Using Human iPSC Models to Determine the Mechanism of Inflammation-Induced Disruption of Dopamine Neurotransmission

项目总结 目前的提议将使用创新的干细胞技术来探索炎症的机制(S) 影响多巴胺(DA)神经传递，特别关注炎症对突触前DA的影响 水泡。炎症被认为在约30%的抑郁症患者中起着关键的病理生理作用，而且 与奖赏回路的影响有关，导致动机不足，最终导致快感丧失。快感缺乏症 是抑郁症的核心和致残症状，抑郁症是全球残疾的主要原因。研究项目： 实验动物和人类表明，大脑炎症的主要靶点之一是大脑中的多巴胺。 腹侧纹状体，一个皮质下的大脑区域，已被证明是唯一可以接触到外周的区域 炎症介质，包括白介素6，通过破坏血脑屏障。活体内 非人灵长类和啮齿类动物的微透析证实了炎性细胞因子的作用 包括IL-6在内的研究减少了人类细胞外DA的可获得性和释放，以及神经成像研究 在注射炎症细胞因子干扰素-α后，纹状体DA转换率降低。 然而，目前尚不清楚的是炎症的具体细胞和分子机制。 IL-6等细胞因子可干扰DA的神经传递。我们的初步发现表明，在体外IL-6治疗 直接表达IL-6受体的人诱导多能干细胞(HiPSC)来源的DA神经元 减少DA的可获得性并下调突触囊泡相关通路的基因表达 功能。这些效应发生在没有细胞毒性的情况下，基因表达的变化是 由美国食品和药物管理局批准的药物巴利替尼逆转，该药物由埃默里公司开发，通过 抑制Janus激酶1和2。在当前的项目中，我们的目标是进一步阐明 IL-6通过人IPSC来源的DA神经元影响DA神经传递。具体来说，我们将确定 IL-6对海马神经元DA中巴利替尼存在或不存在时突触囊泡功能的影响 神经元(目标1)。我们将进一步确定IL-6对DRD2基因选择性剪接的影响，这在 TURN可调节DA神经元的突触囊泡功能(目标2)。这些研究还将包括DA神经元 表达与抑郁症相关的DRD2变异体rs1076560，该变异体与交替DRD2相关 拼接。综上所述，拟议的实验将为使用 干细胞技术揭示炎症影响DA神经传递、脱落的机制 关于精神障碍中DA系统缺陷的致病机制的新认识，同时提供了 药物开发平台与埃默里的药物发现流水线保持一致。

PROJECT SUMMARY The current proposal will use innovative stem cell technology to explore the mechanism(s) by which inflammation affects dopamine (DA) neurotransmission, with a special focus on inflammation's effects on presynaptic DA vesicles. Inflammation is believed to play a pivotal pathophysiologic role in ~30% of depressed patients and is associated with effects on reward circuitry, leading to motivational deficits and ultimately anhedonia. Anhedonia is a core and disabling symptom of depression, which is the leading cause of disability worldwide. Studies in laboratory animals and humans indicate that one of the major targets of inflammation in the brain is DA in the ventral striatum, a subcortical brain region that has been shown to be uniquely accessible to peripheral inflammatory mediators including interleukin (IL)-6 through disruption in the blood brain barrier. In vivo microdialysis in non-human primates and rodents demonstrate that administration of inflammatory cytokines including IL-6 reduces extracellular DA availability and release, and neuroimaging studies in humans demonstrate reduced striatal DA turnover following administration of the inflammatory cytokine interferon-alpha. What remains unknown, however, are the specific cellular and molecular mechanisms by which inflammatory cytokines such as IL-6 disrupt DA neurotransmission. Our preliminary findings indicate that in vitro IL-6 treatment of human induced pluripotent stem cell (hiPSC)-derived DA neurons (which express IL-6 receptors) directly decreases DA availability and downregulates gene expression in pathways associated with synaptic vesicular function. These effects occurred in the absence of cellular toxicity, and the gene expression changes were reversed by baricitinib, an FDA-approved drug that was developed at Emory and blocks IL-6 signaling through inhibition of Janus Kinases 1 and 2. In the current project, we aim to further elucidate the mechanisms by which IL-6 affects DA neurotransmission using human iPSC-derived DA neurons. Specifically, we will determine the impact of IL-6 on synaptic vesicular function in the presence or absence of baricitinib in hiPSC-derived DA neurons (Aim 1). We will further determine the impact of IL-6 on alternative splicing of the DRD2 gene, which in turn can regulate synaptic vesicular function in DA neurons (Aim 2). These studies will also include DA neurons expressing the depression-associated DRD2 variant rs1076560, which is associated with alternate DRD2 splicing. Taken together, the proposed experiments will provide an important proof of principle for the use of stem cell technology to reveal the mechanisms of inflammation's effects on DA neurotransmission, shedding new light on pathogenic mechanisms underlying DA system deficits in psychiatric disorders, while providing a platform for drug development aligned with Emory's drug discovery pipeline.