Optogenetic Approach to Cardiotoxicity Screening: Simultaneous Voltage and Calcium Imaging Under Paced Conditions

Abstract

We have developed an in vitro cardiotoxicity assay using genetically encoded, engineered protein sensors and actuators (Dempsey et al., J Pharmacol Toxicol Methods, doi: 10.1016/j.vascn.2016.05.003 , 2016). The assay, based on the Optopatch platform for all-optical electrophysiology (Hochbaum et al., Nat Methods 11:825–833, 2014), incorporates simultaneous measurement of the action potential (AP) wave-form and Ca 2+ transient (CT) in human-induced pluripotent stem cell (hiPSC)-derived cardiomyocytes (CMs) under light-controlled, optogenetic pacing. A syncytium of hiPSC-derived CMs is prepared in a mixed monolayer culture, where a subset of CMs expresses a genetically encoded, dual-function calcium and voltage reporter, CaViar (Hou et al. Front Physiol 5:1–10, 2014), while others express a channelrho-dopsin variant, CheRiff. Optical pacing of the CheRiff-expressing cells synchronizes the syncytium and provides a stable electrophysiological background. In this chapter, we discuss details of the Cardiac Optopatch methodology, including cell preparation, Optopatch imaging, and data analysis. We have used this platform to screen the cardiotoxic effects of compounds, both acutely and chronically; examples will be shown. The Optopatch platform provides a robust assay to measure APs and CTs in hiPSC-CMs. This assay will facilitate comparisons of cell-based assays to human clinical data with the goal of a more accurate in vitro predictor of clinical torsades de pointes (TdP).