嗅球抑制性微环路在阿尔茨海默病早期嗅觉障碍中的作用及治疗意义

Impairments in the sense of smell have been found to develop prior to the appearance of cognitive dysfunction and be correlating with cognitive decline in patients with Alzheimer's disease (AD). Understanding the progress and therapies of olfactory damages, particularly in the early stages, may provide potential therapeutic interventions for AD. Previous studies of others and ours suggest that early olfactory dysfunction is mainly caused by neuropathology dysfunction of olfactory bulb (OB), while impairments of local inhibitory microcircuit are responsible for OB dysfunction. Thus, revealing the mechanisms of impaired local inhibitory microcircuit of OB will be helpfull for further understanding the progress of olfactory dysfunction in early AD. According to our previous work, we attempt to use electrophysiology, western blot and immunofluorescence to clarify the pre- and post-synaptic cellular and molecular mechanisms of impaired local inhibitory microcircuit of OB in early AD. We will also examine the therapeutical effects and mechanisms of pharmacogenetic/optogenetic and pharmacological upregulation of local inhibitory microcircuit for olfactory behavioral dysfunction at early stages of AD. In this project, we will selectively activate interneurons and use agonists of GABAA receptors in the OB of APP/PS1 mouse model of AD, and investigate the effects of upregulation of inhibitory GABAergic neurotransmission on olfactory information output of OB, olfactory thresholds and odor discrimination impairments from molecular and cellular, organotypic and circuital to animal behavioral levels. This study will not only be helpfull to ameliorate olfactory dysfunction in the early stage of AD, but also important to provide theoretical basis for guiding the treatment of AD.

嗅觉障碍出现在阿尔茨海默病（AD）患者认知障碍之前，并和认知功能下降密切相关，积极探索早期嗅觉障碍的机制及治疗方法成为有效控制和治疗AD的重要方向。前人和申请者的研究结果表明，AD早期嗅觉障碍主要由嗅球的病理改变和功能异常引起，而嗅球抑制性微环路的损伤是嗅球功能异常的重要原因，因此深入研究嗅球抑制性微环路受损的机制有助于进一步理解AD早期嗅觉障碍。本课题在前期工作基础上，拟用电生理、免疫印迹和免疫荧光等技术研究AD早期嗅球抑制性微环路受损的突触前/后细胞分子机制，并利用药物/光遗传学技术特异性激活嗅球中间神经元和药理学使用GABAA受体激动剂上调AD早期嗅球抑制性微环路的功能，结合行为学实验，从分子、细胞、组织环路以及整体行为等层面研究提高抑制性微环路功能对嗅球输出、气味阈值和嗅觉分辨等嗅觉障碍的治疗意义及机制。本研究结果不仅有助于治疗AD早期嗅觉障碍，还为控制和治疗AD提供重要理论依据。

尽管嗅觉障碍出现在阿尔茨海默病（AD）患者认知障碍之前，并和认知功能下降密切相关，但其机制尚不明确。本研究旨在探讨APP/PS1转基因小鼠早期嗅觉障碍的嗅球内神经机制。嗅觉阈值检测采用习惯化/去习惯化等方法。嗅觉分辨能力采用习惯化/去习惯化和执行/不执行测试。寡聚化Aβ的类型使