Skip to main content
SpringerLink
Log in

Mg2+ modulation of the single-channel properties of KCa3.1 in human erythroleukemia cells

  • Ion channels, receptors and transporters
  • Published:
Pflügers Archiv - European Journal of Physiology Aims and scope Submit manuscript

Abstract

The activity of many ion channels is modulated by ions other than the ones they primarily conduct, with important consequences for cell signalling. In this study, we demonstrate that Mg2+ inhibits the intermediate conductance calcium-activated potassium channel (KCa3.1) in human erythroleukemia cells via two distinct mechanisms. Firstly, intracellular Mg2+ blocks this channel via a rapid, voltage-dependent mechanism that leads to a reduction of the channel's unitary current. We show that this block involves interactions which are well described by the Woodhull model. Secondly, we found that Mg2+ reduces the open probability of the channel. By analysing the channel kinetics, we found that this reduction in open probability is at least partly due to a reduction in the rate of channel opening from the closed state, a finding that can be accounted for if Mg2+ competes with Ca2+ for the activation site. Consistent with this interpretation, we find that the decline in relative NPo observed in the presence of 5 mM Mg2+ could be significantly reduced by increasing the free Ca2+ concentration.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5

Similar content being viewed by others

References

  1. Aldrich R, Chandy KG, Grissmer S, Gutman GA, Wei AD, Wulff H (Last modified on 15/10/2009. Accessed on 11/01/2013) Calcium-activated potassium channels, introductory chapter. IUPHAR database (IUPHAR-DB), http://www.iuphar-db.org/DATABASE/FamilyIntroductionForward?familyId=69

  2. Aoki K, Kosakai K, Yoshino M (2008) Monoaminergic modulation of the Na+-activated K+ channel in Kenyon cells isolated from the mushroom body of the cricket (Gryllus bimaculatus) brain. J Neurophysiol 100:1211–1222

    Article  CAS  PubMed  Google Scholar 

  3. Ataga KI, Smith WR, De Castro LM, Swerdlow P, Saunthararajah Y, Castro O, Vichinsky E, Kutlar A, Orringer EP, Rigdon GC, Stocker JW (2008) Efficacy and safety of the Gardos channel blocker, senicapoc (ICA-17043), in patients with sickle-cell anemia. Blood 111:3991–3997

    Article  CAS  PubMed  Google Scholar 

  4. Colquhoun D (1994) Practical analysis of single channel records. In: Ogden DC (ed) Microelectrode techniques. The Plymouth workshop handbook, 2nd edn. The Company of Biologists Ltd, Cambridge, pp 101–139

    Google Scholar 

  5. Cox DH, Cui J, Aldrich RW (1997) Separation of gating properties from permeation and block in mslo large conductance Ca-activated K+ channels. J Gen Physiol 109:633–646

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  6. Diaz F, Wallner M, Stefani E, Toro L, Latorre R (1996) Interaction of internal Ba2+ with a cloned Ca2+-dependent K+ (hslo) channel from smooth muscle. J Gen Physiol 107:399–407

    Article  CAS  PubMed  Google Scholar 

  7. Ferguson WB (1991) Competitive Mg2+ block of a large-conductance, Ca2+-activated K+ channel in rat skeletal muscle. J Gen Physiol 98:163–181

    Article  CAS  PubMed  Google Scholar 

  8. Garcia-Sancho J, Sanchez A, Herreros B (1982) All-or-none response of the Ca2+-dependent K+ channel in inside-out vesicles. Nature 296:744–746

    Article  CAS  PubMed  Google Scholar 

  9. Garfinkel L, Altschuld RA, Garfinkel D (1986) Magnesium in cardiac energy metabolism. J Mol Cell Cardiol 18:1003–1013

    Article  CAS  PubMed  Google Scholar 

  10. Grygorczyk R, Schwarz W (1985) Ca2+-activated K+ permeability in human erythrocytes: modulation of single-channel events. Eur Biophys J 12:57–65

