Modulation of Nicotine Reward-Associated Behaviors by MicroRNAs

说明(申请人提供)：烟草使用造成的不良健康后果是全世界可预防的死亡的主要原因，每年造成约600万人死亡。烟草的成瘾成分是尼古丁，一种三级生物碱，结合并激活烟碱型乙酰胆碱受体(NAChRs)，这是一种配体门控离子通道，通常由内源性神经递质乙酰胆碱(ACh)激活。神经元nAChRs是由不同的受体亚基组合而成的五聚体，不同的亚基组合赋予受体亚型不同的亲和力和功能。在哺乳动物神经元nAChRs中已鉴定出11个亚基，即α2-α7、α9、α10和？2-？4。有趣的是，长期接触尼古丁或香烟会导致大脑中nAChRs的上调，包括与奖赏和成瘾有关的中皮质边缘多巴胺能(DAer)通路中的结构。虽然还不完全清楚，但尼古丁诱导的nAChRs上调被认为是通过改变神经网络而导致成瘾的，可能导致对尼古丁的耐受性增加或敏感性改变。虽然已经提出了许多关于nAChR上调的机制，但人们普遍认为这种现象是多种形式的转录后调控造成的。目前，关于microRNAs(MiRNAs)对哺乳动物nAChR亚单位表达的转录后调控知之甚少。microRNAs是一种小的单链RNA分子，起着基因表达的负调控作用。然而，有新的证据表明，在尼古丁暴露的不同类型的啮齿动物组织中，miRNA的表达减少。此外，最近的研究发现，对包括可卡因在内的各种滥用药物的暴露，miRNA的失调会影响药物的奖赏性质，并改变与成瘾相关的行为。我们最近产生的初步数据表明，一种涉及miRNAs的新的调控机制可能在尼古丁介导的nAChRs上调中发挥作用。我们实验室的初步实验已经鉴定了几个被预测靶向nAChR亚单位基因转录本的miRNA，特别是分别靶向α4和？2转录本的miR-494和miR-542-3p。在目标1中，我将确定在原代培养的中脑神经元中，α4和/或2是否受到miR-494和/或miR-542-3p的调节。在目标2中，我将确定miR-494和/或miR-542-3p是否是小鼠尼古丁奖赏相关行为的调节器。通过这些目的，我希望更好地理解miR-494和miR-542-3p在尼古丁奖赏相关行为中的作用，可能为戒烟辅助药物的开发提供新的靶点。

DESCRIPTION (provided by applicant): Adverse health consequences of tobacco use are the leading cause of preventable mortality worldwide, resulting in approximately 6 million deaths per year. The addictive component of tobacco is nicotine, a tertiary alkaloid that binds and activates nicotinic acetylcholine receptors (nAChRs), ligand-gated ion channels that are normally activated by the endogenous neurotransmitter acetylcholine (ACh). Neuronal nAChRs are pentamers assembled from various combinations of receptor subunits and different subunit combinations confer different affinities and functionalities to the receptor subtypes. Eleven subunits, α2- α7, α9, α10 and ß2-ß4, have been identified in mammalian neuronal nAChRs. Interestingly, chronic nicotine or cigarette smoke exposure results in the upregulation of nAChRs in the brain, including structures within the mesocorticolimbic dopaminergic (DAergic) pathway that is implicated in reward and addiction. While not completely understood, nicotine- induced upregulation of nAChRs is thought to contribute to addiction by altering the neural network, possibly resulting in increased tolerance or altered sensitivity to nicotine. While there are many proposed mechanisms for nAChR upregulation, it is largely believed that multiple forms of posttranscriptional regulation is responsible for this phenomenon. Currently, there is not much known about posttranscriptional regulation of mammalian nAChR subunit expression by microRNAs (miRNAs), small single stranded RNA molecules that function as negative regulators of gene expression. However, there is emerging evidence that miRNA expression is decreased in various rodent tissue types in response to nicotine exposure. In addition, recent studies have found that miRNA dysregulation in response to exposure to various drugs of abuse, including cocaine, can influence rewarding properties of the drug and alter addiction-associated behaviors. We have recently generated preliminary data suggesting that a novel regulatory mechanism involving miRNAs may be at work in the nicotine-mediated upregulation of nAChRs. Preliminary experiments from our lab have identified several miRNAs that are predicted to target nAChR subunit mRNA transcripts, in particular miR-494 and miR-542-3p that target α4 and ß2 transcripts, respectively. In Aim 1, I will determine if α4 and/or ß2 are modulated by miR- 494 and/or miR-542-3p in primary midbrain neuronal cultures. In Aim 2, I will determine if miR-494 and/or miR- 542-3p are modulators of nicotine reward-associated behavior in mice. Through these aims, I hope to achieve a better understanding of the role of miR-494 and miR-542-3p in nicotine reward-associated behaviors, possibly revealing new targets for the development of tobacco cessation aids.