钠钙交换体在缺血预处理产生的心肌保护中的作用及其机制研究
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摘要
众所周知,缺血预处理可以保护心肌,减少缺血再灌注导致的心肌梗死面积,降低心律失常的发生率。然而其具体的机制仍未完全阐明。有研究发现,预处理期间短暂、可逆的胞内钙离子浓度升高与缺血预处理的保护作用密切相关。鉴于心肌缺血再灌注初期,通过激活NCX的反向转运模式使[Ca~(2+)]i升高,引起Ca~(2+)超载及由此导致细胞损伤,而缺血预处理的程序就是多个短暂的缺血再灌注过程,其病理生理条件相似,所以,我们设想NCX的反向转运模式可能参与缺血预处理期间[Ca~(2+)]i升高的调节,从而产生心肌保护作用。目前国内外尚无相关的研究报道。
缺血预处理的下游保护效应分子众多,但保护线粒体结构和功能的完整是缺血预处理保护作用机制中的关键,所以,探讨激活NCX的反向转运模式触发缺血预处理的保护作用的下游作用机制具有重要的意义。线粒体上的两种钾离子通道已经证明是缺血预处理保护作用的关键通道,因此探讨NCX引起[Ca~(2+)]i升高是否通过开放这两种离子通道从而中发挥了心肌保护作用,并且这两种离子通道在其作用机制中所扮演的角色,将为我们深入理解心肌缺血预处理的病理生理过程及其作用机制提供新的思路。
目的
1.明确胞外无钠法对大鼠心室肌细胞Na~+/Ca~(2+)交换体转运模式的影响;
2.观察E4031对NCX反向转运模式的影响,并明确其特异性程度;
3.在单心肌模型上观察缺血预处理期间,激活或阻断NCX的反向转运模式对预处理的心肌保护作用的影响;
4.在整体心脏模型上观察NCX的反向转运模式是否触发缺血预处理的保护作用。
5.观察Mito KATP和Mito KCa两种线粒体钾通道在激活NCX的药物预处理中的作用,并进一步完善IP的作用机制。
方法
1.以常规酶解法分离成年大鼠心肌细胞,选取形态、功能良好的心室肌细胞进行实验观察。应用胞外无钠法(NMDG)检测并通过Fura-2/AM荧光法记录[Ca~(2+)]i的变化。
2.预先给予Ryanodine和Thapsigargin孵育10min以阻断SR功能,分别给予Nifedipine(L-型钙通道阻断剂)和Ni2+、KB-R7943(NCX反向转运模式阻断剂),观察记录[Ca~(2+)]i的升高幅度变化。
3.给予E4031预先孵育后,观察NMDG导致的[Ca~(2+)]i的升高幅度变化,并观察给予KB-R7943对E4031引起的[Ca~(2+)]i变化的影响。
4.单心肌代谢抑制液孵育30min后,给予正常Tyrode液灌洗10min,通过代谢抑制法模拟缺血预处理的保护作用。此后再给予缺血液8min,正常Tyrode液灌洗10min制造缺血再灌注损伤。分别观察E4031预处理能否模拟MIP作用,KB-R7943能否阻断MIP和E4031的保护作用。
5.制备Langendorff离体心脏灌流模型,通过结扎/松开冠脉左前降支制造缺血/再灌注。通过2次5min的缺血再灌注过程模拟缺血预处理作用。以心肌梗死面积和缺血及再灌注初期室性心律失常量化做为评定标准。分别给予不同浓度和给药方式的E4031,观察其对缺血再灌注导致的心肌梗死的影响。进一步预先给予KB-R7943,观察是否阻断IP或E4031预处理的保护作用。
6.我们在给予E4031前,预先给予5-HD(Mito KATP的特异性阻断剂)或Paxilline(Mito KCa的特异性阻断剂)观察是否对E4031的保护作用产生影响,另外,在E4031给药结束后,给予5-HD或Paxilline,持续至再灌注后10min,观察E4031的保护作用是否会被阻断。同样,我们预先给予KB-R7943,以观察其对Diazoxid(eMito KATP的激动剂)或NS1619(Mito KCa的激动剂)保护作用的影响,以心肌梗死面积和缺血及再灌注初期室性心律失常量化做为评定标准。
结果
1.当灌流液换为NMDG液后1min,[Ca~(2+)]i即开始升高,而且随着灌流时间的延长,[Ca~(2+)]i升高幅度逐渐增大;将NMDG溶液中Ca~(2+)浓度由1.25升高至1.8mmol/L时,[Ca~(2+)]i升高幅度进一步增大,用正常Tyrode液冲洗后,[Ca~(2+)]i逐渐恢复,表明胞外无钠法可以准确反映[Ca~(2+)]i的变化。
2.应用L-型钙通道的选择性阻断剂Nifedipine后,NMDG溶液引起的[Ca~(2+)]i升高未见明显改变。而给予Ni2+或KB-R7943,NMDG溶液引起的[Ca~(2+)]i几乎被完全抑制。所以,NMDG溶液引起的[Ca~(2+)]i升高与NCX的反向转运模式激活密切相关。
3.预先给予含10μmol/L E4031的正常Tyrode液10min,可以看到,由NMDG溶液引起的[Ca~(2+)]i上升明显增加,并且使用15μmol/L KB-R7943可以阻断E4031引起的[Ca~(2+)]i变化。这结果表明,E4031通过激活或增强NCX的反向转运模式而引起[Ca~(2+)]i升高。
4.代谢预处理的方法可以模拟缺血预处理,明显改善缺血再灌注后心肌细胞的收缩功能。而在代谢预处理前给予KB-R7943,这种保护作用却明显被减弱。预先给予E4031也可以明显改善心肌细胞缺血再灌注后的收缩功能,而这种保护作用同样可以被KB-R7943明显阻断。
5.在离体灌流心脏上,缺血预处理可以明显减少I/R导致的心肌梗死面积,并且减少在缺血及再灌注初期10min内恶性心律失常的发生,而KB-R7943可以阻断这些保护作用。5μmol/L E4031不论是持续给予10min还是5min间断给予2次,均可以明显减少心肌梗死面积,但间断给予的方式产生的保护作用更加明显,并且KB-R7943也可以阻断其作用。但单独使用KB-R7943却对I/R的损伤没有任何影响。
6.预先给予5-HD/Paxilline或在E4031给药结束后,给予5-HD/Paxilline的两种给药方式均能明显阻断E4031的预处理保护作用,但是Diazoxide/NS1619的药物预处理保护作用却不受预先给予的KB-R7943的影响。
结论
1.应用Fura-2/AM荧光法记录[Ca~(2+)]i浓度变化,建立了通过胞外无钠法对心室肌细胞Na+/Ca~(2+)交换体转运模式的研究方法,并运用该方法明确了E4031可以特异性地激活或增加NCX反向转运模式。
2.通过在单心肌细胞和整体心脏的研究发现,NCX的反向转运模式参与了缺血预处理期间[Ca~(2+)]i升高的调节,从而触发了IP的心肌保护作用。激活NCX反向转运模式的药物预处理可以明显改善I/R导致的心肌收缩功能下降,减少心肌梗死面积并降低室性心律失常的发生。
3.在IP保护机制中,NCX应位于Mito KATP和Mito KCa通道的上游,并且这两种钾通道不但调节NCX反向转运模式激活的作用通路,而且还作为效应分子直接发挥了保护作用。
Ischemic preconditioning (IP) is a phenomenon in which brief exposures of myocardium to ischemia render it more resistant to a subsequent and more severse insult, termed index ischemia. IP protects the heart against infarction and incidence of arrhythmias caused by ischemia and reperfusion. But the underlying mechanisms of IP remain unidentified. It is now known that a transient increase in cytosolic Ca~(2+) during preconditioning contributes to all these protective effects. It has been demonstrated that the reverse mode of Na+/Ca~(2+) exchanger (NCX) is activated during ischemia, contributing to intracellular Ca~(2+) overload and thus causing cardiac injury. In essence, IP are several procedures of short ischemia/reperfusion (I/R) in pathophysiological situation. We therefore hypothesized that increased reverse mode NCX activity during preconditioning may induce cardioprotection.
It has been found many end-effectors of IP. They play the central role in protecting structure and function of Mitochondrion. Many studies have been demonstrated that Mito KATP and Mito KCa act as trigger and/or mediator in IP and pharmacological preconditioning. Based on these finding, we hypothesized that both Mito KATP and Mito KCa channels are involved in the cardioprotective effects of prior stimulation of the reverse mode NCX.
Objective
1. The activity of reverse mode NCX was assessed by withdrawal extracellular Na+ and changes of [Ca~(2+)]i were measured with Fura/AM.
2. To observe the effcets of E4031 on the changes of [Ca~(2+)]i and its specificity for reverse mode NCX.
3. To evaluate the role of reverse mode NCX in the cardioprotection of MIP in rat ventricular myocytes.
4. In perfused isolated rat heart, to observe the effect of prior stimulated reverse mode NCX on infarct size and arrhythmias caused by I/R.
5. To evaluate the hypothesis that Mito KATP and Mito KCa channels are involved in the cardioprotective effects of prior stimulation of the reverse mode NCX.
