Bezprozvanny I, Ehrlich B E
Department of Medicine, University of Connecticut, Farmington 06032-1305.
J Gen Physiol. 1994 Nov;104(5):821-56. doi: 10.1085/jgp.104.5.821.
The conduction properties of inositol (1,4,5)-trisphosphate (InsP3)-gated calcium (Ca) channels (InsP3R) from canine cerebellum for divalent cations and the regulation of the channels by intraluminal Ca were studied using channels reconstituted into planar lipid bilayers. Analysis of single-channel recordings performed with different divalent cations present at 55 mM on the trans (intraluminal) side of the membrane revealed that the current amplitude at 0 mV and the single-channel slope conductance fell in the sequence: Ba (2.2 pA, 85 pS) > Sr (2.0 pA, 77 pS) > Ca (1.4 pA, 53 pS) > Mg (1.1 pA, 42 pS). The mean open time of the InsP3R recorded with Ca (2.9 ms) was significantly shorter than with other divalent cations (approximately 5.5 ms). The "anomalous mole fraction effect" was not observed in mixtures of divalent cations (Mg and Ba), suggesting that these channels are single-ion pores. Measurements of InsP3R activity at different intraluminal Ca levels demonstrated that Ca in the submillimolar range did not potentiate channel activity, and that very high levels of intraluminal Ca (> or = 10 mM) decreased channel open probability 5-10-fold. When InsP3R were measured with Ba as a current carrier in the presence of 110 mM cis potassium, a PBa/PK of 6.3 was estimated from the extrapolated value for the reversal potential. When the unitary current through the InsP3R at 0 mV was measured as a function of the permeant ion (Ba) concentration, the half-maximal current occurred at 10 mM trans Ba. The following conclusions are drawn from these data: (a) the conduction properties of InsP3R are similar to the properties of the ryanodine receptor, another intracellular Ca channel, and differ dramatically from the properties of voltage-gated Ca channels of the plasma membrane. (b) The estimated size of the Ca current through the InsP3R under physiological conditions is 0.5 pA, approximately four times less than the Ca current through the ryanodine receptor. (c) The potentiation of InsP3R by intraluminal Ca in the submillimolar range remains controversial. (d) A quantitative model that explains the inhibitory effects of high trans Ca on InsP3R activity was developed and the kinetic parameters of InsP3R gating were determined.
利用重组到平面脂质双分子层中的通道，研究了犬小脑肌醇（1,4,5）-三磷酸（InsP3）门控钙（Ca）通道（InsP3R）对二价阳离子的传导特性以及腔内Ca对通道的调节作用。对在膜的反式（腔内）侧存在55 mM不同二价阳离子时进行的单通道记录分析表明，0 mV时的电流幅度和单通道斜率电导按以下顺序递减：Ba（2.2 pA，85 pS）> Sr（2.0 pA，77 pS）> Ca（1.4 pA，53 pS）> Mg（1.1 pA，42 pS）。用Ca记录的InsP3R的平均开放时间（2.9 ms）明显短于用其他二价阳离子记录的时间（约5.5 ms）。在二价阳离子（Mg和Ba）混合物中未观察到“异常摩尔分数效应”，表明这些通道是单离子孔。在不同腔内Ca水平下对InsP3R活性的测量表明，亚毫摩尔范围内的Ca不会增强通道活性，而非常高的腔内Ca水平（≥10 mM）会使通道开放概率降低5至10倍。当在110 mM顺式钾存在下用Ba作为电流载体测量InsP3R时，根据反转电位的外推值估计PBa/PK为6.3。当测量0 mV时通过InsP3R的单位电流作为渗透离子（Ba）浓度的函数时，半最大电流出现在10 mM反式Ba处。从这些数据得出以下结论：（a）InsP3R的传导特性类似于另一种细胞内Ca通道——雷诺丁受体的特性，与质膜电压门控Ca通道的特性有显著差异。（b）在生理条件下，通过InsP3R的Ca电流估计大小为0.5 pA，约为通过雷诺丁受体的Ca电流的四分之一。（c）亚毫摩尔范围内腔内Ca对InsP3R的增强作用仍存在争议。（d）建立了一个解释高反式Ca对InsP3R活性抑制作用的定量模型，并确定了InsP3R门控的动力学参数。
