Home About us Contact
|
Home About us Contact | ||
|
|
||
Isolated Cardiac Myocytes (isolated + cardiac_myocyte)
Selected AbstractsComparative Pharmacology of Guinea Pig Cardiac Myocyte and Cloned hERG (IKr) ChannelJOURNAL OF CARDIOVASCULAR ELECTROPHYSIOLOGY, Issue 11 2004CHRISTINA DAVIE Ph.D. Introduction: This study used whole-cell, patch clamp techniques on isolated guinea pig ventricular myocytes and HEK293 cells expressing cloned human ether-a-go-go-related gene (hERG) to examine the action of drugs causing QT interval prolongation and torsades de pointes (TdP) in man. Similarities and important differences in drug actions on cardiac myocytes and cloned hERG IKr channels were established. Qualitative actions of the drugs on cardiac myocytes corresponded with results obtained from Purkinje fibers and measurement of QT interval prolongation in animal and human telemetry studies. Methods and Results: Adult guinea pig ventricular myocytes were isolated by enzymatic digestion. Cells were continuously perfused with Tyrode's solution at 33,35°C. Recordings were made using the whole-cell, patch clamp technique. Action potentials (APs) were elicited under current clamp. Voltage clamp was used to study the effect of drugs on IKr (rapidly activating delayed rectifier potassium current), INa (sodium current), and ICa (L-type calcium current). Dofetilide increased the myocyte action potential duration (APD) in a concentration-dependent manner, with a pIC50 of 7.3. Dofetilide 1 ,M elicited early afterdepolarizations (EADs) but had little affect on ICa or INa. E-4031 increased APD in a concentration-dependent manner, with a pIC50 of 7.2. In contrast, 10 ,M loratadine, desloratadine, and cetirizine had little effect on APD or IKr. Interestingly, cisapride displayed a biphasic effect on myocyte APD and inhibited ICa at 1 ,M. Even at this high concentration, cisapride did not elicit EADs. A number of AstraZeneca compounds were tested on cardiac myocytes, revealing a mixture of drug actions that were not observed in hERG currents in HEK293 cells. One compound, particularly AR-C0X, was a potent blocker of myocyte AP (pIC50 of 8.4). AR-C0X also elicited EADs in cardiac myocytes. The potencies of the same set of drugs on the cloned hERG channel also were assessed. The pIC50 values for dofetilide, E-4031, terfenadine, loratadine, desloratadine, and cetirizine were 6.8, 7.1, 7.3, 5.1, 5.2, and <4, respectively. Elevation of temperature from 22 to 35°C significantly enhanced the current kinetics and amplitudes of hERG currents and resulted in approximately fivefold increase in E-4031 potency. Conclusion: Our study demonstrates the advantages of cardiac myocytes over heterologously expressed hERG channels in predicting QT interval prolongation and TdP in man. The potencies of some drugs in cardiac myocytes were similar to hERG, but only myocytes were able to detect important changes in APD characteristics and display EADs predictive of arrhythmia development. We observed similar qualitative drug profiles in cardiac myocytes, dog Purkinje fibers, and animal and human telemetry studies. Therefore, isolated native cardiac myocytes are a better predictor of drug-induced QT prolongation and TdP than heterologously expressed hERG channels. Isolated cardiac myocytes, when used with high-throughput patch clamp instruments, may have an important role in screening potential cardiotoxic compounds in the early phase of drug discovery. This would significantly reduce the attrition rate of drugs entering preclinical and/or clinical development. The current kinetics and amplitudes of the cloned hERG channel were profoundly affected by temperature, significantly altering the potency of one drug (E-4031). This finding cautions against routine drug testing at room temperature compared to physiologic temperature when using the cloned hERG channel. [source] Effects of labedipinedilol-A, third-generation dihydropyridine-type calcium blocker, on ouabain-induced arrhythmiaDRUG DEVELOPMENT RESEARCH, Issue 1 2008Jhy-Chong Liang Abstract Labedipinedilol-A, a novel dihydropyridine-type calcium antagonist with ,/,-adrenoceptor blocking properties, has been reported to produce a cardioprotective effect against ischemia reperfusion injury in rats. We investigated the protective effects of labedipinedilol-A on ouabain-induced tonotropy and arrhythmias in isolated whole atria, and using patch-clamp techniques to study the underlying mechanism of its antiarrhythmic activity on isolated cardiac myocytes. Labedipinedilol-A (10,µM) suppressed the tonotropic effect of ouabain significantly and prolonged the onset time of extra-systole (arrhythmia) induced by ouabain in isolate atria. In the voltage-clamp study, labedipinedilol-A (1,100,µM) reduced the peak amplitude of sodium inward current (INa) and L-type calcium current (ICa-L), and shifted the current-voltage (I-V) curve upward in a concentration-dependent manner. In contrast, the addition of labedipinedilol-A increased transient outward potassium current (Ito) and inward rectifier potassium current (IK1) significantly. Labedipinedilol-A (10,µM) also effectively depressed the isoproterenol-induced increase in the Ca2+ current. These results show that labedipinedilol-A blocks ICa-L and INa, and increases Ito and IK1. These findings indicate that labedipinedilol-A produces direct cardiac action, probably due to the inhibition of cardiac Na+ and Ca2+ channels. Our results suggest that labedipinedilol-A may reduce the