Methods
1. Single ventricular myocytes were enzymatically isolated and loaded with Fura-2/AM. Withdrawal extracellular Na+ and changes of [Ca~(2+)]i were measured.
2. Cells were pretreated with ryanodine and thapsigargin to block the sarcoplasmic reticulum function. Administered nifedipine (inhibitor of L-type calcium channel) or Ni2+/KB-R7943 (inhibitor of reverse mode NCX) and the changes of [Ca~(2+)]i were measured.
3. pretreated with E4031, the activity of reverse mode NCX was assessed by withdrawal extracellular Na+ and changes of [Ca~(2+)]i were measured with/without KB-R7943.
4. Myocytes were subjected to metabolic inhibition preconditioning for 30 min. E4031 pretreatment was administered to the electrically stimulated ventricular myocytes for 10 min. After that, the cells were perfused with normal Tyrode solution with 0.2% bovine serum albumin for 10 min before they were subjected to severe metabolic inhibition for 8 min followed by reperfusion with normal solution for 10 min. KB-R7943 was given 5min before and during metabolic inhibition preconditioning or E4031 pretreatment.
5. In perfused isolated rat heart, the ischemia or reperfusion by was occluding or loosing the coronary artery. IP was produced by 2 cycles of 5min regional ischemia followed by 5min reperfusion. Assessed the cardioprotection by infarct size and arrhythmia scores. E4031 was perfused with different concentration and for different time. KB-R7943 was perfused for a period of 5 min before the first ischemic episode to 5 min after the second ischemic episode, or E4031 pretreatment.
6. 5-HD (a selective blocker of the mitoKATP), or paxilline (a selective blocker of the mitoKCa ) was either given for 30 min before 30 min of ischemia, or given from 5 min before 30 min of ischemia until 10 min after reperfusion. KB-R7943 was administered from 10 min before to 10 min after one cycle of 10 min treatment with diazoxide (a selective activator of the mitoKATP), or NS1619 (a selective activator of the mitoKCa ). Assessed the cardioprotection by infarct size and arrhythmia scores.
Results
1. [Ca~(2+)]i was increased by the increasing of [Ca~(2+)]o after ventricular myocytes exposed to Na+-free solution (NMDG solution), and returned to baseline following washout.
2. Nifedipine did not affect NMDG solution-induced increases of [Ca~(2+)]i , while both Ni2+ and KB-R7943 almost completely blocked the effect, an indication of the reverse mode NCX.
3. E4031 significantly increased the change of [Ca~(2+)]i upon NMDG, which was also blocked by KB-R7943.
4. In isolated ventricular myocytes, MIP significantly increased the amplitude of twitch contraction. Pretreatment with E4031 also significantly increased the cell contraction, and the effect of MIP or E4031 was abolished by KB-R7943.
5. Preconditioning with two cycles of 5 min ischemia each or pretreatment with E4031, significantly reduced the infarct size, and the effect was abolished by 5μM KB-R7943. Exposure to E4031 at 3 - 5μM for two cycles of 5 min each reduced the infarct size caused by index ischemia in a concentration-dependent manner. Exposure to E4031 at 5μM for one cycle of 10 min also significantly reduced the infarct size, but its effect was significantly weaker than that for twocycles. Similar to IP, preconditioning the heart with E4031 also significantly reduced the arrhythmia scores during index ischemia and reperfusion, and the anti-arrhythmic effect of IP and E-4301 pretreatment were abolished by KB-R7943, which itself had no effect.
6. Treatment of the heart with 5-HD/paxilline, 30 min before ischemia, abolished the beneficial effects of E4031 on infarct size and arrhythmias, and administration of 5-HD/paxilline from 5 min before the 30-min ischemia until 10 min after reperfusion also abolished the cardioprotective effect of E4031. 5-HD/paxilline alone had no effect. Activation of the mito KATP/ KCa channel with Diazoxide/ NS1619 also reduced infarct size and arrhythmias and the protective effects were blocked by 5-HD/paxilline, These effects were not affected by administration of KB-R7943 before index ischemia.
Conclusions
1. The activity of reverse mode NCX was assessed by withdrawal extracellular Na+. It has been demonstrated that the special effcets of E4031 on activation/augmentation of reverse mode NCX.
2. Reverse-mode NCX activation during preconditioning ischemia triggered cardioprotection by increased [Ca~(2+)]i . Prior stimulation of the reverse-mode NCX ameliorated the contraction in rat ventricular myocytes and reduced the infarct size and arrhythmia in perfusion isolated rat heart.
3. Both mitoKATP and mitoKCa channels mediate the protective effects of prior stimulation of the reverse-mode NCX against infarct size and arrhythmias caused by ischemia and reperfusion, and that the NCX is located upstream from both the mitoKATP and mitoKCa channels.
缺血预处理的下游保护效应分子众多,但保护线粒体结构和功能的完整是缺血预处理保护作用机制中的关键,所以,探讨激活NCX的反向转运模式触发缺血预处理的保护作用的下游作用机制具有重要的意义。线粒体上的两种钾离子通道已经证明是缺血预处理保护作用的关键通道,因此探讨NCX引起[Ca~(2+)]i升高是否通过开放这两种离子通道从而中发挥了心肌保护作用,并且这两种离子通道在其作用机制中所扮演的角色,将为我们深入理解心肌缺血预处理的病理生理过程及其作用机制提供新的思路。
目的
1.明确胞外无钠法对大鼠心室肌细胞Na~+/Ca~(2+)交换体转运模式的影响;
2.观察E4031对NCX反向转运模式的影响,并明确其特异性程度;
3.在单心肌模型上观察缺血预处理期间,激活或阻断NCX的反向转运模式对预处理的心肌保护作用的影响;
4.在整体心脏模型上观察NCX的反向转运模式是否触发缺血预处理的保护作用。
5.观察Mito KATP和Mito KCa两种线粒体钾通道在激活NCX的药物预处理中的作用,并进一步完善IP的作用机制。
方法
1.以常规酶解法分离成年大鼠心肌细胞,选取形态、功能良好的心室肌细胞进行实验观察。应用胞外无钠法(NMDG)检测并通过Fura-2/AM荧光法记录[Ca~(2+)]i的变化。
2.预先给予Ryanodine和Thapsigargin孵育10min以阻断SR功能,分别给予Nifedipine(L-型钙通道阻断剂)和Ni2+、KB-R7943(NCX反向转运模式阻断剂),观察记录[Ca~(2+)]i的升高幅度变化。
3.给予E4031预先孵育后,观察NMDG导致的[Ca~(2+)]i的升高幅度变化,并观察给予KB-R7943对E4031引起的[Ca~(2+)]i变化的影响。
4.单心肌代谢抑制液孵育30min后,给予正常Tyrode液灌洗10min,通过代谢抑制法模拟缺血预处理的保护作用。此后再给予缺血液8min,正常Tyrode液灌洗10min制造缺血再灌注损伤。分别观察E4031预处理能否模拟MIP作用,KB-R7943能否阻断MIP和E4031的保护作用。
5.制备Langendorff离体心脏灌流模型,通过结扎/松开冠脉左前降支制造缺血/再灌注。通过2次5min的缺血再灌注过程模拟缺血预处理作用。以心肌梗死面积和缺血及再灌注初期室性心律失常量化做为评定标准。分别给予不同浓度和给药方式的E4031,观察其对缺血再灌注导致的心肌梗死的影响。进一步预先给予KB-R7943,观察是否阻断IP或E4031预处理的保护作用。
6.我们在给予E4031前,预先给予5-HD(Mito KATP的特异性阻断剂)或Paxilline(Mito KCa的特异性阻断剂)观察是否对E4031的保护作用产生影响,另外,在E4031给药结束后,给予5-HD或Paxilline,持续至再灌注后10min,观察E4031的保护作用是否会被阻断。同样,我们预先给予KB-R7943,以观察其对Diazoxid(eMito KATP的激动剂)或NS1619(Mito KCa的激动剂)保护作用的影响,以心肌梗死面积和缺血及再灌注初期室性心律失常量化做为评定标准。
结果
1.当灌流液换为NMDG液后1min,[Ca~(2+)]i即开始升高,而且随着灌流时间的延长,[Ca~(2+)]i升高幅度逐渐增大;将NMDG溶液中Ca~(2+)浓度由1.25升高至1.8mmol/L时,[Ca~(2+)]i升高幅度进一步增大,用正常Tyrode液冲洗后,[Ca~(2+)]i逐渐恢复,表明胞外无钠法可以准确反映[Ca~(2+)]i的变化。
2.应用L-型钙通道的选择性阻断剂Nifedipine后,NMDG溶液引起的[Ca~(2+)]i升高未见明显改变。而给予Ni2+或KB-R7943,NMDG溶液引起的[Ca~(2+)]i几乎被完全抑制。所以,NMDG溶液引起的[Ca~(2+)]i升高与NCX的反向转运模式激活密切相关。
3.预先给予含10μmol/L E4031的正常Tyrode液10min,可以看到,由NMDG溶液引起的[Ca~(2+)]i上升明显增加,并且使用15μmol/L KB-R7943可以阻断E4031引起的[Ca~(2+)]i变化。这结果表明,E4031通过激活或增强NCX的反向转运模式而引起[Ca~(2+)]i升高。
4.代谢预处理的方法可以模拟缺血预处理,明显改善缺血再灌注后心肌细胞的收缩功能。而在代谢预处理前给予KB-R7943,这种保护作用却明显被减弱。预先给予E4031也可以明显改善心肌细胞缺血再灌注后的收缩功能,而这种保护作用同样可以被KB-R7943明显阻断。
5.在离体灌流心脏上,缺血预处理可以明显减少I/R导致的心肌梗死面积,并且减少在缺血及再灌注初期10min内恶性心律失常的发生,而KB-R7943可以阻断这些保护作用。5μmol/L E4031不论是持续给予10min还是5min间断给予2次,均可以明显减少心肌梗死面积,但间断给予的方式产生的保护作用更加明显,并且KB-R7943也可以阻断其作用。但单独使用KB-R7943却对I/R的损伤没有任何影响。
6.预先给予5-HD/Paxilline或在E4031给药结束后,给予5-HD/Paxilline的两种给药方式均能明显阻断E4031的预处理保护作用,但是Diazoxide/NS1619的药物预处理保护作用却不受预先给予的KB-R7943的影响。
结论
1.应用Fura-2/AM荧光法记录[Ca~(2+)]i浓度变化,建立了通过胞外无钠法对心室肌细胞Na+/Ca~(2+)交换体转运模式的研究方法,并运用该方法明确了E4031可以特异性地激活或增加NCX反向转运模式。
2.通过在单心肌细胞和整体心脏的研究发现,NCX的反向转运模式参与了缺血预处理期间[Ca~(2+)]i升高的调节,从而触发了IP的心肌保护作用。激活NCX反向转运模式的药物预处理可以明显改善I/R导致的心肌收缩功能下降,减少心肌梗死面积并降低室性心律失常的发生。
