Fast Optical Investigation of Cardiac Electrophysiology by Parallel Detection in Multiwell Plates

Caterina Credi; Valentina Balducci; U Munagala; C Cianca; S Bigiarini; Antoine A F de Vries; Leslie M Loew; Francesco S Pavone; Elisabetta Cerbai; Laura Sartiani; Leonardo Sacconi

doi:10.3389/fphys.2021.692496

Fast Optical Investigation of Cardiac Electrophysiology by Parallel Detection in Multiwell Plates

Front Physiol. 2021 Sep 3:12:692496. doi: 10.3389/fphys.2021.692496. eCollection 2021.

Authors

Caterina Credi^{1

2}, Valentina Balducci³, U Munagala^{3

4}, C Cianca¹, S Bigiarini¹, Antoine A F de Vries⁵, Leslie M Loew⁶, Francesco S Pavone^{1

2

7}, Elisabetta Cerbai^{1

3}, Laura Sartiani³, Leonardo Sacconi^{1

2}

Affiliations

¹ European Laboratory for Non-linear Spectroscopy, Sesto Fiorentino, Italy.
² National Institute of Optics, National Research Council, Florence, Italy.
³ Department of Neurosciences, Psychology, Drugs and Child Health, University of Florence, Florence, Italy.
⁴ Core Research Laboratory, ISPRO, Florence, Italy.
⁵ Laboratory of Experimental Cardiology, Department of Cardiology, Leiden University Medical Center, Leiden, Netherlands.
⁶ R. D. Berlin Center for Cell Analysis and Modeling, University of Connecticut School of Medicine, Farmington, CT, United States.
⁷ Department of Physics and Astronomy, University of Florence, Sesto Fiorentino, Italy.

Abstract

Current techniques for fast characterization of cardiac electrophysiology employ optical technologies to control and monitor action potential features of single cells or cellular monolayers placed in multiwell plates. High-speed investigation capacities are commonly achieved by serially analyzing well after well employing fully automated fluorescence microscopes. Here, we describe an alternative cost-effective optical approach (MULTIPLE) that exploits high-power LED arrays to globally illuminate a culture plate and an sCMOS sensor for parallel detection of the fluorescence coming from multiple wells. MULTIPLE combines optical detection of action potentials using a red-shifted voltage-sensitive fluorescent dye (di-4-ANBDQPQ) with optical stimulation, employing optogenetic actuators, to ensure excitation of cardiomyocytes at constant rates. MULTIPLE was first characterized in terms of interwell uniformity of the illumination intensity and optical detection performance. Then, it was applied for probing action potential features in HL-1 cells (i.e., mouse atrial myocyte-like cells) stably expressing the blue light-activatable cation channel CheRiff. Under proper stimulation conditions, we were able to accurately measure action potential dynamics across a 24-well plate with variability across the whole plate of the order of 10%. The capability of MULTIPLE to detect action potential changes across a 24-well plate was demonstrated employing the selective K _v 11.1 channel blocker (E-4031), in a dose titration experiment. Finally, action potential recordings were performed in spontaneous beating human induced pluripotent stem cell derived cardiomyocytes following pharmacological manipulation of their beating frequency. We believe that the simplicity of the presented optical scheme represents a valid complement to sophisticated and expensive state-of-the-art optical systems for high-throughput cardiac electrophysiological investigations.

Keywords: action potential; microscopy; multiwell plate; optogenetics; voltage imaging.