Evaluation system of cardiac cells and tissues with combined micro-electrode array based electrical mapping and optical mapping system

随着再生医学在心脏病学中的发展，例如基于培养的心脏组织用于治疗心律失常的生物起搏器，我们需要一个系统来评估心脏细胞、心脏组织以及移植再生组织的整个心脏的功能。1)研制了实时反馈动作电位传播动力学的心脏电刺激系统。在该系统中，心脏电传播通过高空间分辨率和高时间分辨率的光学映射方法进行可视化。实时确定传播的激励波前，并根据传播模式控制电刺激的位置和时间。它可以通过考虑心脏组织的时相(去极化、复极化和静止期)来控制时机和位置来启动通断兴奋。2)我们还开发了培养的心肌细胞和干细胞的测量系统，其中利用光刻技术制作的集成微电极阵列(通道，30μm×30μm，电极间距：130μm)测量了细胞内钙的时间和空间电激活模式，并通过冷却的ccd相机测量了细胞内钙的荧光图像。分离新生大鼠心肌细胞进行培养。体外培养3天后观察细胞机械收缩情况。它生长到培养细胞延展区域的收缩。电活动和机械收缩的同步性也被确认。同时用荧光染料(Fluo-4)测定细胞内钙离子浓度。确定了钙动力学和电活动的同步性。

For progress of regenerative medicine in cardiology, such as bio-pacemaker based on cultured cardiac tissue for arrhythmia therapy, we need a system to evaluate function of cardiac cell, cardiac tissue, and a whole heart where the regenerated tissue is implanted. In this study, combined optical and electrical measurement system of cultured cardiac tissue and whole heart preparation has been developed.1) We have developed a cardiac electric stimulation system with real time feedback of action potential propagation dynamics. In this system, cardiac electrical propagation is visualized by optical mapping method with high spatial and temporal resolution. Propagated excitation wave front is determined in real time, and the location and timing of electrical stimulation is controlled depending on the propagation pattern. It can initiate Make/Break excitation by controlling the timing and location considering the phase (depolarized, repolarized, and resting phase) of the cardiac tissue. The function of the system was confirmed by experiments with Langendorff perfused rabbit heart preparations.2) We have also developed measurement system for cultured cardiac cells and stem cells where temporal and spatial electrical activation patterns are measured by an integrated micro-electrode arrays fabricated with photo lithography (64 channels, 30 μm X 30 μm, electrode spacing: 130 μm), and a fluorescence image of intracellular Ca is measured by a cooled CCD camera. Cardiac cells were separated from neonatal rats and cultured. mechanical contraction was observed after three days in vitro. It grows to contraction of extended areas of cultured cells. Synchronization of the electrical activities and mechanical contraction were also identified. Intracellular calcium concentration was measured by fluorescent dye (fluo-4) simultaneously. Synchronization of calcium dynamics and electrical activities were identified.