Skip to main content
SpringerLink
Log in

Automated Patch Clamp Recordings of Human Stem Cell-Derived Cardiomyocytes

  • Protocol
  • First Online:
Stem Cell-Derived Models in Toxicology

Part of the book series: Methods in Pharmacology and Toxicology ((MIPT))

Abstract

Patch clamp remains the gold standard for studying ion channel activity within cell membranes. Conventional patch clamp is notoriously low throughput and technically demanding making it an unsuitable technique for high-throughput screening (HTS). Automated patch clamp (APC) devices have done much to increase throughput and improve ease of use, particularly when using standard cell line cells such as HEK and CHO. In recent years, however, the use of human-induced pluripotent stem cells (hiPSCs) has become increasingly important, especially for safety screening in response to the Comprehensive In Vitro Proarrhythmia Assay (CiPA) initiative introduced in 2013. The goal of this initiative is to standardize assays, targets, and cell types. One part of the paradigm focuses on the use of APC and hiPSC cardiomyocytes. This chapter describes two automated patch clamp devices recording from up to 8 or 384 cells simultaneously using hiPSC cardiomyocytes. In the voltage clamp mode, voltage-gated Na+ (NaV), Ca2+ (CaV), and K+ (KV) channels could be recorded, and pharmacology using tetracaine, a NaV channel blocker, is described. Additionally, action potentials in the current clamp mode were recorded, and examples are shown including the effect of nifedipine, a CaV channel blocker. Detailed methods are provided for cell culture and harvesting of hiPSCs for use on APC devices. Protocols are also provided for voltage and current clamp recordings on the Patchliner, and voltage clamp experiments on the SyncroPatch 384PE APC instruments.

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

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 139.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

References

  1. Neher E, Sakmann B (1976) Single-channel currents recorded from membrane of denervated frog muscle fibres. Nature 260:799–802. doi:10.1038/260799a0

    Article  CAS  PubMed  Google Scholar 

  2. Dunlop J, Bowlby M, Peri R et al (2008) High-throughput electrophysiology: an emerging paradigm for ion-channel screening and physiology. Nat Rev Drug Discov 7:358–368. doi:10.1038/nrd2552

    Article  CAS  PubMed  Google Scholar 

  3. Farre C, Haythornthwaite A, Haarmann C et al (2009) Port-a-patch and patchliner: high fidelity electrophysiology for secondary screening and safety pharmacology. Comb Chem High Throughput Screen 12:24–37. doi:10.2174/138620709787047966

    Article  CAS  PubMed  Google Scholar 

  4. Brüggemann A, George M, Klau M et al (2003) The NPC © Technology. Assay Drug Dev Technol 1:665–673

    Article  PubMed  Google Scholar 

  5. Brueggemann A, George M, Klau M et al (2004) Ion channel drug discovery and research: the automated Nano-Patch-Clamp © technology. Curr Drug Discov Technol 1:91–96

    Article  CAS  PubMed  Google Scholar 

  6. Brüggemann A, Stoelzle S, George M et al (2006) Microchip technology for automated and parallel patch-clamp recording. Small 2:840–846. doi:10.1002/smll.200600083

    Article  PubMed  Google Scholar 

  7. Jones KA, Garbati N, Zhang H, Large CH (2009) Automated patch clamping using the QPatch. In: Janzen WP, Bernasconi P (eds) High throughput screening: methods and protocols, 2nd edn. Humana Press, a part of Springer Science & Business Media, Totowa, pp 209–223

    Chapter  Google Scholar 

  8. Mathes C, Friis S, Finley M, Liu Y (2009) QPatch: the missing link between HTS and ion channel drug discovery. Comb Chem High Throughput Screen 12:78–95. doi:10.2174/138620709787047948

    Article  CAS  PubMed  Google Scholar 

  9. Tao H, Santa Ana D, Guia A et al (2004) Automated tight seal electrophysiology for assessing the potential hERG liability of pharmaceutical compounds. Assay Drug Dev Technol 2:497–506. doi:10.1089/adt.2004.2.497

    Article  CAS  PubMed  Google Scholar 

  10. Xu J, Guia A, Rothwarf D et al (2003) A benchmark study with SealChipTM planar patch-clamp technology. Assay Drug Dev Technol 1:675–684. doi:10.1089/154065803770381039

