Sinoatrial node myocytes (SAMs) act as cardiac pacemaker cells by firing spontaneous action potentials (APs) that initiate each heartbeat. The funny current, If, is critical for the generation of these spontaneous APs; however, its precise role during the pacemaking cycle remains unresolved. We used the AP-clamp technique to quantify If during the cardiac cycle in mouse SAMs. We found that If is persistently active throughout the sinoatrial AP, with surprisingly little voltage-dependent gating. As a consequence, it carries both inward and outward current around its reversal potential of -30 mV. Despite operating at only 2-5% of its maximal conductance, If carries a substantial fraction of both depolarizing and repolarizing net charge movement during the firing cycle. We also show that β-adrenergic receptor stimulation increases the percentage of net depolarizing charge moved by If, consistent with a contribution of If to the fight-or-flight increase in heart rate. These properties were confirmed by heterologously-expressed HCN4 channels and by mathematical models of If. Modelling further suggested that the slow activation and deactivation of the HCN4 isoform underlie the persistent activity of If during the sinoatrial AP. These results establish a new conceptual framework for the role of If in pacemaking, in which it operates at a very small fraction of maximal activation but nevertheless drives membrane potential oscillations in SAMs by providing substantial driving force in both inward and outward directions.Significance StatementCardiac pacemaker cells trigger each heartbeat by virtue of spontaneous oscillations in their membrane voltage. Although the funny current (If) is critical for these oscillations and for setting heart rate, its precise role remains an enigma because it activates mostly outside of the physiological voltage range and quite slowly relative to the pacemaker cycle. Here we show that If is persistently active in pacemaker cells; once opened, the small fraction of ion channels that conduct If do not re-close. Consequently, If flows both inward and outward to help propel the voltage oscillations and it paradoxically conducts a large fraction of the net charge movement. These results establish a new conceptual framework for the role of If in driving cardiac pacemaking.
