Simultaneous Optical Mapping of Intracellular Free Calcium and Action Potentials from Langendorff Perfused Hearts

Abstract

The cardiac action potential (AP) controls the rise and fall of intracellular free Ca2+ (Cai), and thus the amplitude and kinetics of force generation. Besides excitation‐contraction coupling, the reverse process where Cai influences the AP through Cai‐dependent ionic currents has been implicated as the mechanism underlying QT alternans and cardiac arrhythmias in heart failure, ischemia/reperfusion, cardiac myopathy, myocardial infarction, congenital and drug‐induced long QT syndrome, and ventricular fibrillation. The development of dual optical mapping at high spatial and temporal resolution provides a powerful tool to investigate the role of Cai anomalies in eliciting cardiac arrhythmias. This unit describes experimental protocols to map APs and Cai transients from perfused hearts by labeling the heart with two fluorescent dyes, one to measure transmembrane potential (Vm), the other Cai transients. High spatial and temporal resolution is achieved by selecting Vm and Cai probes with the same excitation but different emission wavelengths, to avoid cross‐talk and mechanical components. Curr. Protoc. Cytom. 49:12.17.1‐12.17.32. © 2009 by John Wiley & Sons, Inc.

The Future of Optical Mapping is Bright

“…11 Two alternatives, photodiode arrays (PDAs) and CMOS cameras offer superior dynamic range and spatio-temporal resolution. Although Hammamatsu and Centronic Ltd have discontinued the manufacture of PDAs, RedShirts still offers a PDA system.…”

“…A main advantage is the dynamic range of PDAs of 10 6 compared to CCDs (10 3 ) or at best 10 4 . 11 In addition, PDA signals can be alternating current coupled with long time constant and background fluorescence can be subtracted with analog circuitry before digitization making it possible to take the full measure of digitization range (14 bit) and achieve considerably greater accuracy than any other photodetector. This feature of PDAs was critical to measure small changes in membrane potential to detect atrioventricular node action potentials 12 and "virtual electrode" and electroporation effects caused by electric stimulation.…”

“…2 Camera technology progressed to CCD sensors but CMOS cameras are a game changer offering considerable advantages compared to CCDs by achieving outstanding acquisition rates (up to 10,000 frames/second), high dynamic range (10 5 ), high spatial resolution, low background noise, and large pixel dimensions. 11,14 Based on our experience with all 3 technologies, PDA, CCD, and CMOS, we would venture to surmise that scientific (large format) CMOS cameras will dominate in research instruments and will replace the relatively inferior CCD technology.…”

Optical Mapping of Action Potentials and Calcium Transients in the Mouse Heart

“…A vertical Langendorff perfusion setup can also be used 10 , but may result in slightly different cardiac mechanics 11 . In addition to CMOS cameras, alternative detectors are also available and can be applied to map V m and CaT simultaneously 12 .…”

The Photodiode Array: A Critical Cornerstone in Cardiac Optical Mapping

“…Choi and G. Salama, and uses two Hamamatsu C4675-103 detectors. Please see the excellent review by Salama et al for more details regarding this system (Salama et al, 2009). The whole-heart guinea pig (GP) Langendorff model was used, where the aorta was cannulated and hung vertically within a surrounding bath of physiological solution referred to as Tyrode's solution.…”