Modulation of Colonic Response to Stress by Peripheral CRF2 Receptors

摘要 IBS患者反复腹痛与排便习惯改变有关。IBS的机制/S是 人们对此知之甚少，也缺乏治疗方法。折衷的应激反应是IBS的关键风险因素。这个 应激的主要介质是促肾上腺皮质激素释放因子(CRF)。在上一次授权期内，我们 研究表明，与CRF1相反，CRF2受体抑制结肠感觉运动反应，并且 CRF2功能受损会加剧结肠对应激的反应。CRF2的调节机制 结肠的应激恢复能力尚不清楚。在初步数据中，我们发现a)CRF2激活结肠 神经元型一氧化氮合酶和一氧化氮的释放，并参与血管活性肠肽抑制结肠运动反应，b)人tau- 过度表达的小鼠对轻度应激和c)CRF2的结肠反应发生了改变，但对CRF1的激活没有影响 抑制应激或CRF诱导的肠道和初级传入神经元tau的磷酸化。基于数据 根据上次的实验和初步数据，我们假设结肠中的CRF2参与了压力应对。 通过激活肠道NO和调节神经元tau发挥作用，tau是一种新的肠道应激途径 CRF2信号的改变将导致对压力的结肠感觉运动反应不适应。这将是 在三个目标下进行了测试。目的1将确定CRF2-NO-VIP三联体的功能相互作用。CRF2 体内介导的nNOS分子活化、NO释放、VIP激活和结肠收缩 在幼年和应激大鼠中进行监测。应激状态下结肠CRF2受体剪接变异体的调控 评估以确定与结肠动力稳态紊乱相关的变异Aim 2将 刻画了一条新的应激-tau-肠道通路，并将剖析应激诱导的效应和机制 大鼠肠神经tau蛋白的调节。结肠肠神经元与tau蛋白磷酸化的同一性 将研究CRF2对结肠反射和应激诱导的tau磷酸化的抑制作用。目标3 将检验应激或CRF诱导的内脏伤害性感觉调制涉及肠道和 脊髓神经元tau的磷酸化。肌间和背根神经节神经元tau和神经元兴奋性 应激大鼠和非应激大鼠的内脏痛将被评估。CRF2是否介导了 抑制内脏伤害性感觉与抑制tau的磷酸化有关 下定决心。将使用肠道和脊髓tau调节以及CRF2(SiRNA)的位点特异性缺失。总的来说， 这些研究将使用药理学、遗传学、组织、细胞和分子工具来剖析其机制。 CRF2受体激活如何抑制应激相关的结肠感觉运动反应。解开这些 CRF2介导的肠道应激反应机制将对理解 应激诱导的肠道感觉运动改变及其在指导未来应激相关新治疗方法中的作用 IBS等疾病。

Abstract IBS patients have recurrent abdominal pain associated with altered bowel habits. The mechanisms/s of IBS is poorly understood and therapies are lacking. A compromised stress response is a key risk factors for IBS. The primary mediator of stress is the peptide cortocotropin releasing factor (CRF). In the last granting period, we have shown that in contrast to CRF1, CRF2 receptors suppress the colonic sensorimotor responses and that a compromised CRF2 function exacerbates the colonic response to stress. The mechanism how CRF2 mediates stress resiliency in the colon is not known. In preliminary data, we show that a) CRF2 activates colonic neuronal NOS and NO release and involves VIP to inhibit colonic motor response, b) human tau- overexpressing mice display altered colonic response to mild stress and c) CRF2 but not CRF1 activation suppresses stress or CRF-induced enteric and primary afferent neuronal tau phosphorylation. Based on data from last grant and the preliminary data, we hypothesize that CRF2 in the colon subserves a stress-coping function through activation of enteric NO and modulation of neuronal tau, a novel enteric stress pathway, and that altered CRF2 signaling will lead to maladaptive colonic sensorimotor responses to stress. This will be tested under three aims. Aim 1 will determine the functional interaction of the CRF2-NO-VIP triad. CRF2 mediated molecular activation of nNOS, NO release, VIP activation and colonic contraction in vivo will be monitored in na¿ve and stressed rats. Regulation of colonic CRF2 receptor splices variants under stress will be assessed to determine variants that are associated with perturbed colonic motor homeostasis Aim 2 will characterize a novel stress-tau-gut pathway and will dissect the effects and mechanism of stress-induced enteric neuronal tau modulation in rats. The identity of colonic enteric neurons with tau phophorylation, the colonic reflex and the suppression of stress-induced tau phophorylation by CRF2 will be investigated. Aim 3 will test the hypothesis that stress or CRF-induced modulation of visceral nociception involves enteric and spinal neuronal tau phosphorylation. Myenteric and DRG neuronal tau and neuronal excitability along with visceral nocicption in stressed and non stressed rats will be assessed. Whether the CRF2 mediated suppression of visceral nociception is associated with the suppression of tau phosphorylation will be determined. Enteric and spinal tau modulation and site specific deletion of CRF2 (siRNA) will be used. Overall, the studies will use pharmacological, genetic, tissue, cellular and molecular tools to dissect the mechanisms how CRF2 receptor activation suppresses stress-related colonic sensorimotor responses. Unraveling these CRF2 mediated mechanisms in the gut response to stress will have significant impact in the understanding of stress-induced gut sensorimotor alteration and in guiding future novel therapeutic approaches to stress related diseases such as IBS.