Rose B, Loewenstein W R
J Membr Biol. 1976 Aug 27;28(1):87-119. doi: 10.1007/BF01869692.
A technique is devised to determine the spatial distribution of the free ionized cytoplasmic calcium concentration ([Ca2+]i) inside a cell: Chironomus salivary gland cells are loaded with aequorin, and hte Ca2+-dependent light emission of the aequorin is scanned with an image-intensifier/television system. With this technique, the [Ca2+]i is determined simultaneously with junctional electrical coupling when Ca2+ is microinjected into the cells, or when the cells are exposed to metabolic inhibitors, Ca-transporting ionophores, or Ca-free medium. Ca microinjections elevating the [Ca2+]i in the junctional locale produce depression of junctional membrane conductance. When the [Ca2+]i elevation is confined to the vicinity of one cell junction, the conductance of that junction alone is depressed; other junctions of the same cell are not affected. The depression sets in as the [Ca2+]i rises in the junctional locale, and reverses after the [Ca2+]i falls to baseline. When the [Ca2+]i elevation is diffuse throughout the cell, the conductances of all junctions of the cell are depressed. The Ca injections produce no detectable [Ca2+]i elevations in cells adjacent to the injected one; the Ca-induced change in junctional membrane permeability seems fast enough to block appreciable transjunctional flow of Ca2+. Control injections of Cl- or K+ do not affect junctional conductance. The Ca injections that elevate [Ca2+]i sufficiently to depress junctional conductance also produce under the usual conditions an increase in nonjunctional membrane conductance and, hence, depolarization. But injections that elevate [Ca2+]i at the junction while largely avoiding nonjunctional membrane cause depression of junctional conductance with little or no depolarization. Moreover, elevations of [Ca2+]i in cells clamped near resting potential produce the depression, too. On the other hand, complete depolarization in K medium does not produce the depression, unless accompanied by [Ca2+]i elevation. Thus, the depolarization is neither necessary nor sufficient for depression of junctional conductance. Treatment with cyanide, dinitrophenol and ionophores X537A or A23187 produces diffuse elevation of [Ca2+]i associated with depression of junctional conductance. Prolonged exposure to Ca-free medium leads to fluctuation in [Ca2+]i where rise and fall of [Ca2+]i correlate respectively with fall and rise in junctional conductance.
设计了一种技术来确定细胞内游离离子化细胞质钙浓度（[Ca2+]i）的空间分布：将海萤发光蛋白加载到摇蚊唾液腺细胞中，并用图像增强器/电视系统扫描海萤发光蛋白的Ca2+依赖性发光。利用这种技术，当将Ca2+微注射到细胞中，或者当细胞暴露于代谢抑制剂、Ca转运离子载体或无Ca培养基时，可以同时测定[Ca2+]i和连接性电偶联。微注射Ca会升高连接部位的[Ca2+]i，从而导致连接膜电导降低。当[Ca2+]i升高局限于一个细胞连接附近时，仅该连接的电导会降低；同一细胞的其他连接不受影响。随着连接部位[Ca2+]i升高，电导降低开始出现，并在[Ca2+]i降至基线后逆转。当[Ca2+]i升高在整个细胞中扩散时，该细胞所有连接的电导都会降低。Ca注射在相邻注射细胞中未检测到[Ca2+]i升高；Ca诱导的连接膜通透性变化似乎足够快，足以阻止Ca2+的明显跨连接流动。对照注射Cl-或K+不影响连接电导。足以降低连接电导的Ca注射在通常情况下也会导致非连接膜电导增加，从而导致去极化。但是在很大程度上避免非连接膜的情况下，在连接部位升高[Ca2+]i的注射会导致连接电导降低，而去极化很小或没有。此外，在接近静息电位钳制的细胞中[Ca2+]i升高也会产生这种降低。另一方面，在K培养基中完全去极化不会产生这种降低，除非伴有[Ca2+]i升高。因此，去极化对于连接电导降低既不是必需的也不是充分的。用氰化物、二硝基苯酚和离子载体X537A或A23187处理会导致[Ca2+]i弥漫性升高，并伴有连接电导降低。长时间暴露于无Ca培养基会导致[Ca2+]i波动，其中[Ca2+]i的升高和降低分别与连接电导的降低和升高相关。
