Functional analysis of the Mrg receptor family in pain sensation

描述(申请人提供)：我们研究的长期目标是了解痛感神经元在正常条件和慢性疼痛状态下在分子水平上的功能。在哺乳动物中，最初对疼痛刺激的检测是由一组感觉神经元执行的，这些神经元被称为伤害性神经元或痛感神经元。在慢性疼痛状态下，伤害性神经元对疼痛刺激变得高度敏感。最近，我们发现了一个G蛋白偶联受体大家族，称为MRGS，专门在痛觉神经元中表达。我们还发现了几个RFamide多肽，包括FMRFamide、Gamma2-MSH和NPFF，它们在异源系统中作为MRGS的配体。我们的电生理学研究表明，FMRFamide可以以MRG依赖的方式显著增加伤害性神经元的兴奋性。我们的行为分析表明，与野生型小鼠相比，MRG基因簇敲除小鼠表现出更长的炎症性疼痛反应。这些数据表明，mRGS在疼痛中起着调节作用。在这里，我们建议利用MRG基因敲除小鼠进一步研究MRGS在痛觉中的作用。目的分析MRG基因缺陷小鼠的疼痛行为表型，以确定MRG在体内是促进还是降低疼痛敏感性，以及它们在慢性疼痛中是否起作用。在AIM II中，我们将研究向野生型小鼠注射MRG配基多肽是否会引起伤害性反应。如果他们这样做了，我们将确定这种影响是否依赖于MRG。我们最近的数据表明，肥大细胞可能表达MRGS的内源性配体。一个很有希望的候选者是哺乳动物多肽NPFf。我们将测试肥大细胞是否表达该肽。如果他们这样做了，我们将检查肥大细胞的脱颗粒是否可以释放多肽。我们还将使用放射性标记的NPFF和Gamma2-MSH通过放射自显影来研究MRG受体结合部位。目的研究MRG配体多肽诱导神经元兴奋性升高的分子机制。许多离子通道参与调节伤害性神经元的兴奋性。我们将使用电生理记录技术来确定MR是否能够调节培养的DRG神经元的离子通道的活动。这项研究将加深我们对MRG在疼痛中的作用的理解，这可能会为开发副作用有限的慢性疼痛的特定新型治疗剂奠定基础。

DESCRIPTION (provided by applicant): The long-term objective of our research is to understand how pain-sensing neurons function at the molecular level under both normal conditions and chronic pain states. In mammals, the initial detection of pain stimuli is carried out by a group of sensory neurons, known as nociceptive neurons or pain-sensing neurons. Under chronic pain states, nociceptive neurons become hypersensitive to pain stimuli. Recently, we have identified a large family of G protein coupled receptors, called Mrgs, expressed specifically in pain-sensing neurons. We also found several RFamide peptides, including FMRFamide, gamma2-MSH and NPFF, acting as ligands for Mrgs in heterologous systems. Our electrophysiological studies indicate that FMRFamide can significantly increase excitability of nociceptive neurons in an Mrg-dependent manner. Our behavioral analysis showed that Mrg gene cluster knockout mice exhibit a prolonged inflammatory pain response as compared to wild-type mice. These data suggest that Mrgs play a modulatory role in pain. Here we propose to study the function of Mrgs in pain sensation further using Mrg knockout mice. Aim I is to analyze pain behavioral phenotypes in Mrg-deficient mice, from which we would like to determine whether Mrgs act to promote or reduce pain sensitivity in vivo and whether they play roles in chronic pain. In Aim II, we will characterize whether the injection of Mrg ligand peptides into wild-type mice can cause nociceptive effects. If they do, we will determine whether such effects are Mrg-dependent. Our recent data suggest that mast cells are likely to express the endogenous ligands for Mrgs. One promising candidate is the mammalian peptide NPFF. We will test whether mast cells express the peptide. If they do, we will check whether the degranulation of mast cells can release the peptide. We will also investigate Mrg receptor binding sites by autoradiography using radiolabeled NPFF and gamma2-MSH. Aim III is to study the molecular mechanisms of neuronal hyperexcitability induced by Mrg ligand peptides. Many ion channels have been implicated in regulating neuronal excitability of nociceptive neurons. We will employ electrophysiological recording techniques to determine whether Mrgs can modulate the activities of the ion channels in cultured DRG neurons. The proposed research will enhance our understanding of Mrg function in pain, which may allow the development of specific novel therapeutic agents for chronic pain with limited side effects.