Molecular Bases for Effects of Nicotine

描述(由申请者提供)：该项目的长期目标是确定慢性尼古丁暴露是否以及如何改变神经系统功能。该项目基于一个中心假设，即长期接触尼古丁会导致不同烟碱型乙酰胆碱受体(NAChR)亚型的数量和功能发生长期变化。这些影响可能与尼古丁依赖、烟草产品的使用、与烟草有关的疾病、预防、限制或停止烟草使用的治疗以及神经和精神疾病的治疗有关。初步研究结果表明，我们的假设是，慢性尼古丁暴露导致(I)nAChR功能持续失活和(Ii)nAChR数量增加，这是通过两个因果和机械上不同的翻译后过程。尼古丁作用的剂量和时间依赖关系被认为是nAChR亚型特有的，其他作用于改变nAChR数量和功能的药物的药理学特征也是如此。这个多层次项目的一个主要目的是确定暴露于尼古丁或相关物质对不同人类nAChR亚型功能的影响。另一个主要目的是确定这些药物对nAChR数量的影响。对于这两个目标，研究将涉及通过模型细胞系自然和/或异源表达的α1β1伽马三角洲(肌肉型)、(α3α5β4-(自主)、α4β2-(脑尼古丁结合)和α7(自主或脑神经毒素结合)nAChR)。将建立尼古丁细胞处理的时间(起效和恢复)和剂量依赖效应。还将使用其他药物单独或与尼古丁联合使用来获得时间和剂量分布，以评估它们是模拟还是阻断尼古丁的影响，nAChR功能活性将通过电生理记录和离子通量分析进行量化。放射配基结合和免疫分析将被用来定量nAChR，并评估它们的亚细胞分布和代谢。NAChR的化学反应和突变研究将用于确定尼古丁和其他药物的作用机制。 这些研究具有重要意义，因为它们将为慢性尼古丁在神经系统功能调节中的作用部位和机制提供新的视角。还将提供对尼古丁依赖的分子基础的见解。将确定受长期尼古丁暴露影响最大的特定nAChR亚型。此外，还将提供对阻断或模仿尼古丁作用的药物种类的见解。总的来说，这些知识将有助于开发治疗情绪、认知和其他神经系统疾病的策略。

DESCRIPTION (provided by applicant): The long-term goal of this project is to establish whether and how chronic nicotine exposure alters nervous system function. The project is based on the central hypothesis that extended exposure to nicotine induces long-lasting changes in numbers and function of diverse nicotinic acetylcholine receptor (nAChR) subtypes. These effects have potential relevance for nicotine dependence, use of tobacco products, tobacco-related diseases, treatments to prevent, limit or cease tobacco use, and therapies for neurological and psychiatric disorders. Preliminary findings suggest our hypothesis that chronic nicotine exposure causes both (i) a persistent inactivation of nAChR function and (ii) an increase in numbers of nAChR via two causally- and mechanistically-distinct, posttranslational processes. Dose- and time-dependence of nicotine's effects are postulated to be nAChR subtype-specific, as are pharmacological profiles for other drugs acting to alter nAChR numbers and function. One principal aim of this multi-layered project is to establish effects of exposure to nicotine or related substances on function of diverse, human nAChR subtypes. The other principal aim is to determine effects of those agents on numbers of nAChR. For both aims, studies will involve alpha1 beta1 gamma delta- (muscle-type), (alpha3 alpha5 beta4- (autonomic), alpha4 beta2-(brain nicotine-binding), and alpha7- (autonomic or brain neurotoxin-binding) nAChR expressed naturally and/or heterologously by model cell lines. Time (onset of effects and recovery)- and dose-dependent effects of cell treatment with nicotine will be established. Time and dose profiles will also be obtained using other drugs alone or in combination with nicotine to assess whether they mimic or block nicotine's effects, nAChR functional activity will be quantified by electrophysiological recording and ion flux assays. Radioligand binding and immuno- assays will be used to quantitate nAChR and to assess their subcellular distribution and metabolism. Chemical reactivity of nAChR and mutational studies will be among those used to establish mechanisms involved in effects of nicotine and other drugs. These studies are significant because they will provide new perspectives on sites and mechanisms of chronic nicotine action in the modulation of nervous system function. Insights will also be provided into molecular bases of nicotine dependence. Specific nAChR subtypes that are affected most powerfully by chronic nicotine exposure will be identified. Insights will also be provided into the kinds of drugs that block or mimic nicotine's actions. Collectively, this knowledge will benefit development of strategies to treat mood, cognitive, and other nervous system disorders.