membrane conduction through inhibition of ionic channels which decrease ouabain-induced arrhythmia. Drug Dev Res 69:26,33, 2008 © 2008 Wiley-Liss, Inc. [source] Calcium and polyamine regulated calcium-sensing receptors in cardiac tissuesFEBS JOURNAL, Issue 12 2003Rui Wang Activation of a calcium-sensing receptor (Ca-SR) leads to increased intracellular calcium concentration and altered cellular activities. The expression of Ca-SR has been identified in both nonexcitable and excitable cells, including neurons and smooth muscle cells. Whether Ca-SR was expressed and functioning in cardiac myocytes remained unclear. In the present study, the transcripts of Ca-SR were identified in rat heart tissues using RT-PCR that was further confirmed by sequence analysis. Ca-SR proteins were detected in rat ventricular and atrial tissues as well as in isolated cardiac myocytes. Anti-(Ca-SR) Ig did not detect any specific bands after preadsorption with standard Ca-SR antigens. An immunohistochemistry study revealed the presence of Ca-SR in rat cardiac as well as other tissues. An increase in extracellular calcium or gadolinium induced a concentration-dependent sustained increase in [Ca2+]i in isolated ventricular myocytes from adult rats. Spermine (1,10 mm) also increased [Ca2+]i. Pre-treatment of cardiac myocytes with thapsigargin or U73122 abolished the extracellular calcium, gadolinium or spermine-induced increase in [Ca2+]i. The blockade of Na+/Ca2+ exchanger or voltage-dependent calcium channels did not alter the extracellular calcium-induced increase in [Ca2+]i. Finally, extracellular calcium, gadolinium and spermine all increased intracellular inositol 1,4,5-triphosphate (IP3) levels. Our results demonstrated that Ca-SR was expressed in cardiac tissue and cardiomyocytes and its function was regulated by extracellular calcium and spermine. [source] Changes in extracellular K+ concentration modulate contractility of rat and rabbit cardiac myocytes via the inward rectifier K+ current IK1THE JOURNAL OF PHYSIOLOGY, Issue 3 2004Ron Bouchard The mechanisms underlying the inotropic effect of reductions in [K+]o were studied using recordings of membrane potential, membrane current, cell shortening and [Ca2+]i in single, isolated cardiac myocytes. Three types of mammalian myocytes were chosen, based on differences in the current density and intrinsic voltage dependence of the inwardly rectifying background K+ current IK1 in each cell type. Rabbit ventricular myocytes had a relatively large IK1 with a prominent negative slope conductance whereas rabbit atrial cells expressed much smaller IK1, with little or no negative slope conductance. IK1 in rat ventricle was intermediate in both current density and slope conductance. Action potential duration is relatively short in both rabbit atrial and rat ventricular myocytes, and consequently both cell types spend much of the duty cycle at or near the resting membrane potential. Rapid increases or decreases of [K+]o elicited significantly different inotropic effects in rat and rabbit atrial and ventricular myocytes. Voltage-clamp and current-clamp experiments showed that the effects on cell shortening and [Ca2+]i following changes in [K+]o were primarily the result of the effects of alterations in IK1, which changed resting membrane potential and action potential waveform. This in turn differentially altered the balance of Ca2+ efflux via the sarcolemmal Na+,Ca2+ exchanger, Ca2+ influx via voltage-dependant Ca2+ channels and sarcoplasmic reticulum (SR) Ca2+ release in each cell type. These results support the hypothesis that the inotropic effect of alterations of [K+]o in the heart is due to significant non-linear changes in the current,voltage relation for IK1 and the resulting modulation of the resting membrane potential and action potential waveform. [source] BLOCK OF Na+ AND K+ CURRENTS IN RAT VENTRICULAR MYOCYTES BY QUINACAINOL AND QUINIDINECLINICAL AND EXPERIMENTAL PHARMACOLOGY AND PHYSIOLOGY, Issue 1-2 2005Michael K Pugsley SUMMARY 1.,The electrophysiological actions of quinacainol were investigated on sodium (INa), transient outward (ito) and sustained-outward plateau (iKsus) potassium currents in rat isolated cardiac myocytes using the whole-cell patch-clamp technique and compared with quinidine. 2.,Quinacainol blocked sodium currents in a concentration-dependent manner and with a potency similar to that of quinidine (mean (±SEM) EC50 50 ± 12 vs 95 ± 25 µmol/L for quinidine and quinacainol, respectively). However, quinacainol had a considerably prolonged onset and recovery from block compared with quinidine. 3.,Neither quinacainol nor quinidine significantly changed the steady state voltage dependence of activation of sodium currents. Quinidine produced a hyperpolarizing shift in the voltage dependence for sodium current inactivation, but no such shift was observed with quinacainol at doses that produced a substantial current block. 4.,Although quinacainol did not effectively block voltage-dependent potassium currents, even at concentrations as high as 1.5 mmol/L, quinidine, at a half-maximal sodium channel-blocking concentration, reduced peak ito current amplitude, increased the rate of inactivation of ito and blocked iKsus. 5.,These results indicate that quinacainol, a quinidine analogue, blocks sodium currents in cardiac myocytes with little effect on ito or iKsus potassium currents, which suggests that quinacainol may be exerting class 1c anti-arrhythmic actions. [source] | ||||||||||