Characterization of the Neural Circuits Underlying the Rapid Antidepressant Effect of Ketamine

描述（由申请人提供）：重度抑郁症给社会带来了巨大的负担，估计五分之一的美国人在一生中都会受到影响，其标志性症状包括快感缺乏、精力不足、注意力不集中以及睡眠和食欲失调。 这种疾病的流行与缺乏快速有效的治疗形成了鲜明对比。 药物治疗或行为治疗通常需要八周或更长时间才能缓解症状，多达一半的患者对初始治疗没有反应。 最近的研究表明，单次低剂量的氯胺酮（一种增加大脑中谷氨酸信号的NMDA拮抗剂）可以在短短几个小时内缓解症状，即使是对传统抗抑郁治疗没有反应的患者。 氯胺酮的新型抗抑郁机制尚未完全了解，并且迫切需要在这一领域进行研究，因为氯胺酮具有副作用（包括解离症状）和滥用潜力，使其不适合广泛使用。初步研究表明，氯胺酮的抗抑郁作用可以通过刺激下边缘前额叶皮层（ilPFC）中的谷氨酸能细胞来重现，该区域已被证明在氯胺酮给药后不久会出现谷氨酸释放高峰。 这一关键发现需要刺激的空间、时间和细胞类型特异性，这是通过将病毒载体注射到ilPFC中来实现的，ilPFC将表达通道视紫红质2（ChR 2），一种光敏蛋白，仅在含有CAMKII的细胞中表达，CAMKII是一种神经元能细胞的标记物。 然后用激光刺激这些细胞，激光的频率和持续时间旨在模拟全身氯胺酮给药的时间过程，激光直接连接到固定在颅骨上的光纤套管。 目的1将调查特定的ilPFC轴突预测，这种影响的基础。重要的是，病毒载体驱动ChR 2在整个细胞中表达，甚至在轴突的远端末端。 为了了解ilPFC内的哪些神经递质能细胞负责抗抑郁作用，将插管 将光引导至从ilPFC接收末端投射的区域之一，ilPFC包括背侧缰核（NAc）、外侧缰核（lHb）和中缝背核（dR），最近的研究表明，它们中的每一个都涉及情绪调节和抗抑郁反应。 目标2将说明全氟化学品国际计划在上述每个领域的影响。 使用免疫组织化学，每个脑区的切片将在刺激后不久用c-Fos抗体（神经活动的标记物）染色，以评估脑区域是否由于来自ilPFC的刺激而变得更活跃或更不活跃。 此外，组织将是 用CAMKII或GAD（一种在中间神经元中发现的蛋白质）的抗体共染色，以评估ilPFC投射突触到哪些细胞类型上。 该项目将增加对快速抗抑郁作用的基础电路的理解，以帮助为广泛的临床使用安全的氯胺酮类药物治疗指明方向。

DESCRIPTION (provided by applicant): Major depressive disorder imposes a large burden on society, afflicting an estimated one in five Americans over their lifetime with its hallmark symptoms of anhedonia, low energy, loss of concentration, and sleep and appetite dysregulation. The prevalence of the disease contrasts with the paucity of rapid, effective treatments. Drug therapies or behavioral therapy typically take eight weeks or more to bring symptom relief, and up to half of patients do not respond to initial treatment. Recent studies have shown that a single low dose of ketamine, an NMDA antagonist that increases glutamate signaling in the brain, can bring about symptom relief in just a few hours, even in patients who do not respond to traditional antidepressant therapies. Ketamine's novel antidepressant mechanism has yet to be fully understood, and there is a pressing need for research in this area because ketamine has side effects (including dissociative symptoms) and abuse potential that make it unsuitable for widespread use. Preliminary studies have shown that ketamine's antidepressant effect can be reproduced by stimulation of glutamatergic cells in the infralimbic prefrontal cortex (ilPFC), an area that has been shown to undergo a spike in glutamate release shortly after ketamine administration. This key finding requires spatial, temporal, and cell-type specificity of stimulation, which is attained by injecting a viral vector into the ilPFC that will express channelrhodopsin 2 (ChR2), a light-sensitive protein, only in cells containing CAMKII, a marker for glutamatergic cells. These cells are then stimulated by a laser, at a frequency and duration intended to mimic the time course of systemic ketamine administration, connected directly to a fiberoptic cannula secured to the skull. Aim 1 will investigate the specific ilPFC axonal projections that underlie this effect. Importantly, the viral vector drives expression of ChR2 throughout the cell, even in the distal terminals of axons. To understand which glutamatergic cells within ilPFC are responsible for the antidepressant effect, the cannula will be placed to direct light to one of the regions that receives terminal projections from the ilPFC, which includes the nucleus accumbens (NAc), lateral habenula (lHb), and dorsal raphe (dR), each of which has been suggested by recent research to be involved in mood regulation and antidepressant response. Aim 2 will characterize the effect of the ilPFC projection within each of these areas. Using immunohistochemistry, slices of each brain area will be stained with a c-Fos antibody, a marker of neural activity, shortly after stimulation to assess if the brain region becomes more or less active as a result of stimulation from the ilPFC. Further, the tissue will be co-stained with antibodies for either CAMKII or GAD, a protein found in interneurons, to assess which cell types the ilPFC projections synapse onto. This project will increase understanding of circuits that underlie a rapid antidepressant effect to help point the way to ketamine-like pharmacotherapies that would be safe for widespread clinical use.