Ryanodine as a probe for the functional state of the skeletal muscle sarcoplasmic reticulum calcium release channel.
Ryanodine as a probe for the functional state of the skeletal muscle sarcoplasmic reticulum calcium release channel.
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发表时间：
1990-05
影响因子：
3.6
通讯作者：
A. Chu;M. Díaz-Muñoz;M. J. Hawkes;K. Brush;S. Hamilton
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文献类型：
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作者：
A. Chu;M. Díaz-Muñoz;M. J. Hawkes;K. Brush;S. Hamilton
本文通过测定[3 H]ryanodine结合动力学、~（45）Ca ~（2+）通量和单通道活性来研究家兔快收缩骨骼肌钙释放通道的调节。调节剂的Ca 2+释放通道（确认在这里与流量和单通道数据）的影响进行了检查[3 H]ryanodine结合到终端脑池囊泡的影响。我们发现，释放通道的激活剂，如腺嘌呤核苷酸（1 mM）和咖啡因（1 mM），提高了[3 H]ryanodine的关联率，而抑制剂，如Mg 2+（1 mM）和钌红（100 nM），降低了关联率。高浓度的兰尼定或钌红，关闭通道，减缓[3 H]兰尼定的解离，这表明在这些浓度下，兰尼定和钌红的抑制作用是由于在一个不同于高亲和力位点的位点结合而发生的，但与高亲和力位点相互作用。我们的数据与一个模型是一致的，在该模型中，高亲和力ryanodine结合位点是在通道的构象敏感区域内，使得打开通道的条件（ATP，咖啡因等）。提高[3 H]兰尼碱到达其结合位点的速率，以及关闭通道的其他条件（兰尼碱和钌红结合到低亲和力位点）减缓[3 H]兰尼碱从高亲和力位点解离。一些条件下，抑制通道活性（高浓度的Mg 2+和Ca 2+）缓慢协会，但不影响解离结合[3 H]ryanodine，这表明一个完全不同的状态下，这是在高浓度的ryanodine或钌红的存在下，无活性的通道。总之，快收缩骨骼肌钙释放通道的功能状态可以通过[3 H]兰尼碱结合动力学的变化来表征。不同的调节剂（激活剂/抑制剂）影响兰尼定结合的不同方面（结合/解离）。
In this paper, we study the modulation of the rabbit fast twitch skeletal muscle calcium release channel by assaying the kinetics of [3H]ryanodine binding, 45Ca2+ flux, and single-channel activity. The effects of modulators of the Ca2+ release channel (confirmed here with both flux and single-channel data) were examined for effects on [3H]ryanodine binding to terminal cisternae vesicles. We find that activators of the release channel, such as adenine nucleotides (1 mM) and caffeine (1 mM), enhance the rate of association of [3H]ryanodine, whereas inhibitors, such as Mg2+ (1 mM) and ruthenium red (100 nM), decrease the rate of association. High concentrations of either ryanodine or ruthenium red, which close the channel, slow the dissociation of [3H]ryanodine, suggesting that at these concentrations the inhibitory effects of both ryanodine and ruthenium red occur as the result of binding at a site distinct from but interacting cooperatively with the high affinity site. Our data are consistent with a model in which the high affinity ryanodine binding site is within a conformationally sensitive area of the channel, such that conditions that open the channel (ATP, caffeine, etc.) enhance the rate at which [3H]ryanodine reaches its binding site and other conditions that close the channel (the binding of ryanodine and ruthenium red to a low affinity site) slow the dissociation of [3H]ryanodine from the high affinity site. Some conditions that inhibit channel activity (high concentrations of Mg2+ and Ca2+) slow association but do not affect dissociation of bound [3H]ryanodine, suggesting a completely different state of the channel from that which is inactive in the presence of high concentrations of ryanodine or ruthenium red. In summary, the functional state of the fast twitch skeletal muscle calcium release channel can be characterized by the changes in the kinetics of [3H]ryanodine binding. Different modulators (activators/inhibitors) affect different aspects of ryanodine binding (association/dissociation).