CALCIUM-ACTIVATED POTASSIUM CHANNELS IN SKELETAL MUSCLE

骨骼肌中钙激活的钾通道

• 批准号: 3466917
• 负责人: ANDREW L BLATZ
• 金额: $ 7.23万
• 依托单位: UT SOUTHWESTERN MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 1988
• 资助国家: 美国
• 起止时间: 1988-12-01 至 1993-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3466917
• 关键词: calcium; gene expression; histochemistry /cytochemistry; laboratory rat; membrane permeability; muscle cells; neural information processing; neurons; potassium channel; spinal cord; striated muscles; synapses; tissue /cell culture; voltage /patch clamp

The ultimate goal of the research described in this proposal is to fully characterize the properties of small conductance, apamin- blocked Ca-activated K channels (SK channels) that are probably responsible for the sustained afterhyperpolarization (AHP) in tissue cultured rat skeletal muscle. The kinetics, permeability properties and block of SK channels will be examined and the roles of SK channels and other Ca-activated K channels in muscle excitation will be determined. The modulation of SK channels by innervation will be studied, particularly addressing whether or not functional synapse formation is required for channel regulation. Currents through single SK channels will be recorded using the patch voltage clamp technique. Distributions of open and shut channel events will be fitted with sums of exponentials. The number of exponential components required to fit these distributions suggests a minimum number of the kinetic states that underlie SK channel activity. The apparent number of Ca ions that bind during activity will be determined by measuring the slopes of Hill plots of the percent of time the channel is open at several different intracellular Ca concentrations. Interval distributions predicted by several kinetic models will be compared with the experimentally observed distributions in order to find the models that are the most consistent with the experimental data. Permeability properties will be examined by several methods which will address the selectivity of SK channels as well as examine possible ion-channel interactions such as multiple ion occupancy. The block by apamin, a toxin that has a very high affinity for SK channels, will be characterized at the single channel level to investigate the mechanism of block. The roles of SK channels and large conductance Ca-activated channels (BK channels) will be examined by intracellular potential recording while, at the same time, monitoring single channel currents through a cell-attached membrane patch. With this technique it will be possible to observe which channels are active during the various phases of the action potential, particularly during the AHP. Neural modulation of SK channels will be studied by monitoring the muscle cells for SK channels and synapse formation in the presence of growing spinal neurons and/or media conditioned by neurons. These studies will enhance our knowledge about how ion channels function, how the channel properties affect the functioning of excitable cells, and give insights about how neurons can regulate the expression of ion channels.

本提案中描述的研究的最终目标是 充分表征小电导的特性，apamin- 阻断 Ca 激活的 K 通道（SK 通道），这可能是 造成持续后超极化（AHP） 组织培养的大鼠骨骼肌。 动力学、渗透性 将检查 SK 通道的属性和块以及角色 肌肉中 SK 通道和其他 Ca 激活的 K 通道 激励将被确定。 SK 通道的调制 将研究神经支配，特别是解决是否 功能性突触的形成是通道调节所必需的。 通过单个 SK 通道的电流将使用 贴片电压钳技术。 开闭分布 通道事件将用指数和来拟合。 这 拟合这些所需的指数分量的数量 分布表明了最小数量的动力学状态 SK 渠道活动的基础。 Ca离子的表观数量 活动期间的结合将通过测量斜率来确定 渠道在几个小时开放的时间百分比的山图 不同的细胞内Ca浓度。 区间分布 几个动力学模型的预测将与 通过实验观察分布以找到模型 与实验数据最为一致。 渗透性能将通过多种方法进行检查 将解决 SK 通道的选择性问题并检查 可能的离子通道相互作用，例如多个离子占据。 apamin 的阻断，这是一种与 SK 具有非常高亲和力的毒素 通道，将在单通道级别进行表征 研究块的机制。 SK渠道的作用和 大电导 Ca 激活通道（BK 通道）将 通过细胞内电位记录进行检查，同时 时间，监测通过细胞附着的单通道电流 膜片。 通过这种技术，可以观察到 在行动的各个阶段哪些通道处于活动状态 潜力，特别是在 AHP 期间。 SK 的神经调节 将通过监测肌肉细胞的 SK 来研究通道 在脊髓生长的情况下通道和突触的形成 神经元和/或受神经元调节的介质。 这些研究将 增强我们对离子通道如何发挥作用、 通道特性影响可兴奋细胞的功能，并且 深入了解神经元如何调节离子的表达 渠道。