High-throughput drug profiling with voltage- and calcium-sensitive fluorescent probes in human iPSC-derived cardiomyocytes

Abstract

Bedut S, Seminatore-Nole C, Lamamy V, Caignard S, Boutin JA, Nosjean O, Stephan JP, Coge F. High-throughput drug profiling with voltage-and calcium-sensitive fluorescent probes in human iPSC-derived cardiomyocytes. Am J Physiol Heart Circ Physiol 311: H44 -H53, 2016. First published May 3, 2016 doi:10.1152/ajpheart.00793.2015.-Cardiomyocytes derived from human embryonic stem cells (hESCs) or induced pluripotent stem cells (hiPSCs) are increasingly used for in vitro assays and represent an interesting opportunity to increase the data throughput for drug development. In this work, we describe a 96-well recording of synchronous electrical activities from spontaneously beating hiPSC-derived cardiomyocyte monolayers. The signal was obtained with a fast-imaging plate reader using a submillisecond-responding membrane potential recording assay, FluoVolt, based on a newly derived voltagesensitive fluorescent dye. In our conditions, the toxicity of the dye was moderate and compatible with episodic recordings for Ͼ3 h. We show that the waveforms recorded from a whole well or from a single cell-sized zone are equivalent and make available critical functional parameters that are usually accessible only with gold standard techniques like intracellular microelectrode recording. This approach allows accurate identification of the electrophysiological effects of reference drugs on the different phases of the cardiac action potential as follows: fast depolarization (lidocaine), early repolarization (nifedipine, Bay K8644, and veratridine), late repolarization (dofetilide), and diastolic slow depolarization (ivabradine). Furthermore, the data generated with the FluoVolt dye can be pertinently complemented with a calcium-sensitive dye for deeper characterization of the pharmacological responses. In a semiautomated plate reader, the two probes used simultaneously in 96-well plates provide an easy and powerful multiparametric assay to rapidly and precisely evaluate the cardiotropic profile of compounds for drug discovery or cardiac safety.

Calcium Transient Assays for Compound Screening with Human iPSC-derived Cardiomyocytes: Evaluating New Tools

“…CaT amplitude was very stable across different measurements of the plate, in contrast to CaT duration which was highly dependent on the conditions of the plate (i.e. temperature, % CO 2 ) and tended to increase during measurement, as also reported previously 18 . This emphasizes the importance of maintaining the plate as closely as possible to incubation conditions during plate readings and designing an assay flow and analysis to avoid detecting false positives.…”

Two dimensional electrophysiological characterization of human pluripotent stem cell-derived cardiomyocyte system

“…High-throughput optical readouts using voltage dyes, calcium dyes, or their combinations have demonstrated the utility of hESC/iPSC-CMs as a model system for such methods647. While our current system configuration cannot be directly utilized in high-throughput analysis, there is no fundamental barrier that precludes the development of such capabilities.…”

Integrating cardiomyocytes from human pluripotent stem cells in safety pharmacology: has the time come?

“…In contrast, Bedut et al . demonstrated reduced beating rates in hiPSC‐CMs at lower doses of ivabradine (1‐2 μM vs 3‐9 μM) (Bedut et al , 2016). Furthermore, when drug concentrations were compared among different hPSC‐CM lines, techniques or studies, the differences appear to be significant and functionally relevant (Kuusela et al , 2016).…”