Functional analysis of the Mrg receptor family in pain sensation

描述（由申请人提供）：我们研究的长期目标是了解在正常条件和慢性疼痛状态下痛觉神经元在分子水平上的功能。在哺乳动物中，对疼痛刺激的最初检测是由一组感觉神经元完成的，这些神经元被称为痛觉神经元或痛觉神经元。在慢性疼痛状态下，痛觉神经元对疼痛刺激变得超敏感。最近，我们已经确定了一个大的G蛋白偶联受体家族，称为Mrgs，在痛觉神经元中特异性表达。我们还发现了几种RFamide肽，包括FMRFamide、gamma2-MSH和NPFF，在异种系统中作为Mrgs的配体。我们的电生理研究表明，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延长的炎症性疼痛反应。这些数据表明Mrgs在疼痛中起调节作用。本研究拟利用Mrg敲除小鼠进一步研究Mrg在痛觉中的功能。目的1是分析mrg缺陷小鼠的疼痛行为表型，从中我们想要确定mrg是否在体内促进或减少疼痛敏感性，以及它们是否在慢性疼痛中发挥作用。在Aim II中，我们将表征向野生型小鼠注射Mrg配体肽是否会引起伤害性作用。如果确实如此，我们将确定这些影响是否依赖于核磁共振成像。我们最近的数据表明肥大细胞可能表达Mrgs的内源性配体。一个有希望的候选者是哺乳动物肽NPFF。我们将测试肥大细胞是否表达这种肽。如果有，我们将检查肥大细胞的脱颗粒是否能释放肽。我们还将使用放射性标记的NPFF和γ - msh通过放射自显影研究Mrg受体结合位点。目的三：研究Mrg配体肽诱导神经元高兴奋性的分子机制。许多离子通道参与了痛觉神经元兴奋性的调节。我们将采用电生理记录技术来确定Mrgs是否可以调节培养DRG神经元中离子通道的活动。这项拟议的研究将增强我们对Mrg在疼痛中的功能的理解，这可能会使我们开发出副作用有限的慢性疼痛特异性新型治疗药物。

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.