Molecular Analysis of 5HT Receptor Mediated Signaling

描述（由申请人提供）： 在哺乳动物中，5-羟色胺（5-HT）受体调节多种生理功能，包括渴求、成瘾和进食行为。鉴于这些受体的重要作用，建立一个简化的模型系统，以确定遗传修饰剂的β-肾上腺素能信号将是一个重要的进展。已知在哺乳动物和果蝇属蛋白质之间存在显著的结构相似性，所述蛋白质包括多巴胺能途径（即，5 HT合成酶、转运蛋白和受体）。与哺乳动物相比，果蝇中5 HT受体介导的功能尚未明确。我们最近已经证明，5 HT调节食物摄入的苍蝇，从而建立了果蝇和哺乳动物之间的功能平行。我们已经表明，5 HT受体激动剂和拮抗剂的管理显着减少和增加食物摄入量，分别。 作为初始目标，我们将确定果蝇5 HT受体亚型中的哪一种调节食物摄入。将建立基于体外细胞的测定来筛选5 HT配体，以确定化合物对四种苍蝇5 HT受体中的每一种的亚型特异性。然后将这些配体用于幼虫食物摄入范例。体外药理学特异性将与体内受体介导的效应相关，以确定哪种5 HT受体亚型是摄食行为的多巴胺能抑制的基础。下一步，将评估相关受体基因下调的苍蝇（即RNAi苍蝇）的食物摄入量变化。最后，这些苍蝇将进行基因表达谱分析，以确定新的遗传修饰剂的多巴胺能信号。在该遗传模型中具有改变的转录的基因将优先用于使用体外和体内测定的表征。

DESCRIPTION (provided by applicant): In mammals, serotonin (5HT) receptors modulate a variety of physiologic functions including craving, addiction, and feeding behavior. Given the important role of these receptors, the establishment of a simplified model system in which to identify genetic modifiers of serotonergic signaling would be an important advance. It is known that significant structural similarities exist between the mammalian and Drosophila proteins that comprise the serotonergic pathway (i.e. 5HT synthetic enzymes, transporters, and receptors). In contrast to mammals, 5HT receptor mediated function in the fly is not well defined. We have recently demonstrated that 5HT modulates food intake in flies thus establishing a functional parallel between Drosophila and mammals. We have shown that the administration of 5HT receptor agonists and antagonists significantly decreases and increases food intake, respectively. As an initial goal, we will determine which of the Drosophila 5HT receptor subtype(s) regulates food intake. In vitro cell based assays will be established to screen 5HT ligands to determine the subtype specificity of the compounds at each of the four fly 5HT receptors. These ligands will then be utilized in a larval food intake paradigm. The in vitro pharmacologic specificity will be correlated with in vivo receptor mediated effects to define which of the 5HT receptor subtype(s) underlies the serotonergic inhibition of feeding behavior. As a next step, flies in which the relevant receptor genetically down-regulated (i.e. RNAi flies) will be assessed for changes in food intake. Finally, these flies will be analyzed with gene expression profiling to identify novel genetic modifiers of serotonergic signaling. Genes with altered transcription in this genetic model will be prioritized for characterization using both in vitro and in vivo assays.