Gérard V, Rouzaire-Dubois B, Dubois J M
Laboratoire de Physiologie Cellulaire, URA CNRS 1121, Université Paris-Sud, Orsay, France.
J Membr Biol. 1994 Jan;137(2):119-25. doi: 10.1007/BF00233481.
The aim of this work was to examine the effects of changes in external K+ concentration (Ko) around its physiological value, of various K+ channels blockers, including internal Cs+, of vacuolar H(+)-ATPase inhibitors and of the protonophore CCCP on the resting potential and the voltage-dependent K+ current of differentiated neuroblastoma x glioma hybrid NG108-15 cells using the whole-cell patch-clamp technique. The results are as follows: (i) under standard conditions (Ko = 5 mM) the membrane potential was -60 +/- 1 mV. It was unchanged when Ko was decreased to 1 mM and was depolarized by 4 +/- 1 mV when Ko was increased to 10 mM. (ii) Internal Cs+ depolarized the membrane by 21 +/- 3 mV. (iii) The internal application of the vacuolar H(+)-ATPase inhibitors N-ethylmaleimide (NEM), NO3- and bafilomycin A1 (BFA) depolarized the membrane by 15 +/- 2, 18 +/- 2 and 16 +/- 2 mV, respectively. (iv) When NEM or BFA were added to the internal medium containing Cs+, the membrane was depolarized by 45 +/- 1 and 42 +/- 2 mV, respectively. (v) The external application of CCCP induced a transient depolarization followed by a prolonged hyperpolarization. This hyperpolarization was absent in BFA-treated cells. The voltage-dependent K+ current was increased at negative voltages and decreased at positive voltages by NEM, BFA and CCCP. Taken together, these results suggest that under physiological conditions, the resting potential of NG108-15 neuroblastoma cells is maintained at negative values by both voltage-dependent K+ channels and an electrogenic vacuolar type H(+)-ATPase.
本研究旨在利用全细胞膜片钳技术，研究胞外钾离子浓度（Ko）在其生理值附近变化、各种钾离子通道阻滞剂（包括胞内Cs+）、液泡H(+)-ATP酶抑制剂以及质子载体CCCP对分化的神经母细胞瘤x胶质瘤杂交NG108-15细胞静息电位和电压依赖性钾电流的影响。结果如下：（i）在标准条件下（Ko = 5 mM），膜电位为-60±1 mV。当Ko降至1 mM时，膜电位不变；当Ko升至10 mM时，膜电位去极化4±1 mV。（ii）胞内Cs+使膜去极化21±3 mV。（iii）液泡H(+)-ATP酶抑制剂N-乙基马来酰亚胺（NEM）、NO3-和巴弗洛霉素A1（BFA）的胞内应用分别使膜去极化15±2、18±2和16±2 mV。（iv）当将NEM或BFA添加到含有Cs+的胞内培养基中时，膜分别去极化45±1和42±2 mV。（v）胞外应用CCCP诱导短暂去极化，随后是长时间的超极化。在BFA处理的细胞中不存在这种超极化。NEM、BFA和CCCP使电压依赖性钾电流在负电压下增加，在正电压下降低。综上所述，这些结果表明，在生理条件下，NG108-15神经母细胞瘤细胞的静息电位通过电压依赖性钾通道和电生性液泡型H(+)-ATP酶维持在负值。
