Blockade of the human ether-a-go-go-related gene (hERG) potassium channel, with a consequent possibility of QT prolongation and increased susceptibility to a characteristic polymorphic ventricular arrhythmia, torsade de pointes, is an important cause of withdrawal of drugs from the market. In the aftermath of recent drug withdrawals, regulatory agencies now require in vitro hERG screening of all pharmaceutical compounds that are targeted for human use. To minimize the potential for failure in later-stage drug development, many pharmaceutical and biotechnology companies have begun to use automated patch clamp systems with higher throughput than conventional manual patch-clamp techniques to conduct routine functional hERG screening during drug discovery and early development. We have optimized an automated patch-clamp hERG screening method for the PatchXpress 7000A system (Molecular Devices, Sunnyvale, CA) using potassium fluoride (KF) in the internal recording solution. In this study we show that (1) the biophysical and pharmacological properties of hERG current recorded with KF are similar to those with standard potassium chloride solutions, (2) use of KF significantly improves the success rate of hERG screening using PatchXpress without compromising data quality, and (3) utilization of KF can significantly increase the throughput of hERG screening with PatchXpress.
Abnormal diastolic currents in ventricular myocytes from spontaneous hypertensive heart failure rats. Am J Physiol Heart Circ Physiol 291: H2192-H2198, 2006. First published June 9, 2006 doi:10.1152/ajpheart.01146.2005.-Hypertension is a common cause of heart failure, and ventricular arrhythmias are a major cause of death in heart failure. The spontaneous hypertension heart failure (SHHF) rat model was used to study altered ventricular electrophysiology in hypertension and heart failure. We hypothesized that a reduction in the inward rectifier K ϩ current (IK1) and expression of pacemaker current (If) would favor abnormal automaticity in the SHHF ventricle. SHHF ventricular myocytes were isolated at 2 and 8 mo of age and during end-stage heart failure (Ն17 mo); myocytes from age-matched rats served as controls. Inward IK1 was significantly reduced at both 8 and Ն17 mo in SHHF rats compared with controls. There was a reduction in inward IK1 due to aging in the controls only at Ն17 mo. We found a significant increase in If at all ages in the SHHF rats, compared with young controls. In controls, there was an age-dependent increase in If. Action potential recordings in the SHHF rats demonstrated abnormal automaticity, which was abolished by the addition of an If blocker (10 M zatebradine). Increased If during hypertension alone or combined increases in If with reduced IK1 during the progression to hypertensive heart failure contribute to a substrate for arrhythmogenesis. aging; pacemaker current; inward rectifier potassium current; abnormal automaticity HYPERTENSION CAN LEAD TO COMPENSATORY hypertrophy in cardiac muscle. Chronically, these compensatory mechanisms can become maladaptive, and cardiomyopathy and heart failure can result (22). Experimental animal studies in hypertrophy and heart failure have shown abnormalities in ion channel function, altered expression and function of proteins involved in excitation-contraction coupling, and an increased propensity for cardiac arrhythmias (16,30,32). Clinically, ventricular arrhythmias occur in 80% of heart failure patients (11). However, the mechanisms of arrhythmogenic electrophysiological remodeling during hypertension and hypertensive heart failure are not fully elucidated.Spontaneous hypertensive heart failure (SHHF) rats develop hypertension at an early age. In contrast to spontaneously hypertensive rats (SHRs), the SHHF rats consistently develop reproducible, hypertensive heart failure in an age-dependent manner (26). Echocardiographic studies in the SHHF rat demonstrate left ventricular dysfunction (left ventricular ejection fraction Ͻ40%) at 17-18 mo of age (31). There is also evidence of progressive cardiac hypertrophy in the SHHF rats, evidenced as an increase in heart weights (36).Hypertension and heart failure can increase the pacemaker current (I f ) in the ventricular myocardium (4 -7). This can pathologically alter the diastolic phase of the action potential and enhance abnormal automaticity. In addition, aging has been shown to increase I f density i...
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