Synaptic Glutamate Receptor Trafficking.

描述（由申请人提供）：CNS中的兴奋性突触释放谷氨酸，谷氨酸作用于两种类型的离子型受体：AMPA受体（AMPAR）和NMDA受体（NMDAR）。有证据表明，与NMDAR相反，AMPAR是高度移动的，并且活动可以迅速改变突触处受体的数量。为了开始了解突触AMPAR调控的分子基础，我们将重点放在共济失调和癫痫小鼠的观星者身上。这只小鼠的小脑颗粒细胞缺乏功能性AMPAR，尽管NMDAR是正常的，兴奋性突触释放正常量的谷氨酸。在目前的Conte赠款期间，我们对stargazin（v-2）的作用进行了一系列研究，stargazin是stargazer小鼠中的突变基因。我们已经发现，stargazin是AMPAR的辅助亚基，不仅控制它们向细胞表面和突触的运输，而且还控制它们的生物物理性质。我们已经确定了总共三个额外的蛋白质（v-3，y-4，v-8），在整个中枢神经系统中表达，可以挽救小脑颗粒细胞中的AMPAR缺陷。我们将这些蛋白质称为跨膜AMPAR调节蛋白（TARP）。我们已经成功地删除了小鼠中每种TARPs的基因。这些突变小鼠将成为RO 1资助中许多拟议实验的基础，这是Conte Grant工作的延续。有四个具体目标。(1)确定v-3在小鼠脑中的作用，（2）确定y-4在小鼠脑中的作用，（3）确定TARP之间的功能差异，以及（4）确定TAPRs是否在CNS中具有AMPAR非依赖性作用。TARP在AMPAR运输、突触传递和可塑性中的作用将主要在海马中进行研究。初步研究表明，y-8在海马中的AMPAR运输中起重要作用，但仍存在大量AMPAR传递。我们还将比较各种TARP修改AMPAR失活和突触传递动力学的能力。我们有证据表明，每一个角色的Tarp发挥（即，表面递送、突触靶向和受体门控）对于每个TARP都是不同的。我们已经发现Y-2/Y-3A/-4，三重KO是致命的，并且新生幼仔完全不能移动。我们将确定为什么脊髓没有功能。这些研究可能会揭示TARP在神经系统中的新作用。考虑到受体运输在突触可塑性中的关键作用，以及在学习和记忆的某些方面的暗示，预计这些研究的结果将具有直接的临床影响。实际上，临床上有前景的AMPAkine部分地通过控制类似于TARP的AMPAR门控的动力学来发挥其作用。此外，TARPs修饰AMPakines的药理学性质，因此代表了药物设计的新靶点。

DESCRIPTION (provided by applicant): Excitatory synapses in the CNS release glutamate, which acts on two types of ionotropic receptors: AMPA receptors (AMPARs) and NMDA receptors (NMDARs). Evidence indicates that AMPARs, in contrast to NMDARs, are highly mobile and that activity can rapidly change the number of receptors at the synapse. To begin to understand the molecular basis underlying the regulation of synaptic AMPARs, we have focused on the ataxic and epileptic mouse stargazer. Cerebellar granule cells in this mouse lack functional AMPARs, although NMDARs are normal and excitatory synapses release normal amounts of glutamate. During the current Conte grant we have carried out a series of studies on the role of stargazin (v-2), the mutated gene in the stargazer mouse. We have found that stargazin is an auxiliary subunit of AMPARs, not only controlling their trafficking to the cell surface and to the synapse, but also controlling their biophysical properties. We have identified a total of three additional proteins (v-3, y-4, v-8) that are expressed throughout the CNS and can rescue the AMPAR defect in cerebellar granule cells. We refer to these proteins as transmembrane AMPAR regulatory proteins (TARPs). We have succeeded in deleting the gene for each of the TARPs in mice. These mutant mice will form the basis for many of the proposed experiments in this RO1 grant, which is a continuation of the work carried out on the Conte Grant. There are 4 Specific Aims. (1) Determine the role of v-3 in the mouse brain, (2) determine the role of y-4 in the mouse brain, (3) determine the functional differences among TARPs, and (4) determine whether TAPRs may have AMPAR-independent roles in the CNS. The role of TARPs in AMPAR trafficking, synaptic transmission and plasticity will be studied primarily in the hippocampus. Preliminary studies indicate that y-8 plays an important role in AMPAR trafficking in the hippocampus, but substantial AMPAR transmission remains. We will also compare the ability of various TARPs to modify the deactivation of AMPARs and the kinetics of synaptic transmission. We have evidence that each of the roles TARPs play (i.e., surface delivery, synaptic targeting and receptor gating) all vary for each of the TARPs. We have found that the Y-2/Y-3A/-4, triple KO is lethal and the newborn pups are completely immobile. We will determine why the spinal cord is nonfunctional. These studies may uncover novel roles for TARPs in the nervous system. Given the critical role that receptor trafficking plays in synaptic plasticity and, by implication in certain aspects of learning and memory, it is anticipated that findings from these studies will have direct clinical impact. Indeed, clinically promising AMPAkines exert their effect, in part, by controlling the kinetics of AMPAR gating similar to TARPs. In addition, TARPs modify the pharmacological properties of AMPAkines and, thus, represent a novel target for drug design.