Optical Mapping of Cardiac Electromechanics

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Cardiac biophysics rests on three fundamental pillars: energy production, electrical excitation, and mechanical work. Studies of cardiac metabolism, bioelectricity, and biomechanics have been very productive at all levels of reduced complexity, but especially at the levels of tissue, cell, and molecule. The three cardiac biophysical pillars work in concert and cannot be fully understood in isolation. However, available experimental methods are limited, restricting our ability to study them simultaneously. We are often forced by experimental reality to reduce the complexity of the cardiac system to study one component at a time.

Optical mapping of fluorescence reporters is a cornerstone of present-day cardiac biophysics research, because it provides valuable information about transmembrane potential, intracellular calcium, mitochondrial inner membrane potential, NADH, and other critical physiological parameters (1). A multitude of metabolic, bioelectric, and biomechanical mechanisms converge to provide a foundation for the function of the working heart, which delivers oxygen and nutrients to the entire body. Excised perfused hearts are valuable preparations for studying cardiac metabolism, bioelectricity, and biomechanics (2, 3). Yet the application of optical mapping to study cardiac mechanics in perfused hearts is limited, because recordings of fluorescence require the suppression of contraction to prevent artifacts caused by the motion of a beating heart. Obviously, this makes the assessment of cardiac mechanics impossible. Furthermore, cardiac contraction is often suppressed in bioelectric studies by blocking the actomyosin ATPase, which drops myocardial oxygen consumption fourfold (4). This dramatic reduction in oxygen consumption significantly slows metabolic kinetics and alters cellular processes that depend upon metabolic rate. This is a particularly important issue in optical mapping studies of ischemia and reperfusion (5, 6).

The limitations of optical mapping are well recognized by the cardiac research community, but the development of new optical mapping technologies that provide for artifact-free mapping of normally contracting hearts has been challenging. Clearly, the technical evolution of optical mapping progresses with a goal of simultaneous imaging of the three fundamental pillars of cardiac biophysics. Examples of recent progress include optical mapping of action potentials, while increasing RV strain (7), and high-speed measurements of epicardial deformation during the cardiac cycle using structured light (8). Structured light imaging provides high spatial and temporal resolution measurements of epicardial deformation in three dimensions: a requirement for accurate assessment of cardiac mechanics. But fast structured light instrumentation is costly, and currently not accessible to most cardiac research laboratories.

In this issue of the Biophysical Journal, Zhang et al. (9) present a new and exciting advancement in the evolution of cardiac optical mapping. They demonstrate simultaneous imaging of epicardial transmembrane potential and contraction (strain) in excised perfused hearts. The novelty of their approach is that it uses hardware components (CCD cameras and LEDs) that are common to most cardiac electrophysiology research laboratories and novel software that computes epicardial strains and optical action potentials. Three high-speed cameras are used: one camera to image cyan and blue-elicited fluorescence of di-4-ANEPPS on alternating camera frames (excitation ratiometry) and two other cameras for binocular imaging of the epicardial surface. In left ventricular (LV) working heart studies, the authors used an additional camera to provide a wider binocular field of view. With binocular imaging, in-plane and out-of-plane contraction is measured by tracking epicardial markers in three dimensions, a significant enhancement over conventional two-dimensional contraction analyses used in previous work (7, 10, 11).

The authors developed software to measure, during the full cardiac cycle, the movement and deformation of triangles formed by the markers. Movement and deformation data were used to compute local epicardial strains and remove motion artifacts in optical action potentials. By tracking the triangles, the cyan and blue-elicited fluorescence could be reconstructed for epicardial tissue within each triangle as the triangle traversed the contraction cycle. A distinct advantage of the author’s approach is that the reconstruction is accomplished without reducing the spatial resolution of the fluorescence data, an important feature for precise mapping of spatial changes in electrophysiological parameters. Optical action potentials free of motion artifacts were then revealed by the ratio of motion-tracked cyan-elicited fluorescence to motion-tracked blue-elicited fluorescence. Ratiometry of these two signals is a significant improvement over other ratiometric approaches that do not maintain registration between camera pixels and epicardial sites during contraction (12, 13). Results demonstrated that motion artifacts were best corrected when there was minimal angular motion. Removal of artifacts resulting from out-of-plane motions was also quite impressive.

Of particular interest is the demonstration from Zhang et al. (9) of mapping the electromechanical function of excised swine LV working hearts. The simultaneously measured strain (percent shortening) values and optical action potentials shown in Fig. 7 of their article are the first electromechanical optical mapping measurements reported for an LV working swine heart. During paced rhythm, LV epicardial strain in the major principal direction averaged close to 10%, consistent with in vivo strains measured from the epicardium of pigs (14). A close inspection of their Fig. 7 B reveals that systole (increased strain) is initiated after phase-zero depolarization and diastole (reduced strain) begins after phase-three repolarization, consistent with the physiology of cardiac excitation-contraction coupling (15). The authors also measured changes in electromechanical function during local ischemia after occluding the left anterior descending coronary artery. As expected, reductions in optical action potential amplitudes and durations were observed in the ischemic zone along with a dramatic loss of contractile shortening. Spatially heterogeneous strain, with regions of positive and negative shortening, was observed in strain maps of the ischemic zone. Negative shortening (up to 5%) in the ischemic zone, indicating significant stretch, was measured during systole. Such unique insights into the electromechanical function of locally ischemic tissue demonstrate the significance of this next-generation optical mapping technology.

Zhang et al. (9) have clearly shown that optical mapping of loaded working hearts for the assessment of electromechanics is now a reality. Their work dramatically elevates the physiological sophistication and clinical relevance of optical mapping, demonstrating the potential to uncover new insights into the mechanisms of mechano-electrical feedback, abnormal strain during local ischemia, arrhythmogenesis, ventricular resynchronization therapy, and many other important problems. Incorporating the imaging of metabolically relevant parameters, such as NADH fluorescence or the fluorescence of an intracellular pH probe, into this technology is eagerly anticipated to provide insight into the remaining fundamental pillar of cardiac biophysics. The future of integrated metabolism-bioelectric-biomechanics studies is bright!

Editor: James Sneyd.