[ESC guidelines on atrial fibrillation 2016 : Summary of the most relevant recommendations and modifications]. / ESC-Leitlinien zum Vorhofflimmern 2016 : Zusammenfassung der wichtigsten Empfehlungen und Neuerungen.

The first European Society of Cardiology (ESC) guidelines on atrial fibrillation (AF) developed in collaboration with the European Association for Cardio-Thoracic Surgery (EACTS) were published in August 2016. These guidelines replace the revised guidelines from 2012 and contain some interesting new aspects. The topics range from the pathophysiology through diagnostics, therapy and stroke prevention up to special clinical situations, such as atrial fibrillation in cardiopathy, sport and pregnancy. Early screening, patient informed consent, individualized therapy and the modification of factors promoting atrial fibrillation are of particular importance. The guidelines recommend the establishment of AF heart teams, containing specialists from various disciplines. The guidelines also underline the importance of non-vitamin K­dependent oral anticoagulants (NOAC) for stroke prevention compared to standard anticoagulants with vitamin K antagonists. For symptomatic and especially paroxysmal atrial fibrillation, the guidelines emphasize the importance of an antiarrhythmic treatment with catheter ablation and/or pharmaceutical antiarrhythmic therapy in addition to a frequency regulating therapy.

Interaction of DPP10a with Kv4.3 channel complex results in a sustained current component of human transient outward current Ito.

The sustained component of the K(+) outward current in human atrial myocytes is believed to be due to the slowly inactivating ultra-rapid potassium current I Kur and not to the fast inactivating transient outward current Ito. Here we provide evidence for contribution of Ito to this late current due to the effects of dipeptidyl peptidase-like protein (DPP) 10 (DPP10a) interacting with Kv4.3 channels. We studied the late current component of Ito in human atrial myocytes and CHO cells co-expressing Kv4.3 or Kv4.3/KChIP2 (control) and DPP proteins using voltage-clamp technique and a pharmacological approach. A voltage dependent and slowly inactivating late current (43% of peak amplitude) could be observed in atrial myocytes. We found a similar current in CHO cells expressing Kv4.3/KChIP2 + DPP10a, but not in cells co-expressing Kv4.3 + DPP or Kv4.3/KChIP2 + DPP6-S. Assuming that DPP10a influences atrial Ito, we detected DPP10 expression of three alternatively spliced mRNAs, DPP10 protein and colocalization of Kv4.3 and DPP10 proteins in human atrial myocytes. DPP10a did not affect properties of expressed Kv1.5 excluding a contribution to the sustained IKur in atrial cells. To test for the contribution of Kv4-based Ito on sustained K(+) outward currents in human atrial myocytes, we used 4-AP to block IKur, in combination with Heteropoda toxin 2 to block Kv4 channels. We could clearly separate an Ito fraction of about 19% contributing to the late current in atrial myocytes. Thus, the interaction of DPP10a, expressed in human atrium, with Kv4.3 channels generates a sustained current component of Ito, which may affect late repolarization phase of atrial action potentials.

Accessory subunits alter the temperature sensitivity of Kv4.3 channel complexes.

In human atrial myocytes the transient outward current I(to) develops a conspicuous faster inactivation with increasing temperatures. Since ß-subunits are known to modulate I(to) current kinetics, we hypothesized that the temperature sensitivity of I(to) is not only determined by the property of the ion-passing α-subunit Kv4.3 but also by its interaction with accessory ß-subunits. We therefore studied the influence of the transmembrane ß-subunits KCNE1, KCNE2 and DPP6 on Kv4.3/KChIP2 channels in CHO cells at room temperature and at physiological temperature. Exposure to 37°C caused a significant acceleration of the channel kinetics, whereas current densities and voltage dependences remained unaltered at 37°C compared to 23°C. However, Kv4.3/KChIP2 channels without transmembrane ß-subunits showed the strongest temperature sensitivity with considerably increased rates of activation and inactivation at 37°C. KCNE2 significantly slowed the current kinetics at 37°C compared to Kv4.3/KChIP2 channels, whereas KCNE1 did not influence the channel properties at both temperatures. Interestingly, the accelerating effects of DPP6 on current kinetics described at 23°C were diminished at physiological temperature, thus at 37°C current kinetics became remarkably similar for channel complexes Kv4.3/KChIP2 with and without DPP6 isoforms. A Markov state model was developed on the basis of experimental measurements to simulate the influence of ß-subunits on Kv4.3 channel complex at both temperatures. In conclusion, the remarkably fast kinetics of the native I(to) at 37°C could be reproduced by co-expressing Kv4.3, KChIP2, KCNE2 and DPP6 in CHO cells, whereas the high temperature sensitivity of human I(to) could be not mimicked.

