Human GRIA variants and neurological diseases: from molecular mechanism to rescue pharmacology

修改后的摘要-摘要 神经精神障碍与影响个人行为、记忆和学习能力的大脑功能障碍有关。这种残疾可能会给个人、他们的家庭和社会带来毁灭性的精神和经济后果。这些障碍的一个子集的分子基础涉及单基因通道病，这个术语用于描述各种离子通道中的致病变异。α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic酸受体(AMPAR)是一种配体门控离子通道，由四个亚基GluA1-4(编码GRIA1-4)组成的四聚体复合体。AMPAR介导兴奋性突触后电流的快速成分。AMPAR的激活模式可以触发突触强度的变化，突触强度被广泛认为是学习和记忆的细胞关联，并在神经元发育中发挥重要作用。继2007年首次报道一种致病的AMPAR变异后，在患有各种神经发育和神经精神问题的患者中发现了大量散布在四个AMPAR亚单位上的人类变异(&gt；200)，包括自闭症和智力残疾。已有研究表明，GRIA2和GRIA3基因分别对自闭症和精神分裂症具有全基因组意义。尽管在识别新的变种方面取得了快速进展，但实际上既没有对变种进行系统的功能分析，也没有对患者可能的治疗方案进行任何评估。我们提出了一系列的功能和药理学实验，将填补我们知识的这一空白，并将确定病相关GRIA2和GRIA3突变的潜在作用机制，这些突变在健康人群中不存在。拟议的实验将探索AMPAR GriA基因的遗传变化如何影响受体和神经元功能。我们的药理学实验结果评估了FDA批准的药物对具有患者特定变体的AMPAR的影响，通过建议缓解这些变体的功能变化的新治疗策略，将促进个性化药物的机会。我们的数据还将为AMPAR功能提供新的功能洞察。 目的1.人类GriA变异体如何影响受体功能？我们评估了错义GRIA2和GRIA3变异体对激动剂效力、电流反应的时间进程和细胞表面受体运输的影响。 目的2.人类GriA变异体如何影响神经功能？我们将评估神经元突触数量、脊椎形态、运输位置(突触和突触外)、自发的mEPSCs，以及通过一组GRIA2和GRIA3变体诱导的神经毒性(细胞活性和树突肿胀)的能力。 目的3.AMPAR通道病最好的治疗方法是什么？对于功能增益变体，我们将测量竞争性拮抗剂、负变构调节剂或通道阻滞剂(包括FDA批准的)的IC50。我们将估计正向调节剂(例如Ampakines)对于功能丧失变体的EC50。

Modified SUMMARY-ABSTRACT Neuropsychiatric disorders are associated with disabilities of brain function that affect individual’s behavior, memory and ability to learn. Such disabilities can carry devastating mental and economic consequences for the individuals, their families, and society. The molecular basis of a subset of these disabilities involves monogenic channelopathies, a term used to describe disease-causing variants in various ion channels. The α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs), ligand-gated ion channels, represent tetrameric complexes comprised of varying combinations of four subunits, GluA1-4 (encoded by GRIA1-4). AMPARs mediate the fast component of excitatory post-synaptic currents. Patterns of AMPAR activation can trigger a change in synaptic strength, which is widely considered to be a cellular correlate of learning and memory, and play an important role in neuronal development. Following the first report on a disease-causing AMPAR variant in 2007, a large number of human variants (>200) scattered across four AMPAR subunits have been identified in patients with various neurodevelopmental and neuropsychiatric problems, including autism and intellectual disability. It has been suggested that GRIA2 and GRIA3 genes have genome-wide significance for autism and schizophrenia, respectively. Despite the rapid advance in identification of new variants, there are neither virtually no systematic functional analyses for the variants nor any evaluation of possible treatment options for the patients. We propose a series of functional and pharmacological experiments that will fill this gap in our knowledge and will determine the mechanisms underlying the effects of 64 disease-associated GRIA2 and GRIA3 variants that do not exist in healthy population. The proposed experiments will explore how the receptor and neuronal function is impacted by genetic changes in AMPAR GRIA genes. The results of our pharmacological experiments assessing the effects of FDA-approved drugs on AMPARs with patient-specific variants will advance opportunities for personalized medicine by suggesting new therapeutic strategies for mitigation of functional changes by these variants. Our data will also provide novel functional insight into the AMPAR function. Aim 1. How do human GRIA variants impact receptor function? We assess the effect of 64 missense GRIA2 and GRIA3 variants on agonist potency, time course of current responses, and cell surface receptor trafficking. Aim 2. How do human GRIA variants influence neuronal function? We will assess neuronal synapse number, spine morphology, trafficking locations (synaptic vs extrasynaptic), spontaneous mEPSCs, and the ability of induced neurotoxicity (cell viability as well as dendritic swelling) by a set of GRIA2 and GRIA3 variants. Aim 3. How can AMPAR channelopathies best be treated? For the gain-of-function variants, we will measure the IC50 for competitive antagonists, negative allosteric modulators, or channel blockers (including FDA-approved). We will estimate the EC50 for positive modulators (e.g. ampakines) for the loss-of-function variants.