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. Bentzen
Ion channels, receptors and transporters

Abstract

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.

Keywords

Ion channel TASK K2P3.1 Atrial fibrillation Pharmacology Electrophysiology 

Notes

Acknowledgments

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 S1Effects 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 S2Time 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 S3Effect 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 S1KCNK3 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 S2Patient demographics showing relevant medical data and medication (TIFF 473 kb)

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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
  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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