Effects of MiRP1 and DPP6 beta-subunits on the blockade induced by flecainide of Kv4.3/KChIP2 channels.

BACKGROUND AND PURPOSE: The human cardiac transient outward potassium current (Ito) is believed to be composed of the pore-forming Kv4.3 alpha-subunit, coassembled with modulatory beta-subunits as KChIP2, MiRP1 and DPP6 proteins. beta-Subunits can alter the pharmacological response of Ito; therefore, we analysed the effects of flecainide on Kv4.3/KChIP2 channels coassembled with MiRP1 and/or DPP6 beta-subunits. EXPERIMENTAL APPROACH: Currents were recorded in Chinese hamster ovary cells stably expressing K(V)4.3/KChIP2 channels, and transiently transfected with either MiRP1, DPP6 or both, using the whole-cell patch-clamp technique. KEY RESULTS: In control conditions, Kv4.3/KChIP2/MiRP1 channels exhibited the slowest activation and inactivation kinetics and showed an 'overshoot' in the time course of recovery from inactivation. The midpoint values (Vh) of the activation and inactivation curves for Kv4.3/KChIP2/DPP6 and Kv4.3/KChIP2/MiRP1/DPP6 channels were approximately 10 mV more negative than Vh values for Kv4.3/KChIP2 and Kv4.3/KChIP2/MiRP1 channels. Flecainide (0.1-100 microM) produced a similar concentration-dependent blockade of total integrated current flow (IC50 approximately 10 microM) in all the channel complexes. However, the IC50 values for peak current amplitude and inactivated channel block were significantly different. Flecainide shifted the Vh values of both the activation and inactivation curves to more negative potentials and apparently accelerated inactivation kinetics in all channels. Moreover, flecainide slowed recovery from inactivation in all the channel complexes and suppressed the 'overshoot' in Kv4.3/KChIP2/MiRP1 channels. CONCLUSIONS AND IMPLICATIONS: Flecainide directly binds to the Kv4.3 alpha-subunit when the channels are in the open and inactivated state and the presence of the beta-subunits modulates the blockade by altering the gating function.

Pathology-specific effects of the IKur/Ito/IK,ACh blocker AVE0118 on ion channels in human chronic atrial fibrillation.

BACKGROUND AND PURPOSE: This study was designed to establish the pathology-specific inhibitory effects of the IKur/Ito/IK,ACh blocker AVE0118 on atrium-selective channels and its corresponding effects on action potential shape and effective refractory period in patients with chronic AF (cAF). EXPERIMENTAL APPROACH: Outward K+-currents of right atrial myocytes and action potentials of atrial trabeculae were measured with whole-cell voltage clamp and microelectrode techniques, respectively. Outward currents were dissected by curve fitting. KEY RESULTS: Four components of outward K+-currents and AF-specific alterations in their properties were identified. Ito was smaller in cAF than in SR, and AVE0118 (10 microM) apparently accelerated its inactivation in both groups without reducing its amplitude. Amplitudes of rapidly and slowly inactivating components of IKur were lower in cAF than in SR. The former was abolished by AVE0118 in both groups, the latter was partially blocked in SR, but not in cAF, even though its inactivation was apparently accelerated in cAF. The large non-inactivating current component was similar in magnitude in both groups, but decreased by AVE0118 only in SR. AVE0118 strongly suppressed AF-related constitutively active IK,ACh and prolonged atrial action potential and effective refractory period exclusively in cAF. CONCLUSIONS AND IMPLICATIONS: In atrial myocytes of cAF patients, we detected reduced function of distinct IKur components that possessed decreased component-specific sensitivity to AVE0118 most likely as a consequence of AF-induced electrical remodelling. Inhibition of profibrillatory constitutively active IK,ACh may lead to pathology-specific efficacy of AVE0118 that is likely to contribute to its ability to convert AF into SR.

Receptor-independent sensitization of the adenylyl cylase after chronic treatment with cyclosporine A.

