Multiphoton detection of dopamine and drug release

多巴胺和药物释放的多光子检测

基本信息

批准号：
8780111
负责人：
EDWIN S LEVITAN
金额：
$ 19.25万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-06-15 至 2016-05-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8780111
关键词：
Affect Amphetamines Animal Model Animals Anti-Anxiety Agents Antipsychotic Agents Brain Clinical Cognition Complex Corpus striatum structure DRD2 gene Dendrites Detection Development Dopamine Dopamine Antagonists Dopaminergic Agents Dose Drug Delivery Systems Drug effect disorder Exocytosis Feasibility Studies Fluorescence Image Life Light Microscope Mediating Microscopy Midbrain structure Moods Mus Neurons Optics Parkinson Disease Pharmaceutical Preparations Pharmacology Photochemistry Property Protocols documentation Reporting Rodent Serotonin Signal Transduction Site Slice Specificity Substantia nigra structure Synapses Testing Tryptamines Vesicle addiction atypical antipsychotic base brain tissue clinical efficacy clinically relevant dopamine transporter dopaminergic neuron dorsal raphe nucleus drug of abuse follow-up insight monoamine neuronal cell body novel novel strategies pars compacta patch clamp public health relevance receptor research study response tool

项目摘要

DESCRIPTION (provided by applicant): Dopamine is important in Parkinson's disease and the action of abused drugs (e.g. amphetamines) and clinical psychiatric drugs (e.g. antipsychotics). Because dopamine cannot be seen directly in living neurons with the light microscope, it is difficult to localize and quantify Ca2+-dependent vesicular and amphetamine- induced nonvesicular dopamine release from the soma, dendrites and terminals in living brain tissue. Likewise, the hypothesis that drugs accumulate in and are released from monoamine vesicles has not been tested. Such corelease of antipsychotic drugs with their target transmitters would result in concentrated drug delivery when and where drug action is needed (i.e., at active dopamine and serotonin synapses) resulting in greater efficacy and specificity. To be able to visualize dopamine and drug dynamics in living neurons, we have been developing new experimental approaches based on multiphoton microscopy in the rodent brain slice. First, we found that the clinically used anxiolytic antipsychotic drug cyamemazine produces visible fluorescence upon multiphoton excitation. Multiphoton imaging in midbrain slices showed that cyamemazine is subject to acidic trapping and Ca2+-dependent release. Second, multiphoton microscopy detected autofluorescence in substantia nigra dopamine neurons. Amphetamine induced dopamine transporter- mediated depletion of this signal. Likewise, depolarization induced Ca2+-dependent depletion. These results support the hypothesis that intrinsic multiphoton autofluorescence is derived from dopamine. This proposal builds on these preliminary results to first determine whether multiphoton microscopy in the brain slice can image a dopamine-derived signal that reveals content and release. Then the hypothesis that there is colocalized release of an antipsychotic drug with its target monoamine transmitters (i.e., dopamine and serotonin) is tested. These experiments will explore new optical approaches for studying dopamine and drugs in living brain tissue. Furthermore, determining whether there is corelease of an anxiolytic antipsychotic drug with serotonin and dopamine would be important for establishing a new paradigm for psychiatric drug action.

描述（由申请人提供）：多巴胺在帕金森氏病以及滥用药物（例如苯丙胺）和临床精神药物（例如抗精神病药）中很重要。由于多巴胺在具有光学显微镜的活神经元中无法直接看到，因此很难定位和量化Ca2+依赖性的水泡和苯丙胺诱导的非氧化型多巴胺从躯体，树突和末端释放在活脑组织中。同样，尚未测试药物积累并从单胺囊泡中释放的假设。这种抗精神病药与目标发射器的抗精神病药在需要药物作用（即在活性多巴胺和5-羟色胺突触中）导致浓度的药物递送，从而导致更大的疗效和特异性。为了能够可视化活神经元中的多巴胺和药物动力学，我们一直在基于啮齿动物脑切片中的多光显微镜检查开发新的实验方法。首先，我们发现临床上使用的抗焦虑抗精神病药甲状腺嗪在多光子激发时会产生可见的荧光。中脑切片中的多光子成像表明，甲状腺嗪受到酸性捕获和Ca2+依赖性释放的约束。其次，多光子显微镜检测到黑质多巴胺神经元的自动荧光。苯丙胺诱导该信号的多巴胺转运蛋白转运蛋白介导的耗竭。同样，去极化诱导Ca2+依赖性耗竭。这些结果支持以下假设：固有多光子自动荧光源自多巴胺。该建议基于这些初步结果，以首先确定脑切片中的多光子显微镜是否可以成像多巴胺衍生的信号，以揭示内容和释放。然后假设抗精神病药与其靶标单胺发射器的共定位释放（即测试了多巴胺和5-羟色胺）。这些实验将探讨研究活脑组织中多巴胺和药物的新光学方法。此外，确定是否存在与5-羟色胺和多巴胺的抗焦虑抗精神病药的释放对于建立新的精神药物作用范式很重要。