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2020 - Automated Dynamic Clamp for Simulation of IK1 in Human Induced Pluripotent Stem Cell–Derived Cardiomyocytes in Real Time Using Patchliner Dynamite8

icon pl   Patchliner and Dynamite8 publication in Current Protocols in Pharmacology (2020)

Authors:
Becker N., Horváth A., De Boer T., Fabbri A., Grad C., Fertig N., George M., Obergrussberger A.

Journal:
Current Protocols in Pharmacology (2020) Volume 88, e70


Abstract:

Current in vitro assays typically poorly predict cardiac liability as they focus on single ion channels overexpressed in cell lines. Human induced pluripotent stem cell–derived cardiomyocytes (hiPSC‐CMs), on the other hand, provide a unique opportunity for drug testing on human cardiomyocytes using high‐throughput systems. However, these cells can differ from adult cardiomyocytes in their ion channel expression and, therefore, electrophysiologic properties. One of the main challenges of hiPSC‐CMs is the physiologic expression of ion channels such as the inward rectifiers (e.g., Kir2.1–2.3), which conduct the cardiac inward rectifier potassium current (IK1). IK1 is one of the primary contributors in maintaining a stable resting membrane potential in cardiac cells, which is essential for excitability. This is only expressed in low levels, or sometimes not at all, in hiPSC‐CMs as shown by patch clamp studies. Dynamic clamp is a method of electronically introducing ion currents (e.g., IK1) into cells to compensate for the lack of endogenous expression, thus offering the potential to record more stable action potentials in hiPSC‐CMs. In this article, we describe the method of using hiPSC‐CMs on an automated patch clamp device (Patchliner) coupled with the automated dynamic clamp add‐on (Dynamite8). We describe protocols for optimized cell handling and harvesting for use on the Patchliner and the steps required for automated execution of experiments and data analysis in dynamic clamp mode. © 2019 by John Wiley & Sons, Inc.
 
Basic Protocol: Recording action potential pharmacology from human induced pluripotent stem cell–derived cardiomyocytes in automated patch clamp combined with dynamic clamp to introduce simulated IK1 and compensate seal resistance
 
Support Protocol 1: Cardiomyocyte plating and culture
 
Support Protocol 2: Cell harvesting and dissociation
 
Alternate Protocol: Recording action potential pharmacology at physiologic temperatures

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