    Article  CAS  PubMed  Google Scholar 

  11. Headrick JP, Willis RJ (1991) Cytosolic free magnesium in stimulated, hypoxic, and underperfused rat heart. J Mol Cardiol 23:991–999

    Article  CAS  Google Scholar 

  12. Hille B (1992) Ionic channels of excitable membranes. Sinauer Associates, Sunderland

    Google Scholar 

  13. Horie M, Irisawa H, Noma A (1987) Voltage-dependent magnesium block of adenosine-triphosphate-sensitive potassium channel in guinea-pig ventricular cells. J Physiol 387:251–272

    CAS  PubMed Central  PubMed  Google Scholar 

  14. Kim SY, Silver MR, DeCoursey TE (1996) I. Ion channels in human THP-1 monocytes. J Membrane Biol 152:117–130

    Article  CAS  Google Scholar 

  15. Ledoux J, Bonev A, Nelson M (2008) Ca2+-activated K+ channels in murine endothelial cells: block by intracellular calcium and magnesium. J Gen Physiol 131:125–135

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  16. Leinders T, van Kleef RDGM, Vijverberg HPM (1992) Distinct metal ion binding sites on Ca2+-activated K+ channels in inside-out patches of human erythrocytes. Biochimica et Biophysica Acta–Biomembranes 1112:75–82

    Article  CAS  Google Scholar 

  17. Lesage F, Guillemare E, Fink M, Duprat F, Lazdunski M, Romey G, Barhanin J (1996) TWIK-1, a ubiquitous human weakly inward rectifying K+ channel with a novel structure. Embo J 15:1004–1011

    CAS  PubMed Central  PubMed  Google Scholar 

  18. Li Y, Gamper N, Shapiro MS (2004) Single-channel analysis of KCNQ K+ channels reveals the mechanism of augmentation by cysteine-modifying reagents. J Neurosci 24:5079–5090

    Article  CAS  PubMed  Google Scholar 

  19. Lu X, Fein A, Feinstein MB, O’Rourke FA (1999) Antisense knock out of the inositol 1,3,4,5-tetrakisphosphate receptor GAP1IP4BP in the human erythroleukemia cell line leads to the appearance of intermediate conductance K(Ca) channels that hyperpolarize the membrane and enhance calcium influx. J Gen Physiol 113:81–95

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  20. Marchenko S, Sage SO (1996) Calcium-activated potassium channels in the endothelium of intact rat aorta. J Physiol 492:53–60

    CAS  PubMed Central  PubMed  Google Scholar 

  21. Martin P, Papayannopoulou T (1982) HEL cells: a new human erythroleukemia cell line with spontaneous and induced globin expression. Science 216:1233–1235

    Article  CAS  PubMed  Google Scholar 

  22. Mortensen M, Smart TG (2007) Single-channel recording of ligand-gated ion channels. Nat Protoc 2:2826–2841

    Article  CAS  PubMed  Google Scholar 

  23. Murphy E, Steenbergen C, Levy LA, Raju B, London RE (1989) Cytosolic free magnesium levels in ischemic heart. J Biol Chem 264:5622–5627

    CAS  PubMed  Google Scholar 

  24. Neher E (1992) Correction for liquid junction potentials in patch clamp experiments. Methods in Enzymol 207:123–131

    Article  CAS  Google Scholar 

  25. Neyton J (1996) A Ba2+ chelator suppresses long shut events in fully activated high-conductance Ca2+-dependent K+ channels. Biophys J 71:220–226

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  26. Romani A (2007) Regulation of magnesium homeostasis and transport in mammalian cells. Arch Biochem Biophys 458:90–102

    Article  CAS  PubMed  Google Scholar 

  27. Romani AMP, Scarpa A (2000) Regulation of cellular magnesium. Front Biosci 5:720–734

    Article  Google Scholar 

  28. Selyanko AA, Hadley JK, Brown DA (2001) Properties of single M-type KCNQ2/KCNQ3 potassium channels expressed in mammalian cells. J Physiol 534:15–24

