STUDIES OF P2X ATP RECEPTORS

配体门控离子通道对大脑功能至关重要。这些 蛋白质介导从一个神经元到下一个神经元的快速信号，通过打开离子通道， 选择性孔在表面膜响应的结合 神经递质，从而引发离子流，导致电兴奋 或抑制。当配体门控通道花费太多或太少时间时 大脑无法正确处理信息。此外，修改 在配体门控通道活性中， 包括癫痫和神经退行性疾病。因此， 了解允许通道正确打开的分子机制。的 这里描述的工作的重点是一类通道， 细胞外ATP，被称为P2X受体。基因编码7 不同的P2X受体在哺乳动物脑中表达。了解 特异性P2X受体在正常脑功能和脑疾病中的作用 由于缺乏亚单位特异性配体而受到阻碍。特别是 P2X2亚基在脑中广泛表达，但其作用尚不清楚。 在目标1中，我们将使用一种新的随机诱变策略来获得信息 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 任何其他战略。特别是，这些实验将提供重要的新的 关于ATP结合位点的信息，这将是巨大的使用， 开发新的更具选择性的药剂 在目标2中，我们将探索ATP能够打开P2X2的机制。 受体通道在野生型和突变受体的详细使用单一的 频道录制这些单通道记录可以让我们测试 不同的受体激活模型。了解P2X2通道如何打开 而close对于理解脑损伤具有潜在的重要意义。 大量证据表明，NMDA类谷氨酸的过度活跃 受体是脑损伤的主要原因。与NMDA受体一样，P2X2受体 形成由低pH和升高的Zn调节的钙渗透通道。 然而，这些调节剂抑制通过NMDA受体的电流，但 增强通过P2X2受体的电流。许多神经元共表达NMDA， P2X2受体。因此，P2X2受体与NMDA受体的比例可能是一个关键， 癫痫发作后大脑环境的变化或 局部缺血导致兴奋性过高或过低。这有重要的影响 因为这些条件导致神经元易受损伤， 以及防止这种损害的策略。

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.