七氟烷预处理和后处理对离体大鼠心脏再灌注心律失常和心肌细胞电生理的影响

发布时间：2018-08-11 11:40

【摘要】：目的： 1、观察七氟烷预处理(Sevoflurane preconditioning, SpreC)和七氟烷后处理(Sevoflurane postconditioning, SpostC)对再灌注心律失常的影响,探索其相关机制； 2、观察SpreC和SpostC对离体大鼠心肌细胞动作电位、L型钙通道电流(L-type calcium current, ICa, L)、快钠通道电流(Fast sodium current,INa)和瞬时外向钾通道电流(Transient outward potassium current, Ito)电生理特性的影响及相关机制； 3、比较SpreC和SpostC对再灌注心律失常和离子通道电生理作用的异同。 方法： 1、取40只SD大鼠心脏建立Langendorff灌注模型, KH液平衡15min后随机分入以下4组,每组10只：(1)时间对照组(Time Control,TC),平衡期后继续灌注KH液75min；(2)缺血再灌注组(Ischemia/reperfusion,I/R),平衡期后灌注KH液20min,随后停灌25min复灌30min制作全心缺血再灌注损伤模型；(3)七氟烷预处理组(SpreC),在全心停灌前以3%七氟烷预处理15min,普通KH液洗脱5min,随后停灌25min,复灌30min；(4)七氟烷后处理组(SpostC),平衡期后灌注普通KH液20min,停灌25min,复灌30min,在复灌之初使用3%七氟烷后处理15min。比较各组血流动力学、冠脉流出液肌钙蛋白I、心肌梗死面积、再灌注心律失常及细胞内钙离子和活性氧水平。 2、取SD大鼠心脏制备Langendorff灌注模型,分组及缺血再灌注方案同第一部分,再灌注结束后分离各组心室肌外膜细胞进行电生理实验。使用标准的全细胞膜片钳技术记录各组心室肌细胞的动作电位振幅、静息膜电位和动作电位时程；记录各组心室肌细胞的ICa,L、INa、Ito,分析其电流-电压曲线、稳态激活曲线、稳态失活曲线和稳态失活后恢复曲线的变化,以了解SpreC和SpostC对大鼠心室肌细胞电生理特性的影响。 结果： 1、血流动力学、心肌损伤和再灌注心律失常相关结果。与I／R组相比,SpreC组和SpostC组能增大再灌注心脏的左室发展压、左室最大收缩／舒张速率和心率,降低左室舒张末期压力,降低冠脉流出液肌钙蛋白I水平,减少心梗面积(P0.05)。SpreC和SpostC减少离体心脏再灌注期间VPB的发生个数、减少VF的发生率、缩短VT和VF的发作时程、降低再灌注心律失常评分(P0.05)。SpreC和SpostC降低再灌注后心肌细胞内钙离子水平和活性氧水平(P0.05)。SpreC和SpostC在上述指标的差异未见统计学意义(P0.05)。 2、心肌细胞电生理相关结果。(1)动作电位：与TC组相比,I／R组的心肌细胞动作电位振幅减小、静息膜电位升高、动作电位时程缩短(P0.05)。与I／R组相比,SpreC和SpostC能增大动作电位振幅、降低静息膜电位、延长动作电位时程(P0.05)。(2)L型钙通道：与TC组相比,I／R组ICa，L的峰值密度降低,半失活电压减小,失活曲线左移,恢复时间常数增大(P0.05)。与I／R组相比,SpreC和SpostC能增加ICa,L的峰值密度,增加ICa,L失活曲线的半失活电压,右移失活曲线,降低ICa，L恢复曲线的时间常数(P0.05)。(3)快钠通道：与TC组相比,I／R组INa的峰值密度降低,稳态失活曲线的斜率因子k降低(P0.05)。与I／R组相比,SpreC和SpostC能增加INa的峰值密度,增加失活曲线的斜率因子k(P0.05)。(4)瞬时外向钾通道：与TC组相比,I／R组Ito的峰值密度降低,半激活电压和半失活电压增大,稳态激活曲线和稳态失活曲线右移(P0.05)。与I／R组比较,SpreC和SpostC能增加Ito的峰值密度,减小半失活电压,左移失活曲线(P0.05)。SpreC和SpostC关于动作电位、ICa,L、INa和Ito相关指标之间的差异无统计学意义(P0.05)。 结论： 1、SpreC和SpostC能改善缺血再灌注心脏的血流动力学,减轻心肌损伤,减少心肌梗死面积,对离体大鼠心脏发挥保护效应。SpreC和SpostC能改善离体大鼠再灌注心律失常的发生及发作。上述作用可能与SpreC和SpostC降低心肌细胞内钙离子和氧自由基水平有关。 2.(1) SpreC和SpostC能减小缺血再灌注损伤对动作电位参数的影响。(2)SpreC和SpostC能增加ICa,L峰值密度,加快失活.,减慢恢复,缓解缺血再灌注损伤造成的变化,提示SpreC和SpostC有利于延长再灌注损伤后心肌细胞的动作电位时程,缓解心肌复极离散度的增大。此外,SpreC和SpostC对L型钙通道的调控有利于维持钙稳态,缓解再灌注损伤后钙超载。(3)SpreC和SpostC能增加再灌注损伤后INa的峰值密度,提示SpreC和SpostC能增加再灌注损伤心肌细胞动作电位的0相上升速率,加快冲动传导,对兴奋性折返性心律失常有一定的预防作用。(4)SpreC和SpostC能增加再灌注损伤心肌细胞Ito的峰值密度,降低半失活电压,加快失活过程,能减少缺血再灌注损伤对动作电位时程的影响,减少复极离散度,减少Ito介导的2相折返和后除极的发生率,从而缓解再灌注心律失常的发生及发作。 3、SpreC和SpostC对再灌注心脏的血流动力学、心肌酶释放水平、心梗面积、再灌注心律失常等方面的影响类似。SpreC和SpostC对动作电位、ICa，，L、INa和Ito的电生理特性的影响类似。
[Abstract]:Objective:
1. To observe the effects of sevoflurane preconditioning (SpreC) and sevoflurane postconditioning (SpostC) on reperfusion arrhythmia and explore its related mechanism.
2. To observe the effects of SpreC and Post C on action potential, L-type calcium current (ICa, L), Fast sodium current (INa) and transient outward potassium channel current (Ito) in isolated rat cardiomyocytes and the related mechanisms.
3, we compared the electrophysiological effects of SpreC and SpostC on reperfusion arrhythmias and ion channels.
Method:
