The publication data currently available has been vetted by Vanderbilt faculty, staff, administrators and trainees. The data itself is retrieved directly from NCBI's PubMed and is automatically updated on a weekly basis to ensure accuracy and completeness.
If you have any questions or comments, please contact us.
Flecainide blocks ryanodine receptor type 2 (RyR2) channels in the open state, suppresses arrhythmogenic Ca2+ waves and prevents catecholaminergic polymorphic ventricular tachycardia (CPVT) in mice and humans. We hypothesized that differences in RyR2 activity induced by CPVT mutations determines the potency of open-state RyR2 blockers like flecainide (FLEC) and R-propafenone (RPROP) against Ca2+ waves in cardiomyocytes. Using confocal microscopy, we studied Ca2+ sparks and waves in isolated saponin-permeabilized ventricular myocytes from two CPVT mouse models (Casq2-/-, RyR2-R4496C+/-), wild-type (c57bl/6, WT) mice, and WT rabbits (New Zealand white rabbits). Consistent with increased RyR2 activity, Ca2+ spark and wave frequencies were significantly higher in CPVT compared to WT mouse myocytes. We next obtained concentration-response curves of Ca2+ wave inhibition for FLEC, RPROP (another open-state RyR2 blocker), and tetracaine (TET) (a state-independent RyR2 blocker). Both FLEC and RPROP inhibited Ca2+ waves with significantly higher potency (lower IC50) and efficacy in CPVT compared to WT. In contrast, TET had similar potency in all groups studied. Increasing RyR2 activity of permeabilized WT myocytes by exposure to caffeine (150 µM) increased the potency of FLEC and RPROP but not of TET. RPROP and FLEC were also significantly more potent in rabbit ventricular myocytes that intrinsically exhibit higher Ca2+ spark rates than WT mouse ventricular myocytes. In conclusion, RyR2 activity determines the potency of open-state blockers FLEC and RPROP for suppressing arrhythmogenic Ca2+ waves in cardiomyocytes, a mechanism likely relevant to antiarrhythmic drug efficacy in CPVT.
Catecholaminergic polymorphic ventricular tachycardia (CPVT) causes sudden cardiac death due to mutations in cardiac ryanodine receptors (RyR2), calsequestrin, or calmodulin. Flecainide, a class I antiarrhythmic drug, inhibits Na(+) and RyR2 channels and prevents CPVT. The purpose of this study is to identify inhibitory mechanisms of flecainide on RyR2. RyR2 were isolated from sheep heart, incorporated into lipid bilayers, and investigated by single-channel recording under various activating conditions, including the presence of cytoplasmic ATP (2 mM) and a range of cytoplasmic [Ca(2+)], [Mg(2+)], pH, and [caffeine]. Flecainide applied to either the cytoplasmic or luminal sides of the membrane inhibited RyR2 by two distinct modes: 1) a fast block consisting of brief substate and closed events with a mean duration of ∼1 ms, and 2) a slow block consisting of closed events with a mean duration of ∼1 second. Both inhibition modes were alleviated by increasing cytoplasmic pH from 7.4 to 9.5 but were unaffected by luminal pH. The slow block was potentiated in RyR2 channels that had relatively low open probability, whereas the fast block was unaffected by RyR2 activation. These results show that these two modes are independent mechanisms for RyR2 inhibition, both having a cytoplasmic site of action. The slow mode is a closed-channel block, whereas the fast mode blocks RyR2 in the open state. At diastolic cytoplasmic [Ca(2+)] (100 nM), flecainide possesses an additional inhibitory mechanism that reduces RyR2 burst duration. Hence, multiple modes of action underlie RyR2 inhibition by flecainide.
Copyright © 2014 by The American Society for Pharmacology and Experimental Therapeutics.
Ca(2+) waves can trigger ventricular arrhythmias such as catecholaminergic-polymorphic ventricular tachycardia (CPVT). Drugs that prevent Ca(2+) waves may have antiarrhythmic properties. Here, we use permeabilized ventricular myocytes from a CPVT mouse model lacking calsequestrin (casq2) to screen all clinically available class I antiarrhythmic drugs and selected other antiarrhythmic agents for activity against Ca(2+) waves. Casq2-/- myocytes were imaged in line-scan mode and the following Ca(2+) wave parameters analyzed: wave incidence, amplitude, frequency, and propagation speed. IC(50) (potency) and maximum inhibition (efficacy) were calculated for each drug. Drugs fell into 3 distinct categories. Category 1 drugs (flecainide and R-propafenone) suppressed wave parameters with the highest potency (IC(50)<10 μM) and efficacy (>50% maximum wave inhibition). Category 2 drugs (encainide, quinidine, lidocaine, and verapamil) had intermediate potency (IC(50) 20-40 μM) and efficacy (20-40% maximum wave inhibition). Category 3 drugs (procainamide, disopyramide, mexiletine, cibenzoline, and ranolazine) had no significant effects on Ca(2+) waves at the highest concentration tested (100 μM). Propafenone was stereoselective, with R-propafenone suppressing waves more potently than S-propafenone (IC(50): R-propafenone 2 ± 0.2 μM vs. S-propafenone 54 ± 18 μM). Both flecainide and R-propafenone decreased Ca(2+) spark mass and converted propagated Ca(2+) waves into non-propagated wavelets and frequent sparks, suggesting that reduction in spark mass, not spark frequency, was responsible for wave suppression. Among all class I antiarrhythmic drugs, flecainide and R-propafenone inhibit Ca(2+) waves with the highest potency and efficacy. Permeabilized casq2-/- myocytes are a simple in-vitro assay for finding drugs with activity against Ca(2+) waves. This article is part of a Special Issue entitled 'Possible Editorial'.
