Real-time measurements of neurotransmission in Drosophila melanogaster

描述(申请人提供)：本研究的长期目标是研究果蝇-黑腹果蝇中枢神经系统神经递质信号的动力学。该项目的具体目标是开发一种微电极方法来测量完整果蝇神经系统中5-羟色胺的实时释放和清除。果蝇是生物学家常用的模型系统，因为它很容易进行基因突变，并且与更高级的生物同源，包括类似的神经递质系统。苍蝇中枢神经系统(8 NL)的小尺寸阻碍了对神经递质释放和摄取的实时研究。这项研究对人类健康有影响，因为5-羟色胺是一种重要的神经递质，其信号与精神疾病有关，包括抑郁症和药物滥用。然而，调节5-羟色胺浓度的机制还不是很清楚。具体目的是：1.研究果蝇幼虫体内5-羟色胺的实时释放和摄取。具有蓝光敏感通道的转基因果蝇将被用来专门诱导5-羟色胺的释放。这项技术将被用来检验有关可卡因注射对5-羟色胺转运体活性的影响以及遗传操作对神经递质释放的影响的假说。2.比较果蝇5-羟色胺能和多巴胺能信号的调节。重复刺激将被用来评估合成和循环在维持释放方面的重要性。3.果蝇5-羟色胺转运体活性快速检测方法的建立少量的5-羟色胺将被压力喷射到神经索中，然后在微电极上被检测到。这项技术将允许对药物或5-羟色胺转运体突变对5-羟色胺信号影响的假说进行测试。这些实验将产生对苍蝇的分析，这对于筛选调节神经递质水平的遗传因素将是有价值的，从而更好地从根本上了解神经传递的时间进程。公共卫生相关性这项研究将有助于更好地了解5-羟色胺的基本神经生物学，5-羟色胺是一种重要的调节性神经递质。我们将研究药物和基因突变如何影响果蝇神经系统中的5-羟色胺信号。这些研究很重要，因为包括抑郁症和强迫症在内的许多精神疾病都是通过针对5-羟色胺系统的药物来治疗的，但这些药物的作用机制尚不清楚。

DESCRIPTION (provided by applicant): The long-term goal of this research is to study the dynamics of neurotransmitter signaling in the central nervous system of Drosophila melanogaster, the fruit fly. The specific goal of this project is to develop a microelectrode method to measure the real-time release and clearance of serotonin in an intact fruit fly nervous system. The fruit fly is a popular model system for biologists because of the ease of making genetic mutations and its homology with higher order organisms, including similar neurotransmitter systems. The small size of the fly central nervous system (8 nL) has precluded real-time studies of neurotransmitter release and uptake. This study has implications for human health because serotonin is an important neurotransmitter whose signaling is implicated in mental illnesses, including depression and drug abuse. However, the mechanisms for regulation of serotonin concentrations are not well understood. The specific aims are: 1. To characterize real-time, endogenous serotonin release and uptake in individual Drosophila larva. Genetically modified flies with a blue-light sensitive channel will be used to specifically elicit serotonin release. This technique will be used to test hypotheses about the effects of cocaine administration on serotonin transporter activity and the effects of genetic manipulations on neurotransmitter release. 2. To compare the regulation of serotonergic and dopaminergic signaling in Drosophila. Repeated stimulations will be used to assess the importance of synthesis and recycling in maintaining release. 3. Development of a rapid assay for serotonin transporter activity in Drosophila. Small amounts of serotonin will be pressure ejected into the nerve cord and then detected at the microelectrode. This technique will allow tests of hypotheses about the effect of pharmacological agents or mutations of the serotonin transporter on serotonin signaling. These experiments will result in assays for the fly that will be valuable for screening genetic factors that regulate neurotransmitter levels, resulting in a better fundamental understanding of the time course of neurotransmission. PUBLIC HEALTH RELEVANCE This research will lead to a better understanding of the basic neurobiology of serotonin, an important modulatory neurotransmitter. We will study how drugs and genetic mutations affect serotonin signaling in the fruit fly nervous system. These studies are important because many psychiatric disorders, including depression and obsessive compulsive disorder, are treated by drugs that target the serotonergic system, but the mechanism of action of these drugs is not well understood.