Simultaneous measurement and modulation of multiple physiological parameters in the isolated heart using optical techniques

Peter Lee,Peter Kohl,Leslie M Loew,Ping Yan,Christian Bollensdorff,Paul Ewart

doi:10.1007/s00424-012-1135-6

Abstract

Whole-heart multi-parametric optical mapping has provided valuable insight into the interplay of electrophysiological parameters, and this technology will continue to thrive as dyes are improved and technical solutions for imaging become simpler and cheaper. Here, we show the advantage of using improved 2nd-generation voltage dyes, provide a simple solution to panoramic multi-parametric mapping, and illustrate the application of flash photolysis of caged compounds for studies in the whole heart. For proof of principle, we used the isolated rat whole-heart model. After characterising the blue and green isosbestic points of di-4-ANBDQBS and di-4-ANBDQPQ, respectively, two voltage and calcium mapping systems are described. With two newly custom-made multi-band optical filters, (1) di-4-ANBDQBS and fluo-4 and (2) di-4-ANBDQPQ and rhod-2 mapping are demonstrated. Furthermore, we demonstrate three-parameter mapping using di-4-ANBDQPQ, rhod-2 and NADH. Using off-the-shelf optics and the di-4-ANBDQPQ and rhod-2 combination, we demonstrate panoramic multi-parametric mapping, affording a 360° spatiotemporal record of activity. Finally, local optical perturbation of calcium dynamics in the whole heart is demonstrated using the caged compound, o-nitrophenyl ethylene glycol tetraacetic acid (NP-EGTA), with an ultraviolet light-emitting diode (LED). Calcium maps (heart loaded with di-4-ANBDQPQ and rhod-2) demonstrate successful NP-EGTA loading and local flash photolysis. All imaging systems were built using only a single camera. In conclusion, using novel 2nd-generation voltage dyes, we developed scalable techniques for multi-parametric optical mapping of the whole heart from one point of view and panoramically. In addition to these parameter imaging approaches, we show that it is possible to use caged compounds and ultraviolet LEDs to locally perturb electrophysiological parameters in the whole heart.

Highlights

In cardiac tissue, transmembrane voltage (Vm) and intracellular calcium concentration ([Ca2+]i) are arguably two of the most important parameters of interest in basic cardiac electrophysiology research
Recent developments in light-emitting diodes (LEDs) [2], high-speed cameras and optical filter technologies have ushered in a new era of imaging approaches, which is already impacting optical mapping [12]
In terms of high-speed cameras, frame rates have increased to the point where the so called multi-colour imaging technique [38], which uses a single camera and multi-band emission filter to image multiple parameters, can be applied to multi-parametric optical mapping

Summary

Introduction

Transmembrane voltage (Vm) and intracellular calcium concentration ([Ca2+]i) are arguably two of the most important parameters of interest in basic cardiac electrophysiology research. In terms of high-speed cameras, frame rates have increased to the point where the so called multi-colour imaging technique [38], which uses a single camera and multi-band emission filter to image multiple parameters, can be applied to multi-parametric optical mapping. High-speed excitation light switching (as can be achieved with LEDs), coordinated with the camera frame exposure and combined with an appropriate multi-band emission filter provides a simple and cheap multi-parametric optical mapping system. Not using multi-band emission filters, singlecamera detection of ratiometric Vm signals [3] and Vm and [Ca2+]i signals [35], using high-speed and coordinated LED excitation light switching, has been recently reported

Methods

Results

Conclusion