Spatial organization of RYRs and BK channels underlying the activation of STOCs by Ca2+ sparks in airway myocytes
Spatial organization of RYRs and BK channels underlying the activation of STOCs by Ca2+ sparks in airway myocytes
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DOI：
10.1085/jgp.201110626
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发表时间：
2011-08-01
影响因子：
3.8
通讯作者：
ZhuGe, Ronghua
中科院分区：
文献类型：
--
作者：
Lifshitz, Lawrence M.;Carmichael, Jeffrey D.;ZhuGe, Ronghua
短暂的、局部的Ca2+事件以高效率和高保真度介导Ca2+信号，主要是由于Ca2+渗透离子通道和它们的分子靶标之间的密切接近。然而，在大多数情况下，缺乏这两种分子之间空间关系的直接证据，因此，对局部Ca2+信号传导的机制理解是不完整的。在这项研究中，我们使用一种综合的方法来解决这个问题，在一个原型的局部Ca2+信号系统中，由ryanodine受体（RYRs）的开放和由气道平滑肌中Ca2+激活的K+ (BK)通道的开放引起的Ca2+火花和自发瞬态外向电流（STOCs）组成。在333 Hz下获得的STOC和Ca2+火花的生物物理分析表明，这两个事件在时间上密切相关，大约8个ryr打开以产生Ca2+火花，激活类似于15个BK通道以产生0 mV的STOC。双免疫细胞化学和高空间分辨率的三维反褶积显示，ryr和BK通道形成簇，RYR1和RYR2（而不是RYR3）定位在膜附近。利用ryr和BK通道之间的空间关系、Ca2+火花引起的[Ca2+]时空分布以及BK通道的动力学模型，我们估计RYR1或RYR2簇打开引起的平均Ca2+火花作用于BK通道，这些通道随机分布在ryr的600 nm半径内。通过ryr和BK通道的这种空间组织，我们能够在一系列生理膜电位中模拟具有与stos相同显著特征的BK通道电流。因此，本研究通过明确了解RYRs （Ca2+源）和BK通道（Ca2+靶）之间的空间关系，提供了Ca2+火花激活STOCs的机制理解。
Short-lived, localized Ca2+ events mediate Ca2+ signaling with high efficiency and great fidelity largely as a result of the close proximity between Ca2+-permeable ion channels and their molecular targets. However, in most cases, direct evidence of the spatial relationship between these two types of molecules is lacking, and, thus, mechanistic understanding of local Ca2+ signaling is incomplete. In this study, we use an integrated approach to tackling this issue on a prototypical local Ca2+ signaling system composed of Ca2+ sparks resulting from the opening of ryanodine receptors (RYRs) and spontaneous transient outward currents (STOCs) caused by the opening of Ca2+-activated K+ (BK) channels in airway smooth muscle. Biophysical analyses of STOCs and Ca2+ sparks acquired at 333 Hz demonstrate that these two events are associated closely in time, and approximately eight RYRs open to give rise to a Ca2+ spark, which activates similar to 15 BK channels to generate a STOC at 0 mV. Dual immunocytochemistry and 3-D deconvolution at high spatial resolution reveal that both RYRs and BK channels form clusters and RYR1 and RYR2 (but not RYR3) localize near the membrane. Using the spatial relationship between RYRs and BK channels, the spatial-temporal profile of [Ca2+] resulting from Ca2+ sparks, and the kinetic model of BK channels, we estimate that an average Ca2+ spark caused by the opening of a cluster of RYR1 or RYR2 acts on BK channels from two to three clusters that are randomly distributed within an similar to 600-nm radius of RYRs. With this spatial organization of RYRs and BK channels, we are able to model BK channel currents with the same salient features as those observed in STOCs across a range of physiological membrane potentials. Thus, this study provides a mechanistic understanding of the activation of STOCs by Ca2+ sparks using explicit knowledge of the spatial relationship between RYRs (the Ca2+ source) and BK channels (the Ca2+ target).