RNAi-medicated inhibition of BDNF expression and alcohol-drinking behavior

RNAi 抑制 BDNF 表达和饮酒行为

• 批准号: 7340147
• 负责人: QINGSHAN YAN
• 金额: $ 7.75万
• 依托单位: UNIVERSITY OF ILLINOIS AT CHICAGO
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-01-15 至 2008-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7340147
• 关键词: Address; Aftercare; Alcohol consumption; Alcoholism; Alcohols; Animal Model; Animals; Area; Brain; Brain region; Brain-Derived Neurotrophic Factor; Data; Dependency; Development; Dopamine; Drug Addiction; Enzyme-Linked Immunosorbent Assay; Financial compensation; Gene Expression; Gene Silencing; Genes; Genetic; Genetically Modified Animals; Goals; Growth Factor; Hippocampus (Brain); Human; Hyperphagia; Immunohistochemistry; Impairment; In Vitro; Indiana; Infusion procedures; Lentivirus Vector; Link; Masks; Mediating; Messenger RNA; Methods; Mus; Neuraxis; Neuromodulator; Neurons; Neuropeptides; Neurotransmitters; Nucleus Accumbens; Numbers; Pathway interactions; Peptides; Phenotype; Play; Prosencephalon; Proteins; RNA Interference; Rattus; Regulation; Reporting; Research Personnel; Rewards; Role; Seizures; Serotonin; Subfamily lentivirinae; System; Tissues; Tryptophan 5-monooxygenase; Tyrosine 3-Monooxygenase; Universities; Validation; Ventral Tegmental Area; Wistar Rats; alcohol effect; alcohol reward; dopaminergic neuron; drinking behavior; fetal; gene function; in vivo; mesolimbic system; neuronal survival; null mutation; preference; progenitor; research study; small hairpin RNA; tool; vector

The goal of the study is to link brain-derived neurotrophic factor (BDNF) to alcohol-drinking behavior. It is hypothesized that decreased BDNF in the mesolimbic dopamine (DA) pathway, a brain circuit that has been implicated in alcohol's rewarding effects, is associated with high alcohol intakes. This hypothesis is prompted by our finding showing innate deficiencies of BDNF in the nucleus accumbens (NACC) of alcohol-preferring (P) rats compared with alcohol-nonpreferring (NP) rats. Previous studies performed in heterozygous BDNF (+/-) mice have shown the relationship between BDNF and alcohol consumption, but they cannot identify brain area(s) in which BDNF may play a role in regulating alcohol intakes. In addition, the approach of genetically modifying animals for target validation is often limited by developmental adaptation and genetic compensation that may mask the establishment of a clear phenotype. Recently, RNA interference (RNAi) has become a valuable tool for investigating gene function. The applicant has generated a lentiviral vector that is capable of delivering and expressing short hairpin RNA that targets BDNF mRNA. This vector has been proved to be effective in silencing the BDNF gene both in vitro and in vivo in our preliminary study. To determine whether RNAi-mediated inhibitions of BDNF expression in the VTA and/or NACC increase alcohol intakes, this lentivirus and several control substances will be bilaterally infused into the VTA or the NACC of groups of the Wistar rat, the progenitor from which the P/NP rats were originally derived. Then, the effects of the infusion on alcohol consumptions and non-alcohol tastant preference are determined by a two-bottle free-choice method and compared among different treatment groups. BDNF expression at the protein level in each brain area after the treatments is also assessed via the enzyme-linked immunosorbent assay and immunohistochemistry to see whether reductions of BDNF expression are associated with increased alcohol intakes. This study would provide more direct evidence for the role of BDNF in alcohol intakes and specifically addresses whether reductions of BDNF in the mesolimbic DA pathway are associated with increased alcohol consumptions. Since there are no reports regarding silencing the BDNF gene in vivo to date, the establishment of the approach to inhibit BDNF expression, particularly in the mesolimbic DA system of intact animals, would benefit researchers in drug addiction including alcoholism.

这项研究的目的是将脑源性神经营养因子（BDNF）与饮酒行为联系起来。是 假设中脑边缘多巴胺（DA）通路中的BDNF减少，这是一种大脑回路， 与酒精的奖励效应有关，与大量饮酒有关。这一假设是由 我们的发现表明，嗜酒精的伏隔核（NACC）中的脑源性神经营养因子先天缺陷 (P)酒精非偏好（NP）大鼠。先前在杂合子BDNF中进行的研究 （+/-）小鼠已经显示了BDNF和饮酒之间的关系，但他们不能确定 BDNF可能在调节酒精摄入方面发挥作用的脑区。此外， 用于靶标验证的遗传修饰动物通常受到发育适应和遗传特性的限制。 补偿，可能掩盖了明确的表型的建立。最近，RNA干扰（RNAi） 已成为研究基因功能的重要工具。申请人已经产生了慢病毒载体， 能够传递和表达靶向BDNF mRNA的短发夹RNA。这个载体有 在我们的初步研究中，证明了在体外和体内都能有效地沉默BDNF基因。到 确定RNAi介导的BDNF在VTA和/或NACC中表达的抑制是否增加酒精 摄入量，这种慢病毒和几种对照物质将被双侧输注到VTA或NACC， Wistar大鼠组，P/NP大鼠最初来源的祖先。那么， 对酒精消耗和非酒精促味剂偏好的输注通过两瓶 采用自由选择法，并在不同治疗组之间进行比较。BDNF在蛋白水平的表达 还通过酶联免疫吸附测定评估治疗后每个脑区的 免疫组化观察BDNF表达的减少是否与酒精增加有关 摄入量这项研究将为BDNF在酒精摄入中的作用提供更直接的证据， 具体解决了中脑边缘DA通路中BDNF的减少是否与 增加酒精消费。由于没有关于在体内沉默BDNF基因以 迄今为止，已建立了抑制BDNF表达的方法，特别是在中脑边缘DA中 完整的动物系统，将有利于研究药物成瘾，包括酗酒。