The transverse-axial tubular system (tubular system) of cardiomyocytes plays a key role in excitation-contraction coupling. To determine the area of the tubular membrane in relation to the area of the surface membrane, indirect measurements through the determination of membrane capacitances are currently used in addition to microscopic methods. Unlike existing electrophysiological methods based on an irreversible procedure (osmotic shock), the proposed new approach uses a reversible short-term intermittent increase in the electrical resistance of the extracellular medium. The resulting increase in the lumen resistance of the tubular system makes it possible to determine separate capacitances of the tubular and surface membranes. Based on the analysis of the time course of the capacitive current, computational relations were derived to quantify the elements of the electrical equivalent circuit of the measured cardiomyocyte including both capacitances. The exposition to isotonic low-conductivity sucrose solution is reversible which is the main advantage of the proposed approach allowing repetitive measurements on the same cell under control and sucrose solutions. Experiments on rat ventricular cardiomyocytes (n = 20) resulted in the surface and tubular capacitance values implying the fraction of tubular capacitance/area of 0.327 ± 0.018. We conclude that the newly proposed method provides results comparable to the data obtained by the currently used detubulation method and, in addition, by being reversible, allows repeated evaluation of surface and tubular membrane parameters on the same cell.

• Klíčová slova
• Cardiomyocyte, Novel method, Sucrose, Tubular membrane capacitance, Tubular system,
• MeSH
• elektrická vodivost MeSH
• kardiomyocyty * MeSH
• krysa rodu Rattus MeSH
• osmotický tlak MeSH
• sacharosa MeSH
• spřažení excitace a kontrakce * MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• sacharosa MeSH

Sildenafil (Viagra) is a vasodilator mainly used in the treatment of erectile dysfunction. Atrial or ventricular fibrillation may rarely occur as a side effect during sildenafil therapy. Although changes in inward rectifier potassium currents including I K1 are known to contribute to the pathogenesis of fibrillation, the effect of sildenafil on I K1 has not been studied. In experiments, Ba2+ is used as a specific inhibitor of I K1 at high concentrations (usually 100 µM). Being an environmental contaminant, it is also present in the human body; Ba2+ plasmatic concentrations up to 1.5 µM are usually reported in the general population. This study was primarily aimed to investigate changes of I K1 induced by sildenafil in a wide range of concentrations (0.1-100 µM). Additionally, the effect of combination of sildenafil and Ba2+ at selected clinically-relevant concentrations was tested, at 0.1 µM both on I K1 and on the action potential duration (APD). Experiments were performed by the whole-cell patch-clamp technique on enzymatically isolated rat ventricular cardiomyocytes, mostly at 23°C with the exception of APD measurements which were performed at 37°C as well. Sildenafil caused a significant, reversible, and concentration-dependent inhibition of I K1 that did not differ at -50 and -110 mV. Simultaneous application of sildenafil and Ba2+ at 0.1 µM revealed a massive inhibition of both inward and outward components of I K1 (this synergy was missing at other tested combinations). The combined effect at 0.1 µM (45.7 ± 5.7 and 43.0 ± 6.9% inhibition at -50 and -110 mV, respectively) was significantly higher than a simple sum of almost negligible effects of the individual substances and it led to a significant prolongation of APD at both 23 and 37°C. To our knowledge, similar potentiation of the drug-channel interaction has not been described. The observed massive inhibition of I K1 induced by a combined action of the vasodilator sildenafil and environmental contaminant Ba2+ at a low concentration and resulting in a significant APD prolongation may contribute to the genesis of arrhythmias observed in some patients treated with sildenafil.

