Development of the System for Measurement of Intramyocardial Ion Dynamics in Isolated, Perfused Hearts

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It is important to measure intramyocardial ion dynamics for the investigation of physiology and pathophysiology of hearts. The measurements of intracellular ion concentration, the method using fluorescent dyes has advantages including high temporal resolution. However, it has been difficult to apply fluorescent dyes into isolated, perfused hearts. To load dyes to perfused hearts, we developed the system using the iontophoresis technique. In this study, we completed the following items ;1.We optimized the method to produce microelectrodes for iontophoresis of dyes.2.We determined the optimal conditions for the current and the optinal positioning of the electrode.3.We developed the method to stop or attenuate the beating of the hearts. First, the calcium concentration in the perfusate was decreased, and then, potasium concentration was increased or BDM was added to the perfusate.4.In our method, microelectrode was induced above the heart, and iontophoresis was performed from the top of the stage of microscope. In contrast, the ultraviolet beam was introduced to the heart from the bottom of the microscope. Thus, it was necessary to change the orientation of the heart between the procedures, and caused the disorientation. To avoid this, we developed the method to keep the orientation of hearts.Our study indicates the feasibility of the iontophoresis in isolated, perfused hearts, and we accomplished some basic part of the techniques required for this approach. Nevertheless, there are still many remaining problems to make this technique more practical.In addition, we also evaluated the different effects of proteins on Ca^<2+> ion measurements using fluorescent dye or NMR technique.