Schnetkamp P P
Department of Medical Biochemistry, University of Calgary, Alberta, Canada.
Biochemistry. 1995 May 30;34(21):7282-7. doi: 10.1021/bi00021a045.
pH-dependent Ca2+ and Na+ transport via Na-Ca+K exchange was measured in isolated and purified bovine retinal rod outer segments (ROS). Ca2+ transport was measured (1) as 45Ca uptake via reverse Na-Ca+K exchange or via Ca-Ca exchange and (2) as a Ca(2+)-induced rise in cytosolic free Ca2+ concentration via reverse Na-Ca+K exchange measured with the fluorescent Ca(2+)-indicating dye fluo-3. Na+ transport was inferred from a Na(+)-induced lowering of cytosolic free [Ca2+] measured with fluo-3. Both Na+ and Ca2+ influx into ROS mediated by the Na-Ca+K exchanger showed a steep (40-100-fold increase in influx rate when the pH was increased from 6.0 to 8.8) pH dependency when measured at low occupancy of the external transport site with Na+ or Ca2+, respectively. High occupancy of the transport site with either Na+ or Mg2+ caused a dramatic decrease in the pH dependence of either Ca2+ or Na+ influx. These observations corroborate the proposed presence of a common Na+ and Ca2+ transport site of the ROS Na-Ca+K exchanger.
通过钠钙钾交换的pH依赖性钙和钠转运在分离纯化的牛视网膜视杆外段（ROS）中进行了测量。钙转运的测量方法如下：（1）通过反向钠钙钾交换或钙钙交换测量45Ca摄取；（2）使用荧光钙指示染料fluo-3通过反向钠钙钾交换测量钙诱导的胞质游离钙浓度升高。钠转运通过fluo-3测量的钠诱导的胞质游离[Ca2+]降低来推断。当分别在外部转运位点钠或钙低占有率的情况下测量时，由钠钙钾交换器介导的钠和钙流入ROS均显示出陡峭的pH依赖性（当pH从6.0增加到8.8时，流入速率增加40至100倍）。钠或镁对转运位点的高占有率导致钙或钠流入的pH依赖性显著降低。这些观察结果证实了ROS钠钙钾交换器中存在共同的钠和钙转运位点的提议。