Copyright © 2011 Elsevier Ltd. All rights reserved.
Catecholaminergic polymorphic ventricular tachycardia (CPVT) is linked to mutations in the cardiac ryanodine receptor (RyR2) or calsequestrin. We recently found that the drug flecainide inhibits RyR2 channels and prevents CPVT in mice and humans. Here we compared the effects of flecainide and tetracaine, a known RyR2 inhibitor ineffective in CPVT myocytes, on arrhythmogenic Ca(2+) waves and elementary sarcoplasmic reticulum (SR) Ca(2+) release events, Ca(2+) sparks. In ventricular myocytes isolated from a CPVT mouse model, flecainide significantly reduced spark amplitude and spark width, resulting in a 40% reduction in spark mass. Surprisingly, flecainide significantly increased spark frequency. As a result, flecainide had no significant effect on spark-mediated SR Ca(2+) leak or SR Ca(2+) content. In contrast, tetracaine decreased spark frequency and spark-mediated SR Ca(2+) leak, resulting in a significantly increased SR Ca(2+) content. Measurements in permeabilized rat ventricular myocytes confirmed the different effects of flecainide and tetracaine on spark frequency and Ca(2+) waves. In lipid bilayers, flecainide inhibited RyR2 channels by open state block, whereas tetracaine primarily prolonged RyR2 closed times. The differential effects of flecainide and tetracaine on sparks and RyR2 gating can explain why flecainide, unlike tetracaine, does not change the balance of SR Ca(2+) fluxes. We suggest that the smaller spark mass contributes to flecainide's antiarrhythmic action by reducing the probability of saltatory wave propagation between adjacent Ca(2+) release units. Our results indicate that inhibition of the RyR2 open state provides a new therapeutic strategy to prevent diastolic Ca(2+) waves resulting in triggered arrhythmias, such as CPVT.
Copyright 2009 Elsevier Ltd. All rights reserved.
BACKGROUND - Variable function and expression of drug transporters have been proposed as mechanisms contributing to variable response to drug therapy. Block of the HERG channel, encoding IKr, can lead to serious arrhythmias, and a key drug-blocking site in HERG has been identified on the intracellular face of the pore. We begin to advance the hypothesis that active drug uptake enhances IKr block.
METHODS AND RESULTS - Reverse transcriptase-polymerase chain reaction identified expression in the human atrium and ventricle of 14 of 31 candidate drug uptake and efflux transporters, including OCTN1 (SLC22A4), a known uptake transporter of the HERG channel blocker quinidine. In situ hybridization and immunostaining localized OCTN1 expression to cardiomyocytes. The IC50 for quinidine block of IKr in CHO cells transfected with HERG alone was significantly higher than cells transfected with HERG + OCTN1 (0.66 +/- 0.15 microM versus 0.14 +/- 0.06 microM [52% absolute increase in drug block; 95% confidence interval, 0.4-0.64 microM]), and this effect was further potentiated by a common genetic variant of OCTN1, L503F. In the absence of OCTN1, quinidine block could be 91% +/- 5% washed out, but with the transporter, washout was incomplete (57% +/- 6%). OCTN1 coexpression also facilitated HERG block by flecainide and ibutilide, but not erythromycin.
CONCLUSIONS - Coexpression of the organic cation transporter, OCTN1, expressed in human cardiac myocytes, intensifies quinidine-induced HERG block. These findings establish a critical hypothesis that variable drug transporter activity may be a potential risk factor for torsade de pointes.
Catecholaminergic polymorphic ventricular tachycardia (CPVT) is a potentially lethal inherited arrhythmia syndrome in which drug therapy is often ineffective. We discovered that flecainide prevents arrhythmias in a mouse model of CPVT by inhibiting cardiac ryanodine receptor-mediated Ca(2+) release and thereby directly targeting the underlying molecular defect. Flecainide completely prevented CPVT in two human subjects who had remained highly symptomatic on conventional drug therapy, indicating that this currently available drug is a promising mechanism-based therapy for CPVT.
Sudden cardiac death attributable to ventricular tachycardia/fibrillation (VF) remains a catastrophic outcome of myocardial ischemia and infarction. At the same time, conventional antagonist drugs targeting ion channels have yielded poor survival benefits. Although pharmacological and genetic models suggest an association between sodium (Na+) channel loss-of-function and sudden cardiac death, molecular mechanisms have not been identified that convincingly link ischemia to Na+ channel dysfunction and ventricular arrhythmias. Because ischemia can evoke the generation of reactive oxygen species, we explored the effect of oxidative stress on Na+ channel function. We show here that oxidative stress reduces Na+ channel availability. Both the general oxidant tert-butyl-hydroperoxide and a specific, highly reactive product of the isoprostane pathway of lipid peroxidation, E2-isoketal, potentiate inactivation of cardiac Na+ channels in human embryonic kidney (HEK)-293 cells and cultured atrial (HL-1) myocytes. Furthermore, E2-isoketals were generated in the epicardial border zone of the canine healing infarct, an arrhythmogenic focus where Na+ channels exhibit similar inactivation defects. In addition, we show synergistic functional effects of flecainide, a proarrhythmic Na+ channel blocker, and oxidative stress. These data suggest Na+ channel dysfunction evoked by lipid peroxidation is a candidate mechanism for ischemia-related conduction abnormalities and arrhythmias.