Chronic treatment with cyclosporine A (CyA) is often complicated by severe hypertension. If activation of the beta-adrenergic-receptor-linked adenylyl cyclase (AC) system contributes to hypertension is unresolved. Rats were treated with CyA (20 mg kg(-1) day(-1)) for 7 days. beta-adrenergic, muscarinic, and alpha-adrenergic receptors, G-proteins, and the activity of AC were determined in cardiac and pulmonary plasma membranes. The density of cardiac beta-adrenergic receptors, muscarinic receptors, alpha-adrenergic receptors, G(alphas) and, G(alphai) remained unchanged after treatment with CyA. However, CyA increased the responsiveness of AC to different stimulators. The responsiveness of AC was even more pronounced after solubilization and partial purification, suggesting a direct modulation of the enzyme. These data suggest that CyA modulates the activity of the sympathoadrenergic system by a direct, receptor-independent sensitization of AC, suggesting that this pathway contributes to hypertension in patients treated with CyA.

The function of alpha- and beta-adrenoceptors of the saphenous artery in caveolin-1 knockout and wild-type mice.

BACKGROUND AND PURPOSE: Adrenoceptors can associate with cardiac caveolae. To investigate the function of vascular caveolae, adrenoceptor-mediated effects were compared in the saphenous artery of caveolin-1 knockout (cav-1KO) and wild-type (WT) mice. EXPERIMENTAL APPROACH: Electronmicroscopy was used to detect caveolae. Real-Time quantitative PCR was used for adrenoceptor subtypes. Catecholamine-evoked contractions and relaxations were studied in arterial segments. KEY RESULTS: Caveolae were found in arterial smooth muscle from WT but not from cav-1KO mice. Arterial mRNA levels for the adrenoceptors alpha1A, alpha1B, alpha1D, beta1, beta2 and beta3 were similar in cav-1KO and WT. (-)-Noradrenaline contracted cav-1KO (-log EC50M=7.1) and WT (-log EC50M=7.3) arteries through prazosin-sensitive receptors. Maximum (-)-noradrenaline-evoked contractions were greater in cav-1KO than WT arteries. (-)-Isoprenaline relaxed WT arteries (-log EC50M=7.3) more potently than cav-1KO arteries (-log EC50M=6.8); the effects were antagonized partially and similarly by the beta2-selective antagonist ICI118551 (50 nM). The (-)-isoprenaline-evoked relaxation was partially antagonized by the beta1-adrenoceptor-selective antagonist CGP20712 (300 nM) in WT but not cav-1KO arteries. The beta3-adrenoceptor-selective antagonist L748337 (100 nM) partially antagonized the relaxant effects of (-)-isoprenaline in cav-1KO but not in WT arteries. BRL37344 partially relaxed arteries through beta3-adrenoceptors in cav-1KO but not WT. The relaxant effects of BRL37344 were decreased by the NO synthase inhibitor OmegaL-nitroarginine. CONCLUSIONS AND IMPLICATIONS: The function of arterial alpha1- and beta2-adrenoceptors is similar in cav-1KO and WT mice. beta1-adrenoceptor-mediated relaxation in WT is lost in cav-1KO and replaced by the appearance of beta3-adrenoceptors.

The G protein-gated potassium current I(K,ACh) is constitutively active in patients with chronic atrial fibrillation.

BACKGROUND: The molecular mechanism of increased background inward rectifier current (IK1) in atrial fibrillation (AF) is not fully understood. We tested whether constitutively active acetylcholine (ACh)-activated I(K,ACh) contributes to enhanced basal conductance in chronic AF (cAF). METHODS AND RESULTS: Whole-cell and single-channel currents were measured with standard voltage-clamp techniques in atrial myocytes from patients with sinus rhythm (SR) and cAF. The selective I(K,ACh) blocker tertiapin was used for inhibition of I(K,ACh). Whole-cell basal current was larger in cAF than in SR, whereas carbachol (CCh)-activated I(K,ACh) was lower in cAF than in SR. Tertiapin (0.1 to 100 nmol/L) reduced I(K,ACh) in a concentration-dependent manner with greater potency in cAF than in SR (-logIC50: 9.1 versus 8.2; P<0.05). Basal current contained a tertiapin-sensitive component that was larger in cAF than in SR (tertiapin [10 nmol/L]-sensitive current at -100 mV: cAF, -6.7+/-1.2 pA/pF, n=16/5 [myocytes/patients] versus SR, -1.7+/-0.5 pA/pF, n=24/8), suggesting contribution of constitutively active I(K,ACh) to basal current. In single-channel recordings, constitutively active I(K,ACh) was prominent in cAF but not in SR (channel open probability: cAF, 5.4+/-0.7%, n=19/9 versus SR, 0.1+/-0.05%, n=16/9; P<0.05). Moreover, IK1 channel open probability was higher in cAF than in SR (13.4+/-0.4%, n=19/9 versus 11.4+/-0.7%, n=16/9; P<0.05) without changes in other channel characteristics. CONCLUSIONS: Our results demonstrate that larger basal inward rectifier K+ current in cAF consists of increased IK1 activity and constitutively active I(K,ACh). Blockade of I(K,ACh) may represent a new therapeutic target in AF.