    Article  CAS  PubMed  Google Scholar 

  11. Stoelzle S, Obergrussberger A, Brüggemann A et al (2011) State-of-the-art automated patch clamp devices: heat activation, action potentials, and high throughput in ion channel screening. Front Pharmacol 2:1–11. doi:10.3389/fphar.2011.00076

    Article  Google Scholar 

  12. Obergrussberger A, Haarmann C, Rinke I et al (2014) Automated patch clamp analysis of nAChα7 and NaV 1.7 channels. Curr Protoc Pharmacol 65:11.13.1–11.13.48. doi:10.1002/0471141755.ph1113s65

    Article  Google Scholar 

  13. Schroeder K, Neagle B, Trezise DJ, Worley J (2003) IonWorks HT: a new high-throughput electrophysiology measurement platform. J Biomol Screen 8:50–64. doi:10.1177/1087057102239667

    Article  CAS  PubMed  Google Scholar 

  14. Finkel A, Wittel A, Yang N et al (2006) Population patch clamp improves data consistency and success rates in the measurement of ionic currents. J Biomol Screen 11:488–496. doi:10.1177/1087057106288050

    Article  CAS  PubMed  Google Scholar 

  15. Gillie DJ, Novick SJ, Donovan BT et al (2013) Development of a high-throughput electrophysiological assay for the human ether-à-go-go related potassium channel hERG. J Pharmacol Toxicol Methods 67:33–44. doi:10.1016/j.vascn.2012.10.002

    Article  CAS  PubMed  Google Scholar 

  16. Kuryshev YA, Brown AM, Duzic E, Kirsch GE (2014) Evaluating state dependence and subtype selectivity of calcium channel modulators in automated electrophysiology assays. Assay Drug Dev Technol 12:110–119. doi:10.1089/adt.2013.552

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  17. Scheel O, Himmel H, Rascher-Eggstein G, Knott T (2011) Introduction of a modular automated voltage-clamp platform and its correlation with manual human Ether-à-go-go related gene voltage-clamp data. Assay Drug Dev Technol 9:600–607. doi:10.1089/adt.2010.0352

    Article  CAS  PubMed  Google Scholar 

  18. Spencer CI, Li N, Chen Q et al (2012) Ion channel pharmacology under flow: automation via well-plate microfluidics. Assay Drug Dev Technol 10:313–324. doi:10.1089/adt.2011.414

    Article  CAS  PubMed  Google Scholar 

  19. Obergrussberger A, Stölzle-Feix S, Becker N et al (2015) Novel screening techniques for ion channel targeting drugs. Channels 9:367–375. doi:10.1080/19336950.2015.1079675

    Article  PubMed  PubMed Central  Google Scholar 

  20. Obergrussberger A, Brüggemann A, Goetze TA et al (2015) Automated patch clamp meets high-throughput screening: 384 cells recorded in parallel on a planar patch clamp module. J Lab Autom. doi:10.1177/2211068215623209

    PubMed  Google Scholar 

  21. Sager PT, Gintant G, Turner JR et al (2014) Rechanneling the cardiac proarrhythmia safety paradigm: a meeting report from the Cardiac Safety Research Consortium. Am Heart J 167:292–300. doi:10.1016/j.ahj.2013.11.004

    Article  PubMed  Google Scholar 

  22. Fermini B, Hancox JC, Abi-Gerges N et al (2015) A new perspective in the field of cardiac safety testing through the comprehensive in vitro proarrhythmia assay paradigm. J Biomol Screen. doi:10.1177/1087057115594589

    PubMed  Google Scholar 

  23. Cavero I, Holzgrefe H (2014) Comprehensive in vitro proarrhythmia assay, a novel in vitro/in silico paradigm to detect ventricular proarrhythmic liability: a visionary 21st century initiative. Expert Opin Drug Saf 13:745–758. doi:10.1517/14740338.2014.915311

    Article  CAS  PubMed  Google Scholar 

  24. Becker N, Stoelzle S, Göpel S et al (2013) Minimized cell usage for stem cell-derived and primary cells on an automated patch clamp system. J Pharmacol Toxicol Methods 68:82–87. doi:10.1016/j.vascn.2013.03.009

