Lou Qing, Li Wenwen, Efimov Igor R
Department of Biomedical Engineering, Washington University in St. Louis.
J Vis Exp. 2011 Sep 13(55):3160. doi: 10.3791/3160.
Optical imaging and fluorescent probes have significantly advanced research methodology in the field of cardiac electrophysiology in ways that could not have been accomplished by other approaches(1). With the use of the calcium- and voltage-sensitive dyes, optical mapping allows measurement of transmembrane action potentials and calcium transients with high spatial resolution without the physical contact with the tissue. This makes measurements of the cardiac electrical activity possible under many conditions where the use of electrodes is inconvenient or impossible(1). For example, optical recordings provide accurate morphological changes of membrane potential during and immediately after stimulation and defibrillation, while conventional electrode techniques suffer from stimulus-induced artifacts during and after stimuli due to electrode polarization(1). The Langendorff-perfused rabbit heart is one of the most studied models of human heart physiology and pathophysiology. Many types of arrhythmias observed clinically could be recapitulated in the rabbit heart model. It was shown that wave patterns in the rabbit heart during ventricular arrhythmias, determined by effective size of the heart and the wavelength of reentry, are very similar to that in the human heart(2). It was also shown that critical aspects of excitation-contraction (EC) coupling in rabbit myocardium, such as the relative contribution of sarcoplasmic reticulum (SR), is very similar to human EC coupling(3). Here we present the basic procedures of optical mapping experiments in Langendorff-perfused rabbit hearts, including the Langendorff perfusion system setup, the optical mapping systems setup, the isolation and cannulation of the heart, perfusion and dye-staining of the heart, excitation-contraction uncoupling, and collection of optical signals. These methods could be also applied to the heart from species other than rabbit with adjustments to flow rates, optics, solutions, etc. Two optical mapping systems are described. The panoramic mapping system is used to map the entire epicardium of the rabbit heart(4-7). This system provides a global view of the evolution of reentrant circuits during arrhythmogenesis and defibrillation, and has been used to study the mechanisms of arrhythmias and antiarrhythmia therapy(8,9). The dual mapping system is used to map the action potential (AP) and calcium transient (CaT) simultaneously from the same field of view(10-13). This approach has enhanced our understanding of the important role of calcium in the electrical alternans and the induction of arrhythmia(14-16).
光学成像和荧光探针极大地推动了心脏电生理学领域研究方法的发展，这些方法是其他途径无法实现的(1)。通过使用钙敏和电压敏染料，光学标测能够在不与组织进行物理接触的情况下，以高空间分辨率测量跨膜动作电位和钙瞬变。这使得在许多使用电极不方便或不可能的情况下测量心脏电活动成为可能(1)。例如，光学记录能够提供刺激和除颤期间及之后膜电位的准确形态变化，而传统电极技术在刺激期间及之后会因电极极化而受到刺激诱导伪迹的影响(1)。Langendorff灌注兔心脏是研究人类心脏生理学和病理生理学最深入的模型之一。临床上观察到的许多类型的心律失常都可以在兔心脏模型中重现。研究表明，兔心脏在室性心律失常期间的波形模式，由心脏的有效大小和折返波长决定，与人类心脏非常相似(2)。还表明，兔心肌兴奋 - 收缩（EC）偶联的关键方面，如肌浆网（SR）的相对贡献，与人类EC偶联非常相似(3)。在此，我们介绍Langendorff灌注兔心脏光学标测实验的基本步骤，包括Langendorff灌注系统设置、光学标测系统设置、心脏的分离和插管、心脏的灌注和染料染色、兴奋 - 收缩解偶联以及光学信号的采集。这些方法经过流速、光学、溶液等方面的调整后，也可应用于兔以外物种的心脏。本文描述了两种光学标测系统。全景标测系统用于标测兔心脏的整个心外膜(4 - 7)。该系统提供了心律失常发生和除颤期间折返回路演变的全局视图，并已用于研究心律失常机制和抗心律失常治疗(8,9)。双标测系统用于从同一视野同时标测动作电位（AP）和钙瞬变（CaT）(10 - 13)。这种方法增强了我们对钙在电交替和心律失常诱导中的重要作用的理解(14 - 16)。
