Application Note

Channel:Cells:
Tools:hERG
HEK293Port-a-Patch®
External Perfusion

Characterization of hERG (HEK293 cells) onNanion‘s Port-a-Patch®
The electrophysiology team at Nanion Technologies GmbH, Munich.
Cells were supplied by Millipore, USA

Summary

Results

The hERG gene (KCNH2) encodes a potassium ion chan-Figure 1 shows current responses of an individual cell to anel responsible for the repolarizing IKr current in the car-typical hERG voltage protocol. Potentials were stepped

diac action potential (Sanguinetti et al., 1995).

from the holding potential (-80 mV) for 500 ms to +40 mV
followed by a 500 ms step to voltages ranging from

Abnormalities in this channel may lead to either Long QT-120 mV to +40 mV (20 mV steps), before stepping backSyndrome (LQT2) (with loss-of-function mutations) or Shortto holding.

QT syndrome (with gain-of-function mutations), both po-Figure 2 shows the average current-voltage relationshiptentially fatal cardiac arrhythmia, due to repolarization

for recordings equivalent to the one shown in Figure 1. The

disturbances of the cardiac action potential.

averaged mean current at -40 mV of all recorded cells
was 919 ± 57 pA (n=54).

Given the importance of this channel in maintaining cardiac function, it has become an important target in compound safety screening.
A large range of therapeutic agents with diverse chemical structures have been reported to induce long QT syndrome. These include antihistamines (e.g. terfenadine),
gastrointestinal prokinetic agents (e.g. cisapride) andothers.
In this report we present data that were collected on thePort-a-Patch®. Cells (HEK293 stably expressing hERG, supplied by Millipore) were tested. Current amplitudes, IVs
and cisapride as well as quinidine dose-response curves
were analyzed.

Download more Application Notes from www.nanion.de

Figure 1:
Representative current responses of an individual cell to a hERG IV voltage protocol (for details see text above).

Application Note

Figure 4:Shown are raw current responses in the presence of increasing cisapride
concentrations as indicated.

Figure 2:
Average normalized IV (n = 54) for HEK293 cells expressing hERG.

Figure 5:Shown are average dose-response curves for cisapride quinidine.

Figure 3:
Development of the peak current at -40 mV over time after thawing.
Each day between three and five cells were measured.

Since cell lines permanently expressing hERG sometimes
have rather small currents it was tested whether the culturing time after thawing had an effect on the current amplitudes. Results are shown in Figure 3.

With Nanion‘s External Perfusion System also cumulative
does-response curves were collected for quinidine andcisapride. Raw current responses in the presence of different quinidine concentrations are shown in Figure 4. Average dose-response curves for quinidine and cisapride
are shown in Figure 5. From these, IC50s of 267 ± 68 nM and31.5 ± 6.2 nM were determined for quinidine and cisapride, respectively.

Methods

Electrophysiology

Cells

Whole cell patch clamp recordings were conducted ac-HEK293 cells permanently transfected with hERG weresupplied by Millipore.

cording to Nanion’s standard procedure for the Port-aPatch®. Currents were elicited using a voltage step froma holding potential of -80 mV to +40 mV for 500 ms, thenCell culture

to the test potential for 500 ms and back to holding. Puls-Cells were cultured and harvested according to Nan-es were elicited every 20 seconds. External solution ex-ion‘s standard cell culture protocol.

changes were done with Nanion‘s External Perfusion System.

Nanion Technologies GmbHphone +
 49 89 218997972

Erzgiessereistr. 4fax80335 Munich, Germany

www.nanion.de • info@nanion.de+49 89 218997960