Photoactivatable ligands for nicotinic optopharmacology

项目摘要 长期接触烟草制品中的尼古丁会导致许多健康后果（肺癌， 肺气肿、高血压等）每年有超过600万人因此死亡复发率很高， 那些试图戒烟的人，以及寻求促进戒烟的药物疗法， 适度有效。因此，对于治疗尼古丁的更有效的策略存在显著未满足的需求。 依赖制定这样的战略需要更详细地了解生物 导致尼古丁成瘾的机制。与nAChR机制研究相关的一个基本目标是 更好地了解nAChR在单个神经内离散部位的位置和活性 细胞虽然一些基础研究已经开始描述nAChR亚细胞分布， 目前没有合理的方法在亚细胞水平上功能性地询问nAChR。这意味着我们 目前无法确定尼古丁介导的nAChRs功能改变是否 发生在树突、轴突、突触前末梢或神经元胞体中。回答这个关键问题是 该领域完全理解尼古丁依赖的分子和细胞基础绝对需要。 在这里，我们提出了一个R21/R33分阶段创新项目，直接解决我们能力中的这些关键差距， 来研究天然的乙酰胆碱受体R21阶段将开发和表征光活化尼古丁（PA-Nic） 用于尼古丁“释放”实验的化合物。R21目标1侧重于识别1个或多个 合适的化合物使用紫外/可见光光学方法，而目标2将表征有前途的 化合物使用2光子uncaging技术，这提供了增强的空间分辨率。在R33阶段，我们 建议在发现实验中使用这些创新的化合物和光学方法， 揭示nAChR功能的新细节。R33目标1将涉及关键细胞上nAChR的功能定位 尼古丁依赖的类型。R33 Aim 2将探索这些受体如何使尼古丁参与 在大脑奖励系统神经传递的回路水平调制中。该项目是一个重大的 这是胆碱能生物学领域的技术进步，因为它不仅会产生广泛使用的新工具， 将利用这些工具来揭示尼古丁依赖的新机制细节。

PROJECT SUMMARY Chronic exposure to nicotine in tobacco products results in numerous health consequences (lung cancer, emphysema, hypertension, etc.) and accounts for over 6 million deaths per year. Relapse rates are high among those who attempt to quit smoking, and pharmacotherapies that seek to foster smoking cessation have moderate effectiveness. Thus, there is a significant unmet need for more effective strategies to treat nicotine dependence. Development of such strategies requires a more detailed understanding of the biological mechanisms leading to nicotine addiction. An essential goal related to mechanistic studies on nAChRs is gaining a better understanding of the location and activity of nAChRs in discrete sites within individual nerve cells. Although some basic research studies have begun to describe nAChR subcellular distribution, there is currently no plausible way to functionally interrogate nAChRs at the subcellular level. This means we are currently unable to determine whether the important nicotine-mediated functional alterations in nAChRs occur in dendrites, axons, presynaptic terminals, or in neuronal somata. Answering this key question is absolutely required for the field to fully understand the molecular and cellular basis for nicotine dependence. Here, we propose a R21/R33 phased innovation project that directly addresses these critical gaps in our ability to study native nAChRs. The R21 phase will develop and characterize a photoactivatable nicotine (PA-Nic) compound for use in nicotine “uncaging” experiments. R21 Aim 1 focusses on identification of 1 or more suitable compounds using ultraviolet/visible light optical methods, while Aim 2 will characterize promising compounds using 2-photon uncaging techniques, which offer enhanced spatial resolution. In the R33 phase, we propose to employ these innovative compounds and optical methods in discovery experiments designed to uncover new details about nAChR function. R33 Aim 1 will involve functional mapping of nAChRs on key cell types involved in nicotine dependence. R33 Aim 2 will probe how these receptors enable nicotine to participate in circuit-level modulation of neurotransmission in brain's reward system. This project represents a substantial technical advance for the cholinergic biology field, as it will not only produce new tools for widespread use, but will utilize those tools to uncover new mechanistic details about nicotine dependence.