• Klíčová slova
• arrhythmia, barium, cardiomyocytes, inward rectifier potassium current, sildenafil, synergy,
• Publikační typ
• časopisecké články MeSH

Bronchodilator aminophylline may induce atrial or less often ventricular arrhythmias. The mechanism of this proarrhythmic side effect has not been fully explained. Modifications of inward rectifier potassium (Kir) currents including IK1 are known to play an important role in arrhythmogenesis; however, no data on the aminophylline effect on these currents have been published. Hence, we tested the effect of aminophylline (3-100 µM) on IK1 in enzymatically isolated rat ventricular myocytes using the whole-cell patch-clamp technique. A dual steady-state effect of aminophylline was observed; either inhibition or activation was apparent in individual cells during the application of aminophylline at a given concentration. The smaller the magnitude of the control IK1, the more likely the activation of the current by aminophylline and vice versa. The effect was reversible; the relative changes at -50 and -110 mV did not differ. Using IK1 channel population model, the dual effect was explained by the interaction of aminophylline with two different channel populations, the first one being inhibited and the second one being activated. Considering various fractions of these two channel populations in individual cells, varying effects observed in the measured cells could be simulated. We propose that the dual aminophylline effect may be related to the direct and indirect effect of the drug on various Kir2.x subunits forming the homo- and heterotetrameric IK1 channels in a single cell. The observed IK1 changes induced by clinically relevant concentrations of aminophylline might contribute to arrhythmogenesis related to the use of this bronchodilator in clinical medicine.

• Klíčová slova
• Aminophylline, Arrhythmia, Dual effect, Fibrillation, Inward rectifier, Population channel model,
• MeSH
• aminofylin škodlivé účinky   MeSH
• bronchodilatancia škodlivé účinky   MeSH
• draslík farmakologie   MeSH
• draslíkové kanály dovnitř usměrňující * MeSH
• kardiomyocyty fyziologie   MeSH
• krysa rodu Rattus MeSH
• srdeční arytmie MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• aminofylin MeSH
• bronchodilatancia MeSH
• draslík MeSH
• draslíkové kanály dovnitř usměrňující * MeSH

Recent experimental work has revealed unusual features of the effect of certain drugs on cardiac inwardly rectifying potassium currents, including the constitutively active and acetylcholine-induced components of acetylcholine-sensitive current (IKAch). These unusual features have included alternating susceptibility of the current components to activation and inhibition induced by ethanol or nicotine applied at various concentrations, and significant correlation between the drug effect and the current magnitude measured under drug-free conditions. To explain these complex drug effects, we have developed a new type of quantitative model to offer a possible interpretation of the effect of ethanol and nicotine on the IKAch channels. The model is based on a description of IKAch as a sum of particular currents related to the populations of channels formed by identical assemblies of different α-subunits. Assuming two different channel populations in agreement with the two reported functional IKAch-channels (GIRK1/4 and GIRK4), the model was able to simulate all the above-mentioned characteristic features of drug-channel interactions and also the dispersion of the current measured in different cells. The formulation of our model equations allows the model to be incorporated easily into the existing integrative models of electrical activity of cardiac cells involving quantitative description of IKAch. We suppose that the model could also help make sense of certain observations related to the channels that do not show inward rectification. This new ionic channel model, based on a concept we call population type, may allow for the interpretation of complex interactions of drugs with ionic channels of various types, which cannot be done using the ionic channel models available so far.

• MeSH
• acetylcholin farmakologie   MeSH
• biologické modely MeSH
• časové faktory MeSH
• dovnitř usměrňující draslíkové kanály spřažené s G proteiny metabolismus   MeSH
• ethanol farmakologie   MeSH
• gating iontového kanálu účinky léků   MeSH
• nikotin farmakologie   MeSH
• počítačová simulace MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• acetylcholin MeSH
• dovnitř usměrňující draslíkové kanály spřažené s G proteiny MeSH
• ethanol MeSH
• nikotin MeSH

Inward rectifier potassium currents (I Kir,x) belong to prominent ionic currents affecting both resting membrane voltage and action potential repolarization in cardiomyocytes. In existing integrative models of electrical activity of cardiac cells, they have been described as single current components. The proposed quantitative model complies with findings indicating that these channels are formed by various homomeric or heteromeric assemblies of channel subunits with specific functional properties. Each I Kir,x may be expressed as a total of independent currents via individual populations of identical channels, i.e., channels formed by the same combination of their subunits. Solution of the model equations simulated well recently observed unique manifestations of dual ethanol effect in rat ventricular and atrial cells. The model reflects reported occurrence of at least two binding sites for ethanol within I Kir,x channels related to slow allosteric conformation changes governing channel conductance and inducing current activation or inhibition. Our new model may considerably improve the existing models of cardiac cells by including the model equations proposed here in the particular case of the voltage-independent drug-channel interaction. Such improved integrative models may provide more precise and, thus, more physiologically relevant results.

