阿魏酸钠与胺碘酮对家兔心室肌细胞离子通道电流作用的对比实验研究
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摘要
第一部分家兔耐钙心肌细胞急性分离及电生理记录的方法学探讨
目的:细胞标本制备的质量是膜片钳实验成功的关键,本研究探讨通过改变细胞外液成分获取适于电生理研究的耐钙心室肌细胞及L-型钙通道电流(I_(Ca-L))、延迟整流钾电流(I_K)记录的方法。
方法:家兔心室肌细胞急性分离采用Langendorff灌流装置及酶解消化分离技术,心脏灌流前实验组细胞外液为正常台氏液,对照组细胞外液为无钙台氏液,观察细胞外液中的钙离子对获取适于电生理记录的单个耐钙心肌细胞的影响;应用全细胞膜片钳技术,实验组分别以含BaCl_2的台氏液及N-甲基-D-葡萄糖胺(NMDG)溶液作为细胞外液记录I_(Ca-L)及I_K,对照组为正常台氏液。
结果:与对照组相比较,实验组心室肌细胞存活率和耐钙心肌细胞存活率均明显升高(71.0%±9.7% vs 54.5%±10.1%; 52.0%±7.9% vs 38.5%±10.8%, n=10, P<0.05)。实验组引出的I_(Ca-L)峰值电流大于对照组(15.2±7.1 pA/pF vs 7.4±2.4 pA/pF, n=10,P<0.05)。实验20 min与实验5 min记录的I_(Ca-L)峰值比较,对照组I_(Ca-L)峰值电流的变化率为36.1%±9.1% (n=10, P<0.05),即I_(Ca-L)峰值电流在实验20 min衰减;实验组I_(Ca-L)峰值电流的变化率为3.0%±4.7% (n=10, P>0.05),即I_(Ca-L)峰值电流幅值稳定。实验组I_K时间依赖性外向电流及尾电流随去极化时间延长增大作用更明显(1.39±0.05pA/pF vs 0.53±0.14 pA/pF, 0.74±0.09 pA/pF vs 0.39±0.13 pA/pF, n=10, P<0.05)。
结论:细胞外液中的钙离子有助于获取耐钙心肌细胞,以含BaCl_2的台氏液及NMDG溶液作为细胞外液易于获得典型而稳定的I_(Ca-L)及I_K。
第二部分阿魏酸钠与胺碘酮对家兔心室肌细胞L-型钙通道电流的影响
目的:探讨阿魏酸钠对家兔心室肌细胞膜L-型钙通道电流(I_(Ca-L))的影响。
方法:酶解法急性分离家兔单个心室肌细胞,以经典的Ⅲ类抗心律失常药物胺碘酮为对照,应用膜片钳全细胞记录技术观察3、10、30、100μmol/L的阿魏酸钠及1、3、10、30μmol/L的胺碘酮对心室肌细胞膜I_(Ca-L)的作用。
结果:阿魏酸钠及胺碘酮均呈浓度依赖性抑制L-型钙电流。3、10、30、100μmol/L的阿魏酸钠对I_(Ca-L)的抑制率分别为11.1%±2.4%、26.9%±6.2%、40.5%±5.0%、61.9%±5.5% (P<0.05);1、3、10、30μmol/L的胺碘酮对I_(Ca-L)的抑制率分别为21.1%±3.8%、32.6%±2.6%、52.6%±4.6%、71.4%±7.0% (P<0.05);半数抑制浓度(IC_(50))分别为32.6μmol/L及9.5μmol/L,阿魏酸钠的抑制作用弱于胺碘酮(P<0.05)。阿魏酸钠对I_(Ca-L)动力学的影响类似于胺碘酮,二者均能使I_(Ca-L)电流-电压曲线上移,未改变I-V曲线形状;使稳态激活曲线右移,失活曲线左移,使激活曲线与失活曲线之间相交的“窗流”区域缩小;并使I_(Ca-L)恢复曲线的t值增加,恢复曲线右移,即减慢钙通道灭活后的恢复过程。
结论:阿魏酸钠对I_(Ca-L)具有浓度依赖性阻滞作用,且使I_(Ca-L)的激活减慢、失活加快,并且失活后的恢复时间延长,阿魏酸钠的抑制作用虽弱于胺碘酮,但具有与胺碘酮相似的阻滞I_(Ca-L)且影响I_(Ca-L)的动力学特性,可能是其抗心律失常作用的电生理机制之一。
第三部分阿魏酸钠与胺碘酮对家兔心室肌细胞钾通道电流的影响
目的:探讨阿魏酸钠对家兔心室肌细胞膜延迟整流钾电流快速与缓慢激活成分(I_(Kr)、I_(Ks))、内向整流钾电流(I_(Kl))、瞬时外向钾电流(I_(to))的影响。
方法:酶解法急性分离家兔单个心室肌细胞,以经典的Ⅲ类抗心律失常药物胺碘酮为对照,采用膜片钳全细胞记录技术观察浓度为3、10、30、100μmol/L的阿魏酸钠对心室肌细胞膜I_(Kr)、I_(Ks)、I_(Kl)及I_(to)的作用。
结果:阿魏酸钠的作用弱于胺碘酮,二者均可浓度依赖性抑制I_(Kr)、I_(ks)时间依赖性外向电流及尾电流(I_(Kr,tail)、I_(Ks, tail)),3、10、30、100μmol/L的阿魏酸钠对I_(Kr, tail)的抑制率为12.1%±2.5%、24.1%±3.0%、47.0%±5.8%及58.5%±8.3%(n=5,P<0.05);对I_(Ks, tail)的抑制率为15.6%±6.4%、27.1%±6.5%、45.6%±5.7%及51.8%±6.6%(n=5,P<0.05),其对I_(Kr, tail)及I_(Ks, tail)的半数抑制浓度(IC_(50))均大于胺碘酮(43.6 vs 3.48μmol/L,44.8 vs 5.11μmol/L)。30、100μmol/L的阿魏酸钠及10、30μmol/L的胺碘酮可使I_(kl)的I-V曲线左移,在-100 mV及-20 mV测试电压下,浓度为100μmol/L的阿魏酸钠对I_(kl)内向电流及外向电流抑制率小于30μmol/L的胺碘酮(24.3%±8.8% vs 29.7%±8.0%; 17.7%±3.3% vs 20.1%±10.6%, n=5, P<0.05)。阿魏酸钠与胺碘酮均不影响I_(to)及其I-V曲线。
结论:阿魏酸钠对复极期钾电流具有阻滞作用,可能是其抗心律失常作用的电生理机制之一。
第四部分阿魏酸钠与胺碘酮对家兔心室肌细胞钠通道电流的影响
目的:探讨阿魏酸钠对家兔心室肌细胞膜钠通道电流(I_(Na))的影响。
方法:酶解法急性分离家兔单个心室肌细胞,以经典的Ⅲ类抗心律失常药物胺碘酮为对照,采用膜片钳全细胞记录技术观察浓度为100μmol/L的阿魏酸钠及10μmol/L胺碘酮对心室肌细胞膜I_(Na)的作用。
结果:10μmol/L胺碘酮使I_(Na)峰值电流由20.9±3.63 pA/pF减为12.0±2.69 pA/pF(n=5, P<0.05),抑制率为42.6%±4.87%;胺碘酮使I_(Na)的I-V曲线上移。100μmol/L阿魏酸钠不改变I_(Na)峰值电流(16.6±4.52 pA/pF vs 15.8±3.21 pA/pF,抑制率为6.12%±5.41%, n=5, P>0.05),且不影响I-V曲线。
结论:阿魏酸钠急性灌流不影响家兔心室肌细胞I_(Na)。
Part one Investigation of the methods for acute isolatingcalcium-tolerant ventricular myocytes andelectrophysiolological recordings
Objective:It is important to obtain viable cells for long period of electrophysiologicalrecording.In this study,we explore the influences of ingredients of extracellularsuperfusate on acute isolation of calcium-tolerant ventricular myocytes and recording ondelayed rectifier potassium (I_K) and L-type calcium current (I_(Ca-L)).
