Combining electrophysiology and contractility recordings for more
complete assessment of hiPSC-CMs
Alison Obergrussberger, Sonja Stoelzle-Feix, Elena Dragicevic, Nadine Becker, Krisztina Juhasz, Ulrich Thomas, LeoDoerr, Markus Rapedius, Claudia Haarmann, Ilka Rinke, Tom Goetze, Matthias Beckler, Michael George, Andrea
Brüggemann and Niels Fertig
Nanion Technologies, Ganghoferstr. 70A, Munich, Germany. Email: info@nanion.deCardiac ion channels in hiPSC-CMs

Introduction
Human induced pluripotent stem cells (hiPSCs) are becoming increasingly
important for cardiac safety testing due to their recapitulation of native behaviour,relative abundance and ease-of-use. We combine automated patch clamp(APC), impedance and extracellular field potential (EFP) measurements to study
hiPSC-derived cardiomyocytes (hiPSC-CMs) from different sources. In line with thecomprehensive in vitro proarrhythmia assay (CiPA) initiative, cardiac ion channels
expressed in heterologous expression systems have been recorded on APC
devices at room temperature and at physiological temperature and the effect ofdifferent pro-arrhythmic compounds on ion channels was investigated. In addition
hiPSC-CMs were used on APC platforms recording from 8 or 384 wells
simultaneously. Voltage-gated Na+ (NaV), Ca2+ (CaV) and K+ (KV) currents wererecorded in voltage-clamp and action potentials recorded in current clamp
mode. Using the dynamic clamp technique coupled with an APC device,electrically modelled IK1 was injected into the cell under current clamp conditions
and this resulted in a more hyperpolarized and stable resting membrane potential.
Within the myocyte phase II study of the CiPA initiative, a device combining
impedance-based contractility and extracellular field potential (EFP) recordings
was used to investigate the effects of different compounds deemed low,intermediate and high risk by the FDA. Different hiPSC-CMs were used in a large
cross-site evaluation and the results are summarised here.

Effect of vandetanib on hERG and NaV1.5:
comparison with hiPSC-CMs
Automated patch
clamp: Patchliner

ramp
100ms-80mV

500ms
3

Negative Control

0-2nA

0.03 µM

2nA

0.3 µM
1Impedance (Ω)

NaV1.540mV-43 µM-60.20.4sCpd0.6Vandetanib

0.250

0.8Platform

Patchliner

hERG
IC50 [µM]Cav1.2
IC50 [µM]

0.23

AB

A Action potentials from Cellartis Cardiomyocytes recorded on the Patchliner and the effect ofsimulated IK1 injected using dynamic clamp. B Action potentials elicited using a 1 ms depolarizing
pulse and 400 pS/pF IK1 injected, the calcium channel agonist BayK8644 prolonged AP durationand the antagonist, nifedipine, shortened AP duration. From: Goversen et al, 2018.

3.210.252
sNav1.5
IC50 [µM]9.90.254

Cross-site comparison of CiPA compounds
using the CardioExcyte 96

0.256

Kv4.3
IC50 [µM]20.3

Impedance (Ω)

0.248

Vandetanib blocks hERG and NaV1.5 expressed in HEK cells in electrophysiology (Patchliner)
experiments (left) and causes arrhythmia in hiPSC-CMs in impedance and EFP recordings(CardioExcyte 96; right). The red arrows indicate arrhythmic events.

Effect of sotalol on hERG: comparison with
hiPSC-CMs
Automated patch
clamp (35°C):
SyncroPatch 384PE

Impedance/EFP:
CardioExcyte 96

Trend = illustrates trend of average FPD data with min and max values pointed out
FPD>10%†††

FPD>30%

FPD> - 10% FPD > -30%

type A arhythmia
any other arrhythmia type

Test article concentrations (µM)

CompoundHIGH RISK

Site / CelltypeTrend

ETPCFree(uM)

