Synaptic mechanisms underlying the rapid antidepressant actions of scopolamine

描述(申请人提供)：严重抑郁障碍(MDD)是世界范围内最流行和最令人衰弱的疾病之一，影响到约17%的人口，并造成巨大的个人和经济负担。目前可用药物的局限性突出了MDD的影响，包括低应答率、治疗耐药性和治疗时间滞后(数周至数月)。这些数据突显了对更有效、更快起效的抗抑郁药的主要未得到满足的需求。最近的研究表明，单一小剂量的东莨菪碱，一种M受体拮抗剂，在治疗耐药患者中产生快速而持久的抗抑郁作用。这种快速作用的机制与典型的单胺再摄取抑制剂完全不同，代表了过去60年来抑郁症领域最重要的发现之一。东莨菪碱快速抗抑郁作用的机制尚未确定，目前的应用解决了这一问题。初步研究发现，在啮齿动物模型中，东莨菪碱能迅速刺激哺乳动物雷帕霉素复合体1靶点(MTORC1)，迅速增加脊髓数目和PFC神经元的功能，并产生快速的抗抑郁行为反应。此外，东莨菪碱的作用被雷帕霉素阻断，表明mTORC1是必需的。根据这些发现，我们推测东莨菪碱的快速抗抑郁作用是通过刺激mTORC1和增加PFC的突触连接而产生的，这种作用是通过谷氨酸的爆发和脑源性神经营养因子(BDNF)的释放而发生的。本申请描述了一种综合的多学科方法，包括分子、生化、电生理、形态、光遗传学和行为学研究，以检验这一假说。目的1利用药理学、病毒载体和突变小鼠相结合的方法，描述东莨菪碱刺激mTORC1的时间过程和区域定位、脊柱密度和行为反应，并确定mTORC1和BDNF的需求。东莨菪碱可以迅速逆转慢性应激导致的突触和行为缺陷的假设也将得到检验。目的2利用微透析和选择性抑制剂，确定谷氨酸-AMPA受体传递在BDNF-mTORC1信号活性依赖刺激和突触形成中的作用。微输注和光遗传学方法将被用来测试PFC亚区谷氨酸传递的作用，并识别快速抗抑郁反应的靶回路。目的3将利用细胞特异性病毒表达的FLOXED shRNA、M1缺失突变体和细胞特异性Cre小鼠，测试谷氨酸爆发是否通过位于GABA中间神经元上的毒扁豆碱-1(M1)受体发生，从而导致谷氨酸传递的抑制。表征东莨菪碱的信号通路将确定快速作用的抗抑郁药物的基本机制，以及更安全、快速作用的药物的新靶点。

DESCRIPTION (provided by applicant): Major depressive disorder (MDD) is one of the most prevalent and debilitating illnesses world wide, affecting ~17 percent of the population and causing enormous personal and economic burden. The impact of MDD is underscored by the limitations of currently available medications, including low response rates, treatment resistance, and therapeutic time-lag (weeks to months). These data highlight a major unmet need for more efficacious and faster-acting antidepressants. Recent studies demonstrate that a single low dose of scopolamine, a muscarinic receptor antagonist, produces rapid and long-lasting antidepressant actions in treatment resistant patients. This rapid action, by a mechanism completely different from typical monoamine reuptake inhibitors, represents one of the most significant findings in the field of depression over the past 60 years. The mechanisms underlying the rapid antidepressant actions of scopolamine have not been identified, and the current application addresses this issue. Preliminary studies have found that scopolamine rapidly stimulates the mammalian target of rapamycin complex 1 (mTORC1), a pathway involved in synaptic protein synthesis, rapidly increases spine number and function of PFC neurons, and produces rapid antidepressant behavioral responses in rodent models. Moreover, the actions of scopolamine are blocked by rapamycin, demonstrating a requirement for mTORC1. Based on these findings, we hypothesize that the rapid antidepressant actions of scopolamine result from stimulation of mTORC1 and increased synaptic connectivity in PFC that occur via a burst of glutamate and release of brain derived neurotrophic factor (BDNF). This application describes an integrated multidisciplinary approach, including molecular, biochemical, electrophysiological, morphological, optogenetic, and behavioral studies to test this hypothesis. Aim 1 will characterize the time course and regional localization of scopolamine-stimulation of mTORC1, spine density, and behavioral responses, and confirm the requirement for mTORC1 and BDNF using a combination of pharmacological, viral vector, and mutant mouse approaches. The hypothesis that scopolamine can rapidly reverse the synaptic and behavioral deficits caused by chronic stress will also be tested. Aim 2 will determine the role of glutamate-AMPA receptor transmission, which has been implicated in activity-dependent stimulation of BDNF-mTORC1 signaling and synapse formation, using microdialysis and selective inhibitors. Microinfusions and optogenetic approaches will be used to test the role of glutamate transmission in subregions of PFC and to identify target circuits that underlie rapid antidepressant responses. Aim 3 will test if the glutamate burst occurs via muscarinic-1 (M1) receptors located on GABA interneurons, resulting in disinhibition of glutamate transmission, using cell specific virally expressed floxed shRNA, M1 deletion mutants, and cell specific Cre mice. Characterization of the signaling pathways for scopolamine will identify fundamental mechanisms for rapid acting antidepressants and novel targets for safer, rapid-acting agents.