Mechanisms of TNFa Enhancement of Nicotinic Receptor Upregulation

摘要 尼古丁通过与神经元烟碱乙酰胆碱相互作用对细胞功能产生影响 受体（nAChR）。这些受体中的一些，特别是由a4 b2亚基组成的那些，对持续的 通过上调过程的配体暴露，如通过高密度的显著增加所定义的， 亲和尼古丁结合位点。上调的生理后果与许多 不同的过程，从成瘾到病理生理过程，有助于早期阶段的 老年痴呆症。一个令人兴奋的和相对较新的作用nAChRs正在出现的相互作用方面， 促炎细胞因子事实上，尼古丁的抗炎特性，通过各种 nAChR现在被广泛报道影响许多炎性病因学疾病。但相对 关于控制炎症的机制：nAChR相互作用知之甚少。的总目标 这个建议是定义细胞内的机制，调节关键的亲， 炎性细胞因子和nAChR亚型的确定组合。 具体目标1.假设：TNF α激活p38 MAPK通路和/或IL-1b激活PKA 途径增强a4 b2表达，并且这些被涉及PI 3 K/Akt信号传导的途径拮抗。在 为此，我们将对TNF α和/或IL-1b启动的细胞内途径进行定义， 尼古丁介导的nAChRa 4 b2上调。 具体目标2.假设：α 4的直接磷酸化和其他nAChR亚基的存在 改变介导a4 b2上调增强的促炎细胞因子信号。这一目标将 确定以下的影响：1）修饰α 4中的PKA磷酸化位点; 2）引入（或缺失）其它 nAChR（特别是α 7或α 5）;和3）确定TNF α或IL-1b细胞内信号传导的作用， 在培养的神经元中增强A4 B2上调。 具体目标3.假设：CNS细胞（如神经元和小胶质细胞（Mg））之间的相互作用 直接的结果，整体炎症：nAChR反应。神经元和小胶质细胞将在 不同的比例来剖析旁分泌，自分泌或外分泌相互作用的相对贡献， 在混合细胞系统中调节TNF α和IL-1b介导的α 4 β 2上调增强。

ABSTRACT Nicotine imparts its effects on cellular function through interaction with neuronal nicotinic acetylcholine receptors (nAChR). Some of these receptors, especially those composed of a4b2 subunits, respond to sustained ligand exposure through the process of upregulation as defined by a substantial increase in the density of high affinity nicotine binding sites. The physiological consequences of upregulation have been linked to many diverse processes ranging from addiction to pathophysiological processes contributing to early phases of Alzheimers disease. An exciting and relatively new role for nAChRs is emerging with regard to the interaction with pro-inflammatory cytokines. In fact, the anti-inflammatory properties of nicotine, acting through various nAChRs, is now widely reported to influence many diseases of inflammatory etiology. However, relatively little is known about the mechanisms controlling the inflammatory:nAChR interaction. The overall goal of this proposal is to define intracellular mechanisms that regulate the interactions between key pro- inflammatory cytokines and defined combinations of nAChR subtypes. SPECIFIC AIM 1. Hypothesis: TNFa-activation of the p38MAPK pathway and/or IL-1b activation of PKA pathways enhance a4b2 expression and these are antagonized by pathways involving PI3K/Akt signaling. In this Aim we will pursue the definition of intracellular pathways initiated by TNFa and/or IL-1b to enhance nicotine-mediated upregulation of nAChRa4b2. SPECIFIC AIM 2. Hypothesis: Direct phosphorylation of a4 and the presence of other nAChR subunits modify the pro-inflammatory cytokine signals mediating enhanced upregulation of a4b2. This Aim will determine the impact of: 1) modifying PKA phosphorylation sites in a4; 2) introducing (or deleting) other nAChRs (specifically a7 or a5); and 3) determining effects of TNFa or IL-1b intracellular signaling leading to enhanced a4b2 upregulation in cultured neurons. SPECIFIC AIM 3. Hypothesis: Interactions between cells of the CNS such as neurons and microglia (Mg) direct the outcome of the overall inflammatory:nAChR response. Neurons and microglia will be co-cultured in different ratios to dissect the relative contribution of paracrine, autocrine or juxtacrine interactions in modulating the TNFa and IL-1b-mediated enhancement of a4b2-upregulation in a mixed cell system.