The patch-clamp experiments was performed as previously described [19 (link),22 (link)]. Briefly, 35,000 atrial iPSC-CMs were plated on glass-bottom Fluoro Dishes and incubated with either isoproterenol (50 nmol/L, Sigma-Aldrich, Taufkirchen, Germany)) or isoproterenol + PF01247324 (1 µmol/L, Sigma-Aldrich, Taufkirchen, Germany)) for 15 min before starting the measurements. The experiments were conducted at room temperature.
Action potential recordings were performed using the whole-cell patch-clamp technique. To elicit action potentials, square current pulses with amplitudes of 0.5–1 nA and durations of 1–5 ms were applied. The stimulation frequency was increased gradually from 0.5 to 2 Hz.
The late sodium current (INaL) was measured using the ruptured-patch whole-cell patch-clamp technique. The pipette used had a resistance ranging from 2 to 3 mega-ohms (MΩ). INaL recordings were performed exclusively in CMs where a seal with a resistance of over 1 giga-ohm (GΩ) was achieved, and the access resistance remained below 7 MΩ. After a stabilization period of 3 min, the iPSC-derived CMs were held at a holding potential of −120 mV and then depolarized to −35 mV for 1000 ms with 10 pulses and a basic cycle length of 2 s. The INaL was quantified as the integral current amplitude between 100 and 500 ms and was normalized to the membrane capacitance.
Action potential recordings were performed using the whole-cell patch-clamp technique. To elicit action potentials, square current pulses with amplitudes of 0.5–1 nA and durations of 1–5 ms were applied. The stimulation frequency was increased gradually from 0.5 to 2 Hz.
The late sodium current (INaL) was measured using the ruptured-patch whole-cell patch-clamp technique. The pipette used had a resistance ranging from 2 to 3 mega-ohms (MΩ). INaL recordings were performed exclusively in CMs where a seal with a resistance of over 1 giga-ohm (GΩ) was achieved, and the access resistance remained below 7 MΩ. After a stabilization period of 3 min, the iPSC-derived CMs were held at a holding potential of −120 mV and then depolarized to −35 mV for 1000 ms with 10 pulses and a basic cycle length of 2 s. The INaL was quantified as the integral current amplitude between 100 and 500 ms and was normalized to the membrane capacitance.
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