适合膜片钳实验的心肌细胞分离方法研究

The study of isolation methods of myocardial cell appropriate for patch clamp experiments

目的:摸索并建立简便可行的适于做膜片钳实验的心肌细胞急性分离方法。方法:采用自行改进的Langendorff心脏灌流装置,用无钙台氏液和酶解液逆行主动脉灌流之后,剪取心肌组织置于KB液中保存以供全细胞膜片钳方式记录电流。结果:分离所得80%～90%的细胞呈杆状,边缘清楚,表面光滑,纹理清晰,可记录到典型的快钠电流。结论:控制好分离心肌细胞的实验条件以及熟练掌握实验操作的方法,是分离出适合于膜片钳实验的心肌细胞的重要因素。

Objective： To explore and establish a simple and reliable isolation method of single myocardial cell in rats for patch clamp experiments. Methods： Hearts were firstly perfused by the Langendorff perfusion apparatus with Ca2＋-free Tyrode＇ s solution and enzyme solution retrogradated arteriae aorta. The myocardial tissues were minced into small pieces in KB solution, and conserved in KB solution. The myocardial cells were supplied to record electric current through the way of whole cell patch clamp. Results： The survival rate of myocardial cells was 80%-90% and the morphous and construction of myocardial cells was fine. The sodium current was recorded successfully. Conclusion： The experimental conditions controlled and proficiency in a good experimental method of operation are important factors, which are appropriate for patch damp experiments in myocardial cells.