Extracellular ATP Triggers Action Potentials in Ventricular Cardiomyocytes

Myocardial ischemia causes a high frequency of ventricular arrhythmias. A long established metabolic response to hypoxia in the heart is the release of ATP. Here we tested the hypothesis that ATP released from cardiomyocytes during acute myocardial injury can contribute to ventricular ectopy by causing delayed afterdepolarizations and triggered action potentials. Isolated mouse ventricular cardiomyocytes were loaded with Fura2-AM to measure intracellular Ca. Spontaneous Ca waves (SCW) and triggered beats (TB) were quantified after a train of field-stimulated Ca transients (for 20s with 3 Hz) in presence of inotropic stimulation with isoproterenol (1 µM) and partial inhibition of the inward rectifier with Ba (5 µM) to facilitate TBs. Application of extracellular ATP (3-50 µM) significantly increased the incidence and frequency of TBs, whereas the rate of SCWs was not significantly changed. Notably, extracellular ATP application generated a different type of TBs that was not preceded by SCW, which were termed ATP-triggered beats (TBATP). Higher concentrations of ATP (> 50 µM) reduced the rates of TBATP. Simultaneous [Ca]i and membrane potential measurements using perforated patch method demonstrated that both regular TBs and TBATP were preceded by a delayed afterdepolarization (DAD) of similar shape. However, in presence of ATP, 34% of DAD that triggered beats occurred WITHOUT concomitant elevations of cytosolic [Ca]. In other words, general DADs and TBs were mediated by the classic Ca-dependent mechanism (i.e., Na-Ca exchange current in response to spontaneous SR Ca release), whereas ATP generated DADs occurred in the absence of the classic Ca-dependent pathway. In isolated Langendorff perfused mouse hearts, adding ATP (30 µM) to the perfusate triggered both atrial and ventricular premature beats. We conclude that low micromolar extracellular ATP activates an inward current that is sufficient to generate DADs and spontaneous action potentials in the absence of spontaneous SR Ca release. Although the molecular identity of ATP-activated inward current remains to be determined, the current study demonstrates that ATP release by acute ischemic stress could facilitate arrhythmia susceptibility via a Ca-independent mechanism.