Cilostamide potentiates more the positive inotropic effects of (-)-adrenaline through beta(2)-adrenoceptors than the effects of (-)-noradrenaline through beta (1)-adrenoceptors in human atrial myocardium.

Activation of both beta(1)- and beta(2)-adrenoceptors increases the contractility of human atrial myocardium through cyclic AMP-dependent pathways. Cyclic AMP is hydrolised by phosphodiesterases, but little is known about which isoenzymes catalyse inotropically relevant cyclic AMP accumulated upon stimulation of beta-adrenoceptor subtypes. We have compared the positive inotropic effects of (-)-noradrenaline and (-)-adrenaline, mediated through beta(1)- and beta(2)-adrenoceptors, respectively, in the absence and presence of the PDE3 inhibitor cilostamide (300 nM) or PDE4 inhibitor rolipram (1 muM) on human atrial trabeculae from non-failing hearts. Cilostamide, but not rolipram, potentiated the effects of both (-)-noradrenaline and (-)-adrenaline. Cilostamide increased the -logEC(50)M of (-)-adrenaline more than of (-)-noradrenaline (P < 0.05), regardless of whether or not the patients had been chronically treated with beta-blockers. The results are consistent with a greater PDE3-catalysed hydrolysis of inotropically relevant cyclic AMP produced through beta(2)-adrenoceptors than beta(1)-adrenoceptors in human atrium.

[New antiarrhythmic drugs for therapy of atrial fibrillation: I. Ion channel blockers]. / Neue Antiarrhythmika in der Therapie des Vorhofflimmerns--I. Ionenkanalblocker.

During the last ten years we have made substantial progress in our understanding of the underlying mechanisms of atrial fibrillation. The high rate associated alterations in electrical and structural properties of the atria, referred to as atrial remodeling, promote the progression of atrial fibrillation. The development of new therapeutic approaches addresses three different directions: (i) prevention of atrial remodeling, especially of structural remodeling; (ii) increase of long-term efficacy of currently used drugs and improvement of their side-effect profile; and (iii) design of atria- and pathology-specific antiarrhythmic drugs without concomitant proarrhythmic effects in the ventricles. The current review outlines the pathophysiology of atrial fibrillation and focuses on electrical remodeling. The properties of new antiarrhythmic drugs for atrial fibrillation are discussed in detail.

An update on atrial fibrillation in 2014: From pathophysiology to treatment.

Atrial fibrillation (AF) is the most frequently encountered cardiac arrhythmia. The trigger for initiation of AF is generally an enhanced vulnerability of pulmonary vein cardiomyocyte sleeves to either focal or re-entrant activity. The maintenance of AF is based on a "driver" mechanism in a vulnerable substrate. Cardiac mapping technology is providing further insight into these extremely dynamic processes. AF can lead to electrophysiological and structural remodelling, thereby promoting the condition. The management includes prevention of stroke by oral anticoagulation or left atrial appendage (LAA) occlusion, upstream therapy of concomitant conditions, and symptomatic improvement using rate control and/or rhythm control. Nonpharmacological strategies include electrical cardioversion and catheter ablation. There are substantial geographical variations in the management of AF, though European data indicate that 80% of patients receive adequate anticoagulation and 79% adequate rate control. High rates of morbidity and mortality weigh against perceived difficulties in management. Clinical research and growing experience are helping refine clinical indications and provide better technical approaches. Active research in cardiac electrophysiology is producing new antiarrhythmic agents that are reaching the experimental clinical arena, inhibiting novel ion channels. Future research should give better understanding of the underlying aetiology of AF and identification of drug targets, to help the move toward patient-specific therapy.

G-Protein beta(3)-subunit 825T allele is associated with enhanced human atrial inward rectifier potassium currents.

