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BACKGROUND AND PURPOSE Although isoprenaline (ISO) is known to activate several ion currents in mammalian myocardium, little is known about the role of action potential morphology in the ISO-induced changes in ion currents. Therefore, the effects of ISO on action potential configuration, L-type Ca2+ current ( ICa), slow delayed rectifier K+ current ( IKs) and fast delayed rectifier K+ current ( IKr) were studied and compared in a frequency-dependent manner using canine isolated ventricular myocytes from various transmural locations. EXPERIMENTAL APPROACH Action potentials were recorded with conventional sharp microelectrodes; ion currents were measured using conventional and action potential voltage clamp techniques. KEY RESULTS In myocytes displaying a spike-and-dome action potential configuration (epicardial and midmyocardial cells), ISO caused reversible shortening of action potentials accompanied by elevation of the plateau. ISO-induced action potential shortening was absent in endocardial cells and in myocytes pretreated with 4-aminopyridine. Application of the IKr blocker E-4031 failed to modify the ISO effect, while action potentials were lengthened by ISO in the presence of the IKs blocker HMR-1556. Both action potential shortening and elevation of the plateau were prevented by pretreatment with the ICa blocker nisoldipine. Action potential voltage clamp experiments revealed a prominent slowly inactivating ICa followed by a rise in IKs, both currents increased with increasing the cycle length. CONCLUSIONS AND IMPLICATIONS The effect of ISO in canine ventricular cells depends critically on action potential configuration, and the ISO-induced activation of IKs- but not IKr- may be responsible for the observed shortening of action potentials. [ABSTRACT FROM AUTHOR]

Aim The aim of this work was to study antagonistic interactions between the effects of various types of Ca2+ channel blockers and isoproterenol on the amplitude of L-type Ca2+ current in canine ventricular cells. Methods Whole-cell version of the patch clamp technique was used to study the effect of isoproterenol on Ca2+ current in the absence and presence of Ca2+ channel-blocking agents, including nifedipine, nisoldipine, diltiazem, verapamil, Co Cl2 and Mn Cl2. Results Five micromolar Nifedipine, 1 μ M nisoldipine, 10 μ M diltiazem, 5 μ M verapamil, 3 m M Co Cl2 and 5 m M Mn Cl2 evoked uniformly a 90-95% blockade of Ca2+ current in the absence of isoproterenol. Isoproterenol (100 n M) alone increased the amplitude of Ca2+ current from 6.8 ± 1.3 to 23.7 ± 2.2 pA/ pF in a reversible manner. Isoproterenol caused a marked enhancement of Ca2+ current even in the presence of nifedipine, nisoldipine, diltiazem and verapamil, but not in the presence of Co Cl2 or Mn Cl2. Conclusion The results indicate that the action of isoproterenol is different in the presence of organic and inorganic Ca2+ channel blockers. Co Cl2 and Mn Cl2 were able to fully prevent the effect of isoproterenol on Ca2+ current, while the organic Ca2+ channel blockers failed to do so. This has to be born in mind when the effects of organic Ca2+ channel blockers are evaluated either experimentally or clinically under conditions of increased sympathetic tone. [ABSTRACT FROM AUTHOR]

