Structure-Function and Signaling of Glutamate Delta 1 in Pain Mechanism

摘要： 疼痛影响了1亿多美国人，产生了包括长期残疾在内的破坏性影响， 精神健康、共病和药物依赖以及巨大的经济负担。可用于治疗的药物 慢性疼痛并不总是有效的，可能会有滥用的倾向。迫切需要确定新的机制 用于治疗慢性疼痛。大量证据支持臂旁杏仁核的可塑性作用 谷氨酸能突触在疼痛的时序化中的作用，但其潜在的机制还不完全清楚。我们有 最近证实了谷氨酸三角洲1受体(GluD1；基因GRID1：UniProt Q9ULK0)及其关键作用。 炎症性和神经病理性臂旁-杏仁核突触中突触源性结合伙伴小脑蛋白1的表达 疼痛。GluD1是离子型谷氨酸受体家族的一员，但它不寻常，因为它缺乏典型的配体门控 离子通道活性，并通过形成跨突触在突触形成/维持中起关键作用 Glud1-Cbln1-neuresin三联体。我们发现GluD1-Cbln1信号在疼痛状态下下调，与之形成对比的是 AMPA受体亚单位表达增加。重要的是，我们发现将重组Cbln1注射到中枢 杏仁核对中枢GluD_1和AMPA受体表达及神经元超兴奋性的影响 杏仁核，也减轻疼痛模型中的厌恶和情感行为。这些结果证明了GluD1的作用- Cbln1信号在疼痛诱导的可塑性中的作用。在进一步的研究中，我们发现重组Cbln1的镇痛作用 被D-丝氨酸抑制。增加D-丝氨酸水平被认为主要是为了加剧明示行为 通过NMDA受体依赖的机制。然而，D-丝氨酸结合到GluD1的配体结合域和 在受体中产生构象变化。最近的研究也报道了GluD1在自然界中的离子通道孔活动 该系统可能由亲代谢性受体激活而触发，并受D-丝氨酸结合的调节。的贡献 D-丝氨酸与GluD1的结合以及GluD1对疼痛机制的潜在离子通道孔活性尚不清楚。的目标是 本研究旨在研究GluD1的结构与功能之间的关系，特别是探讨配体的功能 杏仁核疼痛机制中GluD1的结合域和离子通道孔。提出了两个目标；目标1将 检测D-丝氨酸与GluD1相互作用对重组Cbln1镇痛作用的调节。Aim 2将检查 GluD1的离子通道功能与疼痛机制的关系。这个项目意义重大，因为它将确定一个 神秘的GluD1可能参与慢性疼痛的新机制。预期结果可能会提供 通过靶向配体结合结构域或离子来开发治疗慢性疼痛的新方法的关键信息 GluD_1的通道孔。

Summary: Pain affects more than 100 million Americans producing devastating effects including long-term disabilities, mental health comorbidities and drug dependence as well as enormous economic burden. Available medications for chronic pain are not always effective and may have abuse liability. There is an urgent need to identify novel mechanisms for the treatment of chronic pain. A large body of evidence supports a role of plasticity of parabrachio-amygdala glutamatergic synapses in chronification of pain, but the underlying mechanisms are not fully understood. We have recently demonstrated a critical role of glutamate delta 1 receptor (GluD1; gene GRID1: UniProt Q9ULK0) and its synaptogenic binding partner cerebellin 1 (Cbln1) at the parabrachio-amygdala synapses in inflammatory and neuropathic pain. GluD1 is a member of the ionotropic glutamate receptor family but is unusual because it lacks typical ligand-gated ion channel activity and instead plays a critical role in synapse formation/maintenance by forming a trans-synaptic GluD1-Cbln1-Neurexin triad. We found a downregulation of GluD1-Cbln1 signaling in pain states and a contrasting increase in AMPA receptor subunit expression. Importantly, we found that injection of recombinant Cbln1 into the central amygdala normalized the changes in GluD1 and AMPA receptor expression and neuronal hyperexcitability in the central amygdala and also mitigated averse and affective behaviors in pain models. These results demonstrate a role of GluD1- Cbln1 signaling in pain-induced plasticity. In additional studies we found that the analgesic effect of recombinant Cbln1 was inhibited by D-serine. An increase in D-serine levels has been proposed to exacerbate nocifensive behaviors primarily via a NMDA receptor-dependent mechanism. However, D-serine binds to the ligand binding domain of GluD1 and produces conformational changes in the receptor. Recent studies also report ion channel pore activity of GluD1 in native system potentially trigged by activation of metabotropic receptors and modulated by D-serine binding. The contribution of D-serine binding to GluD1 and potential ion channel pore activity of GluD1 to pain mechanisms is unknown. The goal of the proposed studies is to examine the structure-function relationship of GluD1 specifically probing the function of ligand binding domain and ion channel pore of GluD1 in amygdala pain mechanisms. Two aims are proposed; Aim 1 will examine the regulation of analgesic effect of recombinant Cbln1 by D-serine interaction with GluD1. Aim 2 will examine the ion channel function of GluD1 in relation to pain mechanism. This project is significant because it will identify a novel mechanism by which the enigmatic GluD1 may contribute to chronic pain. The expected outcomes may provide critical information to develop new therapeutic approaches for chronic pain by targeting the ligand binding domain or ion channel pore of GluD1.