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Abnormal ryanodine receptor calcium release induces atrial fibrillation
ABSTRACTS / Journal of Molecular and Cellular Cardiology 42 (2007) S1–S23
drug-induced long QT syndrome. Ranolazine, an anti-ischemic agent inhibits IHERG, and causes a small QT interval prolongation. We investigated the kinetics of ranolazine block of IHERG at 23 °C using voltage-clamp analysis of HERG channels expressed in HEK293 cells. Block of IHERG by ranolazine was reversible and voltage-dependent, but frequency-independent. Block developed rapidly following channel activation, suggesting state-dependence. At 0 mV, the time constants for development of block were 76.6 ± 1.6, 35.8 ± 2.4, and 19.4 ± 1.7 ms with 10, 30 and 100 μM ranolazine (n = 4), respectively. The time course of ranolazine-induced IHERG decay was used to estimate the apparent dissociation constant (14.2 μM). Following repolarization, ranolazine-induced block of IHERG reversed rapidly. At − 80 and − 100 mV, recovery from block followed a monophasic time course with τ values of 204.3 ± 51.5 and 155.0 ± 31.9 ms (n = 5), respectively. Intracellular but not extracellular application of a membrane-impermeable (permanently charged) ranolazine analog caused rapid block of IHERG. Ranolazine antagonized E-4031 block of IHERG, suggesting that both compounds compete for a common binding site. Taken together, the unique ultra-rapid kinetics of block (at positive potentials) and unblock (upon hyperpolarization) of IHERG by ranolazine may explain the observations that: (1) ranolazine causes minimal QT interval prolongation with no reverse use dependence, and (2) pro-arrhythmic events have not been observed during exposures of cardiac myocytes or whole hearts to ranolazine. Keywords: Ranolazine; HERG; Arrhythmia doi:10.1016/j.yjmcc.2007.03.053
Effects of changes in unitary Ca2+ current on Ca2+-induced Ca2+-release from the sarcoplasmic reticulum Moutaz Elkadri, Eric Dubuis, George Hart, Munir Hussain. School of Clinical Sciences, University of Liverpool, L69 3GA, UK L-type Ca2+ current (ICa) is thought to be the main trigger for Ca -induced Ca 2+ -release (CICR) from the sarcoplasmic reticulum (SR) in cardiac muscle. In this study we have used a TTL-controlled piezoelectric device to rapidly switch external solutions to investigate the effects of altering the unitary current through the L-type Ca2+ channel without changing the Ca2+ content of the SR. Myocytes were voltage clamped using amphotericin B and stimulated with 5 conditioning pulses from − 40 mV to 0 mV (0.5 Hz, 35 °C) before applying a test pulse to different potentials. Two-channel theta tubing was used to rapidly switch between control (1.0 mM Ca2+ ) and test solutions, containing either 5.0 or 0.2 mM [Ca2+], immediately after the last conditioning pulse. Increasing [Ca2+]o from 1.0 to 5.0 mM significantly increased ICa amplitude at all the potentials tested e.g. at − 40 to 0 mV, ICa increased to 136 ± 6% (n = 10) following the switch to 5.0 mM [Ca2+]o and 2+
S19
returned to 106 ± 2% of the pre-switch control amplitude upon returning to 1.0 mM [Ca2+]o. However, SR Ca2+ release was not increased at 0 mV but was greater at most of the other potentials tested. Similarly, rapidly lowering [Ca2+]o to 0.2 mM decreased ICa amplitude by 34 ± 5% (n = 10) but had no effect on SR Ca2+ release and only modestly decreased SR Ca2+ release at potentials between 20 and 50 mV. Data show that CICR can be modulated by rapid changes in unitary ICa and that this effect is more apparent at positive potentials, possibly because more channels are open due to a higher open probability at these voltages. Keywords: Sarcoplasmic reticulum; Calcium-induced calcium release; Calcium current doi:10.1016/j.yjmcc.2007.03.054
Abnormal ryanodine receptor calcium release induces atrial fibrillation Subeena Sood, Marco Santonastasi, Haiyun Cheng, Xander H. Wehrens. Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, TX, USA Abnormal calcium release from the sarcoplasmic reticulum (SR) may contribute to arrhythmogenesis in atrial fibrillation (AF). Decreased binding of the ryanodine receptor (RyR2)stabilizing protein FKBP12.6 (calstabin2) has been reported in right atrial appendages of patients in chronic AF. Aim of this study was to examine whether inhibition of SR Ca2+ leak through RyR2 in FKBP12.6-deficient (−/−) mice decreases the vulnerability to AF. Surface and intracardiac electrophysiology studies were performed in wildtype (WT; n = 20) and FKBP12.6−/− (n = 16) mice. Right atrial programmed stimulation was performed before and after injection of 0.5 mg/kg tetracaine (i.p.). Under baseline conditions, there were no differences in electrophysiological (EP) parameters comparing WT and FKBP12.6−/− mice. AF was observed in 60% of FKBP12.6−/− mice compared with 5% of WT mice (P < 0.05). Whereas tetracaine did not significantly alter baseline EP parameters in WT or FKBP12.6−/− mice, tetracaine prevented AF in most FKBP12.6−/− mice vulnerable to AF before treatment. The results of this study show an increased susceptibility to AF in FKBP12.6−/− mice, which suggests that intracellular Ca2+ leak through abnormal ryanodine receptors may initiate or sustain atrial arrhythmias. Treatment with tetracaine to reduce SR Ca2+ leak decreased the vulnerability to AF. These findings suggest prevention of SR Ca2+ leak through RyR2 or enhancing FKBP12.6 binding to RyR2 may constitute a novel pharmacological agent for the treatment of atrial fibrillation. Keywords: Arrhythmias; Calcium handling doi:10.1016/j.yjmcc.2007.03.055
