STUDIES OF P2X ATP RECEPTORS

P2X ATP 受体的研究

基本信息

批准号：
6718943
负责人：
RICHARD IRWIN HUME
金额：
$ 29.99万
依托单位：
UNIVERSITY OF MICHIGAN AT ANN ARBOR
依托单位国家：
美国
项目类别：
财政年份：
2001
资助国家：
美国
起止时间：
2001-03-01 至 2005-11-30
项目状态：
已结题

项目摘要

Ligand-gated ion channels are critical to brain function. These proteins mediate rapid signaling from one neuron to the next, by opening ion selective pores in the surface membrane in response to the binding of neurotransmitter, and thus eliciting ion flows that cause electrical excitation or inhibition. When ligand-gated channels spend too much or too little time open, the brain cannot process information correctly. Furthermore, alterations in ligand-gated channel activity can result in overt neurological disease, including epilepsy and neurodegenerative diseases. It is thus important to understand the molecular mechanisms that allow channels to open properly. The focus of the work described here is on a class of channels gated by extracellular ATP, which are referred to as P2X receptors. Genes encoding seven different P2X receptors are expressed in the mammalian brain. Understanding the role of specific P2X receptors in normal brain function and in brain disease has been impeded by the absence of subunit specific ligands. In particular, the P2X2 subunit is widely expressed in the brain, but its role is unclear. In Goal 1 we will use a novel random mutagenesis strategy to gain information about the structure and function of P2X receptors. This approach should generate a large number of new mutant receptors that could not be identified by any other strategy. In particular, these experiments will provide important new information about the ATP binding site, which will be of tremendous use in developing new, more selective agents. In Goal 2, we will explore the mechanism by which ATP is able to open P2X2 receptor channels in wild type and mutant receptors in detail using single channel recording. These single channel recordings will allow us to test distinct models of receptor activation. Understanding how P2X2 channels open and close is potentially of great significance for understanding brain injury. A great deal of evidence suggests that overactivity of NMDA-class glutamate receptors is a major cause of brain injury. Like NMDA receptors, P2X2 receptors form calcium permeable channels that are modulated by low pH and elevated Zn. However, these modulators inhibit current through NMDA receptors, but potentiate current through P2X2 receptors. Many neurons co-express NMDA and P2X2 receptors. Thus the ratio of P2X2 receptors to NMDA receptors may be a key determinant of whether changes in the brain environment following seizures or ischemia result in hyper or hypoexcitability. This has important ramifications for which neurons are susceptible to damage as a result of these conditions, and for strategies to try to prevent this damage.

配体门控离子通道对大脑功能至关重要。这些蛋白质通过打开离子介导从一个神经元到下一个神经元的快速信号传导表面膜上的选择性孔响应结合神经递质，从而引发引起电激发的离子流或抑制。当配体门控通道花费太多或太少的时间时打开后，大脑无法正确处理信息。此外，改动配体门控通道活性可导致明显的神经系统疾病，包括癫痫和神经退行性疾病。因此重要的是了解允许通道正常打开的分子机制。这这里描述的工作重点是一类由细胞外 ATP，称为 P2X 受体。编码七的基因哺乳动物大脑中表达不同的 P2X 受体。了解特定 P2X 受体在正常脑功能和脑疾病中的作用由于缺乏亚基特异性配体而受到阻碍。特别是， P2X2亚基在大脑中广泛表达，但其作用尚不清楚。在目标 1 中，我们将使用一种新颖的随机诱变策略来获取信息关于P2X受体的结构和功能。这种方法应该产生大量新的突变受体，这些受体无法通过任何其他策略。特别是，这些实验将提供重要的新有关 ATP 结合位点的信息，这将在以下方面发挥巨大作用：开发新的、更具选择性的药物。在目标2中，我们将探索ATP能够打开P2X2的机制使用单通道详细分析野生型和突变型受体的受体通道频道录音。这些单通道录音将允许我们测试受体激活的不同模型。了解 P2X2 通道如何打开接近对于理解脑损伤具有潜在的重要意义。大量证据表明 NMDA 类谷氨酸的过度活性受体是脑损伤的主要原因。与 NMDA 受体一样，P2X2 受体形成受低 pH 值和升高 Zn 调节的钙渗透通道。然而，这些调节剂通过 NMDA 受体抑制电流，但增强通过 P2X2 受体的电流。许多神经元共同表达 NMDA 和 P2X2 受体。因此P2X2受体与NMDA受体的比例可能是关键癫痫发作后大脑环境是否发生变化的决定因素缺血导致兴奋性过度或低下。这有重要的影响神经元容易因这些情况而受到损伤，以及尝试防止这种损害的策略。