Whole‐Cell Configuration of the Patch‐Clamp Technique in the hERG Channel Assay to Predict the Ability of a Compound to Prolong QT Interval

2012 ◽  
Vol 57 (1) ◽  
Author(s):  
Sonia Goineau ◽  
Christophe Legrand ◽  
Guillaume Froget
Keyword(s):  
Patch Clamp ◽  
Qt Interval ◽  
Herg Channel ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Clamp Technique ◽  
Cell Configuration

Whole cell sodium conductance of principal cells freshly isolated from rat cortical collecting duct

1995 ◽  
Vol 269 (3) ◽  
pp. C791-C796 ◽  
Author(s):  
J. K. Bubien
Keyword(s):  
Patch Clamp ◽  
Collecting Duct ◽  
Cortical Collecting Duct ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Cyclic Monophosphate ◽  
Principal Cells ◽  
Clamp Technique ◽  
Duct Cells ◽  
Collecting Duct Cells

Cortical collecting duct fragments were manually dissected from 6-wk-old Sprague-Dawley rats. The fragments were enzymatically digested (collagenase A) into single cells, washed, and resuspended in serum-free RPMI 1640. Individual cells were examined electrophysiologically using the whole cell patch-clamp technique. Two morphologically distinct cell types were present in the cell suspension. Small round cells that had a capacitance of 7 pF and larger oval cells with a capacitance of 29 pF were consistently observed. Whole cell electrophysiological examination revealed that the small round cells had virtually no plasma membrane ionic conductance, whereas both inward and outward currents were observed in the larger oval-type cells. Also, superfusion of 250 pM arginine vasopressin specifically increased the inward conductance of only the larger cells. The effect could be completely inhibited by 2 microM amiloride or 100 mumol of the Rp diastereomer of 8-(4-chlorophenylthio)-adenosine 3',5'-cyclic monophosphate (a specific adenosine 3',5'-cyclic monophosphate inhibitor). These findings are consistent with the hypothesis that the larger cells are principal cells and the smaller cells are intercalated cells and directly demonstrate that an amiloride-sensitive whole cell conductance is readily observable in freshly isolated cortical collecting duct cells. Thus the whole cell configuration of the patch-clamp technique appears to be well suited for assessing cellular mechanisms that regulate the ionic conductances of cortical collecting duct cells.

scholarly journals Baro-excited neurons in the caudal ventrolateral medulla (CVLM) recorded using the whole-cell patch-clamp technique

2011 ◽  
Vol 35 (5) ◽  
pp. 500-506 ◽  
Author(s):  
Naoki Oshima ◽  
Hiroo Kumagai ◽  
Kamon Iigaya ◽  
Hiroshi Onimaru ◽  
Akira Kawai ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Ventrolateral Medulla ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Caudal Ventrolateral Medulla ◽  
Clamp Technique ◽  
Cell Patch Clamp ◽  
Whole Cell Patch Clamp

Membrane properties of mesopontine cholinergic neurons studied with the whole-cell patch-clamp technique: implications for behavioral state control

1992 ◽  
Vol 68 (4) ◽  
pp. 1359-1372 ◽  
Author(s):  
A. Kamondi ◽  
J. A. Williams ◽  
B. Hutcheon ◽  
P. B. Reiner
Keyword(s):  
Patch Clamp ◽  
Cholinergic Neurons ◽  
Membrane Properties ◽  
Type I ◽  
Type Ii ◽  
Patch Clamp Technique ◽  
Whole Cell ◽  
Type Iii ◽  
Clamp Technique ◽  
Whole Cell Patch Clamp

1. The whole-cell patch-clamp technique was used to study the membrane properties of identified cholinergic and noncholinergic laterodorsal tegmental neurons in slices of rat brain maintained in vitro. 2. On the basis of their expression of the transient outward potassium current IA and the transient inward calcium current IT, three classes of neurons were observed: type I neurons exhibited a large IT; type II neurons exhibited a prominent IA; and type III neurons exhibited both IA and IT. 3. Combining intracellular deposition of biocytin with NADPH diaphorase histochemistry revealed that the vast majority of type III neurons were cholinergic, whereas only a minority of type I and type II neurons were cholinergic. Thus mesopontine cholinergic neurons possess intrinsic ionic currents capable of inducing burst firing. 4. Delineation of the intrinsic membrane properties of identified mesopontine cholinergic neurons, in concert with recent results regarding the responses of these neurons to neurotransmitter agents, has led us to present a unifying and mechanistic hypothesis of brain stem cholinergic function in the control of behavioral states.

