Collective Ryanodine Receptor Operation at Release Sites

兰尼定受体(RyRs)介导钙诱导的肌浆网钙释放(CICR)。这个 SR有离散的钙释放部位，每个都有一簇RyR。在舒张期，单RYR很少开放， 但当一个人这样做时，它释放的钙可能会点燃局部的RyR间CICR(火花)。异常频繁或 大的火花会引起传播的钙波、室性心动过速和心脏性猝死。 RYR在释放地点相互依赖地运行。不变地和动态地打开集群中的任何RyR (在空间和时间上)改变局部[Ca]‘S(胞浆和胞内)。这些[Ca]变化可能会激活邻近的 雷亚尔。释放点的簇状RyR几何结构和CICR固有的正反馈构成了一个前所未有的 目前存在引发钙波的风险。因此，故障安全释放站点的运行需要保持当地的CICR处于受控状态。为 几十年来，各种单一RyR水平的机制(失活、适应、腔内钙调节)被提出 以解释释放点矛盾的CICR稳定性。这些单RyR过程最终被发现 是不够的。我们的初步研究表明，原因是所需的CICR负面控制产生于 RYR的集体运行，而不是每个RYR相互独立地运作。集体-RyR操作在 发布站点本质上仍然是一个黑匣子，尽管我们从Single-RyR那里知道了很多关于它的组件 结构--功能到火花。 以前的模拟很重要地揭示了“基本”钙释放的细节。但是，这些应用了一些强有力的 简单化。例如，单RyR门控是复杂的，这在过去是没有被捕捉到的。普普通通 泳池隔间模型被使用(现在仍然是)，所有的RYR总是“看到”相同的[Ca]，但这是字面上的 消除了推动集体-RyR的现象(动态纳米级时空[Ca]梯度) 在放行地点进行操作。因此，我们的基本原则是，既然我们有所需的部件，我们必须精心 在没有过去不切实际的假设的情况下，把它们组合起来。为此，我们开发了一种创新的混合动力车 实验/计算方法。我们的初步研究已经揭示了以前未知的集体-RyR CICR控制机制，如有害的自然流失(有助于终止CICR)、RyR招募偏见(这是有效的 以缩小RyRICR事件之间的距离)和长期封闭的阻力(这将CICR的局部传播限制在区域内 站点)。这里检验的假设是：舒张期SR患者的正常故障安全稳定性和致心律失常不稳定性 释放部位由集体-RyR控制机制控制，可以由RyR- 靶向药物。而且，具体目标是1)确定集体-RyR控制机制，以促进 舒张期稳定性正常的人心室肌浆网钙释放部位和2)在人的集体RyR控制 致心律失常的舒张期释放部位和试验RyR靶向药物作为潜在的治疗集体对照 稳定器。这项研究将为理解集合式RyR的运行提供坚实的机制基础 SR钙释放部位等有望改变我们对致心律失常的SR钙释放部位的认识。

Ryanodine receptors (RyRs) mediate Ca-induced Ca release (CICR) from the sarcoplasmic reticulum (SR). The SR has discrete Ca release sites, each with a cluster of RyRs. During diastole, single -RyRs open infrequently, but when one does, the Ca it releases may ignite localized inter-RyR CICR (a spark). Abnormally frequent or large sparks can evoke propagating Ca waves, ventricular tachycardia and sudden cardiac death. RyRs operate inter-dependently at release sites. Opening of any RyR in a cluster invariably and dynamically (spatially & temporally) alters local [Ca]’s (cytosolic & intra-SR). These [Ca] changes may activate neighboring RyRs. The clustered RyR-geometry at release sites and the inherent positive feedback of CICR pose an ever- present risk of evoking a Ca wave. So, fail-safe release site operation requires keeping local CICR in check. For decades, various single RyR-level mechanisms (inactivation, adaptation, luminal Ca regulation) were proposed to explain the paradoxical CICR stability of release sites. These single-RyR processes were ultimately found to be insufficient. Our pilot studies indicate the reason is the required CICR negative control arises from the collective operation of RyRs, not each RyR acting independently of one another. Collective-RyR operation at release sites is still essentially a black box even though we know a lot about its components, from single-RyR structure-function to sparks. Previous simulations importantly revealed details of “elementary” Ca release. But, these applied some strong simplifications. For example, single-RyR gating is complex and this was just not captured in the past. Common pool compartment models were used (and still are), where all RyRs always “see” the same [Ca], but this literally eliminates the very phenomenon (dynamic nano-scale spatiotemporal [Ca] gradients) that drives collective-RyR operation at release sites. Thus, our foundational principle is, as we have the parts needed, we must meticulously assemble them without the unrealistic assumptions of the past. To this end, we developed an innovative hybrid experimental/computational approach. Our pilot studies have already revealed previously unknown collective-RyR CICR control mechanisms like pernicious attrition (which helps terminates CICR), RyR recruitment bias (that works to shrink inter-RyRCICR events) and long-closed resistance(whichlimits the local spread of CICR within arelease site). The hypothesis tested here is: The normal fail-safe stability and arrhythmogenic instability of diastolic SR Ca release sites are governed by collective-RyR control mechanisms that can be therapeutically manipulated by RyR- targeted drugs. And, the specific aims are 1) Identify collective-RyR control mechanisms that promote the diastolic stability normal human ventricular SR Ca release sites and 2) Define collective-RyR control at human arrhythmogenic diastolic release sites and test RyR-targeted drugs as potential therapeutic collective control stabilizers. This study will provide a strong mechanistic foundation for understanding collective -RyR operation at SR Ca release sites and so promises to transform our knowledge of arrhythmogenic SR Ca release sites.