Methods:Single ventricular myocytes of rabbits were obtained enzymatically byretrograde Langendorff perfusing apparatus.Before Langendorff perfusing,theextracellular superfusate was Tyrode's solution in experiment group and calcium-freeTyrode's solution in control group.The effect of extracellular superfusate Ca~(2+) oncalcium-tolerant ventricular myocytes available for electrophysiological recordings wereinvestigated.Whole-cell patch clamp recording technique was used to record the changes ofI_K and I_(Ca-L) in experimental groups following the administration of extracellular with BaCl_2and N-methyl-D-glu-camine (NMDG) in experiment group and extracellular Tyrode'ssolution in control group.
Results:The survival rates of ventricular myocytes and calcium-tolerant myocyteswere significantly higher in experimental groups than in control group (71.0%±9.7% vs54.5%±10.1%,52.0%±.7.9% vs 38.5%±10.8%,n=10,P＜0.05).In experimental group, peak I_(Ca-L) was elicited stably and larger than in control group (15.2±7.1 pA/pF vs 7.4±2.4pA/pF,n=10,P＜0.05).As compared 20 min with 5 min,the rate of change in peak I_(Ca-L) was36.1%±9.1% in control group (n=10,P＜0.05),and 3.0%±4.7% in experimental group(n=10,P＞0.05).The time-dependent outward current and tail current of I_K were higher inexperimental group than in control group (1.39±0.05 pA/pF vs 0.53±0.14 pA/pF,0.74±0.09 pA/pF vs 0.39±0.13 pA/pF,n=10,P＜0.05).
Conclusion:Extracellular superfusate Ca~(2+) is helpful for isolating singlecalcium-tolerant cardiomyocytes.With BaCl_2 and NMDG in extracellular superfusate,theionic currents of I_(Ca-L) and IK were elicited typically and easily.
Part two Effects of sodium ferulate and amiodarone on L-typecalcium channel current in isolated rabbit ventricular myocytes
Objective:To study the effect of sodium ferulate on L-type calcium current (I_(Ca-L)) inisolated rabbit ventricular myocytes.
Methods:Compared with amiodarone- a typical classⅢantiarrhythmic drug,singleventricular myocytes of rabbits were isolated enzymatically and whole-cell patch clamprecording technique was used to record the changes of I_(Ca-L) following the administration ofsodium ferulate at 3,10,30 and 100μmol/L.
Results:Both sodium ferulate and amiodarone could block the I_(Ca-L) which hadconcentration dependence,however,sodium ferulate inferior to amiodarone (P＜0.05).PeakI_(Ca-L) was reduced 21.1%±3.8%,32.6%±2.6%,52.6%±4.6% and 71.4%±7.0% (n=5,P＜0.05) respectively at amiodarone 1,3,10 and 30μmol/L.In contrast,Peak I_(Ca-L) wasreduced 11.1%±2.4%,26.9%±6.2%,40.5%±5.0% and 61.9%±5.5% (n=5,P＜0.05)respectively at sodium ferulate 3,10,30 and 100μmol/L.IC_(50) (the concentration for half-maximal block) was 32.6μmol/L and 9.5μmol/L.The current-voltage curve movedupward,stead-state activated curve moved right and stead-state deactivated curve movedleft,furthermore,they delayed the recovery time course of I_(Ca-L) from inactive state.
Conclusion:Sodium ferulate block L-type calcium channel,slow it's activate,rapidit's inactivate and delay the recovery time.These may be the related mechanisms of theantiarrhythmic effects.
Part three Effects of sodium ferulate and amiodarone on potassiumchannel currents in isolated rabbit ventricular myocytes
Objective:To study the effect of sodium ferulate on rapidly and slowly activatingcomponent of delayed rectifier potassium current (I_(Kr),I_(Ks)),inwardly rectified potassiumcurrent (I_(K1)) and transient outward potassium current (I_(to))in isolated rabbit ventricularmyocytes.
Methods:Compared with amiodarone - a typical classⅢantiarrhythmic drug,singleventricular myocytes of rabbits were isolated enzymatically and whole-cell patch clamprecording technique was used to record the changes of I_(Kr),I_(Ks),I_(K1) and I_(to) following theadministration of sodium ferulate at 3,10,30 and 100μmol/L.
