Deciphering the neurocircuits that initiate counterregulation

破译启动反调节的神经回路

• 批准号: 9085534
• 负责人: Jonathan Nicholas Flak
• 金额: $ 8.97万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-09-01 至 2016-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9085534
• 关键词: Ablation; Advisory Committees; Aggressive behavior; Appearance; Area; Blood Glucose; Brain; Brain Stem; Cells; Cessation of life; Cholecystokinin; Cholecystokinin B Receptor; Data; Deoxyglucose; Development; Diabetes Mellitus; Educational process of instructing; Educational workshop; Emergency department visit; Event; Exposure to; Faculty; Failure; Foundations; Fright; Glucose; Goals; Health; Hormonal; Hyperglycemia; Hypoglycemia; Incidence; Insulin; Insulin-Dependent Diabetes Mellitus; Learning; Leptin; Life; Link; Mediating; Metabolic; Metabolism; Michigan; Microdialysis; Modeling; Mus; Nature; Neurons; Neurosciences; Output; Paper; Patients; Pharmacogenetics; Plague; Play; Population; Positioning Attribute; Publishing; Recurrence; Regulation; Research; Research Personnel; Resources; Risk; Role; Satiation; Severities; Shock; Signal Transduction; Site; Stimulus; Sympathetic Nervous System; System; Teacher Professional Development; Techniques; Testing; Therapeutic; Therapeutic Intervention; United States; Universities; Work; blood glucose regulation; career; cost; counterregulation; design; diabetic; diabetic patient; effective therapy; glucose disposal; glucose production; hypoglycemia unawareness; in vivo; insulin secretagogues; molecular marker; neural circuit; novel; optogenetics; parabrachial nucleus; post-doctoral training; public health relevance; relating to nervous system; research study; response; skills; targeted treatment; tool; ventromedial hypothalamic nucleus

 DESCRIPTION (provided by applicant): Approximately 29.1 million patients in the United States suffer from diabetes (9.3% of the population) costing 245 billion dollars a year, which have increased 40% in the last five years. These staggering figures underscore the importance of developing more effective and potentially cheaper therapeutics for diabetes treatment. While the goal of diabetes treatment is normoglycemia, most therapeutics (especially insulin and insulin secretagogues) carry with them a significant risk of potentially life-threatening hypoglycemia; this risk increases with the intensity of therapy. Hypoglycemic shock is the cause of 282,000 emergency room visits a year and the death of about 1 in every 20 patients with type 1 diabetes. In addition to the financial burden and risk associated with hypoglycemic episodes, the risk of hypoglycemia limits diabetes treatment. Low glucose in the brain during hypoglycemia triggers the counter-regulatory response (CRR), activating the sympathetic nervous system (SNS) and hormonal response that stimulate glucose production. Repeated exposure to hypoglycemia promotes hypoglycemia-associated autonomic failure (HAAF) and hypoglycemia unawareness, increasing the risk and severity of hypoglycemic events. Hence, it will be crucial to understand the systems that mediate the CRR to hypoglycemia so that we may identify potential targets for therapeutic intervention. My goal is to become an independent investigator studying the neurocircuits that initiate the counterregulatory responses to hypoglycemia. I plan to use the techniques outlined in this proposal to reveal the neurocircuitry of counterregulation, as well as novel factors within this circuitry that can regulate these responses. These techniques (TRAP-seq, optogenetics, and in vivo microdialysis) in conjunction with previously gathered tools, will provide me with a strong set of skills for direct circuit analysis. In addition to the technical development outlined in this proposal, significant professional development will take place as I transition into an independent faculty position. The University of Michigan offers a host of resources, in the form of workshops, seminars, and short courses provided by the Postdoctoral Association, Center for Research Learning and Teaching, Office of Faculty Affairs and Faculty Development, among others. I will also receive guidance from my career advisory committee, who will assist me in finding a position that I will be successful at as an independent investigator. As the foundation of postdoctoral training (T32 and F32 support), I have been studying leptin action via brainstem sites (primarily the parabrachial nucleus (PBN)). These areas contain significant populations of leptin responsive neurons that, until recently, have been essentially unstudied. We found that CCK- expressing, leptin and glucose-inhibited PBN neurons project to the ventromedial hypothalamic nucleus to initiate counterregulatory responses to hypoglycemia, revealed by responses to insulin, 2-deoxyglucose, and during a hyperinsulinemic/ hypoglycemic clamp. Remotely activating these neurons revealed that these neurons can sufficiently drive hyperglycemia and associated homonal changes and remotely inhibiting these neurons blunt counterregulation. Following these studies that yielded exciting results published in a Nature Neuroscience paper, we followed the circuit down to the ventromedial hypothalamic nucleus (VMN) to study the downstream neurons and decipher this neurocircuit that induces pronounced hyperglycemia. The VMN links a variety of metabolic parameters (including glucose concentrations) to SNS outflow and other parameters of the CRR. Many VMN neurons connect to brain areas essential to glucose homeostasis. Distinct, but intermingled, subsets of VMN neurons mediate different effects, however. Indeed, not only do neurons that increase glucose production as part of the CRR reside in the VMN, but the VMN also contains neurons that promote glucose disposal and thus decrease circulating glucose. The intermingling of essentially oppositely-acting neurons within the VMN and the lack of molecular markers to define CRR-specific subtypes of VMN neurons has hampered our ability to study and understand the VMN neurons that contribute to the CRR. Since the VMN contains substantial CCK receptor B (Cckbr) (aka CCK-2-receptor) and the PBN neurons that drive hyperglycemia express CCK, we generated Cckbrcre mice to study CCK-responsive VMN cells. Our preliminary data demonstrate that Cckbrcre identifies a subpopulation of VMN neurons, and that pharmacogenetic activation of these VMNCckbr neurons increases blood glucose. The proposed experiments test the hypothesis that low glucose activates VMNCckbr neurons, these neurons project to regions that mediate SNS outflow and glucose production, and VMNCckbr neurons play a central role in the CRR to hypoglycemia and the onset/appearance of HAAF. In addition to potentially revealing a neural system underlying the CRR, these studies will define the regulation, circuitry and function of a novel subpopulation of neurons in the metabolically-important VMN, thereby increasing our understanding of brain systems that control metabolism. By understanding the underlying mechanisms of both the CRR and HAAF, we will potentially determine patients who are susceptible to HAAF and carefully administer treatment, while also aggressively treating patients without such a risk. We may also be able to develop therapeutics to co-administer with insulin and insulin secretagogues or design better therapeutics that will lower blood glucose without inducing diabetic shock and HAAF.

