pH-dependent inhibition of K2P3.1 prolongs atrial refractoriness in whole hearts

  • Mark A. Skarsfeldt
  • Thomas A. Jepps
  • Sofia H. Bomholtz
  • Lea Abildgaard
  • Ulrik S. Sørensen
  • Emilie Gregers
  • Jesper H. Svendsen
  • Jonas G. Diness
  • Morten Grunnet
  • Nicole Schmitt
  • Søren-Peter Olesen
  • Bo H. BentzenEmail author
Ion channels, receptors and transporters


In isolated human atrial cardiomyocytes, inhibition of K2P3.1 K+ channels results in action potential (action potential duration (APD)) prolongation. It has therefore been postulated that K2P3.1 (KCNK3), together with K2P9.1 (KCNK9), could represent novel drug targets for the treatment of atrial fibrillation (AF). However, it is unknown whether these findings in isolated cells translate to the whole heart. The purposes of this study were to investigate the expression levels of KCNK3 and KCNK9 in human hearts and two relevant rodent models and determine the antiarrhythmic potential of K2P3.1 inhibition in isolated whole-heart preparations. By quantitative PCR, we found that KCNK3 is predominantly expressed in human atria whereas KCNK9 was not detectable in heart human tissue. No differences were found between patients in AF or sinus rhythm. The expression in guinea pig heart resembled humans whereas rats displayed a more uniform expression of KCNK3 between atria and ventricle. In voltage-clamp experiments, ML365 and A293 were found to be potent and selective inhibitors of K2P3.1, but at pH 7.4, they failed to prolong atrial APD and refractory period (effective refractory period (ERP)) in isolated perfused rat and guinea pig hearts. At pH 7.8, which augments K2P3.1 currents, pharmacological channel inhibition produced a significant prolongation of atrial ERP (11.6 %, p = 0.004) without prolonging ventricular APD but did not display a significant antiarrhythmic effect in our guinea pig AF model (3/8 hearts converted on A293 vs 0/7 hearts in time-matched controls). These results suggest that when K2P3.1 current is augmented, K2P3.1 inhibition leads to atrial-specific prolongation of ERP; however, this ERP prolongation did not translate into significant antiarrhythmic effects in our AF model.


Ion channel TASK K2P3.1 Atrial fibrillation Pharmacology Electrophysiology 



We thank Amer Mujezinovic (Dept. of Biomedical Sciences, University of Copenhagen) for technical assistance and Drs Susanne Holme, Jens Juel Thiis and Jens Lund (University Hospital Copenhagen, Rigshospitalet) for their help in biopsy extraction. We thank the University of Kansas Specialized Chemistry Center (Grant number U54HG005031) for synthesizing ML365. This study was supported by the Innovation Fund Denmark and The Danish National Research Foundation Centre for Cardiac Arrhythmia.

Compliance with ethical standards

Conflict of interest

No conflict of interest.

Supplementary material

424_2015_1779_MOESM1_ESM.eps (6.6 mb)
Supplementary Figure S1 Effects of K2P3.1 inhibitors on atrial K+ channels. Representative current traces before (black) and after (red) application of 3 μM ML365 (left) and A293 (right). Kir2.1 (IK1), Kir3.1/Kir3.4 (IKAch), KV7.1/KCNE1 (IKs), KV1.5 (IKur) and KV4.3/KChIP2 (Ito) were recorded using two-electrode voltage-clamp on Xenopus laevis oocytes, whereas KCa2.3 (IKCa) and KV11.1 (IKr) were recorded using automated patch clamping. Currents were elicited by the voltage protocol shown (EPS 6755 kb)
424_2015_1779_MOESM2_ESM.eps (360 kb)
Supplementary Figure S2 Time matched controls and effects of pH on guinea pig hearts. Guinea pig atrial ERP (a), atrial (b) and ventricular APD (c) measured (before black circles and after white circles) application of DMSO, pH 7.8, n = 6. Comparison of atrial ERP (d), atrial APD (e) an ventricular APD (f) of values in guinea pig hearts perfused at pH 7.4 (black circles) and pH 7.8 (open circles), n = 7. Atrial ERP: Unpaired t-test, p = 0.0568 (EPS 359 kb)
424_2015_1779_MOESM3_ESM.eps (116 kb)
Supplementary Figure S3 Effect of K2P3.1 inhibition on aERP in the presence of acetylcholine. Guinea pig atrial ERP measured at pH 7.8, baseline (black square boxes) followed by application of 1 μM ACh (white square boxes) for 15 minutes and finally 3 μM A293 + 1 μM ACh for 30 minutes (white circles). Paired t-test, **p = 0.0021, n = 5. (EPS 115 kb)
424_2015_1779_MOESM4_ESM.tiff (474 kb)
Supplementary Table S1 KCNK3 and KCNK9 primer sequences used for qPCR analysis in rat, guinea pig and human heart tissue (TIFF 473 kb)
424_2015_1779_MOESM5_ESM.tiff (474 kb)
Supplementary Table S2 Patient demographics showing relevant medical data and medication (TIFF 473 kb)