BACKGROUND: A C825T polymorphism was recently identified in the human gene encoding for the beta(3)-subunit of heterotrimeric G proteins. The 825T allele is associated with a splice variant of Gbeta(3) and enhanced signal transduction. We hypothesized that patients carrying the 825T allele exhibit the modified Gbeta(3) phenotype. The resulting enhancement of signal transduction should be detectable in the Gbetagamma-dimer-mediated acetylcholine-stimulated K(+) current (I(K,ACh)). METHODS AND RESULTS: Seventy patients undergoing cardiac surgery were genotyped for the C825T polymorphism. In right atrial myocytes from these patients, the inward rectifier K(+) currents (I(K1), I(K,ACh)) were studied with the whole-cell patch-clamp technique. Background current I(K1) was measured with depolarizing ramp pulses and quantified as inward current at -100 mV; mean amplitudes were (pA/pF) 4.98+/-0.49 (n=30/93 patients/cells) in patients with CC genotype, 4.25+/-0.36 (n=31/121 patients/cells) with TC, and 7. 46+/-1.14 (n=9/32 patients/cells; P<0.05) with TT. Conversely, mean I(K,ACh), which is maximally activated by carbachol (2 micromol/L), was reduced in patients with TT genotype (pA/pF, 4.30+/-1.33, n=9/27 patients/cells; P<0.05) compared with the other 2 groups (6.56+/-0. 54, n=30/80 and 6.16+/-0.45, n=31/117 patients/cells, for CC and TC genotype, respectively). Essentially similar results were obtained with adenosine (1 mmol/L). CONCLUSIONS: We found an association between the Gbeta(3) 825T allele and amplitude of human atrial I(K1) and I(K,ACh). Increased background current density in TT carriers could shorten action potential duration and may be due to I(K,ACh) being constitutively active in this genotype.

L-type Ca2+ current downregulation in chronic human atrial fibrillation is associated with increased activity of protein phosphatases.

BACKGROUND: Although downregulation of L-type Ca2+ current (I(Ca,L)) in chronic atrial fibrillation (AF) is an important determinant of electrical remodeling, the molecular mechanisms are not fully understood. Here, we tested whether reduced I(Ca,L) in AF is associated with alterations in phosphorylation-dependent channel regulation. METHODS AND RESULTS: We used whole-cell voltage-clamp technique and biochemical assays to study regulation and expression of I(Ca,L) in myocytes and atrial tissue from 148 patients with sinus rhythm (SR) and chronic AF. Basal I(Ca,L) at +10 mV was smaller in AF than in SR (-3.8+/-0.3 pA/pF, n=138/37 [myocytes/patients] and -7.6+/-0.4 pA/pF, n=276/86, respectively; P<0.001), though protein levels of the pore-forming alpha1c and regulatory beta2a channel subunits were not different. In both groups, norepinephrine (0.01 to 10 micromol/L) increased I(Ca,L) with a similar maximum effect and comparable potency. Selective blockers of kinases revealed that basal I(Ca,L) was enhanced by Ca2+/calmodulin-dependent protein kinase II in SR but not in AF. Norepinephrine-activated I(Ca,L) was larger with protein kinase C block in SR only, suggesting decreased channel phosphorylation in AF. The type 1 and type 2A phosphatase inhibitor okadaic acid increased basal I(Ca,L) more effectively in AF than in SR, which was compatible with increased type 2A phosphatase but not type 1 phosphatase protein expression and higher phosphatase activity in AF. CONCLUSIONS: In AF, increased protein phosphatase activity contributes to impaired basal I(Ca,L). We propose that protein phosphatases may be potential therapeutic targets for AF treatment.

Molecular basis of downregulation of G-protein-coupled inward rectifying K(+) current (I(K,ACh) in chronic human atrial fibrillation: decrease in GIRK4 mRNA correlates with reduced I(K,ACh) and muscarinic receptor-mediated shortening of action potentials.

BACKGROUND: Clinical and experimental evidence suggest that the parasympathetic nervous system is involved in the pathogenesis of atrial fibrillation (AF). However, it is unclear whether changes in G-protein-coupled inward rectifying K(+) current (I(K,ACh)) contribute to chronic AF. METHODS AND RESULTS: In the present study, we used electrophysiological recordings and competitive reverse-transcription polymerase chain reaction to study changes in I(K,ACh) and the level of the I(K,ACh) GIRK4 subunit in isolated human atrial myocytes and the atrial tissue of 39 patients with sinus rhythm and 24 patients with chronic AF. The density of I(K,ACh) was approximately 50% smaller in myocytes from patients with AF compared with those in sinus rhythm, and this was accompanied by decreased levels of GIRK4 mRNA. The current density of the inward rectifying K(+) current (I(K1)) was 2-fold larger during AF than in sinus rhythm, in correspondence with an increase in Kir2.1 mRNA. The larger I(K1) in AF is consistent with more negative membrane potentials in right atrial trabeculae from AF patients. Moreover, action potential duration was reduced in AF, and the action potential shortening produced by muscarinic receptor stimulation was attenuated, indicating that the changes of I(K1) and I(K,ACh) were functionally relevant. CONCLUSIONS: Chronic human AF induces transcriptionally mediated upregulation of I(K1) but downregulation of I(K,ACh) and attenuates the muscarinic receptor-mediated shortening of atrial action potentials. This suggests that atrial myocytes adapt to a chronically high rate by downregulating I(K,ACh) to counteract the shortening of the atrial effective refractory period due to electrical remodeling.