3.在IP保护机制中,NCX应位于Mito KATP和Mito KCa通道的上游,并且这两种钾通道不但调节NCX反向转运模式激活的作用通路,而且还作为效应分子直接发挥了保护作用。
Ischemic preconditioning (IP) is a phenomenon in which brief exposures of myocardium to ischemia render it more resistant to a subsequent and more severse insult, termed index ischemia. IP protects the heart against infarction and incidence of arrhythmias caused by ischemia and reperfusion. But the underlying mechanisms of IP remain unidentified. It is now known that a transient increase in cytosolic Ca~(2+) during preconditioning contributes to all these protective effects. It has been demonstrated that the reverse mode of Na+/Ca~(2+) exchanger (NCX) is activated during ischemia, contributing to intracellular Ca~(2+) overload and thus causing cardiac injury. In essence, IP are several procedures of short ischemia/reperfusion (I/R) in pathophysiological situation. We therefore hypothesized that increased reverse mode NCX activity during preconditioning may induce cardioprotection.
It has been found many end-effectors of IP. They play the central role in protecting structure and function of Mitochondrion. Many studies have been demonstrated that Mito KATP and Mito KCa act as trigger and/or mediator in IP and pharmacological preconditioning. Based on these finding, we hypothesized that both Mito KATP and Mito KCa channels are involved in the cardioprotective effects of prior stimulation of the reverse mode NCX.
Objective
1. The activity of reverse mode NCX was assessed by withdrawal extracellular Na+ and changes of [Ca~(2+)]i were measured with Fura/AM.
2. To observe the effcets of E4031 on the changes of [Ca~(2+)]i and its specificity for reverse mode NCX.
3. To evaluate the role of reverse mode NCX in the cardioprotection of MIP in rat ventricular myocytes.
4. In perfused isolated rat heart, to observe the effect of prior stimulated reverse mode NCX on infarct size and arrhythmias caused by I/R.
5. To evaluate the hypothesis that Mito KATP and Mito KCa channels are involved in the cardioprotective effects of prior stimulation of the reverse mode NCX.
Methods
1. Single ventricular myocytes were enzymatically isolated and loaded with Fura-2/AM. Withdrawal extracellular Na+ and changes of [Ca~(2+)]i were measured.
2. Cells were pretreated with ryanodine and thapsigargin to block the sarcoplasmic reticulum function. Administered nifedipine (inhibitor of L-type calcium channel) or Ni2+/KB-R7943 (inhibitor of reverse mode NCX) and the changes of [Ca~(2+)]i were measured.
3. pretreated with E4031, the activity of reverse mode NCX was assessed by withdrawal extracellular Na+ and changes of [Ca~(2+)]i were measured with/without KB-R7943.
4. Myocytes were subjected to metabolic inhibition preconditioning for 30 min. E4031 pretreatment was administered to the electrically stimulated ventricular myocytes for 10 min. After that, the cells were perfused with normal Tyrode solution with 0.2% bovine serum albumin for 10 min before they were subjected to severe metabolic inhibition for 8 min followed by reperfusion with normal solution for 10 min. KB-R7943 was given 5min before and during metabolic inhibition preconditioning or E4031 pretreatment.
5. In perfused isolated rat heart, the ischemia or reperfusion by was occluding or loosing the coronary artery. IP was produced by 2 cycles of 5min regional ischemia followed by 5min reperfusion. Assessed the cardioprotection by infarct size and arrhythmia scores. E4031 was perfused with different concentration and for different time. KB-R7943 was perfused for a period of 5 min before the first ischemic episode to 5 min after the second ischemic episode, or E4031 pretreatment.
6. 5-HD (a selective blocker of the mitoKATP), or paxilline (a selective blocker of the mitoKCa ) was either given for 30 min before 30 min of ischemia, or given from 5 min before 30 min of ischemia until 10 min after reperfusion. KB-R7943 was administered from 10 min before to 10 min after one cycle of 10 min treatment with diazoxide (a selective activator of the mitoKATP), or NS1619 (a selective activator of the mitoKCa ). Assessed the cardioprotection by infarct size and arrhythmia scores.
Results
1. [Ca~(2+)]i was increased by the increasing of [Ca~(2+)]o after ventricular myocytes exposed to Na+-free solution (NMDG solution), and returned to baseline following washout.
2. Nifedipine did not affect NMDG solution-induced increases of [Ca~(2+)]i , while both Ni2+ and KB-R7943 almost completely blocked the effect, an indication of the reverse mode NCX.
3. E4031 significantly increased the change of [Ca~(2+)]i upon NMDG, which was also blocked by KB-R7943.
4. In isolated ventricular myocytes, MIP significantly increased the amplitude of twitch contraction. Pretreatment with E4031 also significantly increased the cell contraction, and the effect of MIP or E4031 was abolished by KB-R7943.
5. Preconditioning with two cycles of 5 min ischemia each or pretreatment with E4031, significantly reduced the infarct size, and the effect was abolished by 5μM KB-R7943. Exposure to E4031 at 3 - 5μM for two cycles of 5 min each reduced the infarct size caused by index ischemia in a concentration-dependent manner. Exposure to E4031 at 5μM for one cycle of 10 min also significantly reduced the infarct size, but its effect was significantly weaker than that for twocycles. Similar to IP, preconditioning the heart with E4031 also significantly reduced the arrhythmia scores during index ischemia and reperfusion, and the anti-arrhythmic effect of IP and E-4301 pretreatment were abolished by KB-R7943, which itself had no effect.
6. Treatment of the heart with 5-HD/paxilline, 30 min before ischemia, abolished the beneficial effects of E4031 on infarct size and arrhythmias, and administration of 5-HD/paxilline from 5 min before the 30-min ischemia until 10 min after reperfusion also abolished the cardioprotective effect of E4031. 5-HD/paxilline alone had no effect. Activation of the mito KATP/ KCa channel with Diazoxide/ NS1619 also reduced infarct size and arrhythmias and the protective effects were blocked by 5-HD/paxilline, These effects were not affected by administration of KB-R7943 before index ischemia.
Conclusions
1. The activity of reverse mode NCX was assessed by withdrawal extracellular Na+. It has been demonstrated that the special effcets of E4031 on activation/augmentation of reverse mode NCX.
2. Reverse-mode NCX activation during preconditioning ischemia triggered cardioprotection by increased [Ca~(2+)]i . Prior stimulation of the reverse-mode NCX ameliorated the contraction in rat ventricular myocytes and reduced the infarct size and arrhythmia in perfusion isolated rat heart.
3. Both mitoKATP and mitoKCa channels mediate the protective effects of prior stimulation of the reverse-mode NCX against infarct size and arrhythmias caused by ischemia and reperfusion, and that the NCX is located upstream from both the mitoKATP and mitoKCa channels.
引文
1. Murry CE, Jennings RB, Reimer KA, 1986. Preconditioning with ischemia: a delay of lethal cell injury in ischemic myocardium. Circulation, 74 (5) :1124-1136.
2. Marber MS, Latchman DS, Walker JM, et al. 1993. Cardiac stress protein elevation 24 hours after brief ischemia or heat stress is associated with resistance to myocardial infarction. Circulation. 88 (3) :1264-1272.
3. Yellon DM, Baxter GF, Garcia-Dorado D, et al. 1998. Ischaemic preconditioning: present position and future directions. Cardio Res. 37 (1) :21-33.