Results Sodium ferulate was inferior to amiodarone,though both of them could blockthe time-dependent outward current and tail current of I_(Kr) and I_(Ks) which had concentrationdependence.Peak tail currents of I_(Kr) were reduced 12.1%±2.5%,24.1%±3.0%,47.0%±5.8%,58.5%±8.3% (n=5,P＜0.05) respectively,while,peak tail currents of I_(Ks) werereduced 15.6%±6.4%,27.1%±6.5%,45.6%±5.7%,51.8%±6.6% (n=5,P＜0.05)respectively at sodium ferulate 3,10,30 and 100μmol/L.IC_(50) (the concentration for half-maximal block) were higher than amiodarone respectively (43.6 vs 3.48μmol/L,44.8 vs5.11μmol/L).Compared with amiodarone (30μmol/L),sodium ferulate (100μmol/L)decreased I_(k1) by 24.3%±8.8% vs 29.7%±8.0% at test potential of -100 mV (n=5,P＜0.05),At -20mV,I_(K1) decreased by 17.7%±3.3% vs 20.1%±10.6% (n=5,P＜0.05).Thecurrent-voltage curve of I_(K1) moved left by sodium ferulate (30,100μmol/L) andamiodarone (10,30μmol/L) acute superfusion.However,neither of them alter I_(to) and it'scurrent-voltage curve.
Conclusion:Sodium ferulate block potassium channels,these may be the relatedmechanisms of the antiarrhythmic effects.
Part four Effects of sodium ferulate and amiodarone on sodiumchannel currents in isolated rabbit ventricular myocytes
Objective:To study the effect of sodium ferulate on sodium current (I_(Na)) in isolatedrabbit ventricular myocytes.
Methods:Compared with amiodarone- a typical classⅢantiarrhythmic drug,singleventricular myocytes of rabbits were isolated enzymatically and whole-cell patch clamprecording technique was used to record the changes of I_(Na).
Results:The amplitude of peak I_(Na) was decreased by acute amiodarone (30μmol/L)superfusion (20.9±3.63 pA/pF vs 12.0±2.69 pA/pF,n=5,P＜0.05),the inhibition was42.6%±4.87%,and current-voltage curves of I_(Na) moved upward.However,sodiumferulate (100μmol/L) superfusion had no effect on I_(Na) (16.6±4.52 pA/pF vs 15.8±3.21pA/pF,n=5,P＜0.05 ),the inhibition was 6.12%±5.41%,neither did sodium ferulate altercurrent-voltage curves of I_(Na) ( n=5,P＞0.05 ).
Conclusion:Sodium ferulate had no effect on sodium channel current in Langendorff-perfused rabbit heart.
目的:细胞标本制备的质量是膜片钳实验成功的关键,本研究探讨通过改变细胞外液成分获取适于电生理研究的耐钙心室肌细胞及L-型钙通道电流(I_(Ca-L))、延迟整流钾电流(I_K)记录的方法。
方法:家兔心室肌细胞急性分离采用Langendorff灌流装置及酶解消化分离技术,心脏灌流前实验组细胞外液为正常台氏液,对照组细胞外液为无钙台氏液,观察细胞外液中的钙离子对获取适于电生理记录的单个耐钙心肌细胞的影响;应用全细胞膜片钳技术,实验组分别以含BaCl_2的台氏液及N-甲基-D-葡萄糖胺(NMDG)溶液作为细胞外液记录I_(Ca-L)及I_K,对照组为正常台氏液。
结果:与对照组相比较,实验组心室肌细胞存活率和耐钙心肌细胞存活率均明显升高(71.0%±9.7% vs 54.5%±10.1%; 52.0%±7.9% vs 38.5%±10.8%, n=10, P<0.05)。实验组引出的I_(Ca-L)峰值电流大于对照组(15.2±7.1 pA/pF vs 7.4±2.4 pA/pF, n=10,P<0.05)。实验20 min与实验5 min记录的I_(Ca-L)峰值比较,对照组I_(Ca-L)峰值电流的变化率为36.1%±9.1% (n=10, P<0.05),即I_(Ca-L)峰值电流在实验20 min衰减;实验组I_(Ca-L)峰值电流的变化率为3.0%±4.7% (n=10, P>0.05),即I_(Ca-L)峰值电流幅值稳定。实验组I_K时间依赖性外向电流及尾电流随去极化时间延长增大作用更明显(1.39±0.05pA/pF vs 0.53±0.14 pA/pF, 0.74±0.09 pA/pF vs 0.39±0.13 pA/pF, n=10, P<0.05)。
结论:细胞外液中的钙离子有助于获取耐钙心肌细胞,以含BaCl_2的台氏液及NMDG溶液作为细胞外液易于获得典型而稳定的I_(Ca-L)及I_K。
第二部分阿魏酸钠与胺碘酮对家兔心室肌细胞L-型钙通道电流的影响
目的:探讨阿魏酸钠对家兔心室肌细胞膜L-型钙通道电流(I_(Ca-L))的影响。
方法:酶解法急性分离家兔单个心室肌细胞,以经典的Ⅲ类抗心律失常药物胺碘酮为对照,应用膜片钳全细胞记录技术观察3、10、30、100μmol/L的阿魏酸钠及1、3、10、30μmol/L的胺碘酮对心室肌细胞膜I_(Ca-L)的作用。
结果:阿魏酸钠及胺碘酮均呈浓度依赖性抑制L-型钙电流。3、10、30、100μmol/L的阿魏酸钠对I_(Ca-L)的抑制率分别为11.1%±2.4%、26.9%±6.2%、40.5%±5.0%、61.9%±5.5% (P<0.05);1、3、10、30μmol/L的胺碘酮对I_(Ca-L)的抑制率分别为21.1%±3.8%、32.6%±2.6%、52.6%±4.6%、71.4%±7.0% (P<0.05);半数抑制浓度(IC_(50))分别为32.6μmol/L及9.5μmol/L,阿魏酸钠的抑制作用弱于胺碘酮(P<0.05)。阿魏酸钠对I_(Ca-L)动力学的影响类似于胺碘酮,二者均能使I_(Ca-L)电流-电压曲线上移,未改变I-V曲线形状;使稳态激活曲线右移,失活曲线左移,使激活曲线与失活曲线之间相交的“窗流”区域缩小;并使I_(Ca-L)恢复曲线的t值增加,恢复曲线右移,即减慢钙通道灭活后的恢复过程。
