Schofield G G, Mason M J
Department of Physiology, Tulane University Medical School, 1430 Tulane Avenue, New Orleans, Louisiana 70112, USA.
J Membr Biol. 1996 Oct;153(3):217-31. doi: 10.1007/s002329900125.
We have characterized a Ca2+ current activated by depletion of intracellular Ca2+ stores (capacitative Ca2+ entry current) as a first step to investigate the mechanisms underlying communication between the intracellular Ca2+ stores and the plasma membrane Ca2+ permeability. Whole cell currents in response to voltage ramps from -125 to +60 mV from a holding potential of -40 mV were recorded in rat basophilic leukemia cells (RBL-1 cells) in solutions designed to optimize detection of a Ca2+ current. An inwardly rectifying current could be activated upon dialysis of the cell interior with pipette solutions devoid of Ca2+ and containing 20 mm BAPTA, a procedure expected to passively deplete intracellular Ca2+ stores. The current was maximally activated within 2 min, was sensitive to extracellular Ca2+ concentration and was abolished by removal of extracellular Ca2+. The current was markedly reduced in the presence of Ni2+ or La3+. The pathway activated by this protocol was permeant to Ba2+, displaying complex permeability characteristics at negative potentials. A small inward Mn2+ current consistent with a finite permeability of the pathway to Mn2+ was detected. In contrast Ni2+ displayed no detectable current carrying ability. Extracellular Na+ permeated the pathway in the absence of extracellular Ca2+. Under conditions designed to reduce passive depletion of intracellular Ca2+ stores, a Ca2+ current indistinguishable from that described above was activated by addition of ionomycin. This observation is consistent with the activation of the Ca2+ influx pathway occurring as a result of events associated with depletion of intracellular Ca2+ stores. Importantly, application of extracellular Ni2+ in the presence of ionomycin irreversibly inhibited the current. The presence of an inwardly rectifying K+ current in RBL cells could confound studies of the capacitative Ca2+ entry current when recorded using pipette solutions devoid of K+ since this current would be inward over the voltage range used to investigate the capacitative Ca2+ entry current. This study compares an inward rectifying K+ current and the capacitative Ca2+ entry current in RBL cells and highlights some similarities and differences between the two currents. The results demonstrate that caution should be exercised in interpreting recordings made using extracellular solutions containing even modest amounts of K+ when studying the capacitative Ca2+ entry current in RBL cells.
作为研究细胞内钙库与质膜钙通透性之间通讯机制的第一步，我们已对一种由细胞内钙库耗竭激活的钙电流（容量性钙内流电流）进行了特性描述。在旨在优化钙电流检测的溶液中，记录了来自大鼠嗜碱性白血病细胞（RBL-1细胞）的全细胞电流，该电流是在从-40 mV的 holding 电位到-125至+60 mV的电压斜坡刺激下产生的。当用不含钙且含有20 mM BAPTA的移液管溶液透析细胞内部时，可激活一种内向整流电流，该操作预期会被动耗尽细胞内钙库。该电流在2分钟内达到最大激活，对细胞外钙浓度敏感，并且在去除细胞外钙后消失。在存在Ni2+或La3+时，电流明显降低。此方案激活的途径对Ba2+有通透性，在负电位下表现出复杂的通透性特征。检测到一小股内向Mn2+电流，这与该途径对Mn2+的有限通透性一致。相比之下，Ni2+没有显示出可检测到的载流能力。在没有细胞外钙的情况下，细胞外Na+可通过该途径。在旨在减少细胞内钙库被动耗竭的条件下，加入离子霉素可激活一种与上述电流无法区分的钙电流。这一观察结果与由于细胞内钙库耗竭相关事件导致的钙内流途径激活一致。重要的是，在离子霉素存在的情况下应用细胞外Ni2+会不可逆地抑制该电流。当使用不含K+的移液管溶液记录时，RBL细胞中内向整流K+电流的存在可能会混淆对容量性钙内流电流的研究，因为在用于研究容量性钙内流电流的电压范围内，该电流会向内流动。本研究比较了RBL细胞中的内向整流K+电流和容量性钙内流电流，并突出了这两种电流之间的一些异同。结果表明，在研究RBL细胞中的容量性钙内流电流时，在解释使用含有适量K+的细胞外溶液进行的记录时应谨慎。