4. Bolli R. 2000. The late phase of preconditioning. Circ Res. 87 (11) :972-983.
5. Latimer R, Murali B. 2003. Ischemic preconditioning: helping or hindering the natural protection. Middle East J Anesthesiol. 17 (1) : 83-89.
6. 李杰, 张尔永, 王兰兰,等. 2003. 缺血预处理对心肌的保护作用.中国胸心血管外科临床杂志. 10 (2) :120-123.
7. Wu ZK, Iivainen T, Pehkonen E. 2003. Perioperative and postoperative arrhythmia in three-vessel coronary artery disease patients and antiarrhythmic effects of ischemic preconditioning. Eur J Cardio Surg. 23 (4) : 578-584.
8. Tanoue Y, Herijgers P , Meuris B , et al. 2002. Ischemic preconditioning reduces unloaded myocardial oxygen consumption in an in-vivo sheep model . Cardio Res. 55 (3): 633-641
9. MATSUDA T,TAKUMA K,BABA A. 1997. Na+-Ca2+ exchanger: physiologyand pharmacology. Jpn J Pharmacol. 74:l-20.
10. NICOLL DA,LONGONI S,PHILIPSON KD. 1990. Molecular cloning and functional expression of the cardiac sarcolemmal Na+-Ca2+ exchanger. Science. 250:562-565.
11. LI Z,NICOLL DA,PHILIPSON KD. 1994. cloning of the NCX2 isoform of the plasma membrane Na+-Ca2+ exchanger. J Biol Chem. 269:17434-17439.
12. NICOLL DA,QUEDNAU BD,PHILIPSON KDl. 1996. Cloning of a third manmalian Na+-Ca2+ exchanger,NCX3. J Biol Chem. 271:24914-24921.
13. KIEVAL RS,BLOCH RJ,LEDERER WJl. 1992. Immunofluorescence localization of the Na+-Ca2+ exchanger in heart cells. Am J Physiol. 263:C545-C550.
14. HE Z,PHILIPSON KD,HILGEMANN DW. 1998. Cloning, expression, and characterization of the squid Na+-Ca2+ exchanger (NCX-SQ1). J Gen Physiol. 111:857 一 873.
15. SCHWARZ EM,BENZER S. 1997. Calx,a Na+-Ca2+ exchanger gene of Drosophila melanogaster. Proc Natl Acad Sci USA. 94:10249-10254.
16. SHIGEKAWA M,IWAMOTO T. 2001. Cardiac Na+-Ca2+ exchanger: Molecular and pharmacological Aspects.Cric Res. 88:864-876.
17. NICOLL DA,PHILIPSON KD.1991. Molecular studies of the cardiac sarcolemmal sodium-calcium exchanger. Ann NY Acad Sci. 639:181-188.
18. BARCENAS-RUIZ L,BEUCHELMAN DJ,WIER WG. 1987. Sodium-calcium exchanger in heart: membrane currents and change in [Ca2+]i .Scinece. 238:1720-1722.
19. HILGEMANN DW,NICOLL DA,PHILIPSON KD.1991. Charge movement during Na+ translocation by native and cloned cardiac Na+-Ca2+ exchanger. Nature. 352(6337):715-718.
20. BERS DM. 2002. Cardiac excitation-contraction coupling. Nature. 415:198-205.
21. MATSUOKA S. 2004. Forefront of Na+-Ca2+ exchang studies: RegulationKineticsof Na+-Ca2+ exchanger. J Pharmacol Sci. 96:12-14.
22. BLAUSTEIN MP,LEDERER WJ. 1999. Sodium/Calcium Exchangers: Its Physiological Implications. Physiol Rev. 79:763-854.
23. SHIGEKAWA M,IWAMOTO T. 2001. Cardiac Na+-Ca2+ exchanger. Molecular and pharmacological aspects. Circ Res. 88:86-4876.
24. IWAMOTO T,WATANO T,SHIGEKAWA M. 1996. A novel isothiourea derivative selectively inhibits the reverse mode of Na+-Ca2+ exchange in cells expressing NCX1. J Biol Chem. 271:22391-2397.
25. WATANO T,KIMURA J,MORITA T. 1996. A novel antagonist, No.7943,of the Na+-Ca2+ exchange current in guinea-pig cardiac ventricular cells. Br J Pharmacol. 119:555-63.
26. IWAMOTO T,KITA S. 2004. Development and application of Na+-Ca2+ exchange inhibitors. Mol Cell Biochem. 259:157-161.
27. MATSUDA T,ARAKAWA N,TAKUMA K. 2001. SEA0400,a novel and selection inhibitor of the Na+-Ca2+ exchange, attenuates reperfusion injury in the in vitro and in vivo cerebral ischemic models. J Pharmacol Exp Ther. 298:24-9256.
28. IWAMOTO T,INOUE Y,ITO K. 2004. The exchanger inhibitory peptide region-dependent inhibition of Na+-Ca2+ exchange by SN-6, a novel benzyloxyphenyl derivative. Mol Pharmacol. 66:45-55.
29. HILGEMANN DW, 1990, Regulation and deregulation of cardiac Na+-Ca2+ exchange in giant excised sarcolemmal membrane patches. Nature. 344(6263):242-245
30. MOTSUOKA S,HILGEMANN DW. 1994. Inactivation of outward Na+-Ca2+ exchange current in guinea-pig ventricular myocytes. J Physiol. 476(3):443-58.
31. MOTSUOKA S,NICOLL DA,HE Z. 1997. Regulation of cardiac Na+-Ca2+exchange by the endogenous XIP regionh. J Gen physiol. 109(2):273-56.
32. DIPOLO R,BERBERIAN G, DELGADO D. 1997. A novel 13 kDa cytoplasmic soluble protein is required for the nucleotide (MgATP) modulation of the Na+-Ca2+ exchange in squid nerve fibers. FEBS Lett. 401(l):6-10.
33. PHLIPSON KD,NICOLL DA. 1993. Molecular and kinetic aspects of sodium-calcium exchanger. Int Rev Cytol. 137C:199-227.
34. HILGMANN DW,BALL R. 1996. Regulation of cardiac Na+-Ca2+ exchanger and KATP potassium channels by PIP2. Seinece. 273:956-959.
35. DOERING AE,LEDERE WJ. 1994. The action of Na+ as a cofactor in the inhibition by cytoplasmic protons of the cardiac Na+-Ca2+ exchanger in the guinea-pig. J Physiol. 480:9-20.
36. NIGGLI E,LEDERER WJ. 1991. Molecular operations of the sodium-calcium exchanger revealed by conformation currents. Nature. 349:621-624.
37. KAPPL M,HARTUNG K. 1996. Rapid charge translocation by cardiac Na+-Ca2+ exchanger after a Ca2+ concentration jump. Biophys J. 71:2473-2455.
38. WRIGHT AR, REES SA,VANDENBERG JI. 1995. Extracellular osmotic pressure modulates sodium-calcium exchange in isolated guinea-pig ventricular myocytes. J Physlol. 488:293-301.
39. WEBER CR, PIACENTINO V, GINSBURG KS. 2002. Na+-Ca2+ exchange current and submembrane Ca2+ during the cardiac action potential. Circ Res. 90:182-189.
40. LINASK KK,HAN MD,ARTMAN M. 2001. Sodium-calcium exchange (NCX1) and calcium modulation: NCX protein expression patterns and regulation of early heart development. Dev dyn. 221:249-264.
41. HATEM SN, SHAM JS, MORAD M. 1994. Enhanced Na+-Ca2+ exchangeractivity in cardiomyopathic Syrian hamster. Circ Res. 74:253-261.
42. POGWIZD SM,SCHLOTTHAUER K,LI L. 2001. Arrhythmogenesis and contractile dysfunction in heart failure: role of sodium-calcium exchanger, inward rectifier potassium current, and residual beta-adrenergic responsiveness. Circ Res. 88:1159-1167.
43. GAUGHAN JP,FURUKAWA S, JEEVANANDAM V. 1999. Sodium/calcium exchange contributes to contraction and relaxation in failed human ventricular myocytes. Am J Physiol. 277:H714-724.
44. BENARDEAU A,HATEM SN,MARTIN CR. 1996. Contribution of Na+-Ca2+ exchanger to action potential of human atrial myocyte. Am J Physio. 271:H1151-1161
45. 贾宏钧,王钟林,杨期东,等. 离子通道心血管病理生理与心脑血管疾病基础与临床[M ]. 第一版,北京:人民卫生出版社, 2001. 25269.
46. Liu Y, Ren G, O’Rourke B. 2001. Pharmacological comparison of native mitochondrial KATP channels with molecularly defined surface KATP channels. Mol Pharmacol. 59 (2) : 2252230.
47. Seharaseyon J, Ohler A, Sasaki N. 2000. Molecular composition of mitochondrial ATP-sensitive potassium channels probed by viral Kir gene transfer. J Mol Cell Cardiol. 32 (11) : 1923-1930.
48. Sato T, SasakiN, Seharaseyon J. 2000. Selective pharmacological agents implicate mitochondrial but not sarcolemmal KATP channels in ischemic cardiop rotection. Circulation. 101 (20) : 2418-2423.