结论:阿魏酸钠对I_(Ca-L)具有浓度依赖性阻滞作用,且使I_(Ca-L)的激活减慢、失活加快,并且失活后的恢复时间延长,阿魏酸钠的抑制作用虽弱于胺碘酮,但具有与胺碘酮相似的阻滞I_(Ca-L)且影响I_(Ca-L)的动力学特性,可能是其抗心律失常作用的电生理机制之一。
第三部分阿魏酸钠与胺碘酮对家兔心室肌细胞钾通道电流的影响
目的:探讨阿魏酸钠对家兔心室肌细胞膜延迟整流钾电流快速与缓慢激活成分(I_(Kr)、I_(Ks))、内向整流钾电流(I_(Kl))、瞬时外向钾电流(I_(to))的影响。
方法:酶解法急性分离家兔单个心室肌细胞,以经典的Ⅲ类抗心律失常药物胺碘酮为对照,采用膜片钳全细胞记录技术观察浓度为3、10、30、100μmol/L的阿魏酸钠对心室肌细胞膜I_(Kr)、I_(Ks)、I_(Kl)及I_(to)的作用。
结果:阿魏酸钠的作用弱于胺碘酮,二者均可浓度依赖性抑制I_(Kr)、I_(ks)时间依赖性外向电流及尾电流(I_(Kr,tail)、I_(Ks, tail)),3、10、30、100μmol/L的阿魏酸钠对I_(Kr, tail)的抑制率为12.1%±2.5%、24.1%±3.0%、47.0%±5.8%及58.5%±8.3%(n=5,P<0.05);对I_(Ks, tail)的抑制率为15.6%±6.4%、27.1%±6.5%、45.6%±5.7%及51.8%±6.6%(n=5,P<0.05),其对I_(Kr, tail)及I_(Ks, tail)的半数抑制浓度(IC_(50))均大于胺碘酮(43.6 vs 3.48μmol/L,44.8 vs 5.11μmol/L)。30、100μmol/L的阿魏酸钠及10、30μmol/L的胺碘酮可使I_(kl)的I-V曲线左移,在-100 mV及-20 mV测试电压下,浓度为100μmol/L的阿魏酸钠对I_(kl)内向电流及外向电流抑制率小于30μmol/L的胺碘酮(24.3%±8.8% vs 29.7%±8.0%; 17.7%±3.3% vs 20.1%±10.6%, n=5, P<0.05)。阿魏酸钠与胺碘酮均不影响I_(to)及其I-V曲线。
结论:阿魏酸钠对复极期钾电流具有阻滞作用,可能是其抗心律失常作用的电生理机制之一。
第四部分阿魏酸钠与胺碘酮对家兔心室肌细胞钠通道电流的影响
目的:探讨阿魏酸钠对家兔心室肌细胞膜钠通道电流(I_(Na))的影响。
方法:酶解法急性分离家兔单个心室肌细胞,以经典的Ⅲ类抗心律失常药物胺碘酮为对照,采用膜片钳全细胞记录技术观察浓度为100μmol/L的阿魏酸钠及10μmol/L胺碘酮对心室肌细胞膜I_(Na)的作用。
结果:10μmol/L胺碘酮使I_(Na)峰值电流由20.9±3.63 pA/pF减为12.0±2.69 pA/pF(n=5, P<0.05),抑制率为42.6%±4.87%;胺碘酮使I_(Na)的I-V曲线上移。100μmol/L阿魏酸钠不改变I_(Na)峰值电流(16.6±4.52 pA/pF vs 15.8±3.21 pA/pF,抑制率为6.12%±5.41%, n=5, P>0.05),且不影响I-V曲线。
结论:阿魏酸钠急性灌流不影响家兔心室肌细胞I_(Na)。
Part one Investigation of the methods for acute isolatingcalcium-tolerant ventricular myocytes andelectrophysiolological recordings
Objective:It is important to obtain viable cells for long period of electrophysiologicalrecording.In this study,we explore the influences of ingredients of extracellularsuperfusate on acute isolation of calcium-tolerant ventricular myocytes and recording ondelayed rectifier potassium (I_K) and L-type calcium current (I_(Ca-L)).
Methods:Single ventricular myocytes of rabbits were obtained enzymatically byretrograde Langendorff perfusing apparatus.Before Langendorff perfusing,theextracellular superfusate was Tyrode's solution in experiment group and calcium-freeTyrode's solution in control group.The effect of extracellular superfusate Ca~(2+) oncalcium-tolerant ventricular myocytes available for electrophysiological recordings wereinvestigated.Whole-cell patch clamp recording technique was used to record the changes ofI_K and I_(Ca-L) in experimental groups following the administration of extracellular with BaCl_2and N-methyl-D-glu-camine (NMDG) in experiment group and extracellular Tyrode'ssolution in control group.
Results:The survival rates of ventricular myocytes and calcium-tolerant myocyteswere significantly higher in experimental groups than in control group (71.0%±9.7% vs54.5%±10.1%,52.0%±.7.9% vs 38.5%±10.8%,n=10,P＜0.05).In experimental group, peak I_(Ca-L) was elicited stably and larger than in control group (15.2±7.1 pA/pF vs 7.4±2.4pA/pF,n=10,P＜0.05).As compared 20 min with 5 min,the rate of change in peak I_(Ca-L) was36.1%±9.1% in control group (n=10,P＜0.05),and 3.0%±4.7% in experimental group(n=10,P＞0.05).The time-dependent outward current and tail current of I_K were higher inexperimental group than in control group (1.39±0.05 pA/pF vs 0.53±0.14 pA/pF,0.74±0.09 pA/pF vs 0.39±0.13 pA/pF,n=10,P＜0.05).
Conclusion:Extracellular superfusate Ca~(2+) is helpful for isolating singlecalcium-tolerant cardiomyocytes.With BaCl_2 and NMDG in extracellular superfusate,theionic currents of I_(Ca-L) and IK were elicited typically and easily.