Aim: The aim of the present study was to give a parametric description of the most important K+ currents flowing during canine ventricular action potential. Methods: Inward rectifier K+ current (IK1), rapid delayed rectifier K+ current ( IKr), and transient outward K+ current ( Ito) were dissected under action potential clamp conditions using BaCl2, E-4031, and 4-aminopyridine, respectively. Results: The maximum amplitude of Ito was 3.0 ± 0.23 pA/pF and its integral was 29.7 ± 2.5 fC/pF. The current peaked 4.4 ± 0.7 ms after the action potential upstroke and rapidly decayed to zero with a time constant of 7.4 ± 0.6 ms. IKr gradually increased during the plateau, peaked 7 ms before the time of maximum rate of repolarization ( V ) at −54.2 ± 1.7 mV, had peak amplitude of 0.62 ± 0.08 pA/pF, and integral of 57.6 ± 6.7 fC/pF. IK1 began to rise from −22.4 ± 0.8 mV, peaked 1 ms after the time of V at −58.3 ± 0.6 mV, had peak amplitude of 1.8 ± 0.1 pA/pF, and integral of 61.6 ± 6.2 fC/pF. Good correlation was observed between peak IK1 and V ( r = 0.93) but none between IKr and V . Neither IK1 nor IKr was frequency-dependent between 0.2 and 1.66 Hz. Congruently, IKr failed to accumulate in canine myocytes at fast driving rates. Conclusion: Terminal repolarization is dominated by IK1, but action potential duration is influenced by several ion currents simultaneously. As Ito was not active during the plateau, and neither IK1 nor IKr was frequency-dependent, other currents must be responsible for the frequency dependence of action potential duration at normal and slow heart rates in canine ventricular cells. [ABSTRACT FROM AUTHOR]

Current density, the membrane current value divided by membrane capacitance (C m ), is widely used in cellular electrophysiology. Comparing current densities obtained in different cell populations assume that C m and ion current magnitudes are linearly related, however data is scarce about this in cardiomyocytes. Therefore, we statistically analyzed the distributions, and the relationship between parameters of canine cardiac ion currents and C m , and tested if dividing original parameters with C m had any effect. Under conventional voltage clamp conditions, correlations were high for I K1 , moderate for I Kr and I Ca,L , while negligible for I Ks . Correlation between I to1 peak amplitude and C m was negligible when analyzing all cells together, however, the analysis showed high correlations when cells of subepicardial, subendocardial or midmyocardial origin were analyzed separately. In action potential voltage clamp experiments I K1, I Kr and I Ca,L parameters showed high correlations with C m . For I NCX , I Na,late and I Ks there were low-to-moderate correlations between C m and these current parameters. Dividing the original current parameters with C m reduced both the coefficient of variation, and the deviation from normal distribution. The level of correlation between ion currents and C m varies depending on the ion current studied. This must be considered when evaluating ion current densities in cardiac cells., (© 2024. The Author(s).)

The present study was designed to test the hypothesis that the selectivity of blocking the late Na + current (I NaL ) over the peak Na + current (I NaP ) is related to the fast offset kinetics of the Na + channel inhibitor. Therefore, the effects of 1 µM GS967 (I NaL inhibitor), 20 µM mexiletine (I/B antiarrhythmic) and 10 µM quinidine (I/A antiarrhythmic) on I NaL and I NaP were compared in canine ventricular myocardium. I NaP was estimated as the maximum velocity of action potential upstroke (V + max ). Equal amounts of I NaL were dissected by the applied drug concentrations under APVC conditions. The inhibition of I NaL by mexiletine and quinidine was comparable under a conventional voltage clamp, while both were smaller than the inhibitory effect of GS967. Under steady-state conditions, the V + max block at the physiological cycle length of 700 ms was 2.3% for GS967, 11.4% for mexiletine and 26.2% for quinidine. The respective offset time constants were 110 ± 6 ms, 456 ± 284 ms and 7.2 ± 0.9 s. These results reveal an inverse relationship between the offset time constant and the selectivity of I NaL over I NaP inhibition without any influence of the onset rate constant. It is concluded that the selective inhibition of I NaL over I NaP is related to the fast offset kinetics of the Na + channel inhibitor.

