成年小鼠心室肌细胞分离及L型钙电流记录

Isolation of Ventricular Myocytes in Adult Mice and Observation of L-Type Ca^(2+) Current

目的:寻找一种成年小鼠心肌细胞分离的简便、可行且稳定的方法,以获得耐钙心肌细胞并记录心室肌细胞L型钙电流(IL-Ca)。方法:采用Langerdorff逆行主动脉灌流、无钙台氏液+胶原酶Ⅱ消化法分离成年小鼠单个心室肌细胞;利用含钙细胞外液复钙法筛选耐钙心肌细胞;并采用全细胞膜片钳技术记录心室肌细胞L型钙电流。结果:利用该方法分离心肌细胞可获得90%活细胞;复钙后细胞存活率60%-70%;并可记录到典型L型钙电流。结论:无钙台氏液+胶原酶Ⅱ消化法简单可行且稳定,可分离大量活性较佳且耐钙心肌细胞,可用于心肌细胞离子通道电流记录,从而为心脏电生理研究提供有力的技术支持。

Objective：To explore a simple,reliable and stable method of isolation of single cardiomyocyte cells in adult mice so as to obtain calcium-resistant cardiomyocytes to record L-type Ca^2＋current.Methods：Retrograde perfusion of hearts was performed in arteriae by the Langendorff perfusion apparatus and calcium-free Tyrode solution＋collagenaseⅡsolution digestive method was used to obtain single ventricular cells.Then the cells were exposed to calcium solution to select calciumresistant cardiomyocytes.And the L-type Ca^2＋current was recorded by the whole cell patch clamp.Results：The survival rate of Cardiomyocytes was 90% by collagenaseⅡsolution digestive method and remained 60%-70% after recalcification.And typical L-type Ca^2＋current was recorded.Conclusion：Calcium-free Tyrode solution＋collagenaseⅡ method is a simple,feasible and stable method to isolate large numbers of calcium-resistant cardiomyocytes with better activity,which are appropriate for myocardial ion channel currents and provide strong technical support for the heart electrophysiological studies.