Part two Effects of sodium ferulate and amiodarone on L-typecalcium channel current in isolated rabbit ventricular myocytes
Objective:To study the effect of sodium ferulate on L-type calcium current (I_(Ca-L)) inisolated rabbit ventricular myocytes.
Methods:Compared with amiodarone- a typical classⅢantiarrhythmic drug,singleventricular myocytes of rabbits were isolated enzymatically and whole-cell patch clamprecording technique was used to record the changes of I_(Ca-L) following the administration ofsodium ferulate at 3,10,30 and 100μmol/L.
Results:Both sodium ferulate and amiodarone could block the I_(Ca-L) which hadconcentration dependence,however,sodium ferulate inferior to amiodarone (P＜0.05).PeakI_(Ca-L) was reduced 21.1%±3.8%,32.6%±2.6%,52.6%±4.6% and 71.4%±7.0% (n=5,P＜0.05) respectively at amiodarone 1,3,10 and 30μmol/L.In contrast,Peak I_(Ca-L) wasreduced 11.1%±2.4%,26.9%±6.2%,40.5%±5.0% and 61.9%±5.5% (n=5,P＜0.05)respectively at sodium ferulate 3,10,30 and 100μmol/L.IC_(50) (the concentration for half-maximal block) was 32.6μmol/L and 9.5μmol/L.The current-voltage curve movedupward,stead-state activated curve moved right and stead-state deactivated curve movedleft,furthermore,they delayed the recovery time course of I_(Ca-L) from inactive state.
Conclusion:Sodium ferulate block L-type calcium channel,slow it's activate,rapidit's inactivate and delay the recovery time.These may be the related mechanisms of theantiarrhythmic effects.
Part three Effects of sodium ferulate and amiodarone on potassiumchannel currents in isolated rabbit ventricular myocytes
Objective:To study the effect of sodium ferulate on rapidly and slowly activatingcomponent of delayed rectifier potassium current (I_(Kr),I_(Ks)),inwardly rectified potassiumcurrent (I_(K1)) and transient outward potassium current (I_(to))in isolated rabbit ventricularmyocytes.
Methods:Compared with amiodarone - a typical classⅢantiarrhythmic drug,singleventricular myocytes of rabbits were isolated enzymatically and whole-cell patch clamprecording technique was used to record the changes of I_(Kr),I_(Ks),I_(K1) and I_(to) following theadministration of sodium ferulate at 3,10,30 and 100μmol/L.
Results Sodium ferulate was inferior to amiodarone,though both of them could blockthe time-dependent outward current and tail current of I_(Kr) and I_(Ks) which had concentrationdependence.Peak tail currents of I_(Kr) were reduced 12.1%±2.5%,24.1%±3.0%,47.0%±5.8%,58.5%±8.3% (n=5,P＜0.05) respectively,while,peak tail currents of I_(Ks) werereduced 15.6%±6.4%,27.1%±6.5%,45.6%±5.7%,51.8%±6.6% (n=5,P＜0.05)respectively at sodium ferulate 3,10,30 and 100μmol/L.IC_(50) (the concentration for half-maximal block) were higher than amiodarone respectively (43.6 vs 3.48μmol/L,44.8 vs5.11μmol/L).Compared with amiodarone (30μmol/L),sodium ferulate (100μmol/L)decreased I_(k1) by 24.3%±8.8% vs 29.7%±8.0% at test potential of -100 mV (n=5,P＜0.05),At -20mV,I_(K1) decreased by 17.7%±3.3% vs 20.1%±10.6% (n=5,P＜0.05).Thecurrent-voltage curve of I_(K1) moved left by sodium ferulate (30,100μmol/L) andamiodarone (10,30μmol/L) acute superfusion.However,neither of them alter I_(to) and it'scurrent-voltage curve.
Conclusion:Sodium ferulate block potassium channels,these may be the relatedmechanisms of the antiarrhythmic effects.
Part four Effects of sodium ferulate and amiodarone on sodiumchannel currents in isolated rabbit ventricular myocytes
Objective:To study the effect of sodium ferulate on sodium current (I_(Na)) in isolatedrabbit ventricular myocytes.
Methods:Compared with amiodarone- a typical classⅢantiarrhythmic drug,singleventricular myocytes of rabbits were isolated enzymatically and whole-cell patch clamprecording technique was used to record the changes of I_(Na).
Results:The amplitude of peak I_(Na) was decreased by acute amiodarone (30μmol/L)superfusion (20.9±3.63 pA/pF vs 12.0±2.69 pA/pF,n=5,P＜0.05),the inhibition was42.6%±4.87%,and current-voltage curves of I_(Na) moved upward.However,sodiumferulate (100μmol/L) superfusion had no effect on I_(Na) (16.6±4.52 pA/pF vs 15.8±3.21pA/pF,n=5,P＜0.05 ),the inhibition was 6.12%±5.41%,neither did sodium ferulate altercurrent-voltage curves of I_(Na) ( n=5,P＞0.05 ).
Conclusion:Sodium ferulate had no effect on sodium channel current in Langendorff-perfused rabbit heart.