Late sodium current (I Na,late ) is an important inward current contributing to the plateau phase of the action potential (AP) in the mammalian heart. Although I Na,late is considered as a possible target for antiarrhythmic agents, several aspects of this current remained hidden. In this work, the profile of I Na,late , together with the respective conductance changes (G Na,late ), were studied and compared in rabbit, canine, and guinea pig ventricular myocytes using the action potential voltage clamp (APVC) technique. In canine and rabbit myocytes, the density of I Na,late was relatively stable during the plateau and decreased only along terminal repolarization of the AP, while G Na,late decreased monotonically. In contrast, I Na,late increased monotonically, while G Na,late remained largely unchanged during the AP in guinea pig. The estimated slow inactivation of Na + channels was much slower in guinea pig than in canine or rabbit myocytes. The characteristics of canine I Na,late and G Na,late were not altered by using command APs recorded from rabbit or guinea pig myocytes, indicating that the different shapes of the current profiles are related to genuine interspecies differences in the gating of I Na,late . Both I Na,late and G Na,late decreased in canine myocytes when the intracellular Ca 2+ concentration was reduced either by the extracellular application of 1 µM nisoldipine or by the intracellular application of BAPTA. Finally, a comparison of the I Na,late and G Na,late profiles induced by the toxin of Anemonia sulcata (ATX-II) in canine and guinea pig myocytes revealed profound differences between the two species: in dog, the ATX-II induced I Na,late and G Na,late showed kinetics similar to those observed with the native current, while in guinea pig, the ATX-II induced G Na,late increased during the AP. Our results show that there are notable interspecies differences in the gating kinetics of I Na,late that cannot be explained by differences in AP morphology. These differences must be considered when interpreting the I Na,late results obtained in guinea pig.

Aims: Heart failure with reduced ejection fraction (HFrEF) is a disease with high mortality and morbidity. Recent positive inotropic drug developments focused on cardiac myofilaments, that is, direct activators of the myosin molecule and Ca 2+ sensitizers for patients with advanced HFrEF. Omecamtiv mecarbil (OM) is the first direct myosin activator with promising results in clinical studies. Here, we aimed to elucidate the cellular mechanisms of the positive inotropic effect of OM in a comparative in vitro investigation where Ca 2+ -sensitizing positive inotropic agents with distinct mechanisms of action [EMD 53998 (EMD), which also docks on the myosin molecule, and levosimendan (Levo), which binds to troponin C] were included., Methods: Enzymatically isolated canine cardiomyocytes with intact cell membranes were loaded with Fura-2AM, a Ca 2+ -sensitive, ratiometric, fluorescent dye. Changes in sarcomere length (SL) and intracellular Ca 2+ concentration were recorded in parallel at room temperature, whereas cardiomyocyte contractions were evoked by field stimulation at 0.1 Hz in the presence of different OM, EMD, or Levo concentrations., Results: SL was reduced by about 23% or 9% in the presence of 1 μM OM or 1 μM EMD in the absence of electrical stimulation, whereas 1 μM Levo had no effect on resting SL. Fractional sarcomere shortening was increased by 1 μM EMD or 1 μM Levo to about 152%, but only to about 128% in the presence of 0.03 μM OM. At higher OM concentrations, no significant increase in fractional sarcomere shortening could be recorded. Contraction durations largely increased, whereas the kinetics of contractions and relaxations decreased with increasing OM concentrations. One-micromole EMD or 1 μM Levo had no effects on contraction durations. One-micromole Levo, but not 1 μM EMD, accelerated the kinetics of cardiomyocyte contractions and relaxations. Ca 2+ transient amplitudes were unaffected by all treatments., Conclusions: Our data revealed major distinctions between the cellular effects of myofilament targeted agents (OM, EMD, or Levo) depending on their target proteins and binding sites, although they were compatible with the involvement of Ca 2+ -sensitizing mechanisms for all three drugs. Significant part of the cardiotonic effect of OM relates to the prolongation of systolic contraction in combination with its Ca 2+ -sensitizing effect., (© 2023 The Authors. ESC Heart Failure published by John Wiley & Sons Ltd on behalf of European Society of Cardiology.)

