Schnetkamp P P
Department of Medical Biochemistry, Faculty of Medicine, University of Calgary, AB, Canada.
Biochem Cell Biol. 1996;74(4):535-9. doi: 10.1139/o96-457.
The process of Ca2+ homeostasis is of prime importance to all cells because of the ubiquitous role of cytoplasmic Ca2+ as an intracellular messenger and the cytotoxicity of sustained elevated cytosolic Ca2+ concentrations. Two classes of plasma membrane proteins are responsible for maintaining cytosolic free Ca2+ in the submicromolar range against a very large electrochemical Ca2+ gradient across the plasma membrane, the ATP-driven Ca2+ pump and Na-Ca exchangers. Two types of Na-Ca exchangers are known, the 3Na:1Ca exchangers found in heart, brain, kidney, and most other tissues and the 4Na:1Ca+ 1K exchanger found in retinal rod and cone photoreceptors. Functional expression of Na-Ca(/K) exchangers is most often measured as 45Ca uptake in Na(+)-loaded cells or as Na-Ca exchange currents with the giant excised patch technique. In this study, two functional assays used to detect expression of the bovine heart Na-Ca exchanger in CHO cells are described. Both assays are based on measurements of cytosolic free Ca2+ with the fluorescent Ca(2+)-indicating dye fluo-3 and should be equally applicable in the study of functional expression of both Na-Ca and Na-Ca/K exchanger clones.
由于细胞质Ca2+作为细胞内信使的普遍作用以及细胞质Ca2+浓度持续升高的细胞毒性，Ca2+稳态过程对所有细胞都至关重要。两类质膜蛋白负责将细胞质游离Ca2+维持在亚微摩尔范围内，以对抗跨质膜的非常大的电化学Ca2+梯度，即ATP驱动的Ca2+泵和钠钙交换体。已知有两种类型的钠钙交换体，在心脏、大脑、肾脏和大多数其他组织中发现的3Na:1Ca交换体，以及在视网膜视杆和视锥光感受器中发现的4Na:1Ca + 1K交换体。钠钙（/钾）交换体的功能表达最常通过测量钠负载细胞中的45Ca摄取或使用巨膜片钳技术测量钠钙交换电流来衡量。在本研究中，描述了用于检测牛心脏钠钙交换体在CHO细胞中表达的两种功能测定方法。这两种测定方法均基于使用荧光Ca(2+)指示染料fluo-3测量细胞质游离Ca2+，并且在研究钠钙和钠钙/钾交换体克隆的功能表达时应同样适用。