Rb+ Flux through hERG Channels Affects the Potency of Channel Blocking Drugs: Correlation with Data Obtained Using a High-Throughput Rb+ Efflux Assay

2004 ◽  
Vol 9 (7) ◽  
pp. 588-597 ◽  
Author(s):  
Saman Rezazadeh ◽  
J. Christian Hesketh ◽  
David Fedida
Keyword(s):  
Patch Clamp ◽  
High Throughput ◽  
Screening Method ◽  
Herg Channel ◽  
Patch Clamp Technique ◽  
Clamp Technique ◽  
Channel Inactivation ◽  
Channel Blocking ◽  
Low Sensitivity ◽  
Herg Channels

The nonradioactive Rb+ efflux assay has become a reliable and efficient high-throughput hERG screening method, but it is limited by its low sensitivity for potent hERG blockers. Using the patch clamp technique, the authors found that the low sensitivity is due in part to the use of Rb+ as the permeating cation in the assay. The affinities of the drugs measured by patch clamp technique in the presence of Rb+ were 3- to 10-fold lower than when measured by the same method in the presence of K+ ions. The apparent affinity of the drugs decreased even further when monitored bytheRb+ efflux assay. It was also observed that Rb+ had minimal effects on the activation properties of channels while there was a significant change in the half-inactivation potential. This voltage shift reduces hERG channel inactivation at efflux assay potentials, and will reduce the affinity of hERG-blocking drugs that bind to inactivated states of the channel. In combination with the effects of elevated extracellular ion concentrations, it is likely that Rb+ modulation of hERG channel inactivation is largely responsible for the reduced drug potencies observed in the Rb+ efflux assay.

scholarly journals Ginsenoside Rg3 Alleviates Antithyroid Cancer Drug Vandetanib-Induced QT Interval Prolongation

2021 ◽  
Vol 2021 ◽  
pp. 1-14
Author(s):  
Juan Zhang ◽  
Dan Luo ◽  
Fang Li ◽  
Zhiyi Li ◽  
Xiaoli Gao ◽  
...  
Keyword(s):  
Patch Clamp ◽  
Action Potential ◽  
Cardiac Arrhythmia ◽  
Qt Interval ◽  
K Channel ◽  
Ginsenoside Rg3 ◽  
Interval Prolongation ◽  
Patch Clamp Technique ◽  
Clamp Technique ◽  
Qt Interval Prolongation

Inhibition of human ether-a-go-go-related gene (hERG) potassium channel is responsible for acquired long QT syndromes, which leads to life-threatening cardiac arrhythmia. A multikinase inhibitor, vandetanib, prolongs the progression-free survival time in advanced medullary thyroid cancer. However, vandetanib has been reported to induce significant QT interval prolongation, which limits its clinical application. Some studies have showed that ginsenoside Rg3 decelerated hERG K(+) channel tail current deactivation. Therefore, in this study, we aim to confirm whether ginsenoside Rg3 targeting hERG K(+) channel could be used to reverse the vandetanib-induced QT interval prolongation. Electrocardiogram (ECG) and monophasic action potential (MAP) were recorded using electrophysiology signal sampling and analysis system in Langendorff-perfused rabbit hearts. The current clamp mode of the patch-clamp technique was used to record transmembrane action potential. The whole-cell patch-clamp technique was used to record the hERG K+ current. In Langendorff-perfused hearts, vandetanib prolonged the QT interval in a concentration-dependent manner with an IC50 of 1.96 μmol/L. In human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), vandetanib significantly prolonged the action potential duration at 50%, 70%, and 90% repolarization (APD50, APD70, and APD90). In stable transfected human hERG gene HEK293 cells, vandetanib caused concentrate-dependent inhibition in the step and tail currents of hERG. As expected, ginsenoside Rg3 relieved vandetanib-induced QT interval prolongation in Langendorff-perfused heart and reversed vandetanib-induced APD prolongation in hiPSC-CMs. Furthermore, ginsenoside Rg3 alleviated vandetanib-induced hERG current inhibition and accelerated the process of the channel activation. Ginsenoside Rg3 may be a promising cardioprotective agent against vandetanib-induced QT interval prolongation through targeting hERG channel. These novel findings highlight the therapeutic potential of ginsenoside to prevent vandetanib-induced cardiac arrhythmia.

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