Khan K N, Skepper J N, Hockaday A R, Burgess A J, Huang C L
Phyisiogical Laboratory, Cambridge UK.
J Muscle Res Cell Motil. 2000 Jan;21(1):79-90. doi: 10.1023/a:1005618720122.
The effect of loop diuretics at concentrations known to influence cellular water entry coupled to Na-K-Cl co-transport, upon the vacuolation and detubulation following osmotic shock, was investigated in amphibian skeletal muscles. These were exposed to a glycerol-Ringer solution (18 min), an isotonic Ca2+/Mg2+ Ringer solution and cooling. Adding bumetanide (1.0 and 2.0 microM) to these solutions sharply reduced the incidence of detubulation, assessed by abolition or otherwise of action potential after-depolarisations, from 93.9 +/- 4.7% (n = 6) to 5.0 +/- 1.1% (n = 4: mean +/- SEM: 2.0 microM bumetanide). It dramatically reduced the number and fraction of muscle volume occupied by tubular vacuoles, measured using confocal microscopy, from 60.3 +/- 4.3% (n = 10) to 9.0 +/- 1.1% (n = 35). The incidence of large horseradish peroxidase-lined tubular vacuoles, viewed using electronmicroscopy, similarly was reduced with 2 microM bumetanide in the glycerol-Ringer solution. Bumetanide acted through cellular volume adjustments early in the detubulation protocol. Thus, it exerted its maximum effect when added to the glycerol-Ringer, rather than the Ca2+/Mg2+ Ringer solution. Furthermore, whereas fibre diameters measured using scanning electron microscopy returned to normal during glycerol treatment relative to those of control fibres left in isotonic Ringer, addition of 2.0 microM bumetanide in the glycerol Ringer left markedly smaller fibre diameters. Finally equipotent concentrations of the chemically distinct loop diuretics. furosemide and ethacrynic acid similarly influenced detubulation. These findings implicate Na-K-Cl co-transport in the water entry into muscle fibres that would be expected following introduction of extracellular glycerol. This might then enable the subsequent Na-K-ATPase dependent water extrusion that produces the tubular distension (vacuolation) and detachment (detubulation) following glycerol withdrawal, phenomena also observed in muscular dystrophy.
在两栖类骨骼肌中，研究了已知会影响与钠 - 钾 - 氯共转运相关的细胞水进入的浓度下，袢利尿剂对渗透休克后的空泡化和去管化的影响。将这些肌肉暴露于甘油 - 林格溶液（18分钟）、等渗钙/镁林格溶液并进行冷却。向这些溶液中加入布美他尼（1.0和2.0微摩尔），通过动作电位后去极化的消除与否来评估，去管化的发生率从93.9±4.7%（n = 6）急剧降低至5.0±1.1%（n = 4：平均值±标准误：2.0微摩尔布美他尼）。使用共聚焦显微镜测量，它显著减少了管状空泡占据的肌肉体积的数量和比例，从60.3±4.3%（n = 10）降至9.0±1.1%（n = 35）。在甘油 - 林格溶液中，使用电子显微镜观察到的大的辣根过氧化物酶内衬的管状空泡的发生率，同样在2微摩尔布美他尼作用下降低。布美他尼在去管化过程早期通过细胞体积调节起作用。因此，当添加到甘油 - 林格溶液中时，它发挥出最大效果，而不是添加到钙/镁林格溶液中。此外，虽然使用扫描电子显微镜测量的纤维直径在甘油处理期间相对于留在等渗林格溶液中的对照纤维恢复正常，但在甘油林格溶液中添加2.0微摩尔布美他尼会使纤维直径明显更小。最后，化学性质不同的袢利尿剂呋塞米和依他尼酸的等效浓度同样影响去管化。这些发现表明钠 - 钾 - 氯共转运参与了在引入细胞外甘油后预期会发生的水进入肌肉纤维的过程。这可能随后使得依赖钠 - 钾 - ATP酶的水排出能够发生，从而在甘油撤出后产生管状扩张（空泡化）和分离（去管化），这些现象在肌肉营养不良中也有观察到。
