【作者】 易丹；

【导师】 肖桂林；

【作者基本信息】 中南大学 ， 急诊医学， 2010， 硕士

【摘要】 研究背景心肌细胞上存在多种离子通道,如ⅠNa,ⅠCa,Ⅰkl,Ⅰkr和Ⅰks等,每种离子通道都有其特定的功能和特性。各个离子通道间相互协调,维持心肌细胞及心脏的正常舒缩功能。当某些药物作用于这些离子通道后,可诱发心律失常。随着膜片钳技术的发展和成熟,现在开展了大量的关于研究药物在离子通道上的最佳作用靶点,以证实药物引起心律失常的及抗心律失常作用机制。乌头碱中毒损害的靶器官主要为心血管系统,引起的严重的心律失常是致死的主要原因,主要表现为不同形式的心律失常。而临床治疗上尚无特效解毒药治疗乌头碱中毒。在前期工作中,通过大量动物实验和临床治疗观察,结果显示双黄连治疗乌头碱中毒疗效显著。双黄连能快速解救乌头碱导致的心律失常可能与离子通道有关,但其对离子通道的作用尚无报道,为了进一步了解双黄连对乌头碱的解毒机制,本实验通过全细胞膜片钳技术来观察双黄连对乌头碱导致的心室肌细胞L-型钙通道电流变化的影响。目的1.根据乌头碱毒理特点,观察乌头碱对心室肌细胞L-型钙通道的影响；2.观察双黄连粉针剂对乌头碱导致大鼠心室肌细胞L-型钙通道的影响,探讨双黄连对抗乌头碱导致的心律失常的在离子通道水平的药理作用机制；为临床用药提供进一步的理论依据。方法本实验通过原代培养的大鼠心室肌细胞,将实验分为正常组,乌头碱组,及双黄连干预组。应用全细胞膜片钳技术纪录正常培养大鼠心室肌细胞L-型钙通道电流；1umol／L乌头碱作用下大鼠心室肌细胞钙离子通道电流的变化；洗去乌头碱,给与100mg／L双黄连粉针剂干预,5分钟后观察和记录L-型钙通道电流的变化情况。结果在钳制电压-40mv,刺激电压+10mv条件下,1umol／L乌头碱作用下Ⅰca-L电流峰值从(-1104.00±252.3085)PA增加至(-3163.44±272.1169)PA(P＜0.05),加入双黄连后,峰值回复到(-1512.57±488.0761)pA(P<0.05)。乌头碱作用下电流-电压曲线下移,双黄连干预后使下移的曲线上抬,但激活电位、峰电位和Ⅰ-Ⅴ曲线的形状,翻转电位不变。乌头碱作用下L-Ca通道电流稳态激活曲线稍向左移,双黄连干预组,使左移的稳态激活曲线向右移,乌头碱及双黄连的半激活电压分别是(19.4207±2.9148)mv和(23.5792±3.6105)mv,乌头碱及双黄连的斜率分别是(11.1311±1.8980)和(11.4750±3.2999)(P>0.05),无明显统计学差异；对时间效应曲线的影响,乌头碱的上升时间常数是87s左右,洗去后,不能使电流恢复,加入双黄连后,曲线快速下降,约56s左右回复到给药前的状态。结论1.乌头碱对培养大鼠心室肌细胞L-型钙通道电流有明显增加作用。2.双黄连粉针剂可抑制乌头碱导致心肌细胞L-型钙通道电流的增强作用。更多还原

【Abstract】 Effects of shuanghuanglian on the changes of L-type calcium channel current by aconitine of cultured rat ventricular myocytesBackgroundThere are a variety of ion channels on the cardiac cell, such as INa, ICa,Ikl,Ikr and Iks, etc. each ion channel has its own specific features and characteristics. The various ion channels Coordination to maintain normal myocardial systolic and diastolic function. When some drug effects on these ion channels, may lead to arrhythmia. With the patch clamp technique to develop and mature, there are a large number of experimental studies the role of drug-induced arrhythmia mechanisms and the best role of antiarrhythmic targets.Cardiovascular system that is the main target organ damaged by Aconitine poisoning.which cause serious arrhythmia is the main reason of death. But there is no effective medicine treat the aconitine poisoning.In early work, through a large number of animal experiments and clinical observation showed that Shuanghuanglian treat aconitine poisoning have significantly effect.Shuanghuanglian can quickly save aconitine induced arrhythmias may be related to ion channels, in order to further observation the Shuanghuanglian on the detoxification mechanism of aconitine, in this experiment,using whole cell patch clamp technique to observe the effects of shuanghuanglian on the changes of L-type calcium channel current by aconitine of cultured rat ventricular myocytes.Objective1. To determine the effects of aconitine and shuanghuanglian on L-type calcium channels in the ventricular myocytes of the rats.2. To study the pharmacological mechanism of aconitine and Shuanghuanglian intervene after aconitine on the ion channels of myocardium. And the pharmacological mechanism Of Shuanghuanglian against arrhythmia caused by the aconitine.MethodsSingle wentricular myocytes of rats were obtained by primary culture method, Divided into normal group, aconitine group, and shuanghuanglian intervention group.The influence of aconitine and Shuanghuanglian intervene after aconitine on L-type calcium channels was recorded by the technique of whole-cell patch clamp.ResultsAt the test potential of+10mv, aconitine increased L-type calcium current from (-1104.00±252.3085) PA to (-3163.44±272.1169) PA (p < 0.05).Join Shuang huanglian, the Ica-L peak back to (-1512.57±488.0761)pA. Aconitine download the current-voltage curve, but activation potential, peak potential and the shape ofⅠ-Ⅴcurve, reversal potential unchanged. Aconitine and Shuanghuanglian on Ica-L activation curve does not significantly effectoin. Aconitine and Shuanghuanglian there half active potential (V0.5) respectively is (19.4207±2.9148) mv and (17.3374±2.4404) mv (P> 0.05).The steady activation was no statistically significant. The impact on the time effect curve, the rise time of aconitine is 87s or so, wash it away, the current also not recover, but Join Shuanghuanglian after, the curve decrease rapidly (56s) return to the status before administration.Conclusion1. Aconitine in rat ventricular myocytes can significantly enhance the Ica-L..2. Shuanghuanglian can markedly inhibit the increased of Ica-L by aconitine on ventricular myocytes.更多还原