Membrane potential, anion and cation conductances in Ehrlich ascites tumor cell
Membrane potential, anion and cation conductances in Ehrlich ascites tumor cell
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艾利希腹水肿瘤细胞的膜电位、阴离子和阳离子电导
DOI：
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发表时间：
1989
影响因子：
2.4
通讯作者：
F. Jørgensen
中科院分区：
文献类型：
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作者：
I. H. Lambert;E. Hoffmann;F. Jørgensen
在埃利希腹水癌细胞悬液中测定了染料1，1 ′-二丙基氧杂双环菁（DiOC 3-（5））的荧光强度，以监测在不同离子条件下、用阳离子载体处理后和低渗细胞肿胀后的膜电位（Vm）。在无Na+的K+/胆碱+培养基中用短杆菌肽进行校准，即，用KCl和cholineCl代替NaCl的标准培养基，其中钾和胆碱的总和保持恒定在155 mM。通过Lariset等人（Laris，P.C.，Pershadsingh，A.，约翰斯通，R.M.，1976年，生物化学。Biophys. Acta 436：475-488）仅在Cl−通道阻断剂indacrinone（MK 196）存在下有效。发现亲脂性阴离子SCN-的分布作为膜电位的间接估计不适用于本文报道的Vm的快速变化。与DiOC 3-（5）孵育5 min可使Cl-渗透率降低26±5%，NO3-渗透率降低15± 2%，但对K+渗透率无显著影响。在等渗标准NaCl介质中，Vm值（根据染料对阴离子电导的抑制效应进行校正）估计为−61±1 mV，在等渗无Na+胆碱介质中为−78±3 mV，在等渗NaNO 3介质中为−46±1 mV。加入K ~+通道抑制剂奎宁后，细胞膜去极化，悬浮于无Na ~+胆碱介质中时，细胞膜超极化，表明Vm部分由钾离子扩散产生，部分由钠离子扩散产生。埃利希细胞先前已被证明对硝酸盐的渗透性比对氯化物的渗透性更强。用NO3−取代所有细胞和细胞外的Cl−会导致膜的去极化，这表明Vm也是由阴离子产生的，阴离子高于平衡。考虑到之前证明的K+通道的单列行为，埃利希细胞的膜电导估计为K+10.4 μS/cm 2，Na+3.0 μS/cm 2，Cl− 0.6 μS/cm 2，NO3− 8.7 μS/cm 2。添加Ca 2+离子载体A23187导致KCl的净损失和膜的超极化，表明在添加A23187后K+渗透性也超过Cl−渗透性。A23187处理的埃利希细胞中的K+和Cl−电导估计分别为134和30 μS/cm 2。在低渗肿胀的细胞中，膜电位被去极化，证实了低渗暴露后Cl−渗透性的增加超过了K+渗透性的相应增加。在膜电位Vm =EK=ECl=ENa的对照实验中，证明细胞体积变化对荧光信号没有显著影响，显然是因为大的细胞内缓冲能力。根据净Cl-流出量和Vm变化估计，当细胞转移到渗透压浓度为标准培养基一半的培养基中时，Cl-电导增加68倍。从净K+流出量估计，K+电导的相应增加仅为两倍。
SummaryThe fluorescence intensity of the dye 1,1′-dipropyloxadicarbocyanine (DiOC3-(5)) has been measured in suspensions of Ehrlich ascites tumor cells in an attempt to monitor their membrane potential (Vm) under different ionic conditions, after treatment with cation ionophores and after hypotonic cell swelling. Calibration is performed with gramicidin in Na+-free K+/choline+ media, i.e., standard medium in which NaCl is replaced by KCl and cholineCl and where the sum of potassium and choline is kept constant at 155mm. Calibration by the valinomycin “null point” procedure described by Lariset al. (Laris, P.C., Pershadsingh, A., Johnstone, R.M., 1976,Biochim. Biophys. Acta436:475–488) is shown to be valid only in the presence of the Cl−-channel blocker indacrinone (MK196). Distribution of the lipophilic anion SCN− as an indirect estimation of the membrane potential is found not to be applicable for the fast changes inVm reported in this paper. Incubation with DiOC3-(5) for 5 min is demenstrated to reduce the Cl− permeability by 26±5% and the NO3− permeability by 15±2%, while no significant effect of the probe could be demonstrated on the K+ permeability. Values forVm, corrected for the inhibitory effect of the dye on the anion conductance, are estimated at −61±1 mV in isotonic standard NaCl medium, −78±3 mV in isotonic Na+-free choline medium and −46±1 mV in isotonic NaNO3 medium. The cell membrane is depolarized by addition of the K+ channel inhibitor quinine and it is hyperpolarized when the cells are suspended in Na+-free choline medium, indicating thatVm is generated partly by potassium and partly by sodium diffusion. Ehrlich cells have previously been shown to be more permeable to nitrate than to chloride. Substituting NO3− for all cellular and extracellular Cl− leads to a depolarization of the membrane, demonstrating thatVm is also generated by the anions and that anions are above equilibrium. Taking the previously demonstrated single-file behavior of the K+ channels into consideration, the membrane conductances in Ehrlich cells are estimated at 10.4 μS/cm2 for K+, 3.0 μS/cm2 for Na+, 0.6 μS/cm2 for Cl− and 8.7 μS/cm2 for NO3−. Addition of the Ca2+-ionophore A23187 results in net loss of KCl and a hyperpolarization of the membrane, indicating that the K+ permeability exceeds the Cl− permeability also after the addition of A23187. The K+ and Cl− conductances in A23187-treated Ehrlich cells are estimated at 134 and 30 μS/cm2, respectively. The membrane potential is depolarized in hypotonically swollen cells, confirming that the increase in the Cl− permeability following hypotonic exposure exceeds the concommitant increase in the K+ permeability. In control experiments where the membrane potentialVm=EK=ECl=ENa, it is demonstrated that cell volume changes has no significant effect on the fluorescence signal, apparently because of a large intracellular buffering capacity. The increase in the Cl− conductances is 68-fold when cells are transferred to a medium with half the osmolarity of the standard medium, as estimated from the net Cl− efflux and the change inVm. The concommitant increase in the K+ conductance, as estimated from the net K+ efflux, is only twofold.
艾利希腹水肿瘤细胞中 Na 依赖性氨基酸共转运的能量学。
DOI：
10.1016/0005-2736(87)90309-9
发表时间：
1987
期刊：
Biochimica et biophysica acta
影响因子：
--
作者：
Dawson,WD;Smith,TC
通讯作者：
Smith,TC
荧光探针 3,3-二丙基硫二碳花青碘化物对艾氏腹水肿瘤细胞能量代谢的影响。
DOI：
--
发表时间：
1981
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
Smith,TC;Herlihy,JT;Robinson,SC
通讯作者：
Robinson,SC