Chair of Biophysics, University of Rostock, Rostock 18057, Germany.
Sophion-Biolin Scientific, Ballerup 2750, Denmark.
Sci Data. 2017 Apr 25;4:170056. doi: 10.1038/sdata.2017.56.
Human induced pluripotent stem cells can be differentiated into dopaminergic neurons (Dopa.4U). Dopa.4U neurons expressed voltage-gated Na and K channels and showed neuron-like spontaneous electrical activity. In automated patch clamp measurements with suspended Dopa.4U neurons, delayed rectifier K current (delayed K) and rapidly inactivating A-type K current (fast K) were identified. Examination of the fast K current with inhibitors yielded IC values of 0.4 mM (4-aminopyridine) and 0.1 mM (tetraethylammonium). In manual patch clamp measurements with adherent Dopa.4U neurons, fast K current could not be detected, while the delayed K current showed an IC of 2 mM for 4-aminopyridine. The Na channels in adherent and suspended Dopa.4U neurons showed IC values for tetrodotoxin of 27 and 2.9 nM, respectively. GABA-induced currents that could be observed in adherent Dopa.4U neurons could not be detected in suspended cells. Application of current pulses induced action potentials in approx. 70 % of the cells. Our results proved the feasibility of automated electrophysiological characterization of neuronal cells.
人诱导多能干细胞可分化为多巴胺能神经元 (Dopa.4U)。Dopa.4U 神经元表达电压门控钠和钾通道,并表现出类似神经元的自发性电活动。在悬浮的 Dopa.4U 神经元的自动膜片钳测量中,鉴定出延迟整流钾电流 (delayed K) 和快速失活 A 型钾电流 (fast K)。用抑制剂对 fast K 电流的检测得出 IC 值分别为 0.4 mM(4-氨基吡啶)和 0.1 mM(四乙铵)。在贴壁 Dopa.4U 神经元的手动膜片钳测量中,无法检测到 fast K 电流,而延迟 K 电流在 4-氨基吡啶中显示出 2 mM 的 IC 值。贴壁和悬浮的 Dopa.4U 神经元中的钠通道对河豚毒素的 IC 值分别为 27 和 2.9 nM。在贴壁的 Dopa.4U 神经元中可以观察到的 GABA 诱导电流在悬浮细胞中无法检测到。施加电流脉冲可在大约 70%的细胞中诱导动作电位。我们的结果证明了自动电生理特性分析神经元细胞的可行性。