• Klíčová slova
• Cardiomyocytes, Dual effect, Ethanol, I K1, Inward rectifier potassium currents, Quantitative model,
• MeSH
• akční potenciály * MeSH
• alosterická regulace MeSH
• draslíkové kanály dovnitř usměrňující chemie   metabolismus   MeSH
• ethanol farmakologie   MeSH
• kardiomyocyty účinky léků   metabolismus   fyziologie   MeSH
• krysa rodu Rattus MeSH
• modely kardiovaskulární MeSH
• multimerizace proteinu MeSH
• srdce - funkce komor MeSH
• srdeční komory cytologie   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• draslíkové kanály dovnitř usměrňující MeSH
• ethanol MeSH

Nicotine abuse is associated with variety of diseases including arrhythmias, most often atrial fibrillation (AF). Altered inward rectifier potassium currents including acetylcholine-sensitive current I K(Ach) are known to be related to AF pathogenesis. Since relevant data are missing, we aimed to investigate I K(Ach) changes at clinically relevant concentrations of nicotine. Experiments were performed by the whole cell patch clamp technique at 23 ± 1 °C on isolated rat atrial myocytes. Nicotine was applied at following concentrations: 4, 40 and 400 nM; ethanol at 20 mM (∼0.09%). Nicotine at 40 and 400 nM significantly activated constitutively active component of I K(Ach) with the maximum effect at 40 nM (an increase by ∼100%); similar effect was observed at -110 and -50 mV. Changes at 4 nM nicotine were negligible on average. Coapplication of 40 nM nicotine and 20 mM ethanol (which is also known to activate this current) did not show cumulative effect. In the case of acetylcholine-induced component of I K(Ach), a dual effect of nicotine and its correlation with the current magnitude in control were apparent: the current was increased by nicotine in the cells showing small current in control and vice versa. The effect of 40 and 400 nM nicotine on acetylcholine-induced component of I K(Ach) was significantly different at -110 and -50 mV. We conclude that nicotine at clinically relevant concentrations significantly increased constitutively active component of I K(Ach) and showed a dual effect on its acetylcholine-induced component, similarly as ethanol. Synchronous application of nicotine and ethanol did not cause additive effect.

• Klíčová slova
• Acetylcholine-sensitive, Dual effect, Inward rectifier, Nicotine,
• MeSH
• acetylcholin farmakologie   MeSH
• časové faktory MeSH
• dovnitř usměrňující draslíkové kanály spřažené s G proteiny agonisté   účinky léků   MeSH
• ethanol toxicita   MeSH
• hodnocení rizik MeSH
• kardiomyocyty účinky léků   metabolismus   MeSH
• membránové potenciály MeSH
• nikotin toxicita   MeSH
• potkani Wistar MeSH
• srdeční arytmie chemicky indukované   metabolismus   MeSH
• srdeční síně účinky léků   metabolismus   MeSH
• techniky in vitro MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acetylcholin MeSH
• dovnitř usměrňující draslíkové kanály spřažené s G proteiny MeSH
• ethanol MeSH
• nikotin MeSH

