Sleep in Rats: From Conceptualization to Measurement, Analysis and Interpretation

描述（由申请人提供）：我致力于疼痛研究的直接结果是照顾从心脏手术中恢复的非常年幼的儿童，没有足够的镇痛效果。我的目标是证明适当的疼痛管理是生理上的需要。我的论文和早期对大鼠的研究表明，在统计学和生物学上，充分提供围手术期镇痛可显著减弱实验性剖腹手术的体内免疫抑制和肿瘤增强作用，并使探索行为正常化。在我的实验设计中纳入女性是主要的。我发现了一种方法，可以调查在生命过程中反复出现的早期新生儿疼痛经历的长期后果，这使我能够专注于非常年幼的幼鼠，并获得了证据，证明爪针反复戳大鼠幼鼠会导致后爪注射福尔马林（一种常用的补偿性炎症性疼痛刺激）后，幼鼠的情绪发生改变，伤害行为得分更高。雌性在成熟后的疼痛发现中表现得更差。在我最近使用临床前模型研究持续疼痛发展的生物行为易感性的努力中，发现：(1)睡眠限制增加了对照组和患有坐骨炎性神经炎的动物对后爪机械过敏的易感性；(2)反复的社会失败，抑郁症的临床前模型；后爪注射福尔马林似乎会增加伤害性行为得分，这极大地促进了我在疼痛研究项目中加入睡眠领域的紧迫感。我的长期目标是研究使生物体在生命过程中易受持续疼痛发展的因素以及这些关系背后的可能机制。注意力集中的因素是疼痛、睡眠和抑郁，很少被认为是三重合并症。在这个为期12个月的职业发展奖期间，我的短期目标是建立一个关于睡眠的知识库，从概念化到仪器，分析和解释，使其完全融入我的生物行为疼痛研究项目。这个计划的关键要素是完成一门基础的多导睡眠图课程，并发展与我的长期研究目标相关的睡眠文献的工作知识。在我承诺的40%的努力中，最大的一部分是致力于熟练掌握脑电图(EEG)/肌电图（EMG）电极和遥测单元植入，唤醒状态量化和脑电图频谱分析，以及分析睡眠结果的统计方法。约翰霍普金斯大学的研究环境非常好。护理学院的领导热情地支持这一职业发展机会，并致力于我们制定的释放计划。我提议的导师Mark Opp博士是一个很大的优势，他是啮齿动物睡眠方面的知名专家，我和他有合作和指导的历史。他承诺作为导师帮助我实现本文所建议的职业发展，并保证在三次建议访问期间可以完全进入他的实验室，并确认他可以在必要时经常进行咨询。睡眠和抑郁是双向并存的，睡眠中断与抑郁风险增加有关，抑郁与睡眠中断有关。该试点项目的目的是对Fischer 344 （F344）雌性和雄性大鼠的睡眠、神经内分泌和免疫功能的影响进行因果评估，这是一种有前景的临床前抑郁症模型，反复暴露于社交失败。在脑电图/肌电图传递器植入完全恢复后，40只雌性和雄性大鼠在4周内接受12次60分钟的社交失败或新笼子暴露，然后在自己的笼子里呆两周。社会失败暴露包括将F344雄性入侵者放入Long Evans雌雄配对的常驻笼中，或者将F344雌性入侵者在移除Long Evans雄性后立即放入常驻笼中。在入侵者被击败后，在剩余的入侵期间，在常驻笼内的入侵者上方放置一个保护性网状屏障。在这段时间里，新奇的笼养动物被放在一个空笼子里。脑电图/肌电图记录是连续的，在社交失败开始之前，每次社交失败/新笼暴露之前和之后，以及在两周的休息期间进行采样。睡眠结果包括觉醒状态、潜伏期状态和睡眠状态特定的脑电图谱。从社会失败/通过安乐死的新笼子暴露前1周开始，每天监测1%蔗糖与白开水的偏好，以指示快感缺乏。血浆皮质酮水平应在术前基线、第一次和最后一次暴露于社会失败/新笼后以及安乐死时进行评估。安乐死的其他结果包括促炎细胞因子水平，地塞米松抑制脂多糖诱导的全血促炎细胞因子的产生。一般线性混合模型和单变量方差分析将适当地用于分析。重要的方面包括长期睡眠测量、神经内分泌和免疫结果，以及纳入女性。

DESCRIPTION (provided by applicant): My commitment to pain research is a direct result of caring for very young children recovering from cardiac surgery without benefit of adequate analgesia. It has been my goal to causally demonstrate that adequate pain management is a matter of physiologic necessity. My dissertation and early career research studies in rats show that adequate provision of perioperative analgesia statistically and biologically significantly attenuate the in vivo immunosuppressive and tumor-enhancing effects of experimental laparotomy and normalize exploratory behavior. Inclusion of females in my experimental designs is a mainstay. Discovering a means by which to investigate the long-term consequences of repeated early neonatal pain experiences over the lifecourse enabled me to focus on the very young, yielding evidence that repeated paw needle pokes in the rat pup result in altered emotionality and greater nociceptive behavior scores in response to hind paw formalin injection, a commonly used tonic inflammatory pain stimulus. Females fared worse in this latter pain finding at maturity. In my recent efforts using preclinical models to study biobehavioral susceptibility to the development of persistent pain, findings that (1) sleep restriction increases susceptibility to hind paw mechanical hypersensitivity in both controls and animals with sciatic inflammatory neuritis and (2) repeated social defeat, a preclinical model of depression, appears to increase nociceptive behavior scores in response to hind paw formalin injection have greatly contributed to my sense of urgency to add the domain of sleep to my program of pain research. My long-term goal is to study factors that render the organism susceptible to persistent pain development over the lifecourse and possible mechanisms underlying these relationships. The factors of focus are pain, sleep and depression, rarely considered as a triad of comorbidity. My short-term goal for the duration of this 12-month Career Development Award is to build a substantial knowledge base regarding sleep from conceptualization to instrumentation, analysis and interpretation, to enable its full