The Roles of Genetically Distinct Cortical Neuron Types in General-Anesthesia- and Sleep-Induced Slow Waves

项目摘要/摘要。可逆性意识丧失是两个主要医学领域的关键部分： 全身麻醉和睡眠。全身麻醉药和非快速眼动(NREM)睡眠均可诱导 皮层脑电(EEG)中的慢波(0.1-4赫兹)。目前尚不清楚是否产生了慢波 在不同麻醉剂下和在NREM睡眠期间，产生的神经回路活动相同。Dr。 Melonakos的初步数据表明，具有不同分子靶点的麻醉剂具有明显的减慢作用 波动机制(目标1假设)。此外，尽管右旋美托咪定麻醉共享神经回路 对于NREM睡眠，它也可能有明显的直接皮质效应，可能导致不同的慢波活动 (Aim 2假设)。这项研究的目的是通过映射皮质神经活动来检验这些假说。 关于麻醉和非快速眼动睡眠的脑电慢波。为了做到这一点，梅罗纳科斯博士将 学习如何在自由行为的啮齿动物中进行钙成像实验。然后，他将录制钙质图像 从钙/钙调蛋白依赖的蛋白激酶IIa阳性(CaMKIIa+)，小白蛋白阳性(PV+)， 生长抑素阳性(SST+)和血管活性肠肽(VIP+)皮质神经元 麻醉和睡眠引起的慢波。将测试异丙酚、氯胺酮和右美托咪定的麻醉效果。 然后，梅洛纳科斯博士将比较麻醉剂和全身麻醉剂之间的神经活动。 然后睡觉。最后，他将通过(1)观察(1)确定SST+神经元在慢波中的作用(目标3假设) 刺激臂旁核阻断慢波后皮层神经元的活动 (2)在麻醉和睡眠诱导的慢波过程中抑制SST+神经元。 在这个项目的K99阶段，Melonakos博士将由Christa Nehs博士和Emery Brown博士指导， 麻省总医院哈佛医学院麻醉和睡眠神经回路专家及教职员工 MGH医院和麻省理工学院(MIT)。梅洛纳科斯博士还将与梅洛纳科斯博士合作。 Michael Hasselmo(波士顿大学)、Nancy Kopell(波士顿大学)和Daniel Aharoni(波士顿大学 加州、洛杉矶)。他将接受Hasselmo博士和Aharoni博士的钙成像培训，并接受统计学 布朗博士的分析。科佩尔博士将指导梅洛纳科斯博士在假设的慢速范围内确定他的发现的方向 来自计算神经科学领域的波动机制。Melonakos博士还将学习光遗传学 来自Nehs博士和麻省理工学院课程的刺激技术。的导师、合作者和其他成员 MGH社区还将在他接近独立时为他提供专业指导，包括 在基金撰写、同行评议、教学和教师求职方面的培训。科学化、专业化培训 Melonakos博士将使他能够开发一个独立的研究计划来直接研究麻醉药 与间接影响的对比。由此产生的对慢波机制的理解有可能改进协议 用于监测全身麻醉和治疗睡眠障碍，有利于患者的安全和健康。

Project Summary/Abstract. Reversible loss of consciousness is a crucial part of two major medical fields: general anesthesia and sleep. General anesthetics and non-rapid-eye-movement (NREM) sleep both induce slow waves (0.1-4 Hz) in the cortical electroencephalogram (EEG). It is unknown whether slow waves generated with different anesthetic agents and during NREM sleep are generated with the same neural circuit activity. Dr. Melonakos’ preliminary data suggests that anesthetic agents with different molecular targets have distinct slow wave mechanisms (Aim 1 Hypothesis). In addition, although dexmedetomidine anesthesia shares neural circuits with NREM sleep, it may also have distinct direct cortical effects, possibly leading to different slow wave activity (Aim 2 Hypothesis). The purpose of this research is to test these hypotheses by mapping cortical neural activity with respect to the EEG slow waves of both anesthesia and NREM sleep. In order to do this, Dr. Melonakos will learn how to perform calcium imaging experiments in freely behaving rodents. He will then record calcium images from Ca2+/calmodulin-dependent protein kinase IIa-positive (CaMKIIa+), parvalbumin-positive (PV+), somatostatin-positive (SST+), and vasoactive intestinal peptide-positive (VIP+) cortical neurons during anesthesia- and sleep-induced slow waves. Propofol, ketamine, and dexmedetomidine anesthesia will be tested. Dr. Melonakos will then compare the neural activity between the anesthetics and between general anesthesia and sleep. Finally, he will identify the role of SST+ neurons in slow waves (Aim 3 Hypothesis) by (1) looking at the activity of cortical neurons following disruption of slow waves by stimulation of the parabrachial nucleus, an arousal area in the brainstem, and (2) inhibiting SST+ neurons during anesthesia- and sleep-induced slow waves. During the K99 phase of this project, Dr. Melonakos will be mentored by Drs. Christa Nehs and Emery Brown, experts in anesthesia and sleep neurocircuitry and faculty at Harvard Medical School, Massachusetts General Hospital (MGH), and Massachusetts Institute of Technology (MIT). Dr. Melonakos will also collaborate with Drs. Michael Hasselmo (Boston University), Nancy Kopell (Boston University), and Daniel Aharoni (University of California, Los Angeles). He will be trained in calcium imaging by Drs. Hasselmo and Aharoni, and statistical analysis by Dr. Brown. Dr. Kopell will guide Dr. Melonakos as he orients his findings within hypothesized slow wave mechanisms from the field of computational neuroscience. Dr. Melonakos will also learn optogenetics stimulation techniques from Dr. Nehs and in a course at MIT. The mentors, collaborators, and other members of the MGH community will also provide him with professional guidance as he nears independence, including training in grant writing, peer review, teaching, and the faculty job search. The scientific and professional training Dr. Melonakos receives will enable him to develop an independent research program to study anesthetics’ direct vs. indirect effects. The resulting understanding of slow wave mechanisms has potential to improve the protocols used to monitor general anesthesia and treat sleep disorders, thus benefiting patient safety and health.