Skeletal Muscle Ryanodine Receptor Permeation and Self Counter-Ion Flow

骨骼肌 Ryanodine 受体渗透和自反离子流

• 批准号: 7488500
• 负责人: Michael Fill
• 金额: $ 31.18万
• 依托单位: RUSH UNIVERSITY MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-09-01 至 2011-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7488500
• 关键词: Address; Affect; Affinity; Amino Acid Motifs; Amino Acids; Amitriptyline; Antibodies; Appendix; Arts; Attenuated; Binding; Biological Assay; Biological Models; Budgets; Caffeine; Caliber; Calmodulin; Calsequestrin; Cations; Cells; Central Core Myopathy; Characteristics; Charge; Condition; Coupled; Data; Defect; Detection; Dimensions; Disadvantaged; Disease; Disputes; Dose; Electric Capacitance; Electrodes; Event; Experimental Designs; Figs - dietary; Flavoring; Free Energy; Frequencies; Hand; Helix (Snails); Image; Immunoblotting; Ion Channel; Ions; Knowledge; Left; Life; Light; Lipid Bilayers; Localized; Location; Measurement; Measures; Mediating; Mediator of activation protein; Membrane; Membrane Potentials; Methods; Microsomes; Modeling; Molecular; Mus; Muscle; Mutation; Nature; Noise; Operative Surgical Procedures; Oral cavity; Outcome; Pathway interactions; Pharmaceutical Preparations; Phase; Physiological; Pilot Projects; Play; Positioning Attribute; Postdoctoral Fellow; Potassium Channel; Preparation; Principal Investigator; Probability; Process; Property; Proteins; Protocols documentation; Publications; Published Comment; Publishing; Rate; Rattus; Regulation; Relative (related person); Research; Research Personnel; Research Support; Resolution; Risk; Role; RyR1; RyR2; Ryanodine; Ryanodine Receptor Calcium Release Channel; Sarcoplasmic Reticulum; Score; Series; Side; Signal Transduction; Site; Skeletal Muscle; Skeletal system; Structure; Study Section; Suggestion; Tacrolimus Binding Protein 1A; Techniques; Teflon; Testing; Therapeutic Intervention; Thinking; Time; Trees; Triflupromazine; Vesicle; Work; attenuation; base; cell type; computerized data processing; concept; driving force; ear helix; experience; forest; inositol-1,4,5-triphosphate receptor; mouse model; mutant; novel; programs; protein expression; reconstitution; research study; response; size; theories; uptake; voltage; voltage clamp

DESCRIPTION (provided by applicant): Depolarization of skeletal muscle initiates Ca release through type-1 ryanodine receptor (RyR1) Ca release channels in the sarcoplasmic reticulum (SR). Concerted opening of multiple RyR1 channels at discrete SR Ca release sites generates small localized release events called Ca sparks, the elemental unit of SR Ca release in cells. Spark recruitment/summation is what generates the global Ca release transient that drives muscle contractility. Many studies have focused on the onset and termination of the Ca release transient. Fewer have focused on the RyR conduction/selectivity and none (to our knowledge) have studied what physiological ramifications the RyR permeation characteristics of RyR1 have on cellular Ca signaling phenomena like the Ca release transient. The RyR pore is thought to have a structure analogous to that of the bacterial K channel (KcsA). The RyR has a lumenal loop which contains a predicted pore helix and an amino acid motif (GGGIG) identified as a selectivity filter. Unlike the KcsA pore, the RyR pore has high conductance and is poorly selective. Published works as well as our own preliminary data have revealed some key molecular determinants that define the characteristic high conductance, poor selectivity of the RyR1 pore. This includes some naturally occurring mutants associated with central core disease (CCD). This proposal combines this information to define the mechanisms that govern RyR1 permeation under physiological conditions and applies this knowledge to define some key physiological ramifications of the single channel RyR1 permeation process. The following hypothesis is tested. Hypothesis: In cells, the poor selectivity of the RyR1 channel allows multiple ions (Ca, Mg & K) to compete for occupancy of the open pore. While this competition attenuates the net SR Ca efflux through the open pore, it also allows the pore to mediate its own counter ion flux during the Ca release process. This self counter-ion flow effectively clamps local SR membrane potential far from the Ca Nernst potential (ECa) making the trans-SR Ca driving force primarily dependent on the trans-SR Ca concentration gradient. This is physiologically important because it allows the RyR channel to sustain Ca release over an extended period of time (>5 ms or the rise time of Ca transient). The specific aims are: (1) Define how known molecular determinants of RyR1 permeation combine to generate/influence the conductance and selectivity of the open RyR1 pore. (2) Define functional consequences of the relatively poor selectivity of the RyR1 pore; namely attenuation of net SR Ca efflux through the open pore and the capacity of the open pore to carry its own counter ion flux during SR Ca efflux.

描述（由申请人提供）：骨骼肌去极化通过肌浆网（SR）中的1型ryanodine受体（RyR1） Ca释放通道启动Ca释放。在离散的SR Ca释放位点，多个RyR1通道协同打开，产生小的局部释放事件，称为Ca火花，这是细胞中SR Ca释放的基本单位。火花招募/汇总是产生驱动肌肉收缩的全局Ca释放瞬态的原因。许多研究都集中在钙释放瞬态的开始和终止上。很少有人关注RyR的传导/选择性，也没有人（据我们所知）研究RyR1的RyR渗透特性对细胞Ca信号传导现象（如Ca释放瞬态）的生理影响。RyR孔被认为具有类似于细菌K通道（KcsA）的结构。RyR有一个管腔环，其中包含一个预测的孔螺旋和一个氨基酸基序（GGGIG），被确定为选择性过滤器。与KcsA孔不同，RyR孔具有高电导率和低选择性。已发表的研究成果和我们自己的初步数据揭示了一些关键的分子决定因素，这些决定因素决定了RyR1孔的高电导性和低选择性。这包括与中央核心病（CCD）相关的一些自然发生的突变。本提案结合这些信息来定义生理条件下支配RyR1渗透的机制，并应用这些知识来定义单通道RyR1渗透过程的一些关键生理后果。下面的假设得到检验。假设：在细胞中，RyR1通道的低选择性允许多个离子（Ca， Mg和K）竞争开放孔的占用。虽然这种竞争减弱了通过开放孔隙的净SR Ca外排，但它也允许孔隙在Ca释放过程中调解其自身的反离子通量。这种自反离子流有效地限制了远离Ca能势（ECa）的局部SR膜电位，使得反SR Ca驱动力主要依赖于反SR Ca浓度梯度。这在生理上是重要的，因为它允许RyR通道在一段较长的时间内（大约5毫秒或Ca瞬态上升时间）维持Ca释放。具体目标是：(1)定义已知的RyR1渗透的分子决定因素如何结合起来产生/影响打开的RyR1孔的电导和选择性。(2)定义RyR1孔相对较差选择性的功能后果；即通过开放孔的净SR Ca射流的衰减和开放孔在SR Ca射流过程中携带自身反离子通量的能力。