Properties of WT and mutant hERG K⁺ channels expressed in neonatal mouse cardiomyocytes

Abstract

Mutations in humanether-a-go-go-related gene 1 (hERG) are linked to long QT syndrome type 2 (LQT2). hERG encodes the pore-forming ␣-subunits that coassemble to form rapidly activating delayed rectifier K ϩ current in the heart. LQT2-linked missense mutations have been extensively studied in noncardiac heterologous expression systems, where biogenic (protein trafficking) and biophysical (gating and permeation) abnormalities have been postulated to underlie the loss-of-function phenotype associated with LQT2 channels. Little is known about the properties of LQT2-linked hERG channel proteins in native cardiomyocyte systems. In this study, we expressed wild-type (WT) hERG and three LQT2-linked mutations in neonatal mouse cardiomyocytes and studied their electrophysiological and biochemical properties. Compared with WT hERG channels, the LQT2 missense mutations G601S and N470D hERG exhibited altered protein trafficking and underwent pharmacological correction, and N470D hERG channels gated at more negative voltages. The ⌬Y475 hERG deletion mutation trafficked similar to WT hERG channels, gated at more negative voltages, and had rapid deactivation kinetics, and these properties were confirmed in both neonatal mouse cardiomyocyte and human embryonic kidney (HEK)-293 cell expression systems. Differences between the cardiomyocytes and HEK-293 cell expression systems were that hERG current densities were reduced 10-fold and deactivation kinetics were accelerated 1.5-to 2-fold in neonatal mouse cardiomyocytes. An important finding of this work is that pharmacological correction of trafficking-deficient LQT2 mutations, as a potential innovative approach to therapy, is possible in native cardiac tissue.

Phenotypic Correlation of Genetic Mutations with Ventricular Arrhythmias

“…Since the rat neonatal cardiomyocytes may only transiently express the relevant channel, another approach is to use adenoviral or lentiviral vectors containing the cDNA of interest to infect the cells and allow adequate expression for study in a more native system. Comparing the behavior of wild type HERG and KCNQ1 channels with previously characterized deleterious mutants in rat neonatal myocytes has confirmed initial phenotypic characterization (Li et al, 2001;Lin et al, 2010). These groups found that the wild-type and mutant channel behaved generally the same as in cultured cells with some slight differences.…”

Mechanistic Basis for Type 2 Long QT Syndrome Caused by KCNH2 Mutations that Disrupt Conserved Arginine Residues in the Voltage Sensor

“…Slower inactivation kinetics is not expected to cause a loss of I Kr function, but faster deactivation kinetics have been associated with the LQT2 mutations R56Q and ΔY475 (Berecki et al 2005; Chen et al 1999; Lin et al 2010; Nakajima et al 1999). We tested the impact that accelerating I Kr deactivation kinetics 10-fold has on the ventricular APD90 using the OVVR computational model of a human AP pulsed at 1 Hz.…”

Cardiac Sodium Channel Nav1.5 Mutations and Cardiac Arrhythmia

“…Recently, neonatal mouse cardiomyocytes have been used as a mammalian expression system to study ion channel mutations found in patients with LQTs [28]. In this study, fresh isolated cardiomyocytes from neonatal mouses were transiently transfected with the cDNA encoding the channel of interest using Amaxa Nucleotranfection kit (Lonza Inc.).…”