Mohammed Djemai

Optical mapping is a powerful imaging technique widely adopted to measure membrane potential changes and intracellular Ca²⁺ variations in excitable tissues using voltage-sensitive dyes and Ca²⁺ indicators, respectively. This powerful tool has rapidly become indispensable in the field of cardiac electrophysiology for studying depolarization wave propagation, estimating the conduction velocity of electrical impulses, and measuring Ca²⁺ dynamics in cardiac cells and tissues. In addition, mapping these electrophysiological parameters is important for understanding cardiac arrhythmia mechanisms. In this review, we delve into the fundamentals of cardiac optical mapping technology and its applications when applied to hiPSC-derived cardiomyocytes and discuss related advantages and challenges. We also provide a detailed description of the processing and analysis of optical mapping data, which is a crucial step in the study of cardiac diseases and arrhythmia mechanisms for extracting and comparing relevant electrophysiological parameters.

Cardiomyocytes derived from patient-specific induced pluripotent stem cells (iPSC-CMs) successfully reproduce the mechanisms of several channelopathies. However, this approach involve cell reprogramming from somatic tissue biopsies or genomic editing in healthy iPSCs for every mutation found and to be investigated. We aim to knockout (KO) Na_V1.5, the cardiac sodium channel, in a healthy human iPSC line, characterize the model and then, use it to express variants of Na_V1.5. We develop a homozygous Na_V1.5 KO iPSC line able to differentiate into cardiomyocytes with CRISPR/Cas9 tool. The Na_V1.5 KO iPSC-CMs exhibited an organized contractile apparatus, spontaneous contractile activity, and electrophysiological recordings confirmed the major reduction in total Na⁺ currents. The action potentials (APs) exhibited a reduction in their amplitude and in their maximal rate of rise. Voltage optical mapping recordings revealed that the conduction velocity Ca²⁺ transient waves propagation velocities were slow. A wild-type (WT) Na_V1.5 channel expressed by transient transfection in the KO iPSC-CMs restored Na⁺ channel expression and AP properties. The expression of Na_V1.5/delQKP, a long QT type 3 (LQT3) variant, in the Na_V1.5 KO iPSC-CMs showed that dysfunctional Na⁺ channels exhibited a persistent Na⁺ current and caused prolonged AP duration that led to arrhythmic events, characteristics of LQT3.

Cardiac complications such as electrical abnormalities including conduction delays and arrhythmias are the main cause of death in individuals with Myotonic Dystrophy type 1 (DM1). We developed a disease model using iPSC-derived cardiomyocytes (iPSC-CMs) from a healthy individual and two DM1 patients with different CTG repeats lengths and clinical history (DM1-1300 and DM1-300). We confirmed the presence of toxic RNA foci and mis-spliced MBNL1/2 transcripts in DM1 iPSC-CMs. In DM1-1300, we identified a switch in the cardiac sodium channel SCN5A from the adult to the neonatal isoform. The down-regulation of adult SCN5A isoforms is consistent with a shift in the sodium current activation to depolarized potentials observed in DM1-1300. L-type calcium current density was higher in iPSC-CMs from DM1-1300, which is correlated with the overexpression of the Ca_V1.2 transcript and proteins. Importantly, I_Na and I_CaL dysfunctions resulted in prolonged action potentials duration, slower velocities, and decreased overshoots. Optical mapping analysis revealed a slower conduction velocity in DM1-1300 iPSC-CM monolayers. In conclusion, our data revealed two distinct ions channels perturbations in DM1 iPSC-CM from the patient with cardiac dysfunction, one affecting Na⁺ channels and one affecting Ca²⁺ channels. Both have an impact on cardiac APs and ultimately on heart conduction.