Glial-neuronal interactions underlying the molecular feedback between HIV viral p

描述(由申请人提供)：这项研究的长期目标是描绘人类免疫缺陷病毒1型(HIV)阳性个人滥用甲基苯丙胺(METH)等物质增加的可能机制。在这个为期两年的探索性项目中，我们建议专门研究甲基苯丙胺(一种成瘾的精神刺激剂)和HIV-1病毒蛋白在大鼠脑内神经胶质细胞炎症诱导中的协同作用。在中枢神经系统(CNS)中，胶质细胞被HIV-1病毒感染，导致神经炎症。大脑炎症增加与神经退行性疾病和行为障碍有关，如HIV-1诱导的神经变性(HAND)和HIV-1诱导的痴呆(HAD)。此外，艾滋病毒感染者滥用冰毒会导致病毒载量增加和严重的脑部疾病。在这些研究中，我们将使用非传染性HIV-1转基因(HIV-1TG)大鼠和原代细胞培养模型。HIV-1TG大鼠携带Gag/Pol缺失的前病毒基因，并表达9种HIV-1病毒蛋白中的7种。这只HIV-1TG大鼠出现了人类艾滋病毒疾病的临床表现，因此，它模仿了接受抗逆转录病毒治疗的艾滋病毒患者持续存在艾滋病毒-1蛋白所导致的感染。最近，我们利用PCR芯片分析表明，与F344对照组大鼠相比，HIV-1TG大鼠脑内促炎细胞因子IL-1以及CCL2、CCl3和Ccl7趋化因子的增加幅度更大。我们还证明，在HIV-1TG大鼠的前额叶皮质中，多巴胺D1受体(D1R)的表达增加，根据刻板印象的头部运动，这些动物具有更强的冰毒诱导的行为敏感化。根据文献和我们以前的数据，我们假设在HIV-1病毒蛋白存在的情况下，滥用冰毒等物质会增强神经胶质细胞的激活，从而影响神经元，并增加多巴胺系统的活性，从而增加冰毒的摄入量。为了验证这一假设，我们将首先使用免疫组织化学方法检测HIV-1TG大鼠在接触冰毒和不接触冰毒的情况下，大脑中神经胶质细胞和神经细胞的相互作用。然后我们将确定神经递质活性的基因表达变化，特别是多巴胺能系统，以及与神经炎症相关的标记物。我们还将分离HIV-1TG大鼠的原代胶质细胞和神经细胞，并检测体外暴露于冰毒后多巴胺能和其他神经递质途径中基因表达的变化，以及炎症、凋亡和氧化应激标志物的变化。我们的具体目标包括：1)确定冰毒和HIV病毒蛋白对HIV-1TG大鼠脑内神经胶质和神经元激活、神经炎症和神经递质调节的影响；2)利用从HIV-1TG大鼠脑内分离的原代神经胶质和神经元细胞，在体外模型中确定冰毒和HIV病毒蛋白对神经胶质和神经元激活、神经炎症和神经递质调节的影响。据我们所知，这是第一个探索冰毒对胶质-神经元相互作用影响的项目。 以及随后使用HIV-1TG大鼠和从HIV-1TG大鼠分离的原代细胞培养对多巴胺能系统的影响。这项研究的数据将阐明艾滋病毒阳性者滥用冰毒等物质增加的可能细胞和分子机制，并有助于阐明与冰毒滥用和神经艾滋病相关的神经胶质-神经元相互作用。我们的探索性研究还可以带来更好的治疗方法，最终将使艾滋病毒-1患者和药物滥用者受益。因此，建议的研究具有很高的临床相关性，并将显著有助于理解和治疗与药物滥用、艾滋病毒感染和艾滋病相关的神经系统并发症。

DESCRIPTION (provided by applicant): The long-term goal of this research is to delineate the possible mechanisms underlying the increased abuse of substances, such as methamphetamine (METH), by individuals who are human immunodeficiency virus-1 (HIV)-positive. In this two-year exploratory project, we propose to specifically examine the synergistic effects of methamphetamine (METH), an addictive psychostimulant, and HIV-1 viral proteins on glial-neuronal induction of inflammation in the rat brain. In the central nervous system (CNS), the glial cells are infected by the HIV-1 virus, causing neuroinflammation. Increased inflammation in the brain has been correlated with neurodegenerative diseases and behavioral disorders, such as HIV-1 induced neurodegeneration (HAND) and HIV-1 induced dementia (HAD). Moreover, METH abuse in the HIV-infected individual can lead to increased viral loads and severe brain-related disorders. For these studies, we will use both the non-infectious HIV-1 transgenic (HIV-1Tg) rat and primary cell culture models. The HIV-1Tg rat carries a gag/pol deleted provirus gene, and expresses 7 out of 9 HIV-1 viral proteins. This HIV-1Tg rat develops clinical manifestations of human HIV disease, and, thus, mimics the infection that results from the persistent presence of HIV-1 proteins in HIV patients given anti-retroviral therapy. Recently, using PCR array analysis, we showed that the pro- inflammatory cytokine, IL-1¿, as well as the Ccl2, Ccl3, and Ccl7 chemokines, are increased to a greater extent in the brain of the HIV-1Tg rat compared to the F344 control rat. We have also demonstrated that there is elevated dopamine D1 receptor (D1R) expression in the prefrontal cortex of HIV-1Tg rats, and that these animals have greater METH-induced behavioral sensitization, as assessed by stereotypical head movement. Based on the literature and our previous data, we hypothesize that the abuse of substances, such as METH, in the presence of HIV-1 viral proteins, enhances glial activation, which affects neurons, and increases the activity of the dopaminergic system, thereby increasing the intake of METH. To test this hypothesis, we will first examine the glial-neuronal cell interaction in the brain of the HIV-1Tg rat with and without exposure to METH using immunohistochemistry. We will then identify gene expression changes in neurotransmitter activity, in particular, the dopaminergic system, as well as neuroinflammation-related markers. We will also isolate primary glial and neuronal cells from HIV-1Tg rats and examine gene expression changes in the dopaminergic and other neurotransmitter pathways, as well as inflammation, and apoptotic and oxidative stress markers after in vitro exposure to METH. Our specific aims include: 1) To determine the effects of METH and HIV viral proteins on glial and neuronal activation, neuroinflammation, and neurotransmitter modulation in the brain of the HIV-1Tg rat, and 2) To determine the effects of METH and HIV viral proteins on glial and neuronal activation, neuroinflammation, and neurotransmitter modulation in an ex vivo model using primary glial and neuronal cells isolated from the HIV- 1Tg rat brain To our knowledge, this proposal represents the first project to explore the effects of METH on glial-neuronal interactions and the subsequent effects on the dopaminergic system using the HIV-1Tg rat and primary cell cultures isolated from the HIV-1Tg rat. The data from this research will shed light on possible cellular and molecular mechanisms underlying the increased abuse of substances such as METH by HIV-positive individuals, and help to elucidate the glial-neuronal interactions associated with METH abuse and neuroAIDS. Our exploratory studies can also lead to better treatment approaches that will ultimately benefit both HIV-1 patients and substances abusers. Thus, the proposed studies have high clinical relevance and will contribute significantly to the understanding and treatment of neurological complications associated with substance abuse, HIV infection, and AIDS.