1. 40 SD rats were randomly divided into four groups after 15 minutes of KH fluid balance: (1) time control group (TC), after the balance period, KH fluid was continuously perfused for 75 minutes; (2) ischemia / reperfusion group (I / R), after the balance period, KH fluid was perfused for 20 minutes, and then stopped for 25 minutes to reperfusion for 30 minutes. Cardiac ischemia-reperfusion injury model; (3) Sevoflurane preconditioning group (SpreC), before cardiac arrest with 3% sevoflurane preconditioning for 15 minutes, elution of ordinary KH solution for 5 minutes, then stop perfusion for 25 minutes, reperfusion for 30 minutes; (4) Sevoflurane postconditioning group (SpostC), after equilibrium perfusion of ordinary KH solution for 20 minutes, stop perfusion for 25 minutes, reperfusion for 30 minutes, at perfusion at the beginning of reperfusion with 3% sevoflurane postconditioning. 15 min. The hemodynamics, cardiac troponin I, myocardial infarction area, reperfusion arrhythmia, intracellular calcium ion and reactive oxygen species were compared.
2. Langendorff perfusion model was established in SD rat hearts. The ventricular epicardial cells were isolated from each group after reperfusion and electrophysiological experiments were performed. The amplitude of action potential, resting membrane potential and action potential duration of ventricular myocytes were recorded by standard whole cell patch clamp technique. ICa, L, INa, Ito of ventricular myocytes were recorded, and their current-voltage curves, steady-state activation curves, steady-state inactivation curves and recovery curves after steady-state inactivation were analyzed to understand the effects of SpreC and Post-C on the electrophysiological characteristics of ventricular myocytes in rats.
Result:
1. Hemodynamics, myocardial injury and reperfusion arrhythmias. Compared with I/R group, Spre C and Spost C groups increased left ventricular development pressure, maximal left ventricular systolic/diastolic rate and heart rate, decreased left ventricular end-diastolic pressure, decreased coronary effluent troponin I levels, and decreased myocardial infarction area (P 0.05). OstC decreased the number of VPB, the incidence of VF, the duration of VT and VF, and the reperfusion arrhythmia score (P 0.05). SpreC and postC decreased the levels of intracellular calcium and reactive oxygen species (P 0.05). There was no significant difference between SpreC and postC (P 0.0). 5).
2. Relevant electrophysiological results of myocardial cells. (1) Action potential: Compared with TC group, the amplitude of action potential decreased, resting membrane potential increased and action potential duration shortened in I/R group (P 0.05). Compared with I/R group, SpreC and Post C could increase action potential amplitude, decrease resting membrane potential and prolong action potential duration (P 0.05). Track: Compared with TC group, the peak density of ICa and L decreased, the half-inactivation voltage decreased, the inactivation curve shifted to the left, and the recovery time constant increased in I/R group (P 0.05). Compared with I/R group, SpreC and Post C could increase the peak density of ICa and L, increase the half-inactivation voltage of ICa and L inactivation curves, and decrease the time constant of ICa and L recovery curves (P 0.05). 