Monitoring Human-Induced Pluripotent Stem Cell-Derived Cardiomyocytes with Genetically Encoded Calcium and Voltage Fluorescent Reporters

The advent of the human-induced pluripotent stem cell (hiPSC) technology has transformed biomedical research, providing new tools for human disease modeling, drug development, and regenerative medicine. To fulfill its unique potential in the cardiovascular field, efficient methods should be developed for high-resolution, large-scale, long-term, and serial functional cellular phenotyping of hiPSC-derived cardiomyocytes (hiPSC-CMs). To achieve this goal, we combined the hiPSC technology with genetically encoded voltage (ArcLight) and calcium (GCaMP5G) fluorescent indicators. Expression of ArcLight and GCaMP5G in hiPSC-CMs permitted to reliably follow changes in transmembrane potential and intracellular calcium levels, respectively. This allowed monitoring short- and long-term changes in action-potential and calcium-handling properties and the development of arrhythmias in response to several pharmaceutical agents and in hiPSC-CMs derived from patients with different inherited arrhythmogenic syndromes. Combining genetically encoded fluorescent reporters with hiPSC-CMs may bring a unique value to the study of inherited disorders, developmental biology, and drug development and testin

Optogenetic Control of Human Induced Pluripotent Stem Cell‐Derived Cardiac Tissue Models

Background Optogenetics, using light‐sensitive proteins, emerged as a unique experimental paradigm to modulate cardiac excitability. We aimed to develop high‐resolution optogenetic approaches to modulate electrical activity in 2‐ and 3‐dimensional cardiac tissue models derived from human induced pluripotent stem cell (hiPSC)‐derived cardiomyocytes. Methods and Results To establish light‐controllable cardiac tissue models, opsin‐carrying HEK293 cells, expressing the light‐sensitive cationic‐channel CoChR, were mixed with hiPSC‐cardiomyocytes to generate 2‐dimensional hiPSC‐derived cardiac cell‐sheets or 3‐dimensional engineered heart tissues. Complex illumination patterns were designed with a high‐resolution digital micro‐mirror device. Optical mapping and force measurements were used to evaluate the tissues' electromechanical properties. The ability to optogenetically pace and shape the tissue's conduction properties was demonstrated by using single or multiple illumination stimulation sites, complex illumination patterns, or diffuse illumination. This allowed to establish in vitro models for optogenetic‐based cardiac resynchronization therapy, where the electrical activation could be synchronized (hiPSC‐derived cardiac cell‐sheets and engineered heart tissue models) and contractile properties improved (engineered heart tissues). Next, reentrant activity (rotors) was induced in the hiPSC‐derived cardiac cell‐sheets and engineered heart tissue models through optogenetics programmed‐ or cross‐field stimulations. Diffuse illumination protocols were then used to terminate arrhythmias, demonstrating the potential to study optogenetics cardioversion mechanisms and to identify optimal illumination parameters for arrhythmia termination. Conclusions By combining optogenetics and hiPSC technologies, light‐controllable human cardiac tissue models could be established, in which tissue excitability can be modulated in a functional, reversible, and localized manner. This approach may bring a unique value for physiological/pathophysiological studies, for disease modeling, and for developing optogenetic‐based cardiac pacing, resynchronization, and defibrillation approaches