Identification of ligands that bind to the orphan delta2 receptor

描述（由申请人提供）：谷氨酸受体家族由18个基因组成，这些基因可以结合形成数十种不同的异聚体受体复合物。这类受体介导哺乳动物大脑中大多数兴奋性突触传递，并在学习和记忆的细胞模型以及广泛的神经病理中发挥核心作用。在这些基因产物中，两个孤儿受体亚基（delta1和delta2）在序列同源性的基础上被指定为谷氨酸受体家族，尽管没有发现激活配体。delta2亚基在小脑突触中表达，多种电生理研究表明，delta2在小脑发育和突触可塑性中起重要作用。此外，缺乏delta2亚基或表达该受体的组成型活性突变形式（delta2-lurcher）的小鼠表现出不同的运动障碍表型。因此，存在一个悖论，即delta - 2谷氨酸受体亚基被认为在大脑功能中发挥重要作用，但我们对受体的了解却存在真空。我们的合作者最近使用晶体学来确定一个小氨基酸（d -丝氨酸）可以结合到delta2配体结合域，形成一组与谷氨酸结合在其他谷氨酸受体亚基上一致的原子接触。有趣的是，虽然d-丝氨酸与delta2亚基结合，但它在野生型受体中不会引起任何当前反应。然而，在delta2亚基内自然发生的点突变（lurcher突变）会产生构成活性的同质通道，将D-丝氨酸应用于这些通道会抑制流过这些构成活性通道的强直电流。这些数据表明，d -丝氨酸的结合可以导致门控的改变，并提出了delta2亚基与异聚体受体组合的结合可能赋予谷氨酸受体d -丝氨酸的调节位点的可能性。这些新数据为探索delta2受体的结构、功能和药理学提供了前所未有的机会。我们建议在两方面的实验中利用这一最新发现：目标1：鉴定新的配体，有效地激活或抑制δ 2受体。目的2：评估delta2与其他谷氨酸受体亚基的共表达是否增加了d -丝氨酸对受体的敏感性。这些研究的完成将首次对该类受体进行全面的药理学评价，并为研究其在大脑中的功能和调控提供新的工具。公共卫生相关性：我们最近的研究发现，d -丝氨酸可以结合孤儿谷氨酸受体delta2，这为我们提供了一个机会，显著推进我们对该受体功能的理解，尽管在体内的研究结果表明，该受体在小脑发育和突触可塑性中起着重要作用，但该受体的功能已经谜了十多年。这些研究的目的是确定新的和选择性的药理学药物（激活剂和拮抗剂），这可能是了解组织中delta2功能的有用研究工具。此外，我们还将测试delta2亚基是否会改变d -丝氨酸对异多聚谷氨酸受体的作用。这些研究的完成将促进我们对delta2功能的理解，这对小脑中的神经元信号传导、突触可塑性和神经元发育具有重要意义。这些研究也可能为开发针对delta2受体的新治疗药物提供见解。

DESCRIPTION (provided by applicant): The glutamate receptor family is comprised of 18 genes that can combine to form dozens of different heteromeric receptor complexes. This receptor class mediates most excitatory synaptic transmission in the mammalian brain, and plays a central role in cellular models of learning and memory as well as a wide range of neuropathologies. Of these gene products, two orphan receptor subunits (delta1 and delta2) have been designated to be in the glutamate receptor family on the basis of sequence homology even though no activating ligand has been identified. The delta2 subunit is expressed at cerebellar synapses, and a variety of electrophysiological studies suggest that delta2 plays an important role in cerebellar development and synaptic plasticity. In addition, mice that either lack the delta2 subunit or express a constitutively active mutant form of this receptor (delta2-lurcher), show distinct phenotypes with motor disturbances. Thus, a paradox exists whereby the delta2 glutamate receptor subunit is thought to play an important role in brain function yet there is a vacuum regarding what we know about the receptor. Our collaborators have recently used crystallography to determine that a small amino acid (D-serine) can bind to the delta2 ligand binding domain, making a set of atomic contacts that are consistent with glutamate binding at other glutamate receptor subunits. Interestingly, although D-serine binds to the delta2 subunit, it does not induce any current response in wild- type receptors. However, a naturally occurring point mutation within the delta2 subunit (the lurcher mutation) produces constitutively active homomeric channels, and application of D- serine to these channels inhibits the tonic current flowing through these constitutively active channels. These data suggest that binding of D-serine can lead to changes in gating, and raises the possibility that incorporation of the delta2 subunit into heteromeric receptor assemblies may endow glutamate receptors with a regulatory site for D-serine. These new data provide an unprecedented opportunity to explore delta2 receptor structure, function, and pharmacology. We propose to exploit this recent finding in 2 lines of experimentation: Aim 1: Identify new ligands that potently activate or inhibit delta2 receptors. Aim 2: Evaluate whether co-expression of delta2 with other glutamate receptor subunits adds D-serine sensitivity to the receptors. Completion of these studies will provide the first comprehensive pharmacological evaluation of this receptor class, as well as new tools with which to study its function and regulation in brain. PUBLIC HEALTH RELEVANCE: Our recent findings that D-serine can bind to the orphan glutamate receptor delta2 has provided an opportunity to significantly advance our understanding of the function of this receptor, which has been enigmatic for over a decade despite in vivo findings that suggest an important role in cerebellar development and synaptic plasticity. The goal of these studies is to identify novel and selective pharmacological agents (activators and antagonists) that could be useful research tools for understanding delta2 function in tissue. In addition, we will also test whether delta2 subunit alters the effects of D-serine on hetero-mulitmeric glutamate receptors. Completion of these studies will advance our understanding of delta2 function, which holds implications for neuronal signaling, synaptic plasticity, and neuronal development in the cerebellum. These studies may also provide insight into new therapeutic agents that target the delta2 receptor.