MEMBRANE MECHANISMS OF OLFACTORY TRANSDUCTION

嗅觉传导的膜机制

• 批准号: 2678149
• 负责人: FRANK ZUFALL
• 金额: $ 20.06万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 1994
• 资助国家: 美国
• 起止时间: 1994-04-01 至 2001-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2678149
• 关键词: Urodela; biophysics; chemical association; chemical kinetics; chemical models; chemoreceptors; computer simulation; cyclic AMP; cyclic GMP; dendrites; electrophysiology; mathematical model; membrane channels; molecular cloning; olfactions; perfusion; receptor coupling; receptor sensitivity; respiratory epithelium; second messengers; tissue /cell culture; voltage /patch clamp

Olfactory receptor neurons (ORNs) are ideally suited as a model to study the kinetics of a second messenger system since the second messenger cascade is coupled directly to ion channels gated by either cAMP or cGMP (cN channels). Because these channels occupy a critical role in the odor transduction cascade, information about the underlying biochemical events can be gathered by measuring the electrical activity of cN channels. The kinetics of receptor-effector coupling in G-protein signal transduction systems in general is incompletely understood. Previous studies of second messenger kinetics conducted under equilibrium conditions have given rise to a model in which the output is presumed to be proportional to the accumulating concentration of a second messenger product. However, more recent biochemical studies utilizing rapid kinetic measurements suggest that activation of the olfactory G-protein cascade is transient rather than cumulative. The overall goal of this project is to determine whether a pulsatile model of second messenger signalling can explain the known characteristics of the response of olfactory receptor cells to odor application thus solving a long-standing controversy in olfactory transduction. The cN channels under study will be either a native one from salamander ORNs or a recombinant channel from rat olfactory epithelium expressed in a kidney cell line. The novel strategy in these studies will be to mimic a transient second messenger production by using a fast perfusion system and measuring the response of isolated membrane patches containing the cN channel to rapid application of cAMP/cGMP. These responses obtained under non-steady state conditions will be compared to single-channel kinetics obtained in the constant presence of second messenger. Further, we will compare the kinetics of isolated cN channels to the kinetics of the odor response in single ORNs obtained from odor pulses with well-controlled timing and concentration. Our data will lead to a biophysical state model for gating of cN channels providing estimates for the rates constants of association and dissociation of cyclic nucleotides to their target ion channel and eventually these numbers will be included into a comprehensive model for the olfactory second messenger cascade. As our pilot data show, activation of the cN channel could be a rate limiting step in the second messenger cascade. Therefore, the characteristic time course of the odor response may result, not from the biochemical steps of the second messenger system, but from inherent channel gating characteristics. Recent evidence suggested that cN channels could be widely expressed in the brain. Thus it appears possible that cN channels have important roles in modulation of neuronal excitability in the CNS as well and it is therefore anticipated that the results obtained from olfactory cells will provide fundamental information about kinetics of second messenger signalling systems in general.

嗅觉受体神经元（ORN）是理想的研究模型 自第二使者以来的第二个允许系统的动力学 级联直接耦合到由CAMP或CGMP门控的离子通道 （CN通道）。 因为这些渠道在气味中占据关键作用 转导级联，有关基础生化事件的信息 可以通过测量CN通道的电活动来收集。 这 G蛋白信号转导中受体效应偶联的动力学 通常，系统尚不完全理解。 先前的第二次研究 在平衡条件下进行的信使动力学已经产生 对于一个假定输出与成正比的模型 累积的第二信使产品的浓度。 但是，更多 最近利用快速动力学测量的生化研究表明 嗅觉G蛋白级联反应的激活是短暂的，而不是 累积。 该项目的总体目标是确定是否 第二信使信号的脉动模型可以解释已知的 嗅觉受体细胞对气味的反应的特征 应用，从而解决了嗅觉的长期争议 转导。 研究中的CN渠道要么是Salamander的本地通道 来自大鼠嗅觉上皮的ORNS或重组通道 肾细胞系。 这些研究中的新策略将是模仿 通过使用快速灌注系统和 测量包含CN的分离的膜贴片的响应 快速应用CAMP/CGMP的渠道。 这些回答在 将非态状状态与单渠道动力学进行比较 在不断存在的第二使者的情况下获得。 此外，我们会的 比较孤立的CN通道的动力学与气味的动力学 从具有良好控制的气味脉冲获得的单个ORN中的响应 时间和集中。 我们的数据将导致生物物理状态模型 对于CN通道的门控，提供了估计的估计。 循环核苷酸与其靶离的关联和解离 渠道和最终这些数字将包含在一个全面 嗅觉第二信使级联的模型。 正如我们的飞行员数据所示，CN通道的激活可能是一个速率 限制第二个Messenger级联的步骤。 因此， 气味响应的特征时间过程可能会导致 第二信使系统的生化步骤，但来自固有的渠道 门控特征。 最近的证据表明，CN渠道可以 在大脑中被广泛表达。 因此，CN似乎有可能 通道在调节神经元兴奋性中具有重要作用 CNS也可以预期从 嗅觉细胞将提供有关动力学的基本信息 一般而言，第二信使信号系统。