ABT-333 (dasabuvir) is an antiviral agent used in hepatitis C treatment. The molecule, similarly to some inhibitors of hERG channels, responsible for the delayed rectifier potassium current (I Kr ), contains the methanesulfonamide group. Reduced I Kr current leads to long QT syndrome and early afterdepolarizations (EADs), therefore potentially causing life-threatening arrhythmias and sudden cardiac death. Our goal was to investigate the acute effects of ABT-333 in enzymatically isolated canine left ventricular myocardial cells. Action potentials (APs) and ion currents were recorded with a sharp microelectrode technique and whole-cell patch clamp, respectively. Application of 1 μM ABT-333 prolonged the AP in a reversible manner. The maximal rates of phases 0 and 1 were irreversibly decreased. Higher ABT-333 concentrations caused larger AP prolongation, elevation of the early plateau potential, and reduction of maximal rates of phases 0, 1, and 3. EADs occurred in some cells in 3-30 μM ABT-333 concentrations. The 10 μM ABT-333-sensitive current, recorded with AP voltage clamp, contained a late outward component corresponding to I Kr and an early outward one corresponding to transient outward potassium current (I to ). ABT-333 reduced hERG-channel-mediated ion current in a concentration-dependent, partially reversible manner with a half-inhibitory concentration of 3.2 μM. As the therapeutic plasma concentration of ABT-333 can reach the low μM range, ABT-333 application carries a risk of cardiac side effects especially in case of coadministration with strong inhibitors of CYP2C8.

Recent cardiotropic drug developments have focused on cardiac myofilaments. Danicamtiv, the second direct myosin activator, has achieved encouraging results in preclinical and clinical studies, thus implicating its potential applicability in the treatment of heart failure with reduced ejection fraction (HFrEF). Here, we analyzed the inotropic effects of danicamtiv in detail. To this end, changes in sarcomere length and intracellular Ca 2+ levels were monitored in parallel, in enzymatically isolated canine cardiomyocytes, and detailed echocardiographic examinations were performed in anesthetized rats in the absence or presence of danicamtiv. The systolic and diastolic sarcomere lengths decreased; contraction and relaxation kinetics slowed down with increasing danicamtiv concentrations without changes in intracellular Ca 2+ transients in vitro. Danicamtiv evoked remarkable increases in left ventricular ejection fraction and fractional shortening, also reflected by changes in systolic strain. Nevertheless, the systolic ejection time was significantly prolonged, the ratio of diastolic to systolic duration was reduced, and signs of diastolic dysfunction were also observed upon danicamtiv treatment in vivo. Taken together, danicamtiv improves cardiac systolic function, but it can also limit diastolic performance, especially at high drug concentrations.

Cardiac diseases are the leading causes of death, with a growing number of cases worldwide, posing a challenge for both healthcare and research. Therefore, the most relevant aim of cardiac research is to unravel the molecular pathomechanisms and identify new therapeutic targets. Cardiac ryanodine receptor (RyR2), the Ca 2+ release channel of the sarcoplasmic reticulum, is believed to be a good therapeutic target in a group of certain heart diseases, collectively called cardiac ryanopathies. Ryanopathies are associated with the impaired function of the RyR, leading to heart diseases such as congestive heart failure (CHF), catecholaminergic polymorphic ventricular tachycardia (CPVT), arrhythmogenic right ventricular dysplasia type 2 (ARVD2), and calcium release deficiency syndrome (CRDS). The aim of the current review is to provide a short insight into the pathological mechanisms of ryanopathies and discuss the pharmacological approaches targeting RyR2.

The patch clamp technique underwent continual advancement and developed numerous variants in cardiac electrophysiology since its introduction in the late 1970s. In the beginning, the capability of the technique was limited to recording one single current from one cell stimulated with a rectangular command pulse. Since that time, the technique has been extended to record multiple currents under various command pulses including action potential. The current review summarizes the development of the patch clamp technique in cardiac electrophysiology with special focus on the potential applications in integrative physiology., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Horváth, Szentandrássy, Dienes, Kovács, Nánási, Chen-Izu, Izu and Banyasz.)

Late sodium current has long been linked to dysrhythmia and contractile malfunction in the heart. Despite the increasing body of accumulating information on the subject, our understanding of its role in normal or pathologic states is not complete. Even though the role of late sodium current in shaping action potential under physiologic circumstances is debated, it's unquestioned role in arrhythmogenesis keeps it in the focus of research. Transgenic mouse models and isoform-specific pharmacological tools have proved useful in understanding the mechanism of late sodium current in health and disease. This review will outline the mechanism and function of cardiac late sodium current with special focus on the recent advances of the area.