49. SuzukiM, Saito T, Sato T. 2003. Cardioprotective effect of diazoxide is mediated by activation of sarcolemmal but not mitochondrial ATP-sensitive potassium channels in mice. Circulation. 107 (5) : 682-685.
50. Peart J, Headrick JP. 2003. Adenosine-mediated early preconditioning in mouse: protective signaling and concentration dependent effects. Cardiovasc Res. 58 (3) : 589-601.
51. Chen M, Zhou JJ , Kam KW. 2003. Roles of KATP channels in delayed cardioprotection and intracellular Ca2+ in the rat heart as revealed by kappa-opioid receptor stimulation with U50488H. Br J Pharmacol. 140 (4) : 750-758.
52. Das B, Sarkar C. 2003. Cardiomyocyte mitochondrial KATP channels participate in the antiarrhythmic and antiinfarct effects of KATP activators during ischemia and reperfusion in an intact anesthetized rabbitmodel. Pol J Pharmacol. 55(5) : 771-786.
53. Halestrap AP. 1994. Regulation of mitochondrial metabolism through changes in matrix volume. Biochem Soc Trans.22 (2) : 522-529.
54. Gardos G. 1958. The function of calcium in the potassium permeability of human erythrocytes. Biochim. Biophys. Acta. 30: 653–654.
55. AlgerBE, Nicoll RA. 1980. Epileptiform burst afterhyperolarization: calcium-dependent potassium potential in hippocampal CA1 pyramidal cells. Science. 210: 1122–1124.
56. Hotson JR, Prince DA. 1980. A calcium-activated hyperpolarization follows repetitive firing in hippocampal neurons. J Neurophysiol. 43: 409–419.
57. Schwartzkroin PA, Stafstrom CE. 1980. Effects of EGTA on the calcium-activated afterhyperpolarization in hippocampal CA3 pyramidal cells. Science. 210: 1125–1126.
58. Siemen D, Loupatatzis C, Borecky J, Gulbins E, Lang F. 1999. Ca2+-activated K channel of the BK-type in the inner mitochondrialmembrane of a human glioma cell line. Biochem Biophys Res Commun. 257 : 549–554.
59. Xu W, Liu Y, Wang S, McDonald T, Van Eyk JE, Sidor A, O’Rourke B. 2002. Cytoprotective role of Ca2+- activated K+ channels in the cardiac inner mitochondrial membrane. Science. 298: 1029–1033.
60. Ardehali H,Cytoprotective channels in mitochondriaJ Bioenerg Biomembr. 2005 Jun;37(3):171-7.
61. Xiao R P, Spurgeon H A, O’Connor F, et al. 1994. Age-associated changes in β-adrenergic modulation on cardiac excitation-contraction coupling. J Clin Invest. 94: 2051-2059.
62. Ren J, Wold L E. 2001. Measurement of cardiac mechanical function in isolated ventricular myocytes from rats and mice by computerized video-based imaging. Biol Proced Online. 3: 43-53.
63. 牛建立,胡庆生,张宝仁. 1997. 应用F-uar2/AM和图像分析系统测定细胞内游离钙浓度.中华物理医学杂志. 19:92一94.
64. Sipido K R, Callewaert G. 1995. How to measure intracellular Ca2+ in single cardiac cells with fura-2 or indo-1. Cardiovasc Res; 29: 717-726.
65. Li SY, Golden KL, Jiang Y, Wang GJ, Privratsky JR, Zhang X, Eason AR, Culver B, Ren J, 2005. Inhibition of sarco(endo)plasmic reticulum Ca2+-ATPase differentially regulates contractile function in cardiac myocytes from normotensive and spontaneously hypertensive rats: role of Ca2+ regulatory proteins. Cell Biochem. Biophys. 42: 1–12.
66. Satoh H, Ginsburg KS, Qing K, Terada H, Hayashi H, Bers DM, 2000. KB-R7943 block of Ca2+ influx via Na+/Ca2+ exchange does not alter twitches or glycoside inotropy but prevents Ca2+ overload in rat ventricularmyocytes. Circulation 101:1441–1446.
67. Mochizuki S, MacLeod KT. 1997. Effects of hypoxia and metabolic inhibition on increases in intracellular Ca2+ concentration induced by Na+/Ca2+ exchange in isolated guinea-pig cardiacmyocytes. J Mol Cell Cardiol. 29:2979–2987.
68. Nayeem MA, Hess ML, Qian YZ, Loesser KE, Kukreja RC. 1997. Delayed preconditioning of cultured adult rat cardiac myocytes: role of 70- and 90-kDa heat stress proteins. Am J Physiol. 273: H861–H868.
69. Wang GY, Zhou JJ, Shan J,Wong TM. 2001. Protein kinase C-epsilon is a trigger of delayed cardioprotection against myocardial ischemia of kappa-opioid receptor stimulation in rat ventricular myocytes. J Pharmacol Exp Ther. 299: 603–610.
70. Wu S, Li HY, Wong TM. 1999. Cardioprotection of preconditioning by metabolic inhibition in the rat ventricular myocyte. Involvement of kappa-opioid receptor. Circ Res. 84: 1388–1395.
71. Miyawaki H, Ashraf M. 1997. Ca2+ as a mediator of ischemic preconditioning. Circ Res. 80: 790–799.
72. Node K, Kitakaze M, Sato H, Minamino T, Komamura K, Shinozaki Y, Mori H, Hori M. 1997. Role of intracellular Ca2+ in activation of protein kinase C during ischemic preconditioning. Circulation 96:1257–1265.
73. Przyklenk K, Hata K, Kloner RA. 1997. Is calcium a mediator of infarct size reduction with preconditioning in canine myocardium? Circulation 96:1305–1312.
74. 赵彩霞,郑延松,田敏.2001.细胞内游离钙检测方法进展.国外医学临床生物化学与检测学分册. 22(2):71-72.
75. MINTA A, KAO JP, TSIEN RY. 1989. Fluorescent indicators for cytosolic calcium based on rhodamine and fluoresce in chromophores. J Biolchem. 264(14):8171-8178.
76. TERRACCIANO CM, MACLEOD KT. 1997. Effects of lactate on the relative contribution of Ca2+ extrusion mechanisms to relaxation in guinea-pig ventricular myocytes. J Physiol. 500:557-570.
77. ORCHARD CH. 1987. The role of the sarcoplasmic reticulum in the response of ferret and rat heart muscle to acidosis. J Physio. 384:431-449.
78. ALLEN DG, LEE JA,SMITH GL.1989. The consequeneces of simulated ischemia on intercellular Ca2+ and tension in isolated ferret ventricular muscle. J Physiol. 410:297-323.
79. CAIRNS SP,WESTERBLAD H,ALLEN DG. 1993. Changes in myoplasmic pH and calcium concentration during expose to lactate in isolated rat ventricular myocytes. J Physiol. 464:561-574.
80. GAMBASSI G, HANSFORD RG, SOLLOTT SJ. 1993. Effects of acidosison resting cytosolic and mitochondrial Ca in mammaliam myocardium. J Gen Physiol. 102:575-597.
81. CLEVELAND JC. 1996. Ischemia Perconditioning in human and rat ventricular. Am J Physiol. 271:H1786-H1794.
82. OPIE LH. 2003. Preconditioning and metabolic anti-ischaemic agents. Eur Heart J. 24(20):1854-1856.
83. 程芳洲,余细球,唐国华等.2002.缺血预处理对大鼠缺血再灌注心肌细胞凋亡及相关基因蛋白表达的影响.中国组织化学与细胞化学杂志. 11(1):85-88.
84. TERRACCIANO CM,MACLEOD KT. 1994. Effects of acidosis onNa+/Ca2+ exchange and consequences for relaxation in guinea pig cardiac myocytes. Am J Phsiol. 267(2 Pt 2): H477-87.
85. MIYAWAKI H, ZHOU XB,ASHRAF M. 1996. Calcium preconditioning elicits strong protection against ischemia injury via Protein Kinase C signaling pathway. Circ Res. 79:137-146.
86. MURRY CE,RICHARD VJ,REIMER KA. 1990. Ischaemic preconditioning slow energy metabolism and delays ultrastructural damage during a sustained ischemic episodes. Circ Res. 66:913-931.
87. SMITH GB, STEFENELLI T,WU ST. 1996. Rapid adaptation of myocardial calcium homeostasis to short episodes of ischemia in isolated rat hearts. Am Heart J. 131:1106-1112.
88. PIERCE GN,Czubryt MP. 1995. The contribution of ionic imbalance to ischemia/reperfusion-induced injury. J Mol Cell Cardiol. 27:53-63.
89. STEENBERGEN C,FRALIX TA,MURPHY E. 1993. Role of increased cytosolic free calcium concentration in myocardial ischemic injury. Basic Res Cardiol. 88:456-470.
90. ASHRAF M,SULEIMAN J,AHMAD M. 1994. Ca2+ preconditioning elicits a unique protection against the Ca2+ paradox injury in rat heart. Role of adenosine. Fixed. Circ Res. 74:360-367.
91. PRZYKLENK K,HATA K,KLONER RA. 1997. Is calcium a mediator of infarct size reduction with preconditioning in canine myocardium . Circulation. 96: 1305-1312.