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[1]Wang BH, Ou-Yang JP. Pharmacological actions of sodium ferulate in cardiovascular system. Cardiovasc Drug Rev, 2005, 23(2): 161-172
[2]周彤，曾秋棠，张桂清，等．阿魏酸钠与胺碘酮对家免心室肌电生理作用的频率依赖性影响．中国心脏起搏与心电生理杂志，2002，16(1)：54-56
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[1]Wang BH, Ou-Yang JP. Pharmacological actions of sodium ferulate in cardiovascular system. Cardiovasc Drug Rev, 2005, 23(2): 161-172
[2]王卓，韩大英．阿魏酸钠对豚鼠心室肌动作电位的影响．首都医学院学报，1989，10(1)：12-4
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[4]周彤，曾秋棠，张桂清，等．阿魏酸钠与胺碘酮对家免心室肌电生理作用的频率依赖性影响．中国心脏起搏与心电生理杂志，2002，16(1)：54-57
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[8]刘坤，高翔，刘金平，等．阿魏酸钠对人心房肌细胞超快激活延迟整流钾电流的影响．临床心血管病杂志，2007，23(6)：444-446
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[13][Kolettis TM, Agelaki MG, Baltogiannis GG, et al. Comparative effects of acute vs. chronic oral amiodarone treatment during acute myocardial infarction in rats[J].Europace, 2007, 9(11): 1099-1104
[14]Bouter SL, Harchi AE, Marionneau C. Long-term amiodarone administration remodels expression of ion channel transcripts in the mouse heart. Circulation, 2004, 110(11):3028-3035
[15]Yamashita N, Kaku T, Uchino T, et al. Short and long-term amiodarone treatments regulate Ca_v3.2 low voltage activated T-type Ca~(2+) channel through distinct mechanisms . Mol Pharmacol, 2006, 69(5): 1684-1691
[16] Giles WR, Imaizumi Y. Comparison of potassium currents in rabbit atrial and ventricular cells. J physiol, 1988, 405(11): 123-145
[17]Ehrlich JR, Biliczki P, Hohnloser SH, et al. Atrium-selective approaches for the treatment of atrial fibrillation. J Am Coll Cardiol, 2008,51(8): 787-92
[18]Burashn ikov A, Di Diego JM, Zygmunt AC, et al. Atrium-selective sodium channel block as a strategy for suppression of fibrillation. Circulation, 2007,116(8): 1449-1457
[19] 郭继鸿.胺碘酮的现代观点.心电学杂志2007,16(3):215-226
[20]Hancox JC, Patel KCR, Jones JV. Antiarrhythmics-from cell to clinic: past, present,and future.Heart, 2000,84(1): 14-24
[21]Dorian P, Newman D. Rate dependence of the effect of antiarrhythmic drugs delaying cardiac repolarization: an overview . Europace 2000; 2(4): 277-285
[22]Yoshida H, Sugiyama A, Satoh Y, et al. Comparison of the in vivo electro-physiological and proarrhythmic effects of amiodarone with those of a selective classⅢ drug, sematilide, using a canine chronic atrioventricular block model. Circulation,2002, 66(8): 758-762
[23]van Opstal JM, Schoenmakers M, Verduyn SC, et al. Chronic amiodarone evokes no torsade de points arrhythmias despite QT lengthening in a animal model of acquired long-QT syndrome. Circulation, 2001,104(22): 2722-2727
[24]Antzelevitch C , Nesterenko W, Yan GX. Role of M cells in acquired long QT syndrome, U waves, and torsade de points. J Electrocardio, 1995, 28(suppl): 131-138
[25] Moro S, Ferreiro M, Celestino D. In vitro effects of acute amiodarone and dronedarone on Epicardial, Endocardial, and M Cells of the canine ventricle. J Cardiovasc Pharmacol Ther, 2007,12(4): 314-321
[26] 刘泰槰.心肌细胞电生理学.北京:人民卫生出版社, 2003. 59-63
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[28]Vassallo P, Trohman RG. Prescribing amiodarone: an evidence-based review of clinical indications. JAMA, 2008, 298(11): 1312-1322
[29]Schrickel JW, Schwab JO, Yang A, et al. Pro-arrhythmic effects of amiodarone and concomitant rate-control medication. Europace, 2006, 8(6): 403-407
[30]Tafreshi MJ, Rowles J. A review of the investigational antiarrhythmic agent dronedarone. J Cardiovasc Pharmacol Thera, 2007,12(1): 15-26
[31]Waldhauser KM, Torok M, Ha HR. Hepatocellular toxicity and pharmacological effect of amiodarone and amiodarone derivatives. JPET, 2006, 319(3): 1413-1423
[32] Celestino D, Medei E, Moro S, et al. Acute in vitro effects of dronedarone, an iodine-free derivative, and amiodarone, on the rabbit sinoatrial node automaticity: a comparative study. J Cardiovasc Pharmacol Thera, 2007,12(3): 248-257
[33]Kathofer S, Thomas D, Karle C. The novel antiarrhythmic drug dronedarone:comparison with amiodarone. Cardiovasc Drug Rev, 2005, 23(3): 217-230
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[36]Antzelevitch C, Belardinelli L, Zygmunt AW, et al. Electrophysiologic effects of ranolazine. a novel anti-anginal agent with antiarrhythmic properties. Circulation, 2004,110(8): 904-910
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[1]Wang BH, Ou-Yang JP. Pharmacological actions of sodium ferulate in cardiovascular system. Cardiovasc Drug Rev, 2005, 23(2): 161-172
[2]周彤，曾秋棠，张桂清，等．阿魏酸钠与胺碘酮对家免心室肌电生理作用的频率依赖性影响．中国心脏起搏与心电生理杂志，2002，16(1)：54-56
[3]周彤，曾秋棠，张桂清，等．阿魏酸钠抗肾上腺素实验性心律失常作用．中国新药杂志，2003，12(5)：345-347
[4]秦孺子，曾秋棠，李军．阿魏酸钠对卵母细胞Kv1．2外向钾电流的影响．中国心脏起搏与心电生理杂志，2006，20(5)：440-442
[5]Yamashita N, Kaku T, Uchino T, et al. Short and long-term amiodarone treatments regulate Ca_v3.2 low voltage activated T-type Ca~(2+) channel through distinct mechanisms. Mol Pharmacol, 2006, 69(5): 1684-1691
