Spectra-Physics Mai Tai HP Laser

描述（由申请人提供）：神经递质多巴胺在决策、习惯和运动学习以及药物滥用、精神分裂症、注意缺陷多动障碍（ADHD）和帕金森病中起关键作用。这项由三个NIH资助的实验室共同努力的研究，使用新的工具探索了突触前和突触后多巴胺能神经传递的基本机制。该团队包括一个设计和生产光学探针的化学实验室（Sames），一个使用突触水平的光学记录来表征和分析多巴胺突触释放的神经生理学实验室（Sulzer），以及一个使用各种光学方法来阐明多巴胺受体的结构，激活和药理学的分子神经科学实验室（Javitch）。这三个小组在过去五年中一直在密切合作，而这一应用程序正是这些合作的结果。这里描述的新方法使用了第一种光学方法来直接观察神经递质的释放，使用被称为荧光假神经递质（FFN）的光学探针。该团队现在已经开发出第一代比率神经递质探针，提供了一种测量神经末梢内突触囊泡pH值的方法。该探针也已适应于同时测量神经传递和突触囊泡融合在个别突触终端。在突触后侧，额外的探针有望以前所未有的时间分辨率测量受体占有率。虽然我们目前的数据表明，这种新一代的光学探针使用多光子成像在急性脑切片的效用，毫秒级的事件参与神经传递和受体和转运激活需要几乎同时激发两个波长，因此需要两个光源。因此，我们概述了多光子激光器将如何使多个和重要的增强，这个多学科的研究小组的研究。目标1概述了如何突触囊泡融合和神经递质释放将同时记录目标2概述了如何突触囊泡pH值将在活突触终端确定。目标3概述了笼状多巴胺与荧光能量转移将如何确定结构，功能和药理学的D2多巴胺受体在前所未有的动力学分辨率。

DESCRIPTION (provided by applicant): The neurotransmitter dopamine is pivotal in decision making, habit and motor learning as well as in drug abuse, schizophrenia, attention deficit hyperactive disorder (ADHD), and Parkinson's disease. This joint effort from three NIH-funded laboratories explores fundamental mechanisms of both pre- and post-synaptic dopaminergic neurotransmission using novel tools. The team includes a chemistry laboratory (Sames) that designs and produces the optical probes, a neurophysiology laboratory that characterizes and analyzes the synaptic release of dopamine using optical recordings at synaptic level (Sulzer), and a molecular neuroscience laboratory that uses a variety of optical approaches to elucidate the structure, activation, and pharmacology of dopamine receptors (Javitch). The three groups have been collaborating intensively for the past five years, and this application results from these collaborations. The novel approaches described here use the first optical methods developed to directly visualize neurotransmitter release, using optical probes known as fluorescent false neurotransmitters (FFNs). The team has now developed a first generation of ratiometric neurotransmitter probes that provide a means to measure pH in synaptic vesicles within neuronal terminals. The probes have also been adapted to simultaneously measure neurotransmission and synaptic vesicle fusion at individual synaptic terminals. At the postsynaptic side, additional probes promise to measure receptor occupancy at unprecedented temporal resolution. While our current data demonstrate the utility of this new generation of optical probes using multiphoton imaging in the acute brain slice, the millisecond-scale events involved in neurotransmission and receptor and transporter activation require virtually simultaneous excitation at two wavelengths, thus necessitating two light sources. We therefore outline how a multiphoton laser will enable multiple and important enhancements to the research of this multidisciplinary group of research teams. Aim 1 outlines how synaptic vesicle fusion and neurotransmitter release will be recorded simultaneously Aim 2 outlines how synaptic vesicle pH will be determined in living synaptic terminals. Aim 3 outlines how caged dopamine used with fluorescence energy transfer will determine the structure, function, and pharmacology of D2 dopamine receptors at unprecedented kinetic resolution.