Astaxanthin is a lipid-soluble carotenoid influencing lipid metabolism, body weight, and insulin sensitivity. We provide a systematic analysis of acute and chronic effects of astaxanthin on different organs. Changes by chronic astaxanthin feeding were analyzed on general metabolism, expression of regulatory proteins in the skeletal muscle, as well as changes of excitation and synaptic activity in the hypothalamic arcuate nucleus of mice. Acute responses were also tested on canine cardiac muscle and different neuronal populations of the hypothalamic arcuate nucleus in mice. Dietary astaxanthin significantly increased food intake. It also increased protein levels affecting glucose metabolism and fatty acid biosynthesis in skeletal muscle. Inhibitory inputs innervating neurons of the arcuate nucleus regulating metabolism and food intake were strengthened by both acute and chronic astaxanthin treatment. Astaxanthin moderately shortened cardiac action potentials, depressed their plateau potential, and reduced the maximal rate of depolarization. Based on its complex actions on metabolism and food intake, our data support the previous findings that astaxanthin is suitable for supplementing the diet of patients with disturbances in energy homeostasis.

Transient receptor potential melastatin 4 is a unique member of the TRPM protein family and, similarly to TRPM5, is Ca 2+ -sensitive and permeable to monovalent but not divalent cations. It is widely expressed in many organs and is involved in several functions by regulating the membrane potential and Ca 2+ homeostasis in both excitable and non-excitable cells. This part of the review discusses the pharmacological modulation of TRPM4 by listing, comparing, and describing both endogenous and exogenous activators and inhibitors of the ion channel. Moreover, other strategies used to study TRPM4 functions are listed and described. These strategies include siRNA-mediated silencing of TRPM4, dominant-negative TRPM4 variants, and anti-TRPM4 antibodies. TRPM4 is receiving more and more attention and is likely to be the topic of research in the future.

Transient receptor potential melastatin 4 (TRPM4) is a unique member of the TRPM protein family and, similarly to TRPM5, is Ca 2+ sensitive and permeable for monovalent but not divalent cations. It is widely expressed in many organs and is involved in several functions; it regulates membrane potential and Ca 2+ homeostasis in both excitable and non-excitable cells. This part of the review discusses the currently available knowledge about the physiological and pathophysiological roles of TRPM4 in various tissues. These include the physiological functions of TRPM4 in the cells of the Langerhans islets of the pancreas, in various immune functions, in the regulation of vascular tone, in respiratory and other neuronal activities, in chemosensation, and in renal and cardiac physiology. TRPM4 contributes to pathological conditions such as overactive bladder, endothelial dysfunction, various types of malignant diseases and central nervous system conditions including stroke and injuries as well as in cardiac conditions such as arrhythmias, hypertrophy, and ischemia-reperfusion injuries. TRPM4 claims more and more attention and is likely to be the topic of research in the future.

Enhancement of the late sodium current (I NaL ) increases arrhythmia propensity in the heart, whereas suppression of the current is antiarrhythmic. In the present study, we investigated I NaL in canine ventricular cardiomyocytes under action potential voltage-clamp conditions using the selective Na + channel inhibitors GS967 and tetrodotoxin. Both 1 µM GS967 and 10 µM tetrodotoxin dissected largely similar inward currents. The amplitude and integral of the GS967-sensitive current was significantly smaller after the reduction of intracellular Ca 2+ concentration ([Ca 2+ ] i ) either by superfusion of the cells with 1 µM nisoldipine or by intracellular application of 10 mM BAPTA. Inhibiting calcium/calmodulin-dependent protein kinase II (CaMKII) by KN-93 or the autocamtide-2-related inhibitor peptide similarly reduced the amplitude and integral of I NaL . Action potential duration was shortened in a reverse rate-dependent manner and the plateau potential was depressed by GS967. This GS967-induced depression of plateau was reduced by pretreatment of the cells with BAPTA-AM. We conclude that (1) I NaL depends on the magnitude of [Ca 2+ ] i in canine ventricular cells, (2) this [Ca 2+ ] i -dependence of I NaL is mediated by the Ca 2+ -dependent activation of CaMKII, and (3) I NaL is augmented by the baseline CaMKII activity.

