A detailed characterization of the hyperpolarization-activated “funny” current (If) in human-induced pluripotent stem cell (iPSC)–derived cardiomyocytes with pacemaker activity

Abstract

Properties of the funny current (If) have been studied in several animal and cellular models, but so far little is known concerning its properties in human pacemaker cells. This work provides a detailed characterization of If in human-induced pluripotent stem cell (iPSC)–derived pacemaker cardiomyocytes (pCMs), at different time points. Patch-clamp analysis showed that If density did not change during differentiation; however, after day 30, it activates at more negative potential and with slower time constants. These changes are accompanied by a slowing in beating rate. If displayed the voltage-dependent block by caesium and reversed (Erev) at − 22 mV, compatibly with the 3:1 K+/Na+ permeability ratio. Lowering [Na+]o (30 mM) shifted the Erev to − 39 mV without affecting conductance. Increasing [K+]o (30 mM) shifted the Erev to − 15 mV with a fourfold increase in conductance. pCMs express mainly HCN4 and HCN1 together with the accessory subunits CAV3, KCR1, MiRP1, and SAP97 that contribute to the context-dependence of If. Autonomic agonists modulated the diastolic depolarization, and thus rate, of pCMs. The adrenergic agonist isoproterenol induced rate acceleration and a positive shift of If voltage-dependence (EC50 73.4 nM). The muscarinic agonists had opposite effects (Carbachol EC50, 11,6 nM). Carbachol effect was however small but it could be increased by pre-stimulation with isoproterenol, indicating low cAMP levels in pCMs. In conclusion, we demonstrated that pCMs display an If with the physiological properties expected by pacemaker cells and may thus represent a suitable model for studying human If-related sinus arrhythmias.

Desarrollo de un simulador de la corriente funny del nodo sinusal para la enseñanza-aprendizaje

“…Estos resultados concuerdan con los reportados. Una disminución de [Na + ]e hace más negativo E funny (Giannetti et al, 2021). De acuerdo con Frace y colaboradores, un incremento en [K + ]e debe de aumentar la corriente funny (Frace et al, 1992).…”

“…Estos resultados concuerdan con experimentos reportados. Un incremento a 30 mM produce Efunny = -15 mV (Giannetti et al, 2021). El resultado fue un aumento en la amplitud de If (trazo azul, en el osciloscopio superior) (Figura 3).…”

“…Pertenecen a la superfamilia de canales de K + (Saponaro et al, 2021). Estos canales permiten generar potenciales de acción de manera controlada para mantener la frecuencia cardíaca y buen funcionamiento general del corazón (Giannetti et al, 2021;Guerra & Cinca, 2007).…”

Special issue on recent progress with hPSC-derived cardiovascular cells for organoids, engineered myocardium, drug discovery, disease models, and therapy

“…Their findings suggest that early expression of MYL2 may be informative as an early marker of chamber-specific cardiomyocytes. In a separate research article by Giannetti et al [9], the authors perform a comprehensive study of the hyperpolarization-activated "funny" current (I f ) in hiPSC-derived CMs with pacemaker activity. This is a timely publication, as the I f channel is expressed in hPSC-CMs well after the initiation of spontaneous contractions.…”

A model-guided pipeline for drug cardiotoxicity screening with human stem-cell derived cardiomyocytes

“…To verify the finding that quinine blocks If, we used a HEK-293 cell line stably expressing HCN1, the pore domain that conducts If. The HCN1 isoform was chosen because it was recently shown to be present at high densities in iPSC-CMs (Giannetti et al, 2021). Figure 7A…”