褐飞虱5-羟色胺受体作为新型杀虫剂靶标的药理学研究

Serotonin (5-HT) is known for its key role in modulating diverse physiological processes and behaviors by binding various 5-HT receptors (5-HTRs) belonging to the superfamily of G-protein coupled receptors (GPCRs). Because of their crucial roles in the regulation of multiple physiological processes, more than 50% of all marketed drugs are thought to directly or indirectly target GPCRs. However, because less pharmacological data is available for insects GPCRs, we have not yet developed ideal insecticide which acts on insect GPCRs. Although previous studies indicated that pymetrozine selectively inhibits homopteran feeding behaviour, possibly by interfering with the serotonergic system, there is no convincing evidence for that. In this project, we will study the expression pattern of all five 5-HTRs from brown planthopper and construct an efficient expression system for these receptors using DNA cloning, real-time PCR, eukaryotic expression, Ca2+ and cAMP assay methods. On the basis of this, we will finally investigate the signaling and pharmacological characterisation of serotonin receptors in brown planthopper and study whether the 5-HTRs is the molecular targets of pymetrozine. These studies will help us to develop new insecticides which target insect 5-HTRs using the high-throughput drug screening platform and also to understand the molecular targets of pymetrozine in brown planthopper.

5-羟色胺（5-HT）调控昆虫的各种生理与行为过程，其通过结合5-羟色胺受体（5-HTRs）来发挥作用。5-HTRs隶属于G蛋白偶联受体家族（GPCRs），目前已知的药物中，有超过半数以上的药物作用靶标为GPCRs。然而对昆虫GPCRs的药理学研究相对较少，使得现在还没有开发出非常好的作用于昆虫GPCRs的杀虫剂。有研究表明吡蚜酮影响害虫取食行为与5-HT信号系统有关，但缺乏直接证据。本研究拟以水稻害虫褐飞虱5-HTRs为研究对象，运用基因克隆、定量PCR、真核表达、细胞内Ca2+以及cAMP浓度检测等实验手段，获得褐飞虱5种5-HTRs基因，探明其表达模式，搭建各受体的高效表达体系与筛选平台，并在此基础上解析褐飞虱各5-HTRs的药理学性质及其是否受吡蚜酮激活。本研究所得结果，将为基于害虫5-HTRs的杀虫剂研发提供理论依据与高通量筛选平台，亦可为揭示吡蚜酮的分子作用靶标提供帮助。

章鱼胺（Octopamine, OA）是无脊椎动物体内一种重要的生物胺，对应于脊椎动物体内的去甲肾上腺素，参与调控昆虫生长发育、取食、代谢等多种生理行为过程，并使昆虫以合理的方式来应对外界刺激。在昆虫体内，OA主要是通过结合特异性的章鱼胺受体（Octopamine receptors, OARs）来发挥作用。由于OA被认为主要是在无脊椎动物体内发挥重要功能，因此其所对应的受体被认为是新型杀虫剂开发的理想靶标而备受药理学家们关注。本研究中以农业害虫褐飞虱Nilaparvata lugens(Stål)为研究对象，克隆得到了褐飞虱OARs家族的6个基因序列，利用qPCR技术分析了这6个基因在褐飞虱不同发育阶段和成虫不同组织中的表达模式，通过体外真核表达研究了NlOA2B2型受体的药理学特性，包括激动剂和拮抗剂谱，并通过药理学和遗传学RNAi技术明确了NlOA2B2受体参与调控雌性褐飞虱产卵生理行为。