  1. 1.
    Aller MI, Veale EL, Linden A-M, Sandu C, Schwaninger M, Evans LJ, Korpi ER, Mathie A, Wisden W, Brickley SG (2005) Modifying the subunit composition of TASK channels alters the modulation of a leak conductance in cerebellar granule neurons. J Neurosci Off J Soc Neurosci 25:11455–11467. doi: 10.1523/JNEUROSCI.3153-05.2005 CrossRefGoogle Scholar
  2. 2.
    Allessie M, Ausma J, Schotten U (2002) Electrical, contractile and structural remodeling during atrial fibrillation. Cardiovasc Res 54:230–246. doi: 10.1016/S0008-6363(02)00258-4 CrossRefPubMedGoogle Scholar
  3. 3.
    Barth AS, Merk S, Arnoldi E, Zwermann L, Kloos P, Gebauer M, Steinmeyer K, Bleich M, Kääb S, Hinterseer M, Kartmann H, Kreuzer E, Dugas M, Steinbeck G, Nabauer M (2005) Reprogramming of the human atrial transcriptome in permanent atrial fibrillation: expression of a ventricular-like genomic signature. Circ Res 96:1022–1029. doi: 10.1161/01.RES.0000165480.82737.33 CrossRefPubMedGoogle Scholar
  4. 4.
    Chapman CG, Meadows HJ, Godden RJ, Campbell DA, Duckworth M, Kelsell RE, Murdock PR, Randall AD, Rennie GI, Gloger IS (2000) Cloning, localisation and functional expression of a novel human, cerebellum specific, two pore domain potassium channel. Mol Brain Res 82:74–83. doi: 10.1016/S0169-328X(00)00183-2 CrossRefPubMedGoogle Scholar
  5. 5.
    Czirják G, Enyedi P (2002) Formation of functional heterodimers between the TASK-1 and TASK-3 two-pore domain potassium channel subunits. J Biol Chem 277:5426–5432. doi: 10.1074/jbc.M107138200 CrossRefPubMedGoogle Scholar
  6. 6.
    Decher N, Kiper AK, Rolfes C, Schulze-Bahr E, Rinné S (2015) The role of acid-sensitive two-pore domain potassium channels in cardiac electrophysiology: focus on arrhythmias. Pflüg Arch Eur J Physiol 467:1055–1067. doi: 10.1007/s00424-014-1637-5 CrossRefGoogle Scholar
  7. 7.
    Decher N, Wemhöner K, Rinné S, Netter MF, Zuzarte M, Aller MI, Kaufmann SG, Li XT, Meuth SG, Daut J, Sachse FB, Maier SKG (2011) Knock-out of the potassium channel TASK-1 leads to a prolonged QT interval and a disturbed QRS complex. Cell Physiol Biochem Int J Exp Cell Physiol Biochem Pharmacol 28:77–86. doi: 10.1159/000331715 CrossRefGoogle Scholar
  8. 8.
    Diness JG, Sørensen US, Nissen JD, Al-Shahib B, Jespersen T, Grunnet M, Hansen RS (2010) Inhibition of small-conductance Ca2+-activated K+ channels terminates and protects against atrial fibrillation. Circ Arrhythm Electrophysiol 3:380–390. doi: 10.1161/CIRCEP.110.957407 CrossRefPubMedGoogle Scholar
  9. 9.
    Donner BC, Schullenberg M, Geduldig N, Hüning A, Mersmann J, Zacharowski K, Kovacevic A, Decking U, Aller MI, Schmidt KG (2011) Functional role of TASK-1 in the heart: studies in TASK-1-deficient mice show prolonged cardiac repolarization and reduced heart rate variability. Basic Res Cardiol 106:75–87. doi: 10.1007/s00395-010-0128-x CrossRefPubMedGoogle Scholar
  10. 10.
