A Singular Role of I Promoting the Development of Cardiac Automaticity During Cardiomyocyte Differentiation by I -Induced Activation of Pacemaker Current

The inward rectifier potassium current (I) is generally thought to suppress cardiac automaticity by hyperpolarizing membrane potential (MP). We recently observed that I could promote the spontaneously-firing automaticity induced by upregulation of pacemaker funny current (I) in adult ventricular cardiomyocytes (CMs). However, the intriguing ability of I to activate I and thereby promote automaticity has not been explored. In this study, we combined mathematical and experimental assays and found that only I and I, at a proper-ratio of densities, were sufficient to generate rhythmic MP-oscillations even in unexcitable cells (i.e. HEK293T cells and undifferentiated mouse embryonic stem cells [ESCs]). We termed this effect I-induced I activation. Consistent with previous findings, our electrophysiological recordings observed that around 50% of mouse (m) and human (h) ESC-differentiated CMs could spontaneously fire action potentials (APs). We found that spontaneously-firing ESC-CMs displayed more hyperpolarized maximum diastolic potential and more outward I current than quiescent-yet-excitable m/hESC-CMs. Rather than classical depolarization pacing, quiescent mESC-CMs were able to fire APs spontaneously with an electrode-injected small outward-current that hyperpolarizes MP. The automaticity to spontaneously fire APs was also promoted in quiescent hESC-CMs by an I-specific agonist zacopride. In addition, we found that the number of spontaneously-firing m/hESC-CMs was significantly decreased when I was acutely upregulated by Ad-CGI-HCN infection. Our study reveals a novel role of I promoting the development of cardiac automaticity in m/hESC-CMs through a mechanism of I-induced I activation and demonstrates a synergistic interaction between I and I that regulates cardiac automaticity.