Characterization of excitation-contraction coupling in conscious dogs with pacing-induced heart failure.

OBJECTIVE: In isolated cardiac preparations of non-failing hearts from different species, including man, there is a positive force-frequency relation which is reversed into a negative relation in preparation from failing hearts. Whether or not such relations between ventricular function and heart rate hold true in the in situ heart is not clear at present. Mechanical restitution and postextrasystolic potentiation might serve as alternative measures of excitation-contraction coupling. METHODS: Eleven dogs were instrumented with a left ventricular micromanometer, ultrasonic crystals for the measurement of regional wall thickness, two hydraulic occluders around the descending aorta and the inferior caval vein, and left atrial and ventricular pacing leads with a subcutaneous pacemaker. Left ventricular dP/dtmax, as an isovolumic phase index, and systolic wall thickening, as an ejection phase index, were plotted versus heart rate, and heart rate was increased by left atrial pacing from rest to 200 min-1 in increments of 25 min-1. In a subset of dogs, left ventricular filling was controlled and the frequency range expanded by the bradycardic agent UL-FS 49. Measurements were performed in the presence and absence of autonomic blockade (hexamethonium, atropine). Mechanical restitution and postextrasystolic potentiation were determined as normalized dP/dtmax and systolic wall thickening, respectively, of the extra- and postextrasystolic beat versus defined variations of the extrasystolic time interval (250-550 ms). Following control studies, heart failure was induced by rapid left ventricular pacing at 250 min-1 for 20 days +/- 6 (SD) and measurements repeated. Isolated left ventricular trabeculae from non-failing and failing hearts were studied during stimulation at 0.2-4 Hz. RESULTS: Only with filling control and in the absence of autonomic blockade, was there a slightly positive relation between dP/dtmax and heart rate in the control state. Otherwise, the relation of dP/dtmax to heart rate was flat both in the control state and in heart failure. The relation between systolic wall thickening and heart rate in the control state was negative, unless filling was controlled, and it was flat in heart failure. In contrast, the time constants of mechanical restitution and postextrasystolic potentiation were increased significantly with heart failure from 91 +/- 25 (SD) to 164 +/- 13 ms and from 107 +/- 18 to 156 +/- 4 ms, respectively, for dP/dtmax and from 76 +/- 22 to 162 +/- 10 ms and from 101 +/- 17 to 160 +/- 17 ms, respectively, for systolic wall thickening. These time constants were, however, insensitive to UL-FS 49 and autonomic blockade. There was a negative force-frequency relation in left ventricular trabeculae from non-failing hearts at higher calcium concentrations, where it was flat in trabeculae from failing hearts. CONCLUSION: Time constants of mechanical restitution and postextrasystolic potentiation are more sensitive than the steady state relation of ventricular function and heart rate to characterize the impairment of excitation-contraction coupling in heart failure.

Differential effects of the new class III antiarrhythmic agents almokalant, E-4031 and D-sotalol, and of quinidine, on delayed rectifier currents in guinea pig ventricular myocytes.