    Duprat F, Lesage F, Fink M, Reyes R, Heurteaux C, Lazdunski M (1997) TASK, a human background K+ channel to sense external pH variations near physiological pH. EMBO J 16:5464–5471. doi: 10.1093/emboj/16.17.5464 CrossRefPubMedPubMedCentralGoogle Scholar
  11. 11.
    Ellinghaus P, Scheubel RJ, Dobrev D, Ravens U, Holtz J, Huetter J, Nielsch U, Morawietz H (2005) Comparing the global mRNA expression profile of human atrial and ventricular myocardium with high-density oligonucleotide arrays. J Thorac Cardiovasc Surg 129:1383–1390. doi: 10.1016/j.jtcvs.2004.08.031 CrossRefPubMedGoogle Scholar
  12. 12.
    Flaherty DP, Simpson DS, Miller M, Maki BE, Zou B, Shi J, Wu M, McManus OB, Aubé J, Li M, Golden JE (2014) Potent and selective inhibitors of the TASK-1 potassium channel through chemical optimization of a bis-amide scaffold. Bioorg Med Chem Lett 24:3968–3973. doi: 10.1016/j.bmcl.2014.06.032 CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Franz MR (1991) Method and theory of monophasic action potential recording. Prog Cardiovasc Dis 33:347–368CrossRefPubMedGoogle Scholar
  14. 14.
    Franz MR (1994) Bridging the gap between basic and clinical electrophysiology: what can be learned from monophasic action potential recordings? J Cardiovasc Electrophysiol 5:699–710CrossRefPubMedGoogle Scholar
  15. 15.
    Franz MR (1999) Current status of monophasic action potential recording: theories, measurements and interpretations. Cardiovasc Res 41:25–40. doi: 10.1016/S0008-6363(98)00268-5 CrossRefPubMedGoogle Scholar
  16. 16.
    Gaborit N, Le Bouter S, Szuts V, Varro A, Escande D, Nattel S, Demolombe S (2007) Regional and tissue specific transcript signatures of ion channel genes in the non-diseased human heart: regional ion channel subunit gene expression in the human heart. J Physiol 582:675–693. doi: 10.1113/jphysiol.2006.126714 CrossRefPubMedPubMedCentralGoogle Scholar
  17. 17.
    Gaborit N, Steenman M, Lamirault G, Le Meur N, Le Bouter S, Lande G, Léger J, Charpentier F, Christ T, Dobrev D, Escande D, Nattel S, Demolombe S (2005) Human atrial ion channel and transporter subunit gene-expression remodeling associated with valvular heart disease and atrial fibrillation. Circulation 112:471–481. doi: 10.1161/CIRCULATIONAHA.104.506857 CrossRefPubMedGoogle Scholar
  18. 18.
    Hondeghem LM, Snyders DJ (1990) Class III antiarrhythmic agents have a lot of potential but a long way to go. Reduced effectiveness and dangers of reverse use dependence. Circulation 81:686–690. doi: 10.1161/01.CIR.81.2.686 CrossRefPubMedGoogle Scholar
  19. 19.
    Kim Y, Bang H, Kim D (1999) TBAK-1 and TASK-1, two-pore K(+) channel subunits: kinetic properties and expression in rat heart. Am J Physiol 277:H1669–H1678PubMedGoogle Scholar
  20. 20.
    Kiper AK, Rinné S, Rolfes C, Ramírez D, Seebohm G, Netter MF, González W, Decher N (2015) Kv1.5 blockers preferentially inhibit TASK-1 channels: TASK-1 as a target against atrial fibrillation and obstructive sleep apnea? Pflüg Arch Eur J Physiol 467:1081–1090. doi: 10.1007/s00424-014-1665-1 CrossRefGoogle Scholar
