An investigation into the role of the optical detection set-up in the recording of cardiac optical mapping signals: A Monte Carlo simulation study

Photon scattering is known to distort the fluorescence signals recorded from optically mapped cardiac tissue. However, the contribution of the parameters which define the optical detection set-up has not been assessed. In this study, Monte Carlo (MC) simulations of photon scattering within ventricular tissue are combined with a detailed model of a tandem-lens optical detection apparatus to characterise (i) the spatial origin upon emission of photons recorded in voltage-sensitive fluorescence measurements of cardiac electrical activity (using the fluorescent dye di-4-ANEPPS) and how this affects signal distortion, and (ii) the role the detector characteristics could play in modulating signal distortion during uniform illumination and photon emission from tissue depth. Results show that, for the particular excitation/emission wavelengths considered (488 nm and 669 nm, respectively), the dimensions of the scattering volume during uniform illumination extend around 3 times further in the surface recording plane than in depth. As a result, fluorescence recordings during electrical propagation are more distorted when transmembrane potential levels differ predominantly in the surface plane than in depth. In addition, MC simulation results show that the spatial accuracy of the fluorescence signal is significantly limited due to photon scattering, with only a small fraction of the recorded signal intensity originating from tissue beneath the pixel (approximately 11% for a 0.25×0.25 mm pixel). Increasing pixel size increases this fraction, however, it also results in an increase in the scattering volume dimensions, thus reducing the spatial resolution of the optical system, and increasing signal distortion. MC simulations also demonstrate that photon scattering in cardiac tissue limits the ability of optical detection system tuning in accurately locating fluorescent emission from depth. Specifically, our results prove that the focal plane depth that yields maximum signal intensity provides an underestimation of the emission depth. In conclusion, our study demonstrates the potential of MC simulations of photon scattering in guiding the design of optical mapping set-ups to optimise performance under diverse experimental conditions.