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  29. Simons TJB (1976) Calcium-dependent potassium exchange in human red cell ghosts. J Physiol 256:227–244

    CAS  PubMed Central  PubMed  Google Scholar 

  30. Stoneking CJ, Shivakumar O, Nicholson Thomas D, Colledge WH, Mason MJ (2013) Voltage dependence of the Ca2+-activated K+ channel KCa3.1 in human erythroleukemia cells. Am J Physiol 304:C858–C872

    Article  CAS  Google Scholar 

  31. Toyama K, Wulff H, Chandy KG, Azam PG, Saito T, Fujiwara Y, Mattson DL, Das S, Melvin JE, Pratt PF, Hatoum OA, Gutterman DD, Harder DR, Miura H (2008) The intermediate-conductance calcium-activated potassium channel KCa3.1 contributes to atherogenesis in mice and humans. J Clin Invest 118:3025–3037

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  32. Vandenberg CA (1987) Inward rectification of a potassium channel in cardiac ventricular cells depends on internal magnesium ions. Proc Natl Acad Sci USA 84:2560–2564

    Article  CAS  PubMed Central  PubMed  Google Scholar 

  33. Wei AD, Gutman GA, Aldrich R, Chandy KG, Grissmer S, Wulff H (2005) International Union of Pharmacology. LII. Nomenclature and molecular relationships of calcium-activated potassium channels. Pharmacol Rev 57:463–472

    Article  CAS  PubMed  Google Scholar 

  34. Woodhull AM (1973) Ionic blockage of sodium channels in nerve. J Gen Physiol 61:687–708

    Article  CAS  PubMed Central  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Physiology, Development and Neuroscience, University of Cambridge, Downing Street, Cambridge, CB2 3EG, UK

    Colin J. Stoneking & Michael J. Mason

Authors
  1. Colin J. Stoneking
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Michael J. Mason
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Michael J. Mason.

Electronic supplementary material

Below is the link to the electronic supplementary material.

Supplemental Fig. 1

Effect of physiological levels of free Ca2+ on the single-channel I–V relationship in the absence of Mg2+. Excised single-channel patches were exposed to 0.5 or 2 μM free Ca2+ in the absence of Mg2+ and the I–V relationship determined. * significantly different from 2 μM Ca2+ (p < 0.05 as determined by two-way ANOVA, n = 5 for 0.5 μM Ca2+ and n = 6 for 2 μM Ca2+) (PS 506 kb)

Supplemental Fig. 2

Effect of the high affinity Ba2+ chelator (+)-(18-Crown-6)-2,3,11,12-tetracarboxyclic acid (18C6TCA). Excised single-channel patches were exposed to 2 μM free Ca2+ in the presence of 5 mM free Mg2+ and the I–V relationship determined in the absence or presence of 100 μM 18C6TCA. For control and 18C6TCA, n = 7 for –3, +37 and +57 mV and n = 6 for +17 mV. n = 3 and 5 for control and 100 μM 18C6TCA, respectively, at +77 mV. A two-way ANOVA was performed using this dataset with no significant difference detected (p > 0.05). An additional repeated measures two-way ANOVA using only data acquired from patches in which single-channel currents at a given voltage were obtained in both control and during exposure to 18C6TCA was also performed. Data was obtained in seven patches in the presence and absence of 18C6TCA at –3, +37 and +57 mV, six patches at +17 mV and three patches at +77. Again, no significant difference in single-channel currents were detected between control and 18C6TCA exposure (p > 0.05) (PS 490 kb)

Rights and permissions

Reprints and permissions

About this article

Cite this article

Stoneking, C.J., Mason, M.J. Mg2+ modulation of the single-channel properties of KCa3.1 in human erythroleukemia cells. Pflugers Arch - Eur J Physiol 466, 1529–1539 (2014). https://doi.org/10.1007/s00424-013-1375-0

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00424-013-1375-0

Keywords

Search

Navigation