Arrestin Regulation of CRF1 Signaling: Mechanisms for Stress Resilience and PTSD

抑制素对 CRF1 信号传导的调节：压力恢复和 PTSD 的机制

• 批准号: 8440713
• 负责人: RICHARD L HAUGER
• 金额: --
• 依托单位: VA SAN DIEGO HEALTHCARE SYSTEM
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-10-01 至 2016-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8440713
• 关键词: 1-Phosphatidylinositol 3-Kinase; ADRBK2 gene; Acoustics; Acute; Adverse effects; Agonist; Antipsychotic Agents; Anxiety; Arrestins; BCL2L11 gene; Basal Ganglia; Behavior; Behavioral; Binding; Brain; CRF receptor type 1; CRF receptor type 2; Cell model; Cells; Cognitive; Corticotropin-Releasing Hormone; Corticotropin-Releasing Hormone Receptors; Coupled; Coupling; Cyclic AMP; Cyclic AMP-Dependent Protein Kinases; Data; Disease remission; Dopamine D2 Receptor; Endosomes; Environment; Event; Exhibits; Family; Fright; Functional disorder; G Protein-Coupled Receptor Genes; G Protein-Coupled Receptor Signaling; GRK; GTP-Binding Proteins; Gene Deletion; Genes; Goals; Homeostasis; Knock-out; Knockout Mice; Lead; Life; Ligands; Maintenance; Mediating; Mediator of activation protein; Medical; Membrane; Memory; Metabolic; Molecular; Molecular Target; Motor; Mus; Pathway interactions; Patients; Pharmaceutical Preparations; Pharmacologic Substance; Pharmacotherapy; Phosphorylation; Physiological; Play; Post-Traumatic Stress Disorders; Protein Binding; Protein Kinase A Inhibitor; Protein Kinase C Inhibitor; Proteins; Receptor Signaling; Recovery; Recurrence; Reflex action; Regulation; Reporting; Research; Role; Signal Pathway; Signal Transduction; Signal Transduction Pathway; Site; Site-Directed Mutagenesis; Small Interfering RNA; Specificity; Stimulus; Stress; Symptoms; System; Testing; Therapeutic Index; Time; Trauma; Veterans; Wild Type Mouse; Yeasts; attenuation; biological adaptation to stress; clinical efficacy; combat; counterregulation; desensitization; design; drug discovery; improved; insight; neurotransmission; novel; preclinical study; prevent; public health relevance; receptor; receptor function; research study; response; scaffold; stress resilience; urocortin; yeast two hybrid system

DESCRIPTION (provided by applicant): The central CRF system is a critical mediator of behavioral, cognitive, HPA axis, and autonomic responses to stress and trauma. While central CRF system activation is necessary for survival during life-threatening challenges to homeostasis, rapid counterregulation of CRF neurotransmission is equally important for restoring normal physiological function upon threat termination. The CRF/urocortin family of stress ligands activates signaling of the two cloned CRF receptors CRF-R1 and CRF-R2 via Gs-coupled, Gq-coupled and G protein-independent pathways. Preclinical studies including our predators stress paradigm have shown that CRF-R1 signal transduction plays a vital role in the induction and maintenance of a number of behaviors associated with PTSD including stress-induced hyperarousal, hypervigiliance, stimuli avoidance, startle hyperreactivity, persistent anxiety, and consolidation of contextual fear memories. Combat Veterans with PTSD have markers of abnormal CRF system functioning, including elevated cerebrospinal CRF levels and abnormal HPA axis function. Our research has shown that CRF-R1 signaling via the cyclic AMP-protein kinase A pathway is strongly counterregulated by a GRK-¿arrestin2 desensitization mechanism, which inhibits G protein binding and internalizes the membrane receptor into cytosolic endosomes. We have demonstrated that a ¿arrestin2 gene deletion in CRF-R1-expressing cells induces a 5-fold increase in the maximum for CRF-stimulated cyclic AMP accumulation compared to normal Gs-coupled CRF-R1 signaling in wild-type and ¿arrestin1 knockout cells. Our preliminary behavioral experiments found that ¿arrestin2 knockout mice exhibit greater and more prolonged startle hyperreactivity in response to central CRF administration compared to CRF-enhanced startle behavior in controls. We hypothesize that ¿arrestin2 regulation of CRF-R1 signaling is necessary for acute and long-term stress recovery. Conversely, we hypothesize that a ¿arrestin2 deficiency will result in persistent hyperarousal, hypervigiliance, anxiety, and trauma memory due to dysregulated CRF-R1 signal transduction. We will test our hypotheses in the following manner: In Specific Aim #1, we will examine molecular regulation of G protein-coupled and G protein-independent CRF-R1 signaling pathways by ¿arrestin2 mechanisms. We anticipate that ¿arrestin2 will restrain and desensitize Gs- and possibly Gq-coupled signaling after CRF-R1 is phosphorylated by GRK3. We will also determine if ¿arrestin2 will promote alternative CRF-R1 signaling via the PI-3 kinase-Akt cascade independent of G protein coupling. In addition, we will use PDZ scaffold arrays and yeast two-hybrid screens to identify cellular proteins modulating ¿arrestin2 translocation/binding to CRF-R1 and ¿arrestin2-regulated CRF-R1 pathways. In Specific Aim #2, we will determine if ¿arrestin2 counter regulation of G protein-coupled CRF-R1 signaling is critical for stress recovery. We will complete complementary experiments determining the magnitude and time course of CRF- or predator stress-induced anxiety-like defensive behavior in ¿arrestin2 knockout and wild-type mice. We predict that ¿arrestin2 knockout mice will exhibit abnormally enhanced and prolonged anxiety responses to CRF and stress, due to unrestrained Gs-and/or Gq-coupled CRF-R1 signaling. The major goal of our study is to generate important insight into the role of aberrant ¿arrestin2 regulation of CRF-R1 signaling in PTSD pathophysiology. We will also direct our research toward identifying novel molecular targets for drug discovery that will lead to new PTSD treatments with higher efficacy and therapeutic index.

