Department of Physiology, Shiga University of Medical Science;
Department of Physiology, Shiga University of Medical Science.
J Vis Exp. 2021 May 19(171). doi: 10.3791/61866.
In basic research using mouse heart, isolating viable individual cardiomyocytes is a crucial technical step to overcome. Traditionally, isolating cardiomyocytes from rabbits, guinea pigs or rats has been performed via retrograde perfusion of the heart with enzymes using a Langendorff apparatus. However, a high degree of skill is required when this method is used with a small mouse heart. An antegrade perfusion method that does not use a Langendorff apparatus was recently reported for the isolation of mouse cardiomyocytes. We herein report a complete protocol for the improved antegrade perfusion of the excised heart to isolate individual heart cells from adult mice (8 - 108 weeks old). Antegrade perfusion is performed by injecting perfusate near the apex of the left ventricle of the excised heart, the aorta of which was clamped, using an infusion pump. All procedures are carried out on a pre-warmed heater mat under a microscope, which allows for the injection and perfusion processes to be monitored. The results suggest that ventricular and atrial myocytes, and fibroblasts can be well isolated from a single adult mouse simultaneously.
在使用小鼠心脏进行基础研究时，分离有活力的单个心肌细胞是一个关键的技术步骤。传统上，从兔子、豚鼠或大鼠中分离心肌细胞是通过使用 Langendorff 仪器对心脏进行逆行灌注酶来完成的。然而，当用小鼠标本心脏使用这种方法时，需要高度的技巧。最近报道了一种不使用 Langendorff 仪器的顺行灌注方法，用于分离小鼠心肌细胞。我们在此报告了一种完整的方案，用于改进离体心脏的顺行灌注，以从成年小鼠（8-108 周龄）中分离单个心脏细胞。顺行灌注是通过使用输液泵在夹闭主动脉的离体心脏的左心室心尖附近注射灌流液来完成的。所有操作都在预热加热垫上在显微镜下进行，这使得可以监测注射和灌注过程。结果表明，从单个成年小鼠中可以同时很好地分离心室和心房肌细胞以及成纤维细胞。