drug-induced long QT syndrome. Ranolazine, an anti-ischemic agent inhibits IHERG, and causes a small QT interval prolongation. We investigated the kinetics of ranolazine block of IHERG at 23 °C using voltage-clamp analysis of HERG channels expressed in HEK293 cells. Block of IHERG by ranolazine was reversible and voltage-dependent, but frequency-independent. Block developed rapidly following channel activation, suggesting state-dependence. At 0 mV, the time constants for development of block were 76.6 ± 1.6, 35.8 ± 2.4, and 19.4 ± 1.7 ms with 10, 30 and 100 μM ranolazine (n = 4), respectively. The time course of ranolazine-induced IHERG decay was used to estimate the apparent dissociation constant (14.2 μM). Following repolarization, ranolazine-induced block of IHERG reversed rapidly. At − 80 and − 100 mV, recovery from block followed a monophasic time course with τ values of 204.3 ± 51.5 and 155.0 ± 31.9 ms (n = 5), respectively. Intracellular but not extracellular application of a membrane-impermeable (permanently charged) ranolazine analog caused rapid block of IHERG. Ranolazine antagonized E-4031 block of IHERG, suggesting that both compounds compete for a common binding site. Taken together, the unique ultra-rapid kinetics of block (at positive potentials) and unblock (upon hyperpolarization) of IHERG by ranolazine may explain the observations that: (1) ranolazine causes minimal QT interval prolongation with no reverse use dependence, and (2) pro-arrhythmic events have not been observed during exposures of cardiac myocytes or whole hearts to ranolazine. Keywords: Ranolazine; HERG; Arrhythmia doi:10.1016/j.yjmcc.2007.03.053
Effects of changes in unitary Ca2+ current on Ca2+-induced Ca2+-release from the sarcoplasmic reticulum Moutaz Elkadri, Eric Dubuis, George Hart, Munir Hussain. School of Clinical Sciences, University of Liverpool, L69 3GA, UK L-type Ca2+ current (ICa) is thought to be the main trigger for Ca -induced Ca 2+ -release (CICR) from the sarcoplasmic reticulum (SR) in cardiac muscle. In this study we have used a TTL-controlled piezoelectric device to rapidly switch external solutions to investigate the effects of altering the unitary current through the L-type Ca2+ channel without changing the Ca2+ content of the SR. Myocytes were voltage clamped using amphotericin B and stimulated with 5 conditioning pulses from − 40 mV to 0 mV (0.5 Hz, 35 °C) before applying a test pulse to different potentials. Two-channel theta tubing was used to rapidly switch between control (1.0 mM Ca2+ ) and test solutions, containing either 5.0 or 0.2 mM [Ca2+], immediately after the last conditioning pulse. Increasing [Ca2+]o from 1.0 to 5.0 mM significantly increased ICa amplitude at all the potentials tested e.g. at − 40 to 0 mV, ICa increased to 136 ± 6% (n = 10) following the switch to 5.0 mM [Ca2+]o and 2+
S19
returned to 106 ± 2% of the pre-switch control amplitude upon returning to 1.0 mM [Ca2+]o. However, SR Ca2+ release was not increased at 0 mV but was greater at most of the other potentials tested. Similarly, rapidly lowering [Ca2+]o to 0.2 mM decreased ICa amplitude by 34 ± 5% (n = 10) but had no effect on SR Ca2+ release and only modestly decreased SR Ca2+ release at potentials between 20 and 50 mV. Data show that CICR can be modulated by rapid changes in unitary ICa and that this effect is more apparent at positive potentials, possibly because more channels are open due to a higher open probability at these voltages. Keywords: Sarcoplasmic reticulum; Calcium-induced calcium release; Calcium current doi:10.1016/j.yjmcc.2007.03.054
Abnormal ryanodine receptor calcium release induces atrial fibrillation Subeena Sood, Marco Santonastasi, Haiyun Cheng, Xander H. Wehrens. Department of Molecular Physiology and Biophysics, Baylor College of Medicine, Houston, TX, USA Abnormal calcium release from the sarcoplasmic reticulum (SR) may contribute to arrhythmogenesis in atrial fibrillation (AF). Decreased binding of the ryanodine receptor (RyR2)stabilizing protein FKBP12.6 (calstabin2) has been reported in right atrial appendages of patients in chronic AF. Aim of this study was to examine whether inhibition of SR Ca2+ leak through RyR2 in FKBP12.6-deficient (−/−) mice decreases the vulnerability to AF. Surface and intracardiac electrophysiology studies were performed in wildtype (WT; n = 20) and FKBP12.6−/− (n = 16) mice. Right atrial programmed stimulation was performed before and after injection of 0.5 mg/kg tetracaine (i.p.). Under baseline conditions, there were no differences in electrophysiological (EP) parameters comparing WT and FKBP12.6−/− mice. AF was observed in 60% of FKBP12.6−/− mice compared with 5% of WT mice (P < 0.05). Whereas tetracaine did not significantly alter baseline EP parameters in WT or FKBP12.6−/− mice, tetracaine prevented AF in most FKBP12.6−/− mice vulnerable to AF before treatment. The results of this study show an increased susceptibility to AF in FKBP12.6−/− mice, which suggests that intracellular Ca2+ leak through abnormal ryanodine receptors may initiate or sustain atrial arrhythmias. Treatment with tetracaine to reduce SR Ca2+ leak decreased the vulnerability to AF. These findings suggest prevention of SR Ca2+ leak through RyR2 or enhancing FKBP12.6 binding to RyR2 may constitute a novel pharmacological agent for the treatment of atrial fibrillation. Keywords: Arrhythmias; Calcium handling doi:10.1016/j.yjmcc.2007.03.055