The profiles of ion currents during the cardiac action potential can be visualized by the action potential voltage clamp technique. To obtain multiple ion current data from the same cell, the "onion peeling" technique, based on sequential pharmacological dissection of ion currents, has to be applied. Combination of the two methods allows recording of several ion current profiles from the same myocyte under largely physiological conditions. Using this approach, we have studied the densities and integrals of the major cardiac inward (I Ca , I NCX , I Na-late ) and outward (I Kr , I Ks , I K1 ) currents in canine ventricular cells and studied the correlation between them. For this purpose, canine ventricular cardiomyocytes were chosen because their electrophysiological properties are similar to those of human ones. Significant positive correlation was observed between the density and integral of I Ca and I Kr , and positive correlation was found also between the integral of I Ca and I NCX . No further correlations were detected. The Ca 2+ -sensitivity of K + currents was studied by comparing their parameters in the case of normal calcium homeostasis and following blockade of I Ca . Out of the three K + currents studied, only I Ks was Ca 2+ -sensitive. The density and integral of I Ks was significantly greater, while its time-to-peak value was shorter at normal Ca 2+ cycling than following I Ca blockade. No differences were detected for I Kr or I K1 in this regard. Present results indicate that the positive correlation between I Ca and I Kr prominently contribute to the balance between inward and outward fluxes during the action potential plateau in canine myocytes. The results also suggest that the profiles of cardiac ion currents have to be studied under physiological conditions, since their behavior may strongly be influenced by the intracellular Ca 2+ homeostasis and the applied membrane potential protocol., (Copyright © 2021 Elsevier Ltd. All rights reserved.)

Transient receptor potential melastatin 4 (TRPM4) plays an important role in many tissues, including pacemaker and conductive tissues of the heart, but much less is known about its electrophysiological role in ventricular myocytes. Our earlier results showed the lack of selectivity of 9-phenanthrol, so CBA ((4-chloro-2-(2-chlorophenoxy)acetamido) benzoic acid) was chosen as a new, potentially selective inhibitor. Goal: Our aim was to elucidate the effect and selectivity of CBA in canine left ventricular cardiomyocytes and to study the expression of TRPM4 in the canine heart. Experiments were carried out in enzymatically isolated canine left ventricular cardiomyocytes. Ionic currents were recorded with an action potential (AP) voltage-clamp technique in whole-cell configuration at 37 °C. An amount of 10 mM BAPTA was used in the pipette solution to exclude the potential activation of TRPM4 channels. AP was recorded with conventional sharp microelectrodes. CBA was used in 10 µM concentrations. Expression of TRPM4 protein in the heart was studied by Western blot. TRPM4 protein was expressed in the wall of all four chambers of the canine heart as well as in samples prepared from isolated left ventricular cells. CBA induced an approximately 9% reduction in AP duration measured at 75% and 90% of repolarization and decreased the short-term variability of APD 90 . Moreover, AP amplitude was increased and the maximal rates of phase 0 and 1 were reduced by the drug. In AP clamp measurements, CBA-sensitive current contained a short, early outward and mainly a long, inward current. Transient outward potassium current (I to ) and late sodium current (I Na,L ) were reduced by approximately 20% and 47%, respectively, in the presence of CBA, while L-type calcium and inward rectifier potassium currents were not affected. These effects of CBA were largely reversible upon washout. Based on our results, the CBA induced reduction of phase-1 slope and the slight increase of AP amplitude could have been due to the inhibition of I to . The tendency for AP shortening can be explained by the inhibition of inward currents seen in AP-clamp recordings during the plateau phase. This inward current reduced by CBA is possibly I Na,L , therefore, CBA is not entirely selective for TRPM4 channels. As a consequence, similarly to 9-phenanthrol, it cannot be used to test the contribution of TRPM4 channels to cardiac electrophysiology in ventricular cells, or at least caution must be applied.

