Mechanisms controlling AMPA receptor subunit composition

描述（由申请人提供）：谷氨酸是中枢神经系统中的主要兴奋性神经递质。离子型谷氨酸受体的激活有助于突触可塑性、神经元发育、神经病理生理损伤、神经退行性疾病、药物滥用和精神障碍。在谷氨酸受体中，AMPA受体（AMPAR）主要负责快速兴奋性神经传递。AMPAR倾向于优先组装在由GluA 1和GluA 2亚基组成的异聚体复合物中。GluA 2亚基以主导方式控制几种受体性质，包括Ca 2+渗透性、单通道电导、受体动力学和对细胞内多胺的敏感性。因此，亚基组成的改变对突触传递的质量和强度具有显著影响。虽然大多数AMPAR被认为是GluA 1和GluA 2的异聚体，但越来越多的证据表明，突触AMPAR的组成能够向GluA 1同聚体转变。这种转变可以发生在生理和病理生理形式的突触可塑性，包括恐惧，中风，疼痛和药物滥用。因此，了解导致AMPAR组成发生这些变化的机制至关重要。通过将蛋白激酶C（PKC）靶向至GluA 2，与C-激酶1（PICK 1）相互作用的蛋白质被认为稳定GluR 2的PKC磷酸化，其用于发出含GluA 2的AMPAR的内吞作用的信号。因此，PICK 1和PKC被认为是AMPAR组成改变的核心。最近，我们证明了A-激酶锚定蛋白79 kDa（AKAP 79;啮齿动物中的AKAP 150）将PKC靶向GluA 1 AMPAR，以增强这些受体的磷酸化和活性。基于初步的数据，我们提出AKAP 79锚定的PKC调节AMPAR亚基的组成，有利于同型GluA 1受体。此外，我们建议，这两个PKC附属支架相互结合，以最终确定AMPAR亚基组成。为了验证这一假设，我们将使用生物化学，电生理，分子方法，和一种新的成像方法来了解：1）AKAP 79是否有利于同源GluA 1受体通过GluA 1调节位点使用一个简化的异源系统。2)GluA 2上的PICK 1和PKC位点是否有助于AMPAR组成中AKAP 79依赖性的变化，反之，GluA 1调节位点是否有助于AMPAR组成中PICK依赖性的改变。3)AKAP 150锚定的PKC是否是在与突触可塑性相关的许多条件下在神经元中产生同源GluA 1反应的核心。总的来说，这些研究将揭示PKC附属支架对确定AMPAR亚基组成的贡献，并描绘GluA 1和GluA 2 AMPAR亚基上调控位点的相对重要性，这是健康和疾病的关键因素。 公共卫生相关性：快速兴奋性传递主要由谷氨酸受体的AMPA亚型介导。这些受体组成的变化对学习和记忆以及中风、疼痛、恐惧、药物滥用和神经退行性疾病至关重要。这些研究将研究AMPA受体亚基变化的机制，并有助于发现这些疾病的新靶点和治疗策略。

DESCRIPTION (provided by applicant): Glutamate is the major excitatory neurotransmitter in the central nervous system. Activation of ionotropic glutamate receptors contributes to synaptic plasticity, neuronal development, neuropathophysiological insults, neurodegenerative diseases, drug abuse, and psychiatric disorders. Among the glutamate receptors, AMPA receptors (AMPARs) are predominantly responsible for fast excitatory neurotransmission. AMPARs tend to be preferentially assembled in heteromeric complexes composed of GluA1 and GluA2 subunits. GluA2 subunits act in a dominant fashion to control several receptor properties including Ca2+ permeability, single channel conductance, receptor kinetics, and sensitivity to intracellular polyamines. Thus, alterations in subunit composition have a dramatic impact on the quality and strength of synaptic transmission. Although the majority of AMPARs are thought to be heteromers of GluA1 and GluA2, accumulating evidence has highlighted the ability for synaptic AMPAR composition to shift in favor of GluA1 homomers. This shift can occur in response to both physiological and pathophysiological forms of synaptic plasticity including fear, stroke, pain, and drug abuse. Thus, it is critical to understand mechanisms that contribute to these changes in AMPAR composition. By targeting protein kinase C (PKC) to GluA2, protein interacting with C-kinase 1 (PICK1) is thought to stabilize PKC phosphorylation of GluR2 which serves to signal endocytosis of GluA2-containing AMPARs. Thus, PICK1 and PKC are thought to be central for alterations in AMPAR composition. Recently, we demonstrated that A- kinase anchoring protein 79 kDa (AKAP79; AKAP150 in rodents) targets PKC to GluA1 AMPARs to enhance the phosphorylation and the activity of these receptors. Based on preliminary data, we propose that AKAP79- anchored PKC regulates AMPAR subunit composition by favoring homomeric GluA1 receptors. Moreover, we propose that both PKC-affiliated scaffolds work in conjunction with each other to ultimately determine the AMPAR subunit composition. In order to test this hypothesis, we will use biochemical, electrophysiological, molecular methods, and a novel imaging based approach to understand the: 1) Whether AKAP79 favors homomeric GluA1 receptors via GluA1 regulatory sites using a simplified heterologous system. 2) Whether PICK1 and PKC sites on GluA2 contributes to AKAP79-dependent shifts in AMPAR composition and conversely whether GluA1 regulatory sites contribute to PICK-dependent alterations in AMPAR composition. 3) Whether AKAP150-anchored PKC is central towards generating homomeric GluA1 responses in neurons under a number of conditions linked to synaptic plasticity. Collectively, these studies will shed light on the contributions of PKC-affiliated scaffolds towards determining AMPAR subunit composition and delineate the relative importance of regulatory sites on GluA1 and GluA2 AMPAR subunits which are a critical factor for both heath and disease. PUBLIC HEALTH RELEVANCE: Fast excitatory transmission is primarily mediated by the AMPA subtype of glutamate receptors. Changes in the composition of these receptors are critical for learning and memory, as well as for stroke, pain, fear, drug abuse, and neurodegenerative diseases. These studies will examine the mechanisms contributing to AMPA receptor subunit changes and will help to uncover new targets and treatment strategies for these disorders.