Fast Optical Investigation of Cardiac Electrophysiology by Parallel Detection in Multiwell Plates
Anno: 2021
Autori: Credi C., Balducci V., Munagala U., Cianca C., Bigiarini S., de Vries AAF., Loew LM., Pavone F.S., Cerbai E., Sartiani L., Sacconi L.
Affiliazione autori: European Lab Non linear Spect, Sesto Fiorentino, Italy; Natl Inst Optic, Natl Res Council, Florence, Italy; Univ Florence, Dept Neurosci, Psychol, Drugs & Child Hlth, Florence, Italy; Core Res Lab, ISPRO, Florence, Italy; Leiden Univ, Med Ctr, Dept Cardiol, Lab Expt Cardiol, Leiden, Netherlands; Univ Connecticut, Sch Med, R D Berlin Ctr Cell Anal & Modeling, Farmington, CT USA; Univ Florence, Dept Phys & Astron, 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.
Giornale/Rivista: FRONTIERS IN PHYSIOLOGY
Volume: 12 Da Pagina: 692496-1 A: 692496-12
Maggiori informazioni: This work was supported by TelethonItaly (GUP19012).Parole chiavi: optogenetics; voltage imaging; action potential; multiwell plate; microscopyDOI: 10.3389/fphys.2021.692496Citazioni: 8dati da “WEB OF SCIENCE” (of Thomson Reuters) aggiornati al: 2025-05-18Riferimenti tratti da Isi Web of Knowledge: (solo abbonati)