Due to the limited availability of healthy human ventricular tissues, the most suitable animal model has to be applied for electrophysiological and pharmacological studies. This can be best identified by studying the properties of ion currents shaping the action potential in the frequently used laboratory animals, such as dogs, rabbits, guinea pigs, or rats, and comparing them to those of human cardiomyocytes. The authors of this article with the experience of three decades of electrophysiological studies, performed in mammalian and human ventricular tissues and isolated cardiomyocytes, summarize their results obtained regarding the major canine and human cardiac ion currents. Accordingly, L-type Ca 2+ current (I Ca ), late Na + current (I Na-late ), rapid and slow components of the delayed rectifier K + current (I Kr and I Ks , respectively), inward rectifier K + current (I K1 ), transient outward K + current (I to1 ), and Na + /Ca 2+ exchange current (I NCX ) were characterized and compared. Importantly, many of these measurements were performed using the action potential voltage clamp technique allowing for visualization of the actual current profiles flowing during the ventricular action potential. Densities and shapes of these ion currents, as well as the action potential configuration, were similar in human and canine ventricular cells, except for the density of I K1 and the recovery kinetics of I to . I K1 displayed a largely four-fold larger density in canine than human myocytes, and I to recovery from inactivation displayed a somewhat different time course in the two species. On the basis of these results, it is concluded that canine ventricular cells represent a reasonably good model for human myocytes for electrophysiological studies, however, it must be borne in mind that due to their stronger I K1 , the repolarization reserve is more pronounced in canine cells, and moderate differences in the frequency-dependent repolarization patterns can also be anticipated.

Enhancement of the late Na + current (I NaL ) increases arrhythmia propensity in the heart, while suppression of the current is antiarrhythmic. GS967 is an agent considered as a selective blocker of I NaL . In the present study, effects of GS967 on I NaL and action potential (AP) morphology were studied in canine ventricular myocytes by using conventional voltage clamp, action potential voltage clamp and sharp microelectrode techniques. The effects of GS967 (1 µM) were compared to those of the class I/B antiarrhythmic compound mexiletine (40 µM). Under conventional voltage clamp conditions, I NaL was significantly suppressed by GS967 and mexiletine, causing 80.4 ± 2.2% and 59.1 ± 1.8% reduction of the densities of I NaL measured at 50 ms of depolarization, and 79.0 ± 3.1% and 63.3 ± 2.7% reduction of the corresponding current integrals, respectively. Both drugs shifted the voltage dependence of the steady-state inactivation curve of I NaL towards negative potentials. GS967 and mexiletine dissected inward I NaL profiles under AP voltage clamp conditions having densities, measured at 50% of AP duration (APD), of -0.37 ± 0.07 and -0.28 ± 0.03 A/F, and current integrals of -56.7 ± 9.1 and -46.6 ± 5.5 mC/F, respectively. Drug effects on peak Na + current (I NaP ) were assessed by recording the maximum velocity of AP upstroke (V + max ) in multicellular preparations. The offset time constant was threefold faster for GS967 than mexiletine (110 ms versus 289 ms), while the onset of the rate-dependent block was slower in the case of GS967. Effects on beat-to-beat variability of APD was studied in isolated myocytes. Beat-to-beat variability was significantly decreased by both GS967 and mexiletine (reduction of 42.1 ± 6.5% and 24.6 ± 12.8%, respectively) while their shortening effect on APD was comparable. It is concluded that the electrophysiological effects of GS967 are similar to those of mexiletine, but with somewhat faster offset kinetics of V + max block. However, since GS967 depressed V + max and I NaL at the same concentration, the current view that GS967 represents a new class of drugs that selectively block I NaL has to be questioned and it is suggested that GS967 should be classified as a class I/B antiarrhythmic agent.