Molecular Mechanisms of Nicotine Addiction and Extinction

描述（由申请人提供）：主要研究者的长期目标是确定尼古丁成瘾/依赖的分子机制。尼古丁成瘾的一个突出方面是，通常与尼古丁有关的线索对吸烟者的渴望和复发施加了强有力的控制。虽然已经有一些尝试实施线索灭绝来帮助戒烟，但缺乏对线索控制吸烟机制的了解阻碍了进展。本研究旨在阐明场所条件作用范式下小鼠尼古丁成瘾线索控制消失的分子机制。本R01应用验证了cgmp依赖性蛋白激酶亚型II （PKG-II）在中脑皮质边缘多巴胺系统沿线区域是尼古丁诱导的小鼠条件位置偏好（CPP）消退速度的决定因素的总体假设。我们发表的和初步的研究表明：a)尼古丁可以提高纹状体（伏隔核和尾壳核）中cGMP的浓度；b)尼古丁可以上调伏隔核和VTA中PKG的活性，它们分别代表了中皮质边缘多巴胺通路的靶点和起点；c)尼古丁导致纹状体（包括伏隔核）中PKG底物的磷酸化。d)小鼠在无药试验前几天尼古丁CPP增强，随后消失；e) PKG-II敲除（KO）小鼠尼古丁CPP的消失加速。拟议的研究依赖于小鼠大脑中PKG水平的遗传操纵。Specific Aim 1将检验PKG-II水平而非PKG-I水平决定尼古丁CPP消失速率的假设。PKG-I和PKG-II野生型（WT）、杂合型（HT）和KO小鼠将被采用。此外，我们将确定PKG亚型在尼古丁CPP消失过程中发挥作用的确切行为过程，以及PKG亚型在其他成瘾物质的线索控制消失中发挥的一般作用。特异性目标2将确定PKG可能调节尼古丁CPP消失的特定大脑区域。我们将使用慢病毒载体在PKG-II（或PKG-I） KO小鼠中局部恢复PKG-II（或PKG-I）。此外，我们将在灭绝期间尼古丁CPP的维持与沿中脑皮质边缘多巴胺通路的选定脑区域的PKG活动相关联。拟议研究的结果意义重大，因为它将为吸烟者提供可用于设计治疗方案的证据。在特定脑位点中发现PKG亚型与尼古丁成瘾的线索控制有关，这将是提高我们微调分子靶点的能力的重要一步，以防止线索引发的吸烟复发和加速灭绝。如果这种激酶在其他成瘾物质消失过程中的线索控制中被证明是重要的，那么这些研究的结果将对其他形式成瘾的线索触发复发的治疗具有更广泛的意义。最近的估计表明，全世界约有13亿吸烟者，每年有500万人死于烟草使用。目前可用的治疗方法在帮助戒烟方面并不有效，部分原因是人们对尼古丁成瘾消退背后的确切神经机制仍然知之甚少。拟议的研究旨在填补这一知识空白。

DESCRIPTION (provided by applicant): The long-term objective of the principal investigator is to ascertain the molecular mechanisms underlying nicotine addiction/dependence. One salient aspect of nicotine addiction is that cues that are usually associated with nicotine exert a powerful control over craving and relapse in smokers. Although some attempts have been made to implement cue extinction to aid smoking cessation, lack of knowledge regarding the mechanisms of cue control of smoking has hampered progress. The proposed studies are designed to elucidate the molecular mechanisms underlying extinction of cue control of nicotine addiction in the place conditioning paradigm in mice. This R01 application tests the overall hypothesis that cGMP-dependent protein kinase subtype II (PKG-II), in regions along the mesocorticolimbic dopamine system, is a determinant of the rate of extinction of nicotine-induced conditioned place preference (CPP) in mice. Our published and preliminary studies show that a) nicotine elevates the concentration of cGMP in the striatum (nucleus accumbens and caudate-putamen), b) nicotine can up-regulate PKG activity in the nucleus accumbens and the VTA, which represent a target and the origin of the mesocorticolimbic dopamine pathway, respectively, c) nicotine causes phosphorylation of a PKG substrate in the striatum (including the nucleus accumbens), d) mice exhibit intensified nicotine CPP on the first few drug-free test days and extinction thereafter, and e) extinction of nicotine CPP is accelerated in PKG-II knockout (KO) mice. The proposed studies rely on genetic manipulation of PKG levels in the mouse brain. Specific Aim 1 will test the hypothesis that the level of PKG-II, but not PKG-I, determines the rate of extinction of nicotine CPP. PKG-I and PKG-II wild-type (WT), heterozygous (HT), and KO mice will be employed. Moreover, we will determine the exact behavioral processes in which PKG subtypes play a role during extinction of nicotine CPP and the general role played by PKG subtypes in extinction of cue control by other addictive substances. Specific Aim 2 will identify the specific brain region(s) in which PKG may act to regulate extinction of nicotine CPP. We will use a lentiviral vector to locally restore PKG-II (or PKG-I) in PKG-II (or PKG-I) KO mice. Moreover, we will correlate maintenance of nicotine CPP during extinction with PKG activities in selected brain regions along the mesocorticolimbic dopamine pathway. The outcome of the proposed studies is significant, as it will provide evidence that can be used to devise therapeutic options for smokers. Identification of a PKG subtype in specific brain loci, in connection with cue control of nicotine addiction, would be an important step toward enhancing our ability to fine-tune molecular targets to prevent cue-triggered smoking relapse and accelerate extinction. If this kinase turns out to be important for cue control of other addictive substances during extinction, the outcome of these studies will have a much broader implication for treatment of cue-triggered relapses in other forms of addiction. Recent estimates indicate that there are approximately 1.3 billion smokers world-wide and 5 million deaths are attributable to tobacco use annually. Currently available therapies are not effective in aiding cessation, partly because the exact neuronal mechanisms underlying extinction of nicotine addiction are still poorly understood. The proposed studies are designed to fill in this knowledge gap.