92. MIYAWAKI H, ASHRAF M.1997. Ca2+ as a mediator of ischemic preconditioning. Circ Res. 80:790-799.
93. Wang GY, Wu S, Pei JM, Yu XC, Wong TM. 2001. Kappa- but notdelta-opioid receptors mediate effects of ischemic preconditioning on both infarct and arrhythmia in rats. Am J Physiol. 280: H384-391.
94. Curtis MJ, Walker MJ. 1988. Quantification of arrhythmias using scoring systems: an examination of seven scores in an in vivo model of regional myocardial ischaemia. Cardiovasc Res. 22: 656-665.
95. Wang S, Cone J, Liu Y. 2001. Dual roles of mitochondrial KATP channels in diazoxide-mediated protection in isolated rabbit hearts. Am J Physiol. 280: H246-H255.
96. Gross GJ, Peart JN. 2003. KATP channels and myocardial preconditioning: an update. Am J Physiol. 285: H921-H930.
97. Yellon DM, Downey JM. 2003. Preconditioning the myocardium: from cellular physiology to clinical cardiology. Physiol Rev. 83: 1113-1151.
98. Kita H, Miura T, Tsuchida A, Hasegawa T, Shimamoto K. 1998. Suppression of reperfusion arrhythmias by preconditioning is inhibited by an ATP-sensitive potassium channel blocker, 5-hydroxydecanoate, but not by protein kinase C blockers in the rat. J Cardiovasc Pharmacol. 32: 791-797.
99. Vegh A, Parratt JR. 2002. The role of mitochondrial KATP channels in antiarrhythmic effects of ischaemic preconditioning in dogs. Br J Pharmacol. 137:1107-1115.
100. Driamov S, Bellahcene M, Ziegler A, Barbosa V, Traub D, Butz S, Buser PT, Zaugg CE. 2004. Antiarrhythmic effect of ischemic preconditioning during low-flow ischemia. The role of bradykinin and sarcolemmal versus mitochondrial ATP-sensitive K+ channels. Basic Res Cardiol. 99: 299-308.
101. Cao CM, Chen M, Wong TM. 2005. The KCa channel as a trigger for the cardioprotection induced by kappa-opioid receptor stimulation -- itsrelationship with protein kinase C. Br J Pharmacol. 145: 984-991.
102. Shintani Y, Node K, Asanuma H, Sanada S, Takashima S, Asano Y, Liao Y, Fujita M, Hirata A, Shinozaki Y, Fukushima T, Nagamachi Y, Okuda H, Kitakaze M. 2004. Opening of Ca2+-activated K+ channels is involved in ischemic preconditioning in canine hearts. J Mol Cell Cardiol. 37:1213-1218.
103. Gao Q, Zhang SZ, Cao CM, Bruce IC, Xia Q. 2005. The mitochondrial permeability transition pore and the Ca2+-activated K+ channel contribute to the cardioprotection conferred by tumor necrosis factor-alpha. Cytokine 32: 199-205.
104. O'Rourke B. 2004. Evidence for mitochondrial K+ channels and their role in cardioprotection.Circ. Res. 94: 420-432.
105. Ghatta S, Nimmagadda D, Xu X, O'Rourke ST. 2006. Large-conductance, calcium-activated potassium channels: structural and functional implications. Pharmacol Ther. 110: 103-116.
106. O'Rourke B. 2000. Pathophysiological and protective roles of mitochondrial ion channels. J Physiol. (Lond.). 529 Pt 1: 23-36.
107. Akabas MH. 2004. Na+/Ca2+ exchange inhibitors: potential drugs to mitigate the severity of ischemic injury. Mol Pharmacol. 66: 8-10.
108. Reuter H, Henderson SA, Han T, Matsuda T, Baba A, Ross RS, Goldhaber JI, Philipson KD. 2002. Knockout mice for pharmacological screening: testing the specificity of Na+-Ca2+ exchange inhibitors. Circ Res. 91: 90-92.
109. Nishida M, Urushidani T, Sakamoto K, Nagao T. 1999. L-cis diltiazem attenuates intracellular Ca2+ overload by metabolic inhibition in guinea pigmyocytes. Eur J Pharmacol. 385: 225-230.
110. Watano T, Harada Y, Harada K, Nishimura N. 1999. Effect of Na+/Ca2+ exchange inhibitor, KB-R7943 on ouabain-induced arrhythmias in guinea-pigs. Br J Pharmacol. 127: 1846-1850.
2. Marber MS, Latchman DS, Walker JM, et al. 1993. Cardiac stress protein elevation 24 hours after brief ischemia or heat stress is associated with resistance to myocardial infarction. Circulation. 88 (3) :1264-1272.
3. Yellon DM, Baxter GF, Garcia-Dorado D, et al. 1998. Ischaemic preconditioning: present position and future directions. Cardio Res. 37 (1) :21-33.
4. Bolli R. 2000. The late phase of preconditioning. Circ Res. 87 (11) :972-983.
5. Latimer R, Murali B. 2003. Ischemic preconditioning: helping or hindering the natural protection. Middle East J Anesthesiol. 17 (1) : 83-89.
6. 李杰, 张尔永, 王兰兰,等. 2003. 缺血预处理对心肌的保护作用.中国胸心血管外科临床杂志. 10 (2) :120-123.
7. Wu ZK, Iivainen T, Pehkonen E. 2003. Perioperative and postoperative arrhythmia in three-vessel coronary artery disease patients and antiarrhythmic effects of ischemic preconditioning. Eur J Cardio Surg. 23 (4) : 578-584.
8. Tanoue Y, Herijgers P , Meuris B , et al. 2002. Ischemic preconditioning reduces unloaded myocardial oxygen consumption in an in-vivo sheep model . Cardio Res. 55 (3): 633-641
9. MATSUDA T,TAKUMA K,BABA A. 1997. Na+-Ca2+ exchanger: physiologyand pharmacology. Jpn J Pharmacol. 74:l-20.
10. NICOLL DA,LONGONI S,PHILIPSON KD. 1990. Molecular cloning and functional expression of the cardiac sarcolemmal Na+-Ca2+ exchanger. Science. 250:562-565.
11. LI Z,NICOLL DA,PHILIPSON KD. 1994. cloning of the NCX2 isoform of the plasma membrane Na+-Ca2+ exchanger. J Biol Chem. 269:17434-17439.
12. NICOLL DA,QUEDNAU BD,PHILIPSON KDl. 1996. Cloning of a third manmalian Na+-Ca2+ exchanger,NCX3. J Biol Chem. 271:24914-24921.
13. KIEVAL RS,BLOCH RJ,LEDERER WJl. 1992. Immunofluorescence localization of the Na+-Ca2+ exchanger in heart cells. Am J Physiol. 263:C545-C550.
14. HE Z,PHILIPSON KD,HILGEMANN DW. 1998. Cloning, expression, and characterization of the squid Na+-Ca2+ exchanger (NCX-SQ1). J Gen Physiol. 111:857 一 873.
15. SCHWARZ EM,BENZER S. 1997. Calx,a Na+-Ca2+ exchanger gene of Drosophila melanogaster. Proc Natl Acad Sci USA. 94:10249-10254.
16. SHIGEKAWA M,IWAMOTO T. 2001. Cardiac Na+-Ca2+ exchanger: Molecular and pharmacological Aspects.Cric Res. 88:864-876.
17. NICOLL DA,PHILIPSON KD.1991. Molecular studies of the cardiac sarcolemmal sodium-calcium exchanger. Ann NY Acad Sci. 639:181-188.
18. BARCENAS-RUIZ L,BEUCHELMAN DJ,WIER WG. 1987. Sodium-calcium exchanger in heart: membrane currents and change in [Ca2+]i .Scinece. 238:1720-1722.
19. HILGEMANN DW,NICOLL DA,PHILIPSON KD.1991. Charge movement during Na+ translocation by native and cloned cardiac Na+-Ca2+ exchanger. Nature. 352(6337):715-718.
20. BERS DM. 2002. Cardiac excitation-contraction coupling. Nature. 415:198-205.
21. MATSUOKA S. 2004. Forefront of Na+-Ca2+ exchang studies: RegulationKineticsof Na+-Ca2+ exchanger. J Pharmacol Sci. 96:12-14.
22. BLAUSTEIN MP,LEDERER WJ. 1999. Sodium/Calcium Exchangers: Its Physiological Implications. Physiol Rev. 79:763-854.
23. SHIGEKAWA M,IWAMOTO T. 2001. Cardiac Na+-Ca2+ exchanger. Molecular and pharmacological aspects. Circ Res. 88:86-4876.
24. IWAMOTO T,WATANO T,SHIGEKAWA M. 1996. A novel isothiourea derivative selectively inhibits the reverse mode of Na+-Ca2+ exchange in cells expressing NCX1. J Biol Chem. 271:22391-2397.
25. WATANO T,KIMURA J,MORITA T. 1996. A novel antagonist, No.7943,of the Na+-Ca2+ exchange current in guinea-pig cardiac ventricular cells. Br J Pharmacol. 119:555-63.
