Project 2: Pro-inflammatory Cytokines Engaged in Muscle Afferent-Mediated Sympathetic Responsiveness with Femoral Artery Occlusion

项目总结/摘要-项目2 运动升压反射，它产生于收缩骨骼肌，是两种神经反射之一。 引起交感神经和心血管调节运动的机制。这里面的异常 在心血管疾病中，反射影响运动肌肉血流输送和氧气供应。 外周动脉疾病（PAD）是常见的致命性动脉粥样硬化血管疾病的表现。 该项目的主要目标是更好地了解促炎细胞因子（PIC）在控制炎症反应中的作用。 引起运动加压反射的细纤维（III和IV组）肌肉传入神经元的兴奋性， 垫.对于细纤维肌肉传入，我们建议将体外全细胞贴片的力量与联合收割机结合起来， 钳技术，这将确定传入兴奋性的机制，与体内提供的见解 这将决定这些机制如何转化为增加兴奋性。特别 注意具有TTX敏感和抗性Na+通道的传入。对于体外实验， 将通过用逆行标记背根神经节（DRG）神经元来鉴定肌肉传入。 将荧光示踪剂DiI注射到对照大鼠和慢性糖尿病大鼠的小腿三头肌中， 股动脉闭塞此外，分离的背根神经节细胞将通过其瞬时表达进行鉴定 的绿色荧光蛋白，其表达由近端神经元特异性启动子区驱动 TTX敏感性Na+通道（NaV1.7）和TTX抗性Na+通道（NaV1.8）。用于体内 在实验中，将在对照大鼠中检查由肌肉收缩诱发的交感神经反应性， 大鼠股动脉闭塞。在体外和体内实验中，将特别注意 PIC，即IL-6和TNF-α，对传入神经元的膜和放电特性的影响。 此外，细胞因子受体和控制细纤维肌兴奋性的通道之间的相互作用 将检查传入神经。此外，细胞内信号传导途径，参与在兴奋过程中， 将审查太平洋岛屿国家。我们的一般假设是，较高水平的PIC受体诱导肌肉 大鼠PAD模型后肢血流阻断后DRG神经元的变化。这反过来又改变了细胞内的 转导途径和Na+通道的表达和功能，从而导致增强 运动加压反射。预计拟议的实验将提供有关 运动加压反射的传入臂的兴奋性受PIC影响的机制， 垫.

PROJECT SUMMARY/ABSTRACT – PROJECT 2 The exercise pressor reflex, which arises from contracting skeletal muscles, is one of the two neural mechanisms that evoke the sympathetic and cardiovascular adjustments to exercise. The abnormalities in this reflex affect blood flow delivery and oxygen supply to exercising muscles in cardiovascular diseases. Peripheral arterial disease (PAD) is a manifestation of common and lethal atherosclerotic vascular disorders. The major goal of this project is to better understand the role of pro-inflammatory cytokines (PICs) in controlling excitability of the thin fiber (group III & IV) muscle afferent neurons that evoke the exercise pressor reflex in PAD. With respect to thin fiber muscle afferents, we propose to combine the power of in vitro whole-cell patch- clamp techniques, which will determine mechanisms of afferent excitability, with the insights provided by in vivo physiology, which will determine how these mechanisms translate into increased excitability. Particular attention will be paid to afferents that have TTX sensitive and resistant Na+ channels. For in vitro experiments, muscle afferents will be identified by labeling dorsal root ganglion (DRG) neurons with the retrograde fluorescent tracer DiI that has been injected into the triceps surae muscles of control rats and rats with chronic occlusion of the femoral artery. Moreover, the isolated DRG cells will be identified by their transient expression of green fluorescent protein, the expression of which is driven by the proximal neuron specific promoter region of the TTX sensitive Na+ channel (NaV1.7) and TTX-resistant Na+ channel (NaV1.8). For the in vivo experiments, sympathetic responsiveness evoked by muscle contraction will be examined in control rats and rats with femoral artery occlusion. In both in vitro and in vivo experiments, particular attention will be paid to the effects of PICs, namely IL-6 and TNF-α, on the membrane and discharge properties of the afferent neurons. Also, the interplay between cytokine receptors and channels that control excitability of the thin fiber muscle afferents will be examined. Additionally, intracellular signaling pathways that are engaged during excitation by PICs will be examined. Our general hypothesis is that the higher levels of PIC receptors are induced in muscle DRG neurons after hindlimb blood flow occlusion in the rat model of PAD. This in turn alters the intracellular transduction pathways and the expression and function of Na+ channels, thereby leading to the augmented exercise pressor reflex. The proposed experiments are anticipated to provide new information regarding the mechanisms by which the excitability of the afferent arm of the exercise pressor reflex is affected by PICs in PAD.