Whole-cell patch-clamp recordings were made from human ESC-derived serotonergic neurons at 6 weeks. Briefly, the neurons were hold at −70 mV to record the Na+/K+ channel activities, and at 0 mV to record the spontaneous release with voltage-clamp model. For recording action potentials, the cells were held at 0 pA with the current-clamp model, and with the steps of injected currents from −40 pA to + 100 pA. The bath solution consisted of 127 mM NaCl, 1.2 mM KH2PO4, 1.9 mM KCl, 26 mM NaHCO3, 2.2 mM CaCl2, 1.4 mM MgSO4, 10 mM glucose, 290 mM mOsm and 95% O2/5% CO2. Recording pipettes were filled with an intracellular solution containing 20 mM KCl, 121 mM K+-gluconate, 10 mM Na+-HEPES, 10 mM BAPTA, 4 mM Mg2+-ATP pH 7.2 and 290 mOm. An Olympus BX51WI microscope was used to visualize neurons. A MultiClamp 700B amplifer (Axon instruments, Molecular Devices, Sunnyvale, CA, USA) was used to investigate the voltage clamp and current clamp recordings. Signals were filtered at 4 kHZ and sampled at 100 kHz using a Digidata 1322A analog-digital converter (Axon instruments). Data were analyzed with pClamp 9.0 (Axon instruments).
