Abstract 17966: A Near-infrared Fluorescent Voltage-sensitive Dye Allows for Moderate-throughput Electrophysiological Analyses of Human Induced Pluripotent Stem Cell-derived Cardiomyocytes
Introduction: Recent advances now allow for the routine derivation of patient- and disease-specific human iPSC cardiomyocyte (iPSC-CM) models. However, a major impediment to the widespread use of iPSC-CM assays is the low throughput of the currently available electrophysiological tools. The patch clamp technique is too laborious and the inherent phototoxicity of most voltage-sensitive dyes limits their use in single cells.
Hypothesis: We tested the hypothesis that near-infrared fluorescent voltage-sensitive dye di-4-ANBDQBS exhibits minimal phototoxicity and high precision to allow for moderate-throughput analysis of human iPSC-CMs.
Methods: We compared simultaneous electrical and optical action potential (AP) recordings in human iPSC-CM loaded with di-4-ANBDQBS.
Results: Optical APs tracked electrical APs with high precision, generating nearly identical values of action potential duration (APD10, APD50 and APD90) for atrial- and ventricular-like CMs (Table). Fluorescence from a single CM yielded interpretable optical APs (Fig A-C). CMs tolerated repeated laser exposure, with stable optical AP parameters recorded over a 30 min study period. Optical AP recordings appropriately tracked changes in repolarization induced by pharmacological manipulation. Finally, di-4-ANBDQBS allowed for moderate-throughput analyses that facilitated the distinction between atrial- and ventricular-like CM. APD50 values (n=208) were best fit to a Gaussian function with 2 distinct peaks that effectively discriminated atrial- and ventricular-like CMs (Fig D). Overall throughput was increased >10-fold over traditional patch clamp technique.
Conclusions: We conclude that the voltage-sensitive dye di-4-ANBDQBS allows for high precision optical AP measurements that markedly increase the throughput for electrophysiological characterization of human iPSC-CMs. Optical APD50 may be useful to discriminate between atrial- and ventricular-like phenotypes.
Author Disclosures: A. Lopez-Izquierdo: None. M. Warren: None. M. Riedel: None. S. Cho: None. S. Lai: None. R. Lux: None. K.W. Spitzer: None. I. Benjamin: None. M. Tristani-Firouzi: None. C.J. Jou: None.
This research has received full or partial funding support from the American Heart Association
- © 2014 by American Heart Association, Inc.