Opiate and HIV-induced neuropathogenesis in hippocampus: impact on excitatory/inhibitory dysregulation

项目摘要。 近一半的HIV+患者会出现HIV相关的神经认知障碍症状， 阿片类药物成瘾进一步加剧。阿片类药物通过直接（即，兴奋性毒性） 和间接（即，神经胶质炎症）机制。然而，虽然某些大脑区域，如纹状体， 树突肿胀和变性，海马表现出更微妙的病理：短暂的改变， 细胞内钙离子（[Ca 2 +]i）和钠离子（[Na+]i）减少LTP，并降低特定的CA 1中间神经元 亚型，在没有严重神经变性的情况下。我们的初步工作表明，GABAA激动 阻止Tat诱导的[Ca 2 +]i瞬变，而GABAA拮抗作用延长吗啡/Tat介导的[Ca 2 +]i瞬变。 [Ca2+]i升高。我们假设吗啡通过其对神经元的抑制作用加剧HIV诱导的神经元功能障碍。 影响海马体中的抑制性微电路，导致海马体中的净中断。 CA 1锥体细胞的兴奋/抑制平衡和最终的神经元损伤。为了研究这个问题，我们建议 评估GABAA信号传导对CA 1锥体细胞的影响。在目标1中，我们将使用Camk 2a- Cre/TdTomatoflox/flox CA 1锥体细胞报告基因小鼠评估GABA信号传导对HIV-1的影响 达特/吗啡诱导的CA 1锥体细胞离子稳态（Ca 2+和Cl-）、生理、结构和 体外暴露于吗啡、HIV-Tat和GABAA/B激动剂/拮抗剂后的体外存活率。 荧光离子指示剂结果将通过这些细胞的全细胞膜片钳电生理学来证实。 分离的细胞这些观察结果将在人类海马神经元中得到证实，暴露于HIV-Tat或 完整的感染性HIV（HIVBaL）。在目标2中，我们将利用Cre诱导的DREADD（设计受体专用 通过设计药物激活）转基因小鼠与HIV-Tat转基因小鼠杂交，研究 对CA 1锥体神经元的整体中间神经元兴奋或抑制的急性和慢性调制 达特tg模型中的生理和形态结果。我们认为减少GABA能输入 可以进一步HIV-Tat诱导的兴奋性毒性损伤，导致长期的生理和结构改变 海马神经回路中。通过调节该系统内的GABA能增益，我们可以改善达特和 阿片诱导的海马神经变性和功能障碍。通过拟议的 电生理学和化学遗传操作，我们将进一步了解阿片/Tat介导的 海马体内的神经病理学

PROJECT SUMMARY. Nearly half of HIV+ patients will experience symptoms of HIV-associated neurocognitive deficits, which are further exacerbated by opiate drug addiction. Opiate drugs impact neurons through direct (i.e., excitotoxicity) and indirect (i.e., glial inflammation) mechanisms. However, while certain brain regions such as striatum exhibit dendritic swelling and degeneration, the hippocampus exhibits a more subtle pathology: transient alterations in intracellular calcium ([Ca2+]i), and sodium ([Na+]i), reduced LTP, and declines in specific CA1 interneuron subtypes, in the absence of gross neurodegeneration. Our preliminary work revealed that GABAA agonism prevented Tat-induced [Ca2+]i transients in vitro, while GABAA antagonism prolonged morphine/Tat-mediated [Ca2+]i elevation. We hypothesize that morphine exacerbates HIV-induced neuronal dysfunction through its influence on the inhibitory microcircuitry in the hippocampus, leading to a net disruption in the excitatory/inhibitory balance of CA1 pyramidal cells and eventual neuronal injury. To study this, we propose to assess the impact of GABAA signaling on CA1 pyramidal cells. In Aim 1, we will use Camk2a- Cre/TdTomatoflox/flox CA1 pyramidal cell reporter mice to assess the impact of GABA signaling on HIV- Tat/morphine-induced changes in CA1 pyramidal cell ion homeostasis (Ca2+ and Cl-), physiology, structure and survival in vitro following exposure to morphine, HIV-Tat, and GABAA/B agonists/antagonists in vitro. Fluorescent ion indicator results will be confirmed with whole-cell, patch-clamp electrophysiology of these dissociated cells. These observations will be confirmed in human hippocampal neurons, exposed to HIV-Tat or intact, infectious HIV (HIVBaL). In Aim 2, we will utilize Cre-inducible DREADD (Designer receptors exclusively activated by designer drugs) transgenic mice crossed with HIV-Tat transgenic mice to study the impact of acute and chronic modulation of overall interneuron excitation or inhibition on CA1 pyramidal neuron physiological and morphological outcomes in the Tat tg model. We propose that reduced GABAergic input could further HIV-Tat-induced excitotoxic damage, leading to prolonged physiological and structural alterations within hippocampal circuitry. By adjusting GABAergic gain within this system, we could ameliorate Tat and opiate-induced hippocampal neurodegeneration and dysfunction. Through the proposed combination of electrophysiology and chemogenetic manipulation, we will further our understanding of opiate/Tat-mediated neuropathology within the hippocampus.