Opiate abuse and sex steroids influence neuroAIDS pathology

 描述（由申请人提供）：与一般人群相比，人类免疫缺陷病毒（HIV）感染者经历与更大的运动/情绪/认知障碍（统称为"神经艾滋病"）相关的脑部病理学，这些影响在阿片类药物滥用者中加剧。此外，存在性别差异，与滥用阿片类药物的男子相比，滥用阿片类药物的妇女感染艾滋病毒的风险更大;然而，一旦感染，一些妇女可能比男子经历更少的神经艾滋病。我们在小鼠中概括了这些作用，并开始阐明性类固醇激素与阿片类药物相互作用，以保护神经艾滋病。目前的建议将开始揭示性类固醇与阿片类药物和艾滋病毒相互作用的影响和机制，在翻译的方法，利用培养的 鼠和人神经细胞，以及转基因全动物鼠模型。这个独立之路奖（K99/R00）的目标是为Jason巴黎博士提供培训，以辨别性类固醇环境可能影响神经艾滋病病理学的阿片类药物/HIV相互作用的细胞/分子机制。巴黎博士将接受小鼠和人类神经细胞培养和成像技术的高级培训，用于检测亚致死神经元致病性、细胞内离子和线粒体膜完整性。在K99培训阶段，巴黎博士将在Kurt豪瑟博士（初级导师）的指导下工作，以检查性类固醇可能对阿片类药物与HIV-1病毒毒素、达特（Aim 1a）或gp 120（Aim 1b）在小鼠神经共培养物中的协同作用产生的潜在保护作用。将通过免疫细胞化学检测存活/退化的神经元、星形胶质细胞和小胶质细胞，将评估生物细胞素填充的纹状体中型棘神经元（MSN）的亚致死形态变化，并将评估上清液中细胞因子/趋化因子、活性氧和亚硝酸盐的产生。小鼠（目的2a和2b）和人（目的2c）神经培养物中，将评估类固醇对阿片类药物与达特（2a）、gp120（2b）或X4-/R5-嗜性HIV（2c）相互作用的保护作用对MSN形态学、MSN索马和树突内[Ca] i和[Na] i的离子成像的影响，2 ++和线粒体膜不稳定（通过罗丹明123荧光成像）。在R00独立阶段，巴黎博士将使用条件性表达中枢HIV-1达特（Aim 3a）或组成性表达中枢gp120（Aim 3b）的整体动物模型，检查中枢类固醇形成对体内阿片类药物/HIV蛋白相互作用的保护作用。将评估雄性和雌性小鼠的神经AIDS样运动、情感和认知行为。将评估纹状体组织目标1中描述的终点。最后，将在从达特或gp120转基因小鼠获得的鼠神经共培养物中检查类固醇、阿片类药物和HIV蛋白之间重叠的药效学靶点的影响（目的3c）。将用药理学拮抗剂预处理培养物，并按照目的1和2进行检查。这项建议是及时的，提供了新的培训，并将使一个创新和独立的研究计划的发展。

 DESCRIPTION (provided by applicant): Individuals with human immunodeficiency virus (HIV) experience brain pathology associated with greater motor/mood/cognitive disorders (collectively termed "neuroAIDS") than the general population and these effects are exacerbated among opiate abusers. Moreover, gender differences exist with opiate-abusing women at greater risk to contract HIV compared to opiate-abusing men; yet, some women may experience lesser neuroAIDS symptomology than men once infection occurs. We recapitulated these effects in mice and have begun to elucidate the sex steroid hormones that interact with opiates to confer protection to neuroAIDS. The present proposal will begin to reveal the effects and mechanisms of sex steroid interactions with opiates and HIV in a translational approach that utilizes cultured murine and human neural cells, as well as transgenic whole-animal murine models. The goal of this Pathway to Independence Award (K99/R00) is to provide training to Dr. Jason Paris to discern the cellular/molecular mechanisms by which sex steroid milieu may influence opiate/HIV interactions for neuroAIDS pathology. Dr. Paris will receive advanced training in murine and human neural cell culture and imaging techniques for detection of sub lethal neuronal pathogenicity, intracellular ions, and mitochondrial membrane integrity. In the K99 training phase, Dr. Paris will work under the tutelage of Dr. Kurt Hauser (Primary Mentor) to examine the potentially-protective effects that sex steroids may exert over synergistic actions between opiates and the HIV-1 virotoxins, Tat (Aim 1a) or gp120 (Aim 1b) in murine neural co-cultures. Viable/degenerating neurons, astrocytes, and microglia will be detected via immunocytochemistry, biocytin-filled striatal medium spiny neurons (MSNs) will be assessed for sub lethal changes in morphology, and supernatants will be assessed for production of cytokines/ chemokines, reactive oxygen species, and nitrites. In murine (Aims 2a and 2b) and human (Aim 2c) neural cultures, steroid protection against opiate interactions with Tat (2a), gp120 (2b), or X4-/R5-tropic HIV (2c) will be assessed for effects on MSN morphology, ion imaging for [Ca] i and [Na] i within MSN soma and dendrites, 2+ + and mitochondrial membrane destabilization (via Rhodamine 123 fluorescence imaging). In the R00 independent phase, Dr. Paris will examine the protective effects of central steroid formation on opiate/HIV protein interactions in vivo using whole-animal models that conditionally-express central HIV-1 Tat (Aim 3a) or constitutively-express central gp120 (Aim 3b). Male and female mice will be assessed for neuroAIDS-like motor, affective, and cognitive behavior. Striatal tissue will be assessed for endpoints described in Aim 1. Lastly, the influence of pharmacodynamic targets that overlap between steroids, opiates, and HIV proteins will be examined in murine neural co-cultures obtained from Tat- or gp120-transgenic mice (Aim 3c). Cultures will be pretreated with pharmacological antagonists and examined as in Aims 1 and 2. This proposal is timely, provides novel training, and will enable the development of an innovative and independent research program.