[6]Hancox JC. Amiodarone blocks L-type calcium current in single myocytes isolated from the rabbit atrioventricular node. Gen Pharmac, 1997, 29( 3): 429-435
[7]Varro A, Virafg L , Gy.papp J. Comparison of the chronic and acute effects of amiodarone on the calcium and potassium currents in rabbit isolated cardiac myocytes. Br J of Pamarcol, 1996, 117(6): 1181-1186
[8]Tytgat J. How to isolate cardiac myocytes. Cardiovasc Res, 1994, 28(2): 280-283
[9]李泱．离子通道与膜片钳技术．兰州：甘肃文化出版社，2001．98-185
[10]Nattel S. Experimental evidence for proarrhythmic mechanisms of antiarrhythmic drugs. Cardiovasc Res, 1998, 37(3): 567-577
[11]刘泰槰．心肌细胞电生理学．北京：人民卫生出版社，2003．72．-83
[12]杨宝峰．离子通道药理学．北京：人民卫生出版社，2005．185-188
[13]陈修，陈维洲，曾贵云．心血管药理学．第3版．北京：人民卫生出版社，2002． 584-622
[1]Wang BH, Ou-Yang JP. Pharmacological actions of sodium ferulate in cardiovascular system. Cardiovasc Drug Rev, 2005, 23(2): 161-172
[2]王卓，韩大英．阿魏酸钠对豚鼠心室肌动作电位的影响．首都医学院学报，1989，10(1)：12-4
[3]周彤，曾秋棠，张桂清，等．阿魏酸钠与胺碘酮对家免心室肌电生理作用的对比研究．临床心血管病杂志，2002，18(5)：226-228
[4]周彤，曾秋棠，张桂清，等．阿魏酸钠与胺碘酮对家免心室肌电生理作用的频率依赖性影响．中国心脏起搏与心电生理杂志，2002，16(1)：54-57
[5]周彤，曾秋棠，张桂清，等．阿魏酸钠抗肾上腺素实验性心律失常作用[J]．中国新药杂志，2003，12(5)：345-347
[6]刘泰槰．心肌细胞电生理学．北京：人民卫生出版社，2003：89-114
[7]秦孺子，曾秋棠，李军．阿魏酸钠对卵母细胞Kv1．2外向钾电流的影响．中国心脏起搏与心电生理杂志，2006，20(5)：440-442
[8]刘坤，高翔，刘金平，等．阿魏酸钠对人心房肌细胞超快激活延迟整流钾电流的影响．临床心血管病杂志，2007，23(6)：444-446
[9]Lu ZB, Kamiya K, Opthof T, et al. Density and kinetics of I_(Kr) and I_(Kr) in guinea pig and rabbit ventricular myocytes explain different efficacy of I_(Kr) blockade at high heart rate in guinea pig and rabbit: implications for arrhythmogenesis in humans. Circulation, 2001; 104(8): 951-956
[10]Cheng JH, Kamiya K, Liu WR. Heterogeneous distribution of the two components of delayed rectifier K~+ current: a potential mechanism of the proarrhythmic effects of methanesulfonanilideclass Ⅲ agents. Cardiovasc Res, 1999, 43(1): 135-147
[11]Tytgat J. How to isolate cardiac myocytes. Cardiovasc Res, 1994, 28(2): 280-283
[12]李泱．离子通道与膜片钳技术．兰州：甘肃文化出版社，2001：132-185
[13]Tamargo J, Caballero R, G(?)mez, et al. Pharmacology of cardiac potassium channel. Cardiovasc Res, 2004, 62(1): 9-33
[14]Nattel S. Experimental evidence for proarrhythmic mechanisms of antiarrhythmic drugs. Cardiovasc Res, 1998, 37(3): 567-577
[15]Dorian P, Newman D. Rate dependence of the effect of antiarrhythmic drugs delaying cardiac repolarization: an overview. Europace, 2000, 2(4): 277-285
[16]金有豫．药理学．第五版．北京：人民卫生出版社，2001：4-7
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[18]Hancox JC, Patel KCR, Jones JV. Antiarrhythrnics-from cell to clinic: past, present, and future. Heart, 2000, 84(1): 14-24
[1]李泱．离子通道与膜片钳技术．兰州：甘肃文化出版社，2001．5-62
[2]刘泰槰．心肌细胞电生理学．北京：人民卫生出版社，2003．53-71
[3]Chen F, Wetzel GT, Klitzner TS. Acute effects of amiodarone on sodium currents in isolated neonatal ventricular myocytes: comparison with procainamide. Dev Pharmacol Ther, 1992, 19(2-3): 118-130
[4]周彤，曾秋棠，张桂清，等．阿魏酸钠与胺碘酮对家免心室肌电生理作用的对比研究．临床心血管病杂志，2002，18(5)：226-228
[5]Liu H, Sun HY, Lau CP, et al. Regulation of voltage-gated cardiac sodium current by epidermal growth factor receptor kinase in guinea pig ventricular myocytes. J Mol Cell Cardiol 2007, 42(4): 760-768
[6]Pu J, Balser JR, Boyden PA, et al. Lidocaine action on Na~+ currents in ventricular myocytes from the epicardial border zone of the infarcted heart. Circ Res 1998, 83 (4): 431-440
[7]Nattel S. Experimental evidence for proarrhythmic mechanisms of antiarrhythmic drugs. Cardiovasc Res, 1998, 37(3): 567-577
[8]Kodama I, Kamiya K, Toyama J. Cellular electropharmacology of amiodarone. Cardiovasc Res, 1997, 35(1): 13-29
[9]Kamiya k, Nishiyama A, Yasui K, Hojo M, Sanguinetti MC, Kodama I. Short and long-term effects of amiodarone on the two components of cardiac delayed rectifier K current. Circulation, 2001, 103(9): 1317-1321
[10]Varr(?) A, Virafg L, Papp JG. Comparison of the chronic and acute effects of amiodarone on the calcium and potassium currents in rabbit isolated cardiac myocytes. Br J Pharmacol, 1996, 117(6): 1181-1186
[1]Pratt CM, Moy(?) LA.The cardiac arrhythmia suppression trial casting suppression in a different light. Circulation, 1995, 91(1): 245-247
[2]Waldo AL, Carom AJ, Deruyter H. et al. For the SWORD investigators: effect of d-sotalol on mortality in patient with left ventricular dysfuction after recent and remote myocardial infarction. Lancet, 1996, 348(1019): 7-12
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[4]陆再英，钟南山．内科学．第七版．北京：人民卫生出版社，2008．217-220
[5]Kodama I, Kamiya K, Toyama J. Cellular electropharmacology of amiodarone. Cardiovas Res, 1997, 35(1):13-29
[6]中国生物医学工程学会心律分会，中华医学会心血管病学分会，胺碘酮抗心律失常治疗应用指南工作组．胺碘酮抗心律失常治疗应用指南(2008)．中国心脏起搏与心电生理杂志，2008，22(5)：377-385
[7]Nattel S. Experimental evidence for proarrhythmic mechanisms of antiarrhythmic drugs. Cardiovasc Res, 1998, 37(3): 567-577
[8]Chen F, Wetzel GT, Klitzner TS. Acute effects of amiodarone on sodium currents in isolated neonatal ventricular myocytes: comparison with procainamide. Dev Pharmacol Ther, 1992, 19(2-3): 118-130