26. IWAMOTO T,KITA S. 2004. Development and application of Na+-Ca2+ exchange inhibitors. Mol Cell Biochem. 259:157-161.
27. MATSUDA T,ARAKAWA N,TAKUMA K. 2001. SEA0400,a novel and selection inhibitor of the Na+-Ca2+ exchange, attenuates reperfusion injury in the in vitro and in vivo cerebral ischemic models. J Pharmacol Exp Ther. 298:24-9256.
28. IWAMOTO T,INOUE Y,ITO K. 2004. The exchanger inhibitory peptide region-dependent inhibition of Na+-Ca2+ exchange by SN-6, a novel benzyloxyphenyl derivative. Mol Pharmacol. 66:45-55.
29. HILGEMANN DW, 1990, Regulation and deregulation of cardiac Na+-Ca2+ exchange in giant excised sarcolemmal membrane patches. Nature. 344(6263):242-245
30. MOTSUOKA S,HILGEMANN DW. 1994. Inactivation of outward Na+-Ca2+ exchange current in guinea-pig ventricular myocytes. J Physiol. 476(3):443-58.
31. MOTSUOKA S,NICOLL DA,HE Z. 1997. Regulation of cardiac Na+-Ca2+exchange by the endogenous XIP regionh. J Gen physiol. 109(2):273-56.
32. DIPOLO R,BERBERIAN G, DELGADO D. 1997. A novel 13 kDa cytoplasmic soluble protein is required for the nucleotide (MgATP) modulation of the Na+-Ca2+ exchange in squid nerve fibers. FEBS Lett. 401(l):6-10.
33. PHLIPSON KD,NICOLL DA. 1993. Molecular and kinetic aspects of sodium-calcium exchanger. Int Rev Cytol. 137C:199-227.
34. HILGMANN DW,BALL R. 1996. Regulation of cardiac Na+-Ca2+ exchanger and KATP potassium channels by PIP2. Seinece. 273:956-959.
35. DOERING AE,LEDERE WJ. 1994. The action of Na+ as a cofactor in the inhibition by cytoplasmic protons of the cardiac Na+-Ca2+ exchanger in the guinea-pig. J Physiol. 480:9-20.
36. NIGGLI E,LEDERER WJ. 1991. Molecular operations of the sodium-calcium exchanger revealed by conformation currents. Nature. 349:621-624.
37. KAPPL M,HARTUNG K. 1996. Rapid charge translocation by cardiac Na+-Ca2+ exchanger after a Ca2+ concentration jump. Biophys J. 71:2473-2455.
38. WRIGHT AR, REES SA,VANDENBERG JI. 1995. Extracellular osmotic pressure modulates sodium-calcium exchange in isolated guinea-pig ventricular myocytes. J Physlol. 488:293-301.
39. WEBER CR, PIACENTINO V, GINSBURG KS. 2002. Na+-Ca2+ exchange current and submembrane Ca2+ during the cardiac action potential. Circ Res. 90:182-189.
40. LINASK KK,HAN MD,ARTMAN M. 2001. Sodium-calcium exchange (NCX1) and calcium modulation: NCX protein expression patterns and regulation of early heart development. Dev dyn. 221:249-264.
41. HATEM SN, SHAM JS, MORAD M. 1994. Enhanced Na+-Ca2+ exchangeractivity in cardiomyopathic Syrian hamster. Circ Res. 74:253-261.
42. POGWIZD SM,SCHLOTTHAUER K,LI L. 2001. Arrhythmogenesis and contractile dysfunction in heart failure: role of sodium-calcium exchanger, inward rectifier potassium current, and residual beta-adrenergic responsiveness. Circ Res. 88:1159-1167.
43. GAUGHAN JP,FURUKAWA S, JEEVANANDAM V. 1999. Sodium/calcium exchange contributes to contraction and relaxation in failed human ventricular myocytes. Am J Physiol. 277:H714-724.
44. BENARDEAU A,HATEM SN,MARTIN CR. 1996. Contribution of Na+-Ca2+ exchanger to action potential of human atrial myocyte. Am J Physio. 271:H1151-1161
45. 贾宏钧,王钟林,杨期东,等. 离子通道心血管病理生理与心脑血管疾病基础与临床[M ]. 第一版,北京:人民卫生出版社, 2001. 25269.
46. Liu Y, Ren G, O’Rourke B. 2001. Pharmacological comparison of native mitochondrial KATP channels with molecularly defined surface KATP channels. Mol Pharmacol. 59 (2) : 2252230.
47. Seharaseyon J, Ohler A, Sasaki N. 2000. Molecular composition of mitochondrial ATP-sensitive potassium channels probed by viral Kir gene transfer. J Mol Cell Cardiol. 32 (11) : 1923-1930.
48. Sato T, SasakiN, Seharaseyon J. 2000. Selective pharmacological agents implicate mitochondrial but not sarcolemmal KATP channels in ischemic cardiop rotection. Circulation. 101 (20) : 2418-2423.
49. SuzukiM, Saito T, Sato T. 2003. Cardioprotective effect of diazoxide is mediated by activation of sarcolemmal but not mitochondrial ATP-sensitive potassium channels in mice. Circulation. 107 (5) : 682-685.
50. Peart J, Headrick JP. 2003. Adenosine-mediated early preconditioning in mouse: protective signaling and concentration dependent effects. Cardiovasc Res. 58 (3) : 589-601.
51. Chen M, Zhou JJ , Kam KW. 2003. Roles of KATP channels in delayed cardioprotection and intracellular Ca2+ in the rat heart as revealed by kappa-opioid receptor stimulation with U50488H. Br J Pharmacol. 140 (4) : 750-758.
52. Das B, Sarkar C. 2003. Cardiomyocyte mitochondrial KATP channels participate in the antiarrhythmic and antiinfarct effects of KATP activators during ischemia and reperfusion in an intact anesthetized rabbitmodel. Pol J Pharmacol. 55(5) : 771-786.
53. Halestrap AP. 1994. Regulation of mitochondrial metabolism through changes in matrix volume. Biochem Soc Trans.22 (2) : 522-529.
54. Gardos G. 1958. The function of calcium in the potassium permeability of human erythrocytes. Biochim. Biophys. Acta. 30: 653–654.
55. AlgerBE, Nicoll RA. 1980. Epileptiform burst afterhyperolarization: calcium-dependent potassium potential in hippocampal CA1 pyramidal cells. Science. 210: 1122–1124.
56. Hotson JR, Prince DA. 1980. A calcium-activated hyperpolarization follows repetitive firing in hippocampal neurons. J Neurophysiol. 43: 409–419.
57. Schwartzkroin PA, Stafstrom CE. 1980. Effects of EGTA on the calcium-activated afterhyperpolarization in hippocampal CA3 pyramidal cells. Science. 210: 1125–1126.
58. Siemen D, Loupatatzis C, Borecky J, Gulbins E, Lang F. 1999. Ca2+-activated K channel of the BK-type in the inner mitochondrialmembrane of a human glioma cell line. Biochem Biophys Res Commun. 257 : 549–554.
59. Xu W, Liu Y, Wang S, McDonald T, Van Eyk JE, Sidor A, O’Rourke B. 2002. Cytoprotective role of Ca2+- activated K+ channels in the cardiac inner mitochondrial membrane. Science. 298: 1029–1033.
60. Ardehali H,Cytoprotective channels in mitochondriaJ Bioenerg Biomembr. 2005 Jun;37(3):171-7.
61. Xiao R P, Spurgeon H A, O’Connor F, et al. 1994. Age-associated changes in β-adrenergic modulation on cardiac excitation-contraction coupling. J Clin Invest. 94: 2051-2059.
62. Ren J, Wold L E. 2001. Measurement of cardiac mechanical function in isolated ventricular myocytes from rats and mice by computerized video-based imaging. Biol Proced Online. 3: 43-53.
63. 牛建立,胡庆生,张宝仁. 1997. 应用F-uar2/AM和图像分析系统测定细胞内游离钙浓度.中华物理医学杂志. 19:92一94.
64. Sipido K R, Callewaert G. 1995. How to measure intracellular Ca2+ in single cardiac cells with fura-2 or indo-1. Cardiovasc Res; 29: 717-726.
65. Li SY, Golden KL, Jiang Y, Wang GJ, Privratsky JR, Zhang X, Eason AR, Culver B, Ren J, 2005. Inhibition of sarco(endo)plasmic reticulum Ca2+-ATPase differentially regulates contractile function in cardiac myocytes from normotensive and spontaneously hypertensive rats: role of Ca2+ regulatory proteins. Cell Biochem. Biophys. 42: 1–12.
66. Satoh H, Ginsburg KS, Qing K, Terada H, Hayashi H, Bers DM, 2000. KB-R7943 block of Ca2+ influx via Na+/Ca2+ exchange does not alter twitches or glycoside inotropy but prevents Ca2+ overload in rat ventricularmyocytes. Circulation 101:1441–1446.
67. Mochizuki S, MacLeod KT. 1997. Effects of hypoxia and metabolic inhibition on increases in intracellular Ca2+ concentration induced by Na+/Ca2+ exchange in isolated guinea-pig cardiacmyocytes. J Mol Cell Cardiol. 29:2979–2987.
