be through the two-pore domain (K2p), TWIK-related acid-sensitive K ϩ channel 2 (TASK-2). The regulatory volume decrease (RVD), following hypotonic exposure in EATC, is rate limited by IK,vol indicating that inhibition of RVD reflects inhibition of TASK-2. We find that in EATC the tyrosine kinase inhibitor genistein inhibits RVD by 90%, and that the tyrosine phosphatase inhibitor monoperoxo(picolinato)-oxo-vanadate(V) [mpV(pic)] shifted the volume set point for inactivation of the channel to a lower cell volume. Swelling-activated K ϩ efflux was impaired by genistein and the Src kinase family inhibitor 4-amino-5-(4-chloro-phenyl)-7-(t-butyl) pyrazolo [3,4-d]pyrimidine (PP2) and enhanced by the tyrosine phosphatase inhibitor mpV(pic). With the use of the TASK-2 inhibitor clofilium, it is demonstrated that mpV(pic) increased the volumesensitive part of the K ϩ efflux 1.3 times. To exclude K ϩ efflux via a KCl cotransporter, cellular Cl Ϫ was substituted with NO 3 Ϫ . Also under these conditions K ϩ efflux was completely blocked by genistein. Thus tyrosine kinases seem to be involved in the activation of the volumesensitive K ϩ channel, whereas tyrosine phosphatases appears to be involved in inactivation of the channel. Overexpressing TASK-2 in human embryonic kidney (HEK)-293 cells increased the RVD rate and reduced the volume set point. TASK-2 has tyrosine sites, and precipitation of TASK-2 together with Western blotting and antibodies against phosphotyrosines revealed a cell swelling-induced, timedependent tyrosine phosphorylation of the channel. Even though we found an inhibiting effect of PP2 on RVD, neither Src nor the focal adhesion kinase (FAK) seem to be involved. Inhibitors of the epidermal growth factor receptor tyrosine kinases had no effect on RVD, whereas the Janus kinase (JAK) inhibitor cucurbitacin inhibited the RVD by 40%. It is suggested that the cytokine receptor-coupled JAK/STAT pathway is upstream of the swelling-induced phosphorylation and activation of TASK-2 in EATC.cell volume regulation; Janus kinase; volume-sensitive channels

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Swelling-activated potassium currents of Ehrlich ascites tumour cells

Cited by 33 publications

References 14 publications

Cholesterol modulates the volume-regulated anion current in Ehrlich-Lettre ascites cells via effects on Rho and F-actin

Cholesterol modulates the volume-regulated anion current in Ehrlich-Lettre ascites cells via effects on Rho and F-actin

Defective regulatory volume decrease in human cystic fibrosis tracheal cells because of altered regulation of intermediate conductance Ca ²⁺ -dependent potassium channels

Activation of the TASK-2 channel after cell swelling is dependent on tyrosine phosphorylation

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Swelling-activated potassium currents of Ehrlich ascites tumour cells

Cited by 33 publications

References 14 publications

Cholesterol modulates the volume-regulated anion current in Ehrlich-Lettre ascites cells via effects on Rho and F-actin

Cholesterol modulates the volume-regulated anion current in Ehrlich-Lettre ascites cells via effects on Rho and F-actin

Defective regulatory volume decrease in human cystic fibrosis tracheal cells because of altered regulation of intermediate conductance Ca 2+ -dependent potassium channels

Activation of the TASK-2 channel after cell swelling is dependent on tyrosine phosphorylation

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Defective regulatory volume decrease in human cystic fibrosis tracheal cells because of altered regulation of intermediate conductance Ca ²⁺ -dependent potassium channels