Real-time measurements of neurotransmission in Drosophila melanogaster

描述（由申请人提供）：本研究的长期目标是研究果蝇（Drosophila melanogaster）中枢神经系统中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血清素的实时释放和清除。果蝇是一个受生物学家欢迎的模型系统，因为它易于产生基因突变，而且它与高级生物（包括类似的神经递质系统）具有同源性。蝇类中枢神经系统（8nl）的小尺寸妨碍了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，因为血清素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，调节血清素浓度的机制尚不清楚。具体目标是：1。表征实时内源性血清素在个体果蝇幼虫中的释放和摄取。具有蓝光敏感通道的转基因果蝇将被用来特异性地诱导血清素的释放。这项技术将用于测试可卡因对血清素转运体活性的影响以及基因操作对神经递质释放的影响的假设。2. 比较果蝇血清素和多巴胺信号的调节。将使用重复刺激来评估合成和再循环在维持释放中的重要性。3. 果蝇血清素转运体活性快速测定方法的建立。少量的血清素将被压入神经索，然后在微电极上检测到。这项技术将允许测试关于5 -羟色胺转运体突变对5 -羟色胺信号传导的药理作用的假设。这些实验将导致对苍蝇的分析，这将对筛选调节神经递质水平的遗传因素有价值，从而对神经传递的时间过程有更好的基本理解。这项研究将有助于更好地了解血清素（一种重要的调节神经递质）的基本神经生物学。我们将研究药物和基因突变如何影响果蝇神经系统中的血清素信号。这些研究很重要，因为许多精神疾病，包括抑郁症和强迫症，都是通过靶向血清素能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.