Dose 1

Dose 2

type

Dose 3

type

Dose 4

type

100

††
††
††
††
††
††
††
††
††

D,l Sotalol

Site 1, CDI

150.1

1-

10

D,l Sotalol

Site 2, CDI

150.1

1-

10D,l Sotalol

Site 3, CDI

150.1

1-

10D,l Sotalol

Site 1, AXG

150.1

1-10††††††-

D,l Sotalol
D,l Sotalol

Site 2, AXG
Site 4, AXG

15150.1
0.1

11-

1010-

100
100

D,l Sotalol

Site 2, CLS

150.11-

10-

100D,l Sotalol

Site 4, CLS

150.11-

10-

100

D,l Sotalol
Site 2, PLR
INTERMEDIATE RISK:

150.11-10††100Domperidone

Site 2, CDI

0.02

0.003

0.03-

0.3

3

Domperidone

Site 2, AXG

0.02

0.003

0.03-

0.3

Domperidone

Site 2, CLS

0.02

0.003

0.03-

0.3

Domperidone
LOW RISK
Ranolazine

Site 2, PLR

0.020.003

0.03-0.3††††††††3††††††††Site 2, CDI

1.950.11-

10-

100

quiescence

Ranolazine
Ranolazine

Site 3, CDI
Site 2, AXG

1.95
1.95

0.1
0.1

11-

1010-

††-

100
100Ranolazine

Site 2, CLS

1.950.1

1-

10-

100

Ranolazine

Site 2, PLR

1.95

0.11-

10-

100

100
100
100

33quiescence††††

Excerpt of cross-site study using the CardioExcyte 96 with example of high, intermediateand low risk compounds on field potential duration (FPD) in EFP mide hiPSC-CMs. From: Botet al., 2018.

ControlVoltage (µV)

RefSolImpedance (Ω)

35℃
10µM

10µM

300µM
RefSol

RefSolHigh Control

10 µM Sotalol

Fullblock~17min

Cpd

Platform

Highestconc.
[µM]

Sotalol

SyncroPatch
384PE

300

Impedance (Ω)

300µM

Voltage (µV)

0.0

Action potential duration using dynamic clamp on the Patchliner

1 µM Vandetanib
Voltage (µV)

16.8 µM

0iCell® Cardiomyocytes recorded on the Patchliner. (A) Current-voltage relationship of NaV and (B)CaV. Vhalf of activation for NaIV was -29 mV and for CaIV was 6.8 mV. (C) Concentration-responsecurve for tetracaine revealed an IC50 of 2.9 ± 1.4 µM (n = 5). (D) Effect of nifedipine on the action
potential. (E) Screenshot of the SyncroPatch 384PE software during a recording of Pluricyte®
Cardiomyocytes. From: Obergrussberger et al, 2016.

Impedance/EFP:
CardioExcyte 96

Voltage (µV)

hERG

EhERG
IC50 [µM]Cav1.2
IC50 [µM]

Nav1.5
IC50 [µM]

Kv4.3
IC50 [µM]

Kir2.1
IC50 [µM]

KvLQT1KCNE1IC50 [µM]80.2>300>300>300>300>300

Sotalol blocks hERG expressed in HEK cells with an IC50 of 80.2 µM measured at 35°C on theSyncroPatch 384PE (left). Using impedance and EFP measurements of hiPSC-CMs on theCardioExcyte 96 (right), arrhythmic events (red arrows) are detected at low concentrations (10µM).
Cells for this project kindly provided by Ncardia, Cellular Dynamics International and Takara Bio.

Conclusions• Cardiac ion channels expressed in cell lines have been measured on thePatchliner and SyncroPatch 384PE as part of the CiPA Ion Channel WorkStream.
• hiPSC-CMs have been used on the Patchliner and SyncroPatch 384PE,
voltage-gated ion channels have been recorded and pharmacology
performed. Action potentials were recorded in current clamp mode anddynamic clamp combined with the Patchliner.
• hiPSC-derived cardiomyocytes were used on the CardioExcyte 96 as acomplementary tool for safety screening. The effect compounds on ion
channels is often complex and the overall effect in cardiomyocytes on
beating parameters is an important and complementary assay to patch
clamp in heterologous expression systems.

References
Obergrussberger, A, et al., 2016. In: Stem cell-derived models in toxicity. Chapter 4. pp. 57-82
Goversen, B, et al., 2018. Front. Physiol. 8:1094
Bot, C, et al., 2018. J. Pharmacol. Toxicol. Methods. doi: 10.1016/j.vascn.2018.06.006