Alcohol intoxication tends to induce arrhythmias, most often the atrial fibrillation. To elucidate arrhythmogenic mechanisms related to alcohol consumption, the effect of ethanol on main components of the ionic membrane current is investigated step by step. Considering limited knowledge, we aimed to examine the effect of clinically relevant concentrations of ethanol (0.8-80 mM) on acetylcholine-sensitive inward rectifier potassium current I K(Ach). Experiments were performed by the whole-cell patch clamp technique at 23 ± 1 °C on isolated rat and guinea-pig atrial myocytes, and on expressed human Kir3.1/3.4 channels. Ethanol induced changes of I K(Ach) in the whole range of concentrations applied; the effect was not voltage dependent. The constitutively active component of I K(Ach) was significantly increased by ethanol with the maximum effect (an increase by ∼100 %) between 8 and 20 mM. The changes were comparable in rat and guinea-pig atrial myocytes and also in expressed human Kir3.1/3.4 channels (i.e., structural correlate of I K(Ach)). In the case of the acetylcholine-induced component of I K(Ach), a dual ethanol effect was apparent with a striking heterogeneity of changes in individual cells. The effect correlated with the current magnitude in control: the current was increased by eth-anol in the cells showing small current in control and vice versa. The average effect peaked at 20 mM ethanol (an increase of the current by ∼20 %). Observed changes of action potential duration agreed well with the voltage clamp data. Ethanol significantly affected both components of I K(Ach) even in concentrations corresponding to light alcohol consumption.

• Klíčová slova
• Dual effect, Ethanol, Inward rectifier, Kir3.1/3.4, Rat atrial cell model,
• MeSH
• acetylcholin farmakologie   MeSH
• akční potenciály MeSH
• CHO buňky MeSH
• Cricetulus MeSH
• dovnitř usměrňující draslíkové kanály spřažené s G proteiny účinky léků   genetika   metabolismus   MeSH
• ethanol toxicita   MeSH
• hodnocení rizik MeSH
• kardiomyocyty účinky léků   metabolismus   MeSH
• kinetika MeSH
• lidé MeSH
• modely kardiovaskulární MeSH
• morčata MeSH
• počítačová simulace MeSH
• potkani Wistar MeSH
• srdeční arytmie chemicky indukované   metabolismus   patofyziologie   MeSH
• srdeční frekvence účinky léků   MeSH
• srdeční síně účinky léků   metabolismus   patofyziologie   MeSH
• transfekce MeSH
• vztah mezi dávkou a účinkem léčiva MeSH
• zvířata MeSH
• Check Tag
• lidé MeSH
• morčata MeSH
• mužské pohlaví MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• Názvy látek
• acetylcholin MeSH
• dovnitř usměrňující draslíkové kanály spřažené s G proteiny MeSH
• ethanol MeSH

Alcohol consumption may result in electrocardiographic changes and arrhythmias, at least partly due to effects of ethanol on cardiac ionic currents. Contractility and intracellular Ca(2+) dynamics seem to be altered as well. In this study, we integrated the available (mostly animal) experimental data into previously published models of the rat and human ventricular myocytes to assess the share of ionic current components in ethanol-induced changes in AP configuration and cytosolic Ca(2+) transient in ventricular cardiomyocytes. The rat model reproduced well the experimentally observed changes in AP duration (APD) under ethanol (slight prolongation at 0.8 mM and shortening at ≥8 mM). These changes were almost exclusively caused by the ethanol-induced alterations of I K1. The cytosolic Ca(2+) transient decreased gradually with the increasing ethanol concentration as a result of the ethanol-induced inhibition of I Ca. In the human model, ethanol produced a dose-dependent APD lengthening, dominated by ethanol effect on I Kr, the key repolarising current in human ventricles. This effect might contribute to the clinically observed proarrhythmic effects of ethanol in predisposed individuals.

• Klíčová slova
• Action potential, Cardiomyocyte, Ethanol, Human ventricular cell model, Rat ventricular cell model,
• MeSH
• akční potenciály účinky léků   MeSH
• biologické modely MeSH
• ethanol farmakologie   MeSH
• intracelulární prostor účinky léků   metabolismus   MeSH
• kardiomyocyty účinky léků   metabolismus   MeSH
• krysa rodu Rattus MeSH
• lidé MeSH
• počítačová simulace * MeSH
• sarkoplazmatické retikulum účinky léků   metabolismus   MeSH
• srdeční komory cytologie   MeSH
• vápník metabolismus   MeSH
• zvířata MeSH
• Check Tag
• krysa rodu Rattus MeSH
• lidé MeSH
• zvířata MeSH
• Publikační typ
• časopisecké články MeSH
• práce podpořená grantem MeSH
• Názvy látek
• ethanol MeSH
• vápník MeSH