 描述(申请人提供)：美国约有2910万糖尿病患者(占总人口的9.3%)每年花费2450亿美元，在过去五年中增加了40%。这些惊人的数字突显了开发更有效、可能更便宜的糖尿病治疗方法的重要性。虽然糖尿病治疗的目标是血糖正常，但大多数治疗药物(特别是胰岛素和胰岛素促分泌剂)存在潜在威胁生命的低血糖的重大风险；这种风险随着治疗强度的增加而增加。低血糖休克是每年28.2万急诊室就诊的原因，每20名1型糖尿病患者中约有1人死亡。除了与低血糖相关的经济负担和风险外，低血糖的风险还限制了糖尿病的治疗。低血糖期间大脑中的低血糖会触发反调节反应(CRR)，激活交感神经系统(SNS)和刺激葡萄糖产生的激素反应。反复暴露于低血糖会促进低血糖相关的自主神经衰竭(HAAF)和低血糖无意识，增加低血糖事件的风险和严重程度。因此，了解CRR调节低血糖的系统将是至关重要的，这样我们就可以确定潜在的治疗干预靶点。我的目标是成为一名独立的研究员，研究启动对低血糖的反调节反应的神经回路。我计划使用这项提案中概述的技术来揭示逆调节的神经回路，以及该回路中可以调节这些反应的新因素。这些技术(陷阱序列、光遗传学和体内微透析)与以前收集的工具相结合，将为我提供一套强大的直接电路分析技能。除了这份提案中概述的技术发展外，随着我过渡到独立的教职员工职位，重大的专业发展也将发生。密歇根大学提供了大量资源，形式包括由博士后协会、研究性学习和教学中心、教师事务和教师发展办公室等提供的研讨会、研讨会和短期课程。我还将接受我的职业咨询委员会的指导，他们将帮助我找到一个作为一名独立调查员将取得成功的职位。作为博士后培训的基础(T32和F32支持)，我一直在研究瘦素通过脑干部位(主要是臂旁核)的作用。这些区域包含大量瘦素反应神经元，直到最近，这些神经元基本上还没有被研究过。我们发现，CCK表达、瘦素和葡萄糖抑制的PBN神经元投射到下丘脑腹内侧核，启动对低血糖的反向调节反应，表现为对胰岛素、2-脱氧葡萄糖的反应，以及在高胰岛素/低血糖钳夹期间的反应。远程激活这些神经元表明，这些神经元可以充分驱动高血糖和相关的同型变化，并远程抑制这些神经元，钝化反调节。在《自然神经科学》的一篇论文中发表了这些令人兴奋的研究结果后，我们沿着这一回路一直到下丘脑腹内侧核(VMN)，研究下游神经元，并破译导致明显高血糖的神经回路。VMN将各种代谢参数(包括葡萄糖浓度)与SNS流出和CRR的其他参数联系起来。许多VMN神经元连接到大脑中对葡萄糖稳态至关重要的区域。然而，不同但混合的VMN神经元亚群调节不同的效应。事实上，不仅作为CRR的一部分，VMN中存在增加葡萄糖产生的神经元，而且VMN中还包含促进葡萄糖处置从而减少循环血糖的神经元。VMN内本质上相互作用的神经元的混合，以及缺乏分子标记来定义CRR特异性的VMN神经元亚型，阻碍了我们研究和理解参与CRR的VMN神经元的能力。由于VMN含有大量CCK受体B(Cockbr)(又名CCK-2受体)，并且驱动高血糖的PBN神经元表达CCK，因此我们建立了Cockbrcre小鼠来研究CCK反应的VMN细胞。我们的初步数据表明，Cockbrcre识别了VMN神经元的一个亚群，并且这些VMNCockbr神经元的药物发生激活会增加血糖。这些实验验证了低糖激活VMNCck br神经元的假说，这些神经元投射到介导SNS外流和葡萄糖产生的区域，而VMNCck br神经元在低血糖CRR和HAAF的发生/出现中起着中心作用。除了可能揭示CRR背后的神经系统外，这些研究还将定义在代谢上重要的VMN中新的神经元亚群的调节、电路和功能，从而增加我们对控制新陈代谢的大脑系统的理解。通过了解CRR和HAAF的潜在机制，我们将有可能确定哪些患者对HAAF易感，并谨慎地进行治疗，同时积极治疗没有这种风险的患者。我们还可能开发出与胰岛素和胰岛素促分泌剂联合给药的疗法，或者设计出更好的疗法，在不引起糖尿病休克和HAAF的情况下降低血糖。