ELECTROPERMEABILIZATION OF CELL-MEMBRANES - EFFECT OF THE RESTING MEMBRANE-POTENTIAL
ELECTROPERMEABILIZATION OF CELL-MEMBRANES - EFFECT OF THE RESTING MEMBRANE-POTENTIAL
复制标题
DOI：
10.1016/s0006-291x(05)80206-2
复制
发表时间：
1990-10-15
影响因子：
3.1
通讯作者：
CHOCK, PB
中科院分区：
文献类型：
--
作者：
TEKLE, E;ASTUMIAN, RD;CHOCK, PB
电场诱导的细胞膜透化是基因转染和细胞杂交的重要技术1 -4。这一过程的机制研究表明，植物原生质体的荧光指示剂的摄取主要发生在半球面临的正电极5，而在红细胞鬼的探针退出通过半球面临的负电极6。为了调和这些观察对称孔的形成和分子交换机制，电渗已被提出6。鉴于这些有争议的观察结果，我们进行了系统的研究与不同的电场强度，波形和频率的NIH 3 T3细胞的电穿孔。我们的数据显示：（i）细胞膜的对称透化仅发生在双极交流。领域的(ii)当临界膜击穿电位Vc使用单极a.c.场或单个直流方波脉冲时，细胞膜仅在面对正电极的半球处变得透化。(iii)当脉冲诱导的膜电位Vm近似等于或大于固有膜电位（即使用直流或单极交流）时，场）时，观察到不对称透化，其中面对正电极的半球是最可渗透的。(iv)荧光指示剂摄取的速率取决于指示剂的浓度。这些结果表明，细胞膜的电透化受其静息电位的影响，并且电渗不是电穿孔中细胞摄取外源分子的主要机制。
Electric field induced permeabilization of cell membranes is an important technique for gene transfection and cell hybridization1-4. Mechanistic studies of this process revealed that the uptake of fluorescent indicator by plant protoplasts occurs predominantly on the hemisphere facing the positive electrode5, while in erythrocyte ghosts the probes exit through the hemisphere facing the negative electrode6. To reconcile these observations symmetrical pore formation and a mechanism of molecular exchange by electroosmosis has been proposed6. In light of these controversial observations, we conducted a systematic study of electroporation of NIH3T3 cells with varying electric field strength, waveform and frequency. Our data revealed that (i) symmetrical permeabilization of the cell membrane occurs only with bipolar a.c. fields. (ii) When a critical membrane breakdown potential, Vc, is applied using either an unipolar a.c. fields or a single d.c. square pulse, the cell membrane becomes permeabilized only at the hemisphere facing the positive electrode. (iii) When the pulse-induced membrane potential, Vm, is approximately equal to or larger than the intrinsic membrane potential (i.e. using d.c. or unipolar a.c. field), asymmetric permeabilization was observed with the hemisphere facing the positive electrode being most permeable. (iv) The rate of fluorescent indicator uptake is dependent on the concentration of the indicator. These results indicate that electro-permeabilization of cell membranes is affected by its resting potential and that electroosmosis is not the dominant mechanism for the cellular uptake of foreign molecules in electroporation.