前庭核表达的olr81对晕动症易感性的影响及机制

Motion sickness susceptibility (MSS) shows an obvious character of large individual difference. Screening for susceptible individuals is an important method for prevention of motion sickness. However, the mechanism underlying MSS individual difference is still unclear. Our previous study found significant expression of olfactory receptor (OR) olr81 and its downstream G protein Gαolf in rat vestibular nucleus neurons. Previous studies confirmed that OR gene polymorphism correlate with the susceptibility of some diseases. Our further study found that transcription level of olr81 mRNA in vestibular nucleus relates to MSS. These evidences suggested that olr81 might be a key molecule for determination of MSS. In the present project, based on established animal behaviour model for MSS the correlation between olr81 expression level and sex & age variation for MSS will be examined. We will try to prove that the expression level and receptor activity of olr81 will influence individual difference of MSS and then determine the key down stream signal pathway involved in this process. We will further investigate the interaction between olr81 and NMDA or GABAB receptor and the impact on MSS.We will also perform the linkage analysis between OR gene polymorphism and MSS.The purpose for the upper experiments is to clarify the role of olr81 played in determinatin of MSS individual difference and the related mechanisms.

晕动症易感性具有明显个体差异，筛选易感人群是避免晕动症发生的重要手段；但晕动症易感性差异的机制尚不明确；我们的前期实验发现，嗅觉受体olr81及下游G蛋白Gαolf在大鼠前庭核神经元内大量表达；研究证实，嗅觉受体基因多态性与某些疾病的易感性密切相关；我们进一步的实验发现，晕动症易感性与前庭核olr81 mRNA的转录水平有关；提示我们olr81可能是决定晕动症易感性个体差异的关键分子。因此，本研究在建立晕动症易感性判断行为学模型的基础上，观察晕动症易感性的性别、年龄差异及与olr81表达水平的关系；验证前庭核olr81的表达及活性变化对晕动症易感性的影响，确定在此过程中起主要作用的受体下游信号通路；了解olr81与NMDA及GABAB受体在前庭核内的相互作用及对晕动症易感性的影响；分析olr81基因多态性与晕动症易感性的关联性，以阐明前庭核olr81在决定个体晕动症易感性中的作用及机制。

晕动症是人体暴露于异常加速度环境引起的多系统反应，人体对晕动症的易感性具有明显的个体差异；然而晕动症易感性差异的生物学基础尚不清楚，年龄、性别、遗传等因素影响晕动症易感性的分子途径的也不明确。本课题在前期工作的基础上，着重针对前庭核表达的嗅觉受体olr81 在晕动症易感性中的作用及可能机制展开研究。.首先，我们观察了不同性别、日龄大鼠晕动症易感性差异，明确不同性别动物晕动症易感性的日龄变化规律，发现60日龄雄性动物及30日龄雌性动物最为敏感；其旋转刺激后前庭核olr81 mRNA表达水平升高，而下游Gαolf mRNA及蛋白表达水平下降；此外，60日龄雄性晕动症易感动物尾侧前庭核olr81下游PLC(PI3K)/PI3通路活性明显下降，AC/cAMP/PKA通路明显升高，且易感动物的升高幅度大于不易感动物，但雌性动物未观察到该变化。进一步利用miRNA腺病毒表达载体转染尾侧前庭核下调olr81表达，可明显降低雄性易感动物晕动症敏感性；通过免疫荧光结合共聚焦显微镜观察，发现olr81与NMDA受体在尾侧前庭核（内侧核MVe及脊束核SpVe）存在共表达；蛋白质质谱实验发现olr81胞内段可与酪蛋白激酶2（CK2）相互作用，但与NMDA受体亚单位无直接作用，提示olr81可能通过CK2激酶影响NMDA受体的活性。最后，通过测序技术发现人olr81(OR52J3)基因单核苷酸多态性与晕动症易感性相关，涉及3个SNP位点，分别位于上游调控区（rs77237011）及编码区（rs58664826，rs57026471）；钙成像显示，配体（eugenol）处理后，rs57026471位点C→T突变体的受体活性明显增强；原代培养神经细胞过表达olr81 rs57026471 的TT纯合突变体后，其神经细胞膜上NMDA受体NR2亚单位表达增多；且该突变位点与CK2在olr81上的结合位点毗邻，提示该突变可能影响CK2-olr81的相互作用。.综上所述，晕动症易感与前庭核olr81高表达及下游AC/cAMP/PKA信号通路活性增强有关；人群olr81多态性与晕动症易感性亦呈相关性，以上结果可为未来开发针对易感人群的晕动症新型防治手段提供新途径，但olr81基因的rs57026471位点C→T突变如何通过CK2影响NMDA受体活性尚待进一步研究。