OBJECTIVES: The effects of almokalant (4-[3-ethyl[3-(propylsulphinyl)propyl]-amino]-2-hydroxy-propoxy]- benzonitrile), E-4031 (1-[2-(6-methyl-2-pyridyl)-ethyl]-4-(4-methylsulphonyl-amino- benzoyl)piperidine), d-sotalol, and quinidine were investigated on the delayed K+ rectifier current IK. The aim of the study was to compare the drug action on the two components of this current. METHODS: Membrane currents were measured in ventricular myocytes from guinea pig hearts with the whole cell voltage clamp technique. IK was activated during clamp steps from a holding potential of -40 mV to test potentials -30 and +50 mV. The tail current Itail was measured upon stepping back to holding potential. RESULTS: In control experiments. IK and Itail declined spontaneously ("run down"). With 300 ms long test pulses to +50 mV, only d-sotalol (10(-4) M) caused a significant further decrease in IK, whereas all four agents significantly reduced Itail (almokalant 10(-6) M, E-4031 10(-7) M, quinidine 10(-5) M). When tested with 1 s long clamp steps at various potentials almokalant (3 x 10(-6) M), E-4031 (10(-6) M), quinidine (10(-5) M), and d-sotalol (10(-4) M) reduced IK in the potential range between -20 and +40 mV, yielding a bell shaped inward rectifying drug sensitive current. Itail was reduced by almokalant and E-4031 over the whole voltage range with saturation of block positive to +20 mV. Similar reductions with quinidine but not with d-sotalol were also significant. With rest pulses to +50 mV of increasing duration (25 ms-4000 ms), Itail developed with a faster time course than IK and therefore the ratio of Itail/IK declined with pulse duration. With almokalant and E-4031, this ratio became independent of test pulse duration. For 250 ms pulses, Itail/IK was also significantly reduced by d-sotalol and quinidine. CONCLUSION: Inhibition of the rapidly activating inwardly rectifying component of IK is prominent with almokalant and E-4031 and less pronounced with d-sotalol and quinidine. Since inhibition of this component prolongs the cardiac action potential, it should contribute to the antiarrhythmic properties of the agents.

Mechanisms of excitation-contraction coupling studied using the principle of transient perturbation.

We have studied the responses to a brief interruption of a train of steady state beats, namely: (1) a single prolonged depolarisation within the train; (2) a single short interval within the train; (3) a single long interval within the train. These responses are predicted by a two compartment model of intracellular calcium handling. They are characterised by the following phenomena. (1) Prolongation of one depolarisation/action potential in the steady state train causes potentiation of the following beat. We postulate on the basis of the published evidence that this may be due to "reversed" sodium/calcium exchange during late systole leading to extra calcium entry during the prolonged depolarisation. (2) Postextrasystole potentiation is postulated to share this mechanism when a depolarisation (extrasystole) is introduced immediately after one of the steady state depolarisations (single short interval). The postextrasystolic beat is then potentiated. (3) A single short interval during the steady state train also leads to attenuation of contractile force on the beat immediately after the short interval, that is, the extrasystole. Mechanical restitution is the term applied to the recovery of this force with increasing interval. This consists of two phases. The initial rapid phase is ryanodine and caffeine insensitive, indicating possible independence of sarcoplasmic reticular function. We postulate that a "membrane compartment" of internal calcium may be responsible. The second, slower, phase of mechanical restitution is ryanodine and caffeine sensitive, indicating that it is likely to be a property of the sarcoplasmic reticulum.(ABSTRACT TRUNCATED AT 250 WORDS)

Transient outward current in human and rat ventricular myocytes.

OBJECTIVE: The aim was to investigate transient outward currents (I(to)) in single myocytes isolated from human heart muscle specimens which were obtained either from patients in terminal heart failure receiving a transplant or from multiorgan donors whose hearts were not suitable for transplantation. METHODS: Using the whole cell patch clamp technique, depolarisation dependent I(to) was investigated in these myocytes, and its electrophysiological characteristics compared to I(to) of rat myocytes. RESULTS: I(to) was observed in ventricular myocytes isolated from failing and non-failing human hearts. The current density of I(to) was similar in cells from failing and non-failing hearts [at +60 mV: 7.9(SEM 1.0) pA.pF-1, n = 9, and 8.7(1.2) pA.pF-1, n = 8, respectively], but smaller in human than in normal rat myocytes, ie, 8.2(0.7) pA.pF-1 (n = 17) v 19.9(2.8) pA.pF-1 (n = 12, six hearts), respectively. Half maximum activation was found at more positive potentials in human than in rat cells, at +21.2(2.0) v +6.4(1.3) mV. In human myocytes, the fraction of non-inactivating outward current at the end of 300 ms long clamp steps was smaller than in rat cells, ie, 22(5%) of peak I(to) in human (n = 17) and 39(5%) in rat cells (n = 12). The potential of half maximum steady state inactivation of rapidly inactivating I(to) in the presence of 0.1 mM Cd2+ was -21.4(0.7) mV in human (n = 15, five hearts), and -35.3(1.0) mV in rat cells (n = 12, six hearts). The late component of outward current showed no potential dependent inactivation in human cells, but underwent steady state inactivation at all potentials positive to -100 mV in rat myocytes. At -100 mV, recovery of I(to) from inactivation took place with a similar time constant, ie, 18(2) ms (n = 7), 24(2) ms (n = 6), and 25(2) ms (n = 4) in cells from three failing and two non-failing human hearts, and from two normal rat hearts, respectively. CONCLUSIONS: In a limited number of cells, I(to) in human ventricular myocytes shows no dramatic differences between cells derived from failing and non-failing hearts. The characteristics of I(to) in human cells were similar though not identical to I(to) in rat heart cells. This current may be a potential target for antiarrhythmic drug action.

