Orexinergic Modulation of Experimental Sepsis

项目摘要 复杂的生物体通过炎症保护自己免受威胁，炎症部分由大脑控制 神经递质食欲素的通路。在败血症中，炎症变得失调， 功能障碍发展。我们已经表明食欲素能活性在实验性脓毒症（盲肠）中减弱， 小鼠中的结扎和穿孔（CLP））。本项目的主要目标是确定脓毒症如何引起 食欲素能活性的减弱影响器官功能，改变宿主炎症反应， 有助于死亡率，并确定是否纠正食欲素活性降低逆转这些变化。 我们将通过三个具体目标来实现这些目标。 具体目标1。确定CLP诱导的食欲素能活性丧失对器官功能的影响， 病理学和生存。检查自主神经系统对这些变化的贡献。 方法：我们将用ICV食欲素注射液逆转CLP诱导的食欲素能活性丧失，并检查 这是如何改变心脏、肺、肝、肾和免疫功能以及生存率的。我们将使用外围设备 β-肾上腺素能和毒蕈碱阻滞，以确定自主神经系统的作用。此外，本发明还 我们将确定在几个不同的时间点输注ICV食欲素是否能提高14天的存活率。 具体目标2。确定CLP诱导的食欲素能活性丧失对脑胆碱能神经元的影响。 活动和炎症反射。 方法：我们将通过ICV注射食欲素来逆转CLP诱导的食欲素能活性丧失， 确定这如何影响基底前脑毒蕈碱和炎症反射活动。为了实现这一目标，我们 将检查ICV食欲素注射对1）胆碱乙酰转移酶（ChAT）的共定位和 c-fos免疫染色，指示适当脑切片中的胆碱能活性，2）TNFα的血清水平， IL-6和HMGB 1和3）收获的刺激脾细胞培养基中的TNFα、IL-6和HMGB 1水平。. 具体目标3。确定CLP诱导的脑胆碱能活性丧失对食欲素能的影响 活动，因此，对HR，RR，T和垂体激素分泌的改变。 方法：我们将逆转CLP诱导的脑胆碱能活性的损失与咕诺美林，一个中央M1 激动剂，或ICV尼古丁，并确定食欲素活性和HR，RR，T和垂体激素分泌 都恢复了然后我们将研究食欲素拮抗剂Almorexant的作用 证明CLP诱导的食欲素能系统的变化调节器官功能障碍，炎症反应， 反应和生存可能会重新引导调查和治疗范式，将重点放在大脑上， 败血症的主要驱动因素，克服进展的关键障碍。证明我们可以改变这些 通过增强脑胆碱能活性的作用可以提供一种新的临床可行的治疗途径， 在这种致命的，高度流行的疾病。

Project Summary Complex organisms protect themselves against threats via inflammation, which is in part controlled by brain pathways involving the neurotransmitter orexin. In sepsis, inflammation becomes dysregulated and organ dysfunction develops. We have shown that orexinergic activity is attenuated in experimental sepsis (cecal ligation and puncture (CLP) in mice). The main objectives of this project are to identify how sepsis-induced attenuation of orexinergic activity effects organ function, alters the host inflammatory response and contributes to mortality, and to determine if correcting decreased orexinergic activity reverses these changes. We will address these objectives via three Specific Aims. Specific Aim 1. Determine the effects of CLP-induced loss of orexinergic activity on organ function and pathology and on survival. Examine the contribution of the autonomic nervous system to these changes. Approach: We will reverse the CLP-induced loss of orexinergic activity with ICV orexin injection and examine how this alters cardiac, pulmonary, hepatic, renal and immune function, and survival. We will use peripheral β-adrenergic and muscarinic blockade to identify the contribution of the autonomic nervous system. Further, we will determine if an ICV orexin infusion delivered at several different time points improves 14-day survival. Specific Aim 2. Determine the effects of the CLP-induced loss of orexinergic activity on brain cholinergic activity and on the inflammatory reflex. Approach: We will reverse the CLP-induced loss of orexinergic activity with an ICV injection of orexin and determine how this effects basal forebrain muscarinic and inflammatory reflex activity. To accomplish this we will examine the effects of an ICV orexin injection on 1) co-localization of choline-acetyltransferase (ChAT) and c-fos immunostaining, indicative of cholinergic activity, in appropriate brain sections, 2) serum levels of TNFα, IL-6 and HMGB1 and 3) TNFα, IL-6 and HMGB1 levels in the medium of harvested, stimulated splenocytes. . Specific Aim 3. Determine the effects of CLP-induced loss of brain cholinergic activity on orexinergic activity and, as a result, on alterations in HR, RR, T and pituitary hormone secretion. Approach: We will reverse the CLP-induced loss of brain cholinergic activity with xanomeline, a central M1 agonist, or ICV nicotine, and determine if orexinergic activity and HR, RR, T and pituitary hormone secretion are restored. We will then examine the effects of the orexin antagonist almorexant Demonstrating that CLP-induced changes in the orexinergic system modulate organ dysfunction, inflammatory responses and survival could re-direct the investigative and therapeutic paradigm to focus on the brain as a primary driver of sepsis, overcoming a critical barrier to progress. Demonstrating that we can alter these effects by enhancing brain cholinergic activity could provide a new, clinically viable therapeutic avenue for use in this deadly, highly prevalent disorder.