Balnave C D, Allen D G
Department of Physiology, University of Sydney, New South Wales, Australia.
Am J Physiol. 1998 Apr;274(4):C940-6. doi: 10.1152/ajpcell.1998.274.4.C940.
The myoplasmic free Ca2+ concentration ([Ca2+]i) was measured in intact single fibers from mouse skeletal muscle with the fluorescent Ca2+ indicator indo 1. Some fibers were perfused in a solution in which the concentration of Na+ was reduced from 145.4 to 0.4 mM (low-Na+ solution) in an attempt to activate reverse-mode Na+/Ca2+ exchange (Ca2+ entry in exchange for Na+ leaving the cell). Under normal resting conditions, application of low-Na+ solution only increased [Ca2+]i by 5.8 +/- 1.8 nM from a mean resting [Ca2+]i of 42 nM. In other fibers, [Ca2+]i was elevated by stimulating sarcoplasmic reticulum (SR) Ca2+ release with caffeine (10 mM) and by inhibiting SR Ca2+ uptake with 2,5-di(tert-butyl)-1,4-benzohydroquinone (TBQ; 0.5 microM) in an attempt to activate forward-mode Na+/Ca2+ exchange (Ca2+ removal from the cell in exchange for Na+ influx). These two agents caused a large increase in [Ca2+]i, which then declined to a plateau level approximately twice the baseline [Ca2+]i over 20 min. If the cell was allowed to recover between exposures to caffeine and TBQ in a solution in which Ca2+ had been removed, the increase in [Ca2+]i during the second exposure was very low, suggesting that Ca2+ had left the cell during the initial exposure. Application of caffeine and TBQ to a preparation in low-Na+ solution produced a large, sustained increase in [Ca2+]i of approximately 1 microM. However, when cells were exposed to caffeine and TBQ in a low-Na+ solution in which Ca2+ had been removed, a sustained increase in [Ca2+]i was not observed, although [Ca2+]i remained higher and declined slower than in normal Na+ solution. This suggests that forward-mode Na+/Ca2+ exchange contributed to the fall of [Ca2+]i in normal Na+ solution, but when extracellular Na+ was low, a prolonged elevation of [Ca2+]i could activate reverse-mode Na+/Ca2+ exchange. The results provide evidence that skeletal muscle fibers possess a Na+/Ca2+ exchange mechanism that becomes active in its forward mode when [Ca2+]i is increased to levels similar to that obtained during contraction.
使用荧光钙指示剂indo 1测量了来自小鼠骨骼肌的完整单纤维中的肌浆游离钙浓度（[Ca2+]i）。一些纤维在一种溶液中灌注，其中Na+浓度从145.4 mM降至0.4 mM（低钠溶液），试图激活反向模式的Na+/Ca2+交换（Ca2+进入细胞以交换Na+离开细胞）。在正常静息条件下，应用低钠溶液仅使[Ca2+]i从平均静息[Ca2+]i 42 nM增加了5.8±1.8 nM。在其他纤维中，通过用咖啡因（10 mM）刺激肌浆网（SR）Ca2+释放和用2,5-二（叔丁基）-1,4-苯二酚（TBQ；0.5 microM）抑制SR Ca2+摄取来升高[Ca2+]i，试图激活正向模式的Na+/Ca2+交换（Ca2+从细胞中移除以交换Na+内流）。这两种试剂导致[Ca2+]i大幅增加，然后在20分钟内降至约为基线[Ca2+]i两倍的平台水平。如果在去除Ca2+的溶液中，在暴露于咖啡因和TBQ之间让细胞恢复，第二次暴露期间[Ca2+]i的增加非常低，这表明在初始暴露期间Ca2+已离开细胞。将咖啡因和TBQ应用于低钠溶液中的制剂会使[Ca2+]i大幅持续增加约1 microM。然而，当细胞在去除Ca2+的低钠溶液中暴露于咖啡因和TBQ时，未观察到[Ca2+]i的持续增加，尽管[Ca2+]i仍高于正常钠溶液且下降比正常钠溶液慢。这表明正向模式的Na+/Ca2+交换有助于正常钠溶液中[Ca2+]i的下降，但当细胞外Na+较低时，[Ca2+]i的长时间升高可激活反向模式的Na+/Ca2+交换。结果提供了证据，表明骨骼肌纤维具有Na+/Ca2+交换机制，当[Ca2+]i增加到与收缩期间获得的水平相似时，该机制以正向模式变得活跃。
