Control of neuronal ROS generation by membrane potential

描述(改编自申请人提供的摘要)： 越来越明显的是，线粒体在广泛的 一系列急性和慢性神经退行性疾病。最近的这方面的研究 实验室和其他许多人认为线粒体可能是 一种杀死神经元的信号。也许这个信号的最佳候选者是 线粒体产生的活性氧(ROS)。它的作用机制 然而，人们对大脑线粒体产生的ROS知之甚少。此外， 对ROS信号的调节知之甚少。我们建议 探讨脑线粒体和线粒体产生ROS的特点 培养的神经元。在具体目标1中，我们将确定地点(S)在 电子传输链(ETC)是超氧化物的来源，并将 测定ETC抑制离体脑内ROS生成的结果 线粒体。因此，我们将能够根据以下条件对神经退行性疾病建模 先前关于ETC抑制治疗帕金森氏病和 阿尔茨海默氏症。在具体目标2中，我们将研究 调节ROS的产生，并将重点放在改变线粒体的药物上 膜电位和使氧耗与三磷酸腺苷分离的药剂 综合。这些试剂包括钙和解偶联蛋白的底物。 在具体目标3中，我们将使用一系列神经元准备来建立 线粒体在完整细胞中产生ROS的特性 环境。我们将使用标准的神经元培养，通透性神经元和 急性分离的细胞，以及ROS的荧光成像，以建立 线粒体ROS产生的神经元特异性机制。在决赛中 具体目标我们将调查是否逆行运输 体内神经元的线粒体会因氧化损伤而改变。这部小说 这种方法将使我们能够研究细胞线粒体的特性 动态平衡，将提供一个长期的观点，线粒体在 神经退行性变。这些研究将阐明 线粒体和ROS信号在神经元损伤中的作用，并将提供新的 广泛的神经退行性疾病的干预目标。

DESCRIPTION (Adapted from the abstract provided by the applicant): It is becoming increasingly apparent that mitochondria play a critical role in a wide range of acute and chronic neurodegenerative diseases. Recent studies from this laboratory and many others have suggested that mitochondria may be the source of a signal that kills neurons. Perhaps the best candidate for this signal is mitochondrial generation of reactive oxygen species (ROS). The mechanisms of ROS generation by brain mitochondria, however, are poorly understood. Moreover, little is known about the regulation of the ROS signal. We propose to investigate the characteristics of ROS generation by brain mitochondria and cultured neurons. In Specific Aim 1 we will determine the site(s) in the electron transport chain (ETC) that are the source of superoxide, and will determine the consequence of ETC inhibition on ROS generation in isolated brain mitochondria. Thus we will be able to model neurodegenerative disease based on prior reports of ETC inhibition in disorders such as Parkinson's disease and Alzheimer's disease. In Specific Aim 2 we will investigate the mechanisms that regulate ROS generation and will focus on agents that alter the mitochondrial membrane potential and agents that uncouple oxygen consumption from ATP synthesis. These agents include calcium and substrates for uncoupling proteins. In Specific Aim 3 we will use a series of neuron preparations to establish the properties of ROS generation by mitochondria in their intact cellular environment. We will use standard neuronal cultures, permeabilized neurons and acutely dissociated cells, along with fluorescence imaging of ROS, to establish neuron-specific mechanisms of mitochondrial ROS generation. In the final Specific Aim we will investigate whether the retrograde transport of mitochondria by neurons in vivo is altered by oxidative injury. This novel approach will allow us to investigate the properties of cellular mitochondrial homeostasis which will provide a long-term view of the role of mitochondria in neurodegeneration. These studies will illuminate a critical role of mitochondria and ROS signaling in neuronal injury, and will provide novel targets for intervention in a wide range of neurodegenerative disease.