68. Nayeem MA, Hess ML, Qian YZ, Loesser KE, Kukreja RC. 1997. Delayed preconditioning of cultured adult rat cardiac myocytes: role of 70- and 90-kDa heat stress proteins. Am J Physiol. 273: H861–H868.
69. Wang GY, Zhou JJ, Shan J,Wong TM. 2001. Protein kinase C-epsilon is a trigger of delayed cardioprotection against myocardial ischemia of kappa-opioid receptor stimulation in rat ventricular myocytes. J Pharmacol Exp Ther. 299: 603–610.
70. Wu S, Li HY, Wong TM. 1999. Cardioprotection of preconditioning by metabolic inhibition in the rat ventricular myocyte. Involvement of kappa-opioid receptor. Circ Res. 84: 1388–1395.
71. Miyawaki H, Ashraf M. 1997. Ca2+ as a mediator of ischemic preconditioning. Circ Res. 80: 790–799.
72. Node K, Kitakaze M, Sato H, Minamino T, Komamura K, Shinozaki Y, Mori H, Hori M. 1997. Role of intracellular Ca2+ in activation of protein kinase C during ischemic preconditioning. Circulation 96:1257–1265.
73. Przyklenk K, Hata K, Kloner RA. 1997. Is calcium a mediator of infarct size reduction with preconditioning in canine myocardium? Circulation 96:1305–1312.
74. 赵彩霞,郑延松,田敏.2001.细胞内游离钙检测方法进展.国外医学临床生物化学与检测学分册. 22(2):71-72.
75. MINTA A, KAO JP, TSIEN RY. 1989. Fluorescent indicators for cytosolic calcium based on rhodamine and fluoresce in chromophores. J Biolchem. 264(14):8171-8178.
76. TERRACCIANO CM, MACLEOD KT. 1997. Effects of lactate on the relative contribution of Ca2+ extrusion mechanisms to relaxation in guinea-pig ventricular myocytes. J Physiol. 500:557-570.
77. ORCHARD CH. 1987. The role of the sarcoplasmic reticulum in the response of ferret and rat heart muscle to acidosis. J Physio. 384:431-449.
78. ALLEN DG, LEE JA,SMITH GL.1989. The consequeneces of simulated ischemia on intercellular Ca2+ and tension in isolated ferret ventricular muscle. J Physiol. 410:297-323.
79. CAIRNS SP,WESTERBLAD H,ALLEN DG. 1993. Changes in myoplasmic pH and calcium concentration during expose to lactate in isolated rat ventricular myocytes. J Physiol. 464:561-574.
80. GAMBASSI G, HANSFORD RG, SOLLOTT SJ. 1993. Effects of acidosison resting cytosolic and mitochondrial Ca in mammaliam myocardium. J Gen Physiol. 102:575-597.
81. CLEVELAND JC. 1996. Ischemia Perconditioning in human and rat ventricular. Am J Physiol. 271:H1786-H1794.
82. OPIE LH. 2003. Preconditioning and metabolic anti-ischaemic agents. Eur Heart J. 24(20):1854-1856.
83. 程芳洲,余细球,唐国华等.2002.缺血预处理对大鼠缺血再灌注心肌细胞凋亡及相关基因蛋白表达的影响.中国组织化学与细胞化学杂志. 11(1):85-88.
84. TERRACCIANO CM,MACLEOD KT. 1994. Effects of acidosis onNa+/Ca2+ exchange and consequences for relaxation in guinea pig cardiac myocytes. Am J Phsiol. 267(2 Pt 2): H477-87.
85. MIYAWAKI H, ZHOU XB,ASHRAF M. 1996. Calcium preconditioning elicits strong protection against ischemia injury via Protein Kinase C signaling pathway. Circ Res. 79:137-146.
86. MURRY CE,RICHARD VJ,REIMER KA. 1990. Ischaemic preconditioning slow energy metabolism and delays ultrastructural damage during a sustained ischemic episodes. Circ Res. 66:913-931.
87. SMITH GB, STEFENELLI T,WU ST. 1996. Rapid adaptation of myocardial calcium homeostasis to short episodes of ischemia in isolated rat hearts. Am Heart J. 131:1106-1112.
88. PIERCE GN,Czubryt MP. 1995. The contribution of ionic imbalance to ischemia/reperfusion-induced injury. J Mol Cell Cardiol. 27:53-63.
89. STEENBERGEN C,FRALIX TA,MURPHY E. 1993. Role of increased cytosolic free calcium concentration in myocardial ischemic injury. Basic Res Cardiol. 88:456-470.
90. ASHRAF M,SULEIMAN J,AHMAD M. 1994. Ca2+ preconditioning elicits a unique protection against the Ca2+ paradox injury in rat heart. Role of adenosine. Fixed. Circ Res. 74:360-367.
91. PRZYKLENK K,HATA K,KLONER RA. 1997. Is calcium a mediator of infarct size reduction with preconditioning in canine myocardium . Circulation. 96: 1305-1312.
92. MIYAWAKI H, ASHRAF M.1997. Ca2+ as a mediator of ischemic preconditioning. Circ Res. 80:790-799.
93. Wang GY, Wu S, Pei JM, Yu XC, Wong TM. 2001. Kappa- but notdelta-opioid receptors mediate effects of ischemic preconditioning on both infarct and arrhythmia in rats. Am J Physiol. 280: H384-391.
94. Curtis MJ, Walker MJ. 1988. Quantification of arrhythmias using scoring systems: an examination of seven scores in an in vivo model of regional myocardial ischaemia. Cardiovasc Res. 22: 656-665.
95. Wang S, Cone J, Liu Y. 2001. Dual roles of mitochondrial KATP channels in diazoxide-mediated protection in isolated rabbit hearts. Am J Physiol. 280: H246-H255.
96. Gross GJ, Peart JN. 2003. KATP channels and myocardial preconditioning: an update. Am J Physiol. 285: H921-H930.
97. Yellon DM, Downey JM. 2003. Preconditioning the myocardium: from cellular physiology to clinical cardiology. Physiol Rev. 83: 1113-1151.
98. Kita H, Miura T, Tsuchida A, Hasegawa T, Shimamoto K. 1998. Suppression of reperfusion arrhythmias by preconditioning is inhibited by an ATP-sensitive potassium channel blocker, 5-hydroxydecanoate, but not by protein kinase C blockers in the rat. J Cardiovasc Pharmacol. 32: 791-797.
99. Vegh A, Parratt JR. 2002. The role of mitochondrial KATP channels in antiarrhythmic effects of ischaemic preconditioning in dogs. Br J Pharmacol. 137:1107-1115.
100. Driamov S, Bellahcene M, Ziegler A, Barbosa V, Traub D, Butz S, Buser PT, Zaugg CE. 2004. Antiarrhythmic effect of ischemic preconditioning during low-flow ischemia. The role of bradykinin and sarcolemmal versus mitochondrial ATP-sensitive K+ channels. Basic Res Cardiol. 99: 299-308.
101. Cao CM, Chen M, Wong TM. 2005. The KCa channel as a trigger for the cardioprotection induced by kappa-opioid receptor stimulation -- itsrelationship with protein kinase C. Br J Pharmacol. 145: 984-991.
102. Shintani Y, Node K, Asanuma H, Sanada S, Takashima S, Asano Y, Liao Y, Fujita M, Hirata A, Shinozaki Y, Fukushima T, Nagamachi Y, Okuda H, Kitakaze M. 2004. Opening of Ca2+-activated K+ channels is involved in ischemic preconditioning in canine hearts. J Mol Cell Cardiol. 37:1213-1218.
103. Gao Q, Zhang SZ, Cao CM, Bruce IC, Xia Q. 2005. The mitochondrial permeability transition pore and the Ca2+-activated K+ channel contribute to the cardioprotection conferred by tumor necrosis factor-alpha. Cytokine 32: 199-205.
104. O'Rourke B. 2004. Evidence for mitochondrial K+ channels and their role in cardioprotection.Circ. Res. 94: 420-432.
105. Ghatta S, Nimmagadda D, Xu X, O'Rourke ST. 2006. Large-conductance, calcium-activated potassium channels: structural and functional implications. Pharmacol Ther. 110: 103-116.
106. O'Rourke B. 2000. Pathophysiological and protective roles of mitochondrial ion channels. J Physiol. (Lond.). 529 Pt 1: 23-36.
107. Akabas MH. 2004. Na+/Ca2+ exchange inhibitors: potential drugs to mitigate the severity of ischemic injury. Mol Pharmacol. 66: 8-10.
108. Reuter H, Henderson SA, Han T, Matsuda T, Baba A, Ross RS, Goldhaber JI, Philipson KD. 2002. Knockout mice for pharmacological screening: testing the specificity of Na+-Ca2+ exchange inhibitors. Circ Res. 91: 90-92.
109. Nishida M, Urushidani T, Sakamoto K, Nagao T. 1999. L-cis diltiazem attenuates intracellular Ca2+ overload by metabolic inhibition in guinea pigmyocytes. Eur J Pharmacol. 385: 225-230.
110. Watano T, Harada Y, Harada K, Nishimura N. 1999. Effect of Na+/Ca2+ exchange inhibitor, KB-R7943 on ouabain-induced arrhythmias in guinea-pigs. Br J Pharmacol. 127: 1846-1850.
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