  21. 21.
    Kirchhoff JE, Goldin Diness J, Sheykhzade M, Grunnet M, Jespersen T (2015) Synergistic antiarrhythmic effect of combining inhibition of Ca2+-activated K+ (SK) channels and voltage-gated Na+ channels in an isolated heart model of atrial fibrillation. Heart Rhythm 12:409–418. doi: 10.1016/j.hrthm.2014.12.010 CrossRefPubMedGoogle Scholar
  22. 22.
    Kneller J, Zou R, Vigmond EJ, Wang Z, Leon LJ, Nattel S (2002) Cholinergic atrial fibrillation in a computer model of a two-dimensional sheet of canine atrial cells with realistic ionic properties. Circ Res 90:E73–E87CrossRefPubMedGoogle Scholar
  23. 23.
    Kovacs RJ, Bailey JC (1985) Effects of acetylcholine on action potential characteristics of atrial and ventricular myocardium after bilateral cervical vagotomy in the cat. Circ Res 56:613–620CrossRefPubMedGoogle Scholar
  24. 24.
    Limberg SH, Netter MF, Rolfes C, Rinné S, Schlichthörl G, Zuzarte M, Vassiliou T, Moosdorf R, Wulf H, Daut J, Sachse FB, Decher N (2011) TASK-1 channels may modulate action potential duration of human atrial cardiomyocytes. Cell Physiol Biochem Int J Exp Cell Physiol Biochem Pharmacol 28:613–624. doi: 10.1159/000335757 CrossRefGoogle Scholar
  25. 25.
    Medhurst AD, Rennie G, Chapman CG, Meadows H, Duckworth MD, Kelsell RE, Gloger II, Pangalos MN (2001) Distribution analysis of human two pore domain potassium channels in tissues of the central nervous system and periphery. Mol Brain Res 86:101–114. doi: 10.1016/S0169-328X(00)00263-1 CrossRefPubMedGoogle Scholar
  26. 26.
    Nattel S, Harada M (2014) Atrial remodeling and atrial fibrillation recent advances and translational perspectives. J Am Coll Cardiol 63:2335–2345. doi: 10.1016/j.jacc.2014.02.555 CrossRefPubMedGoogle Scholar
  27. 27.
    Putzke C, Wemhöner K, Sachse FB, Rinné S, Schlichthörl G, Li XT, Jaé L, Eckhardt I, Wischmeyer E, Wulf H, Preisig-Müller R, Daut J, Decher N (2007) The acid-sensitive potassium channel TASK-1 in rat cardiac muscle. Cardiovasc Res 75:59–68. doi: 10.1016/j.cardiores.2007.02.025 CrossRefPubMedGoogle Scholar
  28. 28.
    Rensma PL, Allessie MA, Lammers WJ, Bonke FI, Schalij MJ (1988) Length of excitation wave and susceptibility to reentrant atrial arrhythmias in normal conscious dogs. Circ Res 62:395–410CrossRefPubMedGoogle Scholar
  29. 29.
    Rinné S, Kiper AK, Schlichthörl G, Dittmann S, Netter MF, Limberg SH, Silbernagel N, Zuzarte M, Moosdorf R, Wulf H, Schulze-Bahr E, Rolfes C, Decher N (2015) TASK-1 and TASK-3 may form heterodimers in human atrial cardiomyocytes. J Mol Cell Cardiol 81:71–80. doi: 10.1016/j.yjmcc.2015.01.017 CrossRefPubMedGoogle Scholar
  30. 30.