incorporation into my program of biobehavioral pain research. The key elements of this plan are to complete a foundational polysomnography course, and develop a working knowledge of the sleep literature relevant to my long-term research goal. The greatest portion of my committed 40% effort is to be dedicated to developing proficiency in electroencephalogram (EEG)/electromyogram (EMG) electrode and telemetry unit implantation, quantification of arousal states and EEG spectral analysis, and statistical methods in the analysis of sleep outcomes. The research environment at Johns Hopkins University is very strong. The School of Nursing leadership enthusiastically endorses this career development opportunity and is committed to the release plan we have devised. A great strength is proposed mentor Dr. Mark Opp, a renowned expert in rodent sleep with whom I have a history of collaboration and mentoring. He is committed as mentor to enable my career development as proposed herein, assured full access to his laboratory during the three proposed visits, and confirmed his availability for consultation as frequently as necessary. Sleep and depression are bidirectionally comorbid such that disrupted sleep is associated with increased risk for depression and depression is associated with disrupted sleep. The purpose of the proposed pilot project is to causally evaluate the impact of a promising preclinical model of depression, repeated exposures to social defeat, on sleep and neuroendocrine and immune function in Fischer 344 (F344) female and male rats. After full recovery from EEG/EMG transmitter implantation, 40 female and male rats undergo 12 60 min exposures to either social defeat or a novel cage distributed over a 4-week period, and then remain in their home cage for two weeks. A social defeat exposure consists of placing the F344 male intruder into the resident cage of a Long Evans female-male pair, or the F344 female intruder into the resident cage immediately following removal of the Long Evans male. Immediately after defeat of the intruder, a protective mesh barrier is placed over the intruder within the resident cage for the remainder of the intrusion period. Novel cage animals are placed in an empty cage during this time. EEG/EMG recordings are to be continuous, with sampling before social defeat is initiated, before and after each social defeat / novel cage exposure, and during the 2-week rest period. Sleep outcomes include arousal state, latencies to state and sleep-state specific EEG spectral profiles. Sucrose 1% versus plain water preference is monitored daily from 1 week prior to social defeat / novel cage exposure through euthanasia to indicate anhedonia. Plasma corticosterone levels are to be assessed at preoperative baseline, immediately after the first and final exposure to social defeat / novel cage, and at euthanasia. Additional outcomes at euthanasia include pro-inflammatory cytokine levels, and dexamethasone suppression of lipopolysaccharide-induced pro-inflammatory cytokine production in whole blood. General linear mixed models and Univariate ANOVA are to be used as appropriate for analysis. Aspects of significance include long- term sleep measurement, neuroendocrine and immune outcomes, and inclusion of females. PUBLIC HEALTH RELEVANCE: Disrupted sleep, pain affect substantial numbers of individuals and are well known to be co-occurring. The relevance of the proposed Career Development Award is to build a substantial knowledge base regarding sleep in order to add this domain to my program of biobehavioral pain research. The relevance of this proposed award to human health is to discern the contributions and consequences of disrupted sleep as it relates to the development of persistent pain.