描述（由申请人提供）： 中央CRF系统是行为，认知，HPA轴以及对压力和创伤的自主性反应的关键中介。虽然中央CRF系统激活对于威胁性稳态的挑战期间的生存是必要的，但CRF神经传递的迅速反调节对于恢复威胁终止时恢复正常生理功能同样重要。应力配体的CRF/尿科素家族通过GS耦合，GQ耦合和G蛋白独立途径激活两个克隆CRF受体CRF-R1和CRF-R2的信号传导。临床前研究包括我们的捕食者应力范式，表明CRF-R1信号转导在诱导和维持与PTSD相关的许多行为中起着至关重要的作用，包括压力诱导的超出声音，超伴奏，超级预防，刺激性，刺激性避免，惊吓性过度反应性，持久性动画，持久的动画和情境恐惧的模样。具有PTSD的战斗退伍军人具有异常CRF系统功能的标记，包括脑脊CRF水平升高和HPA轴功能异常。我们的研究表明，通过循环AMP蛋白激酶A途径通过GRK-¿Crastin2脱敏机制强烈反调CRF-R1信号传导，该机制抑制G蛋白结合并将膜受体内化为细胞质内体。我们已经证明，与正常的GS偶联的CRF-R1信号传导相比，表达CRF-R1的细胞中的ARTARTIN2基因缺失会影响CRF刺激的环状AMP积累的最大增加5倍，而野生型CRF-R1信号传导则增加了5倍。我们的初步行为实验发现，与对照组中的CRF增强的惊吓行为相比，对中央CRF给药的响应，对中央CRF的给药的逮捕in2敲除小鼠暴露了更长，更长的惊吓性过高的过度反应性。我们假设�CRF-R1信号的逮捕素2对急性和长期压力恢复是必需的。相反，我们假设`hustrin2缺陷将导致持久 过度，高维，动画和创伤记忆因CRF-R1信号通路失调而导致的创伤记忆通过»阻止In2机制。我们预计�crestin2将抑制并脱敏的GS和可能的GQ耦合信号传导在CRF-R1被GRK3磷酸化后。我们还将确定通过PI-3激酶-akt级联反应独立于G蛋白偶联的PI-3激酶-AKT级联反应是否会促进替代CRF-R1信号传导。此外，我们还将使用PDZ支架阵列和酵母两杂交筛选来识别调节的细胞蛋白»抑制In2易位/与CRF-R1的结合/与CRF-R1的结合，并抑制了CRF-R1受阻的CRF-R1途径。在特定的目标＃2中，我们将确定�Matistin2反对G蛋白偶联的CRF-R1信号传导对于应力恢复至关重要。我们将完成确定CRF或捕食者应力引起的类似焦虑的防御行为的幅度和时间过程的完整实验。逮捕了Incout和野生型小鼠。我们预测，由于不受限制的GS和/或GQ耦合的CRF-R1信号传导，逮捕了Incontin2敲除小鼠将显着增强对CRF和压力的焦虑反应和延长焦虑反应。我们研究的主要目的是对异常的作用进行重要洞察力。CRF-R1信号在PTSD病理生理学中调节CRF-R1信号。我们还将指导我们的研究确定新型的药物发现分子靶标，这将导致具有更高效率和治疗指数的新PTSD治疗。