    Schmidt C, Wiedmann F, Langer C, Tristram F, Anand P, Wenzel W, Lugenbiel P, Schweizer PA, Katus HA, Thomas D (2014) Cloning, functional characterization, and remodeling of K2P3.1 (TASK-1) potassium channels in a porcine model of atrial fibrillation and heart failure. Heart Rhythm Off J Heart Rhythm Soc 11:1798–1805. doi: 10.1016/j.hrthm.2014.06.020 CrossRefGoogle Scholar
  31. 31.
    Schmidt C, Wiedmann F, Voigt N, Zhou X-B, Heijman J, Lang S, Albert V, Kallenberger S, Ruhparwar A, Szabó G, Kallenbach K, Karck M, Borggrefe M, Biliczki P, Ehrlich JR, Baczkó I, Lugenbiel P, Schweizer PA, Donner BC, Katus HA, Dobrev D, Thomas D (2015) Upregulation of K2P3.1 K+ current causes action potential shortening in patients with chronic atrial fibrillation. Circulation. doi: 10.1161/CIRCULATIONAHA.114.012657 PubMedCentralGoogle Scholar
  32. 32.
    Schmitt N, Grunnet M, Olesen S-P (2014) Cardiac potassium channel subtypes: new roles in repolarization and arrhythmia. Physiol Rev 94:609–653. doi: 10.1152/physrev.00022.2013 CrossRefPubMedGoogle Scholar
  33. 33.
    Vandesompele J, De Preter K, Pattyn F, Poppe B, Van Roy N, De Paepe A, Speleman F (2002) Accurate normalization of real-time quantitative RT-PCR data by geometric averaging of multiple internal control genes. Genome Biol 3(7):research0034.1–research0034.11Google Scholar
  34. 34.
    Varró A, Lathrop DA, Hester SB, Nánási PP, Papp JG (1993) Ionic currents and action potentials in rabbit, rat, and guinea pig ventricular myocytes. Basic Res Cardiol 88:93–102PubMedGoogle Scholar
  35. 35.
    Zhabyeyev P, Asai T, Missan S, McDonald TF (2004) Transient outward current carried by inwardly rectifying K+ channels in guinea pig ventricular myocytes dialyzed with low-K+ solution. Am J Physiol Cell Physiol 287:C1396–C1403. doi: 10.1152/ajpcell.00479.2003 CrossRefPubMedGoogle Scholar
  36. 36.
    Zou B, Flaherty DP, Simpson DS, Maki BE, Miller MR, Shi J, Wu M, McManus OB, Golden JE, Aubé J, Li M (2010) ML365: development of bis-amides as selective inhibitors of the kcnk3/task1 two pore potassium channel. Probe Rep. NIH Mol. Libr. ProgramGoogle Scholar

Copyright information

© Springer-Verlag Berlin Heidelberg 2015

Authors and Affiliations

  • Mark A. Skarsfeldt
    • 1
  • Thomas A. Jepps
    • 1
  • Sofia H. Bomholtz
    • 1
    • 2
  • Lea Abildgaard
    • 2
  • Ulrik S. Sørensen
    • 2
  • Emilie Gregers
    • 3
    • 4
  • Jesper H. Svendsen
    • 3
  • Jonas G. Diness
    • 2
  • Morten Grunnet
    • 2
  • Nicole Schmitt
    • 1
  • Søren-Peter Olesen
    • 1
  • Bo H. Bentzen
    • 1
    Email author
  1. 1.Department of Biomedical Sciences, Faculty of Health and Medical SciencesUniversity of CopenhagenCopenhagenDenmark
  2. 2.Acesion PharmaCopenhagenDenmark
  3. 3.Laboratory of Molecular Cardiology, Department of Cardiology, The Heart Centre, RigshospitaletUniversity of CopenhagenCopenhagenDenmark
  4. 4.Department of Medicine and Surgery, Faculty of Health and Mediacl SciencesUniversity of CopenhagenCopenhagenDenmark

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