3) Fast sodium channel: Compared with TC group, the peak density of INa and the slope factor K of steady-state inactivation curve decreased in I/R group (P 0.05). Compared with I/R group, SpreC and Post C could increase the peak density of INa and the slope factor K of inactivation curve (P 0.05). (4) Instantaneous outward potassium channel: Compared with TC group, the peak density of Ito in I/R group was decreased and the semi-activated potassium channel was increased. Compared with the I/R group, SpreC and Post C could increase the peak density of Ito, decrease the half-inactivation voltage, and shift the inactivation curve to the left (P 0.05). There was no significant difference between SpreC and Post C on action potential, ICa, L, INa and Ito (P 0.05).
Conclusion:
1. SpreC and postC can improve the hemodynamics of ischemia-reperfusion heart, alleviate myocardial injury, reduce the size of myocardial infarction, and protect isolated rat heart. SpreC and postC can improve the occurrence and attack of reperfusion arrhythmia in isolated rats. These effects may be related to SpreC and postC reducing intracellular calcium ions and oxygen spontaneity. Related to base level.
2. (1) SpreC and postC can reduce the effect of ischemia-reperfusion injury on action potential parameters. (2) SpreC and postC can increase the peak density of ICa and L, accelerate inactivation, slow recovery, and alleviate the changes caused by ischemia-reperfusion injury, suggesting that SpreC and postC can prolong the action potential duration of myocardial cells after reperfusion injury and alleviate myocardial recovery. In addition, the regulation of L-type calcium channel by SpreC and Post C is beneficial to maintain calcium homeostasis and relieve calcium overload after reperfusion injury. (3) SpreC and Post C can increase the peak density of INa after reperfusion injury, suggesting that SpreC and Post C can increase the phase-0 rise rate of action potential and accelerate impulse conduction in myocardial cells after reperfusion injury. (4) SpreC and postC can increase the peak density of Ito, decrease the half-inactivation voltage, accelerate the inactivation process, reduce the effect of ischemia-reperfusion injury on action potential duration, reduce repolarization dispersion, and reduce Ito-mediated phase 2 reentry and depolarization. Thus, the incidence and incidence of reperfusion arrhythmia can be alleviated.
3. The effects of SpreC and Post C on hemodynamics, myocardial enzyme release, myocardial infarction area and reperfusion arrhythmia were similar. The effects of SpreC and Post C on action potential, ICa, L, INa and Ito were similar.
【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2012
【分类号】：R614;R-332

【参考文献】

相关期刊论文 前4条

1 王彤;符岳;方向韶;黄子通;;普通大鼠心肌细胞的分离培养和收缩舒张观察[J];岭南急诊医学杂志;2007年05期

2 廖华;糜涛;涂志业;陈莉莉;郭小梅;;成年大鼠心肌细胞分离方法的改良[J];中国组织工程研究与临床康复;2009年33期

3 丁超,何振山,崔俊玉,杨丽,石巧,胡丽叶,齐书英,刘晓云;兔心肌梗死1周及2月后心室肌细胞瞬间外向钾电流的变化[J];中国病理生理杂志;2004年08期

4 丁超,何振山,齐书英,崔俊玉,杨丽,石巧,胡丽叶;兔急性心肌梗死后二月心室肌细胞钠离子通道活性的变化[J];中国心脏起搏与心电生理杂志;2003年02期

相关博士学位论文 前2条

1 李宁;参松养心胶囊抗心律失常机理研究[D];中国协和医科大学;2007年

2 姚允泰;七氟醚缺血后处理对大鼠离体心脏缺血—再灌注损伤保护的机理研究[D];中国协和医科大学;2010年

本文编号：2176895