[9]Kamiya K, Nishiyama A, Yasui K, et al. Short- and long-term effects of amiodarone on the two components of cardiac delayed rectifier K+ current. Circulation, 2001, 103(9): 1317-1324
[10]Varr(?) A, Virafg L, Papp JG. Comparison of the chronic and acute effects of amiodarone on the calcium and potassium currents in rabbit isolated cardiac myocytes. Br J Pharmacol, 1996, 117(6): 1181-1186
[11]Gray DF, Mihailidou AS, Hansen PS, et al. Amiodarone inhibits the Na~+-K~+ pump in rabbit cardiac myocytes after acute and chronic treatment. JPET, 1998, 284(1): 75-82
[12]Pitt A, Fernandes C, Bewick NL, et al. Chronic amiodarone-induced inhibition of the Na~+-K~+ pump in rabbit cardiac myocytes is thyroid-dependent: comparison with dronedarone. Cardiovasc Res, 2003, 57(1): 101-108
[13][Kolettis TM, Agelaki MG, Baltogiannis GG, et al. Comparative effects of acute vs. chronic oral amiodarone treatment during acute myocardial infarction in rats[J].Europace, 2007, 9(11): 1099-1104
[14]Bouter SL, Harchi AE, Marionneau C. Long-term amiodarone administration remodels expression of ion channel transcripts in the mouse heart. Circulation, 2004, 110(11):3028-3035
[15]Yamashita N, Kaku T, Uchino T, et al. Short and long-term amiodarone treatments regulate Ca_v3.2 low voltage activated T-type Ca~(2+) channel through distinct mechanisms . Mol Pharmacol, 2006, 69(5): 1684-1691
[16] Giles WR, Imaizumi Y. Comparison of potassium currents in rabbit atrial and ventricular cells. J physiol, 1988, 405(11): 123-145
[17]Ehrlich JR, Biliczki P, Hohnloser SH, et al. Atrium-selective approaches for the treatment of atrial fibrillation. J Am Coll Cardiol, 2008,51(8): 787-92
[18]Burashn ikov A, Di Diego JM, Zygmunt AC, et al. Atrium-selective sodium channel block as a strategy for suppression of fibrillation. Circulation, 2007,116(8): 1449-1457
[19] 郭继鸿.胺碘酮的现代观点.心电学杂志2007,16(3):215-226
[20]Hancox JC, Patel KCR, Jones JV. Antiarrhythmics-from cell to clinic: past, present,and future.Heart, 2000,84(1): 14-24
[21]Dorian P, Newman D. Rate dependence of the effect of antiarrhythmic drugs delaying cardiac repolarization: an overview . Europace 2000; 2(4): 277-285
[22]Yoshida H, Sugiyama A, Satoh Y, et al. Comparison of the in vivo electro-physiological and proarrhythmic effects of amiodarone with those of a selective classⅢ drug, sematilide, using a canine chronic atrioventricular block model. Circulation,2002, 66(8): 758-762
[23]van Opstal JM, Schoenmakers M, Verduyn SC, et al. Chronic amiodarone evokes no torsade de points arrhythmias despite QT lengthening in a animal model of acquired long-QT syndrome. Circulation, 2001,104(22): 2722-2727
[24]Antzelevitch C , Nesterenko W, Yan GX. Role of M cells in acquired long QT syndrome, U waves, and torsade de points. J Electrocardio, 1995, 28(suppl): 131-138
[25] Moro S, Ferreiro M, Celestino D. In vitro effects of acute amiodarone and dronedarone on Epicardial, Endocardial, and M Cells of the canine ventricle. J Cardiovasc Pharmacol Ther, 2007,12(4): 314-321
[26] 刘泰槰.心肌细胞电生理学.北京:人民卫生出版社, 2003. 59-63
[27]Wu L, Rajamani S, Shryock JC, et al. Augmentation of late sodium current unmasks the proarrhythmic effects of amiodarone. Cardiovasc Res, 2008, 77(3): 483-488
[28]Vassallo P, Trohman RG. Prescribing amiodarone: an evidence-based review of clinical indications. JAMA, 2008, 298(11): 1312-1322
[29]Schrickel JW, Schwab JO, Yang A, et al. Pro-arrhythmic effects of amiodarone and concomitant rate-control medication. Europace, 2006, 8(6): 403-407
[30]Tafreshi MJ, Rowles J. A review of the investigational antiarrhythmic agent dronedarone. J Cardiovasc Pharmacol Thera, 2007,12(1): 15-26
[31]Waldhauser KM, Torok M, Ha HR. Hepatocellular toxicity and pharmacological effect of amiodarone and amiodarone derivatives. JPET, 2006, 319(3): 1413-1423
[32] Celestino D, Medei E, Moro S, et al. Acute in vitro effects of dronedarone, an iodine-free derivative, and amiodarone, on the rabbit sinoatrial node automaticity: a comparative study. J Cardiovasc Pharmacol Thera, 2007,12(3): 248-257
[33]Kathofer S, Thomas D, Karle C. The novel antiarrhythmic drug dronedarone:comparison with amiodarone. Cardiovasc Drug Rev, 2005, 23(3): 217-230
[34] Singh BN. Classes of antianginal compounds: old and new therapeutic implications. J Cardiovasc Pharmacol Thera, 2003, 8(2): 85-88
[35] Singh BN, Wadhani N. Antiarrhythmic and proarrhythmic properties of QT-prolonging antianginal drugs. J Cardiovasc Pharmacol Thera, 2004, 9(Suppl 1): S85-S97
[36]Antzelevitch C, Belardinelli L, Zygmunt AW, et al. Electrophysiologic effects of ranolazine. a novel anti-anginal agent with antiarrhythmic properties. Circulation, 2004,110(8): 904-910
[37] Antzelevitch C, Belardinelli L, Wu L. Electrophysiologic properties and antiarrhythmic actions of a novel antianginal agent with antiarrhythmic properties. J Cardiovasc Pharmacol Thera, 2004, 9(Suppl 1): S65-S83