L-type calcium current and contractility in ventricular myocytes from mice overexpressing the cardiac beta 2-adrenoceptor.

OBJECTIVES: The reported increase in basal activity of hearts from transgenic mice (TG4) overexpressing the human beta 2-adrenoceptor (beta 2-AR) was explained by spontaneously active beta 2-ARs that stimulate the beta-adrenergic cascade in the absence of an agonist. In order to examine altered myocardial function on a cellular level, we have investigated L-type calcium current (ICa,L) and cell shortening in ventricular myocytes from TG4 hearts. Myocytes from littermates (LM) and wild type animals (WT) served as controls. METHODS: Cardiac beta-AR density was measured by [125I]-iodocyanopindolol binding to ventricular membranes. ICa,L was assessed by standard whole-cell voltage clamp technique. Contractility was measured as cell shortening in ventricular myocytes and as force of contraction in electrically stimulated left atria. RESULTS: Overexpression of beta 2-ARs was confirmed by an almost 400-fold increase in beta-AR density. The beta 1:beta 2-AR ratio in WT mice was 71:29. Myocytes from TG4 and LM mice were similar in size as judged by membrane capacitance and two dimensional cell area. ICa,L amplitude was significantly lower in TG4 than in LM myocytes (with 2 mM [Ca2+]o -4.82 +/- 0.48 vs. -6.56 +/- 0.38 pA/pF, respectively). In TG4 myocytes, the ICa,L response to isoproterenol (1 microM) was almost abolished. Cell shortening was not different in physiological [Ca2+]o, but smaller in maximum [Ca2+]o when comparing TG4 to control myocytes. Basal force of contraction in left atria did not differ between TG4 and LM at any age investigated. In TG4 left atria the inotropic response to isoproterenol was also absent, whereas responses to high [Ca2+]o or dibutyryl-cAMP (1 mM) were present but reduced. The rate of spontaneous beating of right atria was elevated in TG4 mice. CONCLUSIONS: Since only spontaneous beating rate but neither basal ICa,L amplitude nor basal contractile activity were elevated, our data fail to reveal evidence for spontaneously active, stimulating beta 2-ARs in left atrium and ventricle. A contractile deficit unrelated to the beta-adrenoceptor pathway is evident in TG4 myocytes and left atria.

Tachycardia-induced failure alters contractile properties of canine ventricular myocytes.

OBJECTIVE: Rapid cardiac pacing has been used as a model for experimentally-induced cardiomyopathy. However, its relevance to human heart failure is not clear at present because little is known about changes in size and function of ventricular myocytes. We have therefore studied the responses to graded increases in frequency and calcium in canine ventricular myocytes from failing hearts. The aim of our study was to evaluate the resemblance between canine pacing-induced and human end-stage heart failure. METHODS: Myocytes were isolated from the left ventricular wall of dogs that were in heart failure after 6 weeks of pacing at 250 beats/min. Cell shortening was measured by edge detection. RESULTS: Clinical signs of failure included dyspnea, ascites, and heart dilatation; the hemodynamic parameters were: LVdP/dtmax 1613 +/- 149 vs. 4713 +/- 304 mmHg/s in 6 control dogs; LVEDP 17.2 +/- 4.4 vs 5.6 +/- 1.1 mmHg; LV volume 60.5 +/- 6.2 vs. 30-35 ml. Myocytes from failing hearts were longer and thinner than those from controls (from factor: 0.40 +/- 0.01 vs. 0.47 +/- 0.01, P < 0.001, > 30 cells/heart). With 6 mM Ca2+ and at 0.5 Hz, contraction amplitude was significantly attenuated in myocytes from failing hearts: 6.6 +/- 0.9% cell shortening vs. 10.0 +/- 0.8% in controls (P < 0.05). This deficit was exacerbated at higher stimulation rates. Time-to-peak contraction and time-to-50% relaxation were not altered. There was no difference in sensitivity to thapsigargin. CONCLUSION: As with cells from human failing hearts, contraction amplitude showed rate-dependent depression in this animal model, whereas features like slowing of contraction and relaxation and reduced sensitivity to thapsigargin, were not reproduced.