    Article  CAS  PubMed  Google Scholar 

  25. Rajamohan D, Kalra S, Hoang MD et al (2016) Automated electrophysiological and pharmacological evaluation of human pluripotent stem cell-derived cardiomyocytes. Stem Cells Dev 25. doi:10.1089/scd.2015.0253

  26. Haythornthwaite A, Stoelzle S, Hasler A et al (2012) Characterizing human ion channels in induced pluripotent stem cell-derived neurons. J Biomol Screen 17:1264–1272. doi:10.1177/1087057112457821

    Article  PubMed  Google Scholar 

  27. Meijer van Putten RME, Mengarelli I, Guan K et al (2015) Ion channelopathies in human induced pluripotent stem cell derived cardiomyocytes: a dynamic clamp study with virtual IK1. Front Physiol 6:Article 7. doi:10.3389/fphys.2015.00007

  28. Shen JB, Jiang B, Pappano AJ (2000) Comparison of L-type calcium channel blockade by nifedipine and/or cadmium in guinea pig ventricular myocytes. J Pharmacol Exp Ther 294:562–570

    CAS  PubMed  Google Scholar 

  29. Klugbauer N, Lacinova L, Flockerzi V, Hofmann F (1995) Structure and functional expression of a new member of the tetrodotoxin-sensitive voltage-activated sodium channel family from human neuroendocrine cells. EMBO J 14:1084–1090

    CAS  PubMed  PubMed Central  Google Scholar 

  30. Kass RS, Krafte DS (1987) Negative surface charge density near heart calcium channels. Relevance to block by dihydropyridines. J Gen Physiol 89:629–644. doi:10.1085/jgp.89.4.629

    Article  CAS  PubMed  Google Scholar 

  31. Charnet P, Bourinet E, Dubel SJ et al (1994) Calcium currents recorded from a neuronal alpha 1C L-type calcium channel in Xenopus oocytes. FEBS Lett 344:87–90

    Article  CAS  PubMed  Google Scholar 

  32. Sheets MF, Hanck DA (1999) Gating of skeletal and cardiac muscle sodium channels in mammalian cells. J Physiol 514(Pt 2):425–436

    Google Scholar 

Download references

Acknowledgments

We thank Cellular Dynamics International (CDI), Madison, Wisconsin, for the collaboration and for providing us with cardiomyocytes (iCell cardiomyocytes). We also thank Axiogenesis AG, Cologne, Germany, for the collaboration and for providing us with cardiomyocytes (Cor.4U). We also thank Pluriomics for providing us with the Pluricytes.

The authors disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: The work presented here was funded in part by the Bundesministerium fuer Bildung und Forschung (BMBF, grant 01QE1502).

Author information

Authors and Affiliations

  1. Nanion Technologies GmbH, Gabrielenstraße 9, 80636, Munich, Germany

    Alison Obergrussberger, Claudia Haarmann, Sonja Stölzle-Feix, Nadine Becker, Andrea Brüggemann, Michael George & Niels Fertig

  2. Nanion Technologies GmbH, Tokyo Laboratory, Medical Innovation Laboratory, Tokyo Women’s Medical University and Waseda University Joint Institution for Advanced Biomedical Sciences, Shinjyuku-ku, Tokyo, Japan

    Atsushi Ohtsuki

Authors
  1. Alison Obergrussberger
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Claudia Haarmann
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Sonja Stölzle-Feix
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Nadine Becker
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Atsushi Ohtsuki
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Andrea Brüggemann
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Michael George
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Niels Fertig
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Alison Obergrussberger .

Editor information

Editors and Affiliations

  1. Axion BioSystems, Inc., Atlanta, Georgia, USA

    Mike Clements

  2. Cell Applications, GE Healthcare, Cardiff, United Kingdom

    Liz Roquemore

Rights and permissions

Reprints and permissions

Copyright information

© 2017 Springer Science+Business Media New York

About this protocol

Cite this protocol

Obergrussberger, A. et al. (2017). Automated Patch Clamp Recordings of Human Stem Cell-Derived Cardiomyocytes. In: Clements, M., Roquemore, L. (eds) Stem Cell-Derived Models in Toxicology. Methods in Pharmacology and Toxicology. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-6661-5_4

Download citation

  • DOI: https://doi.org/10.1007/978-1-4939-6661-5_4

  • Published:

  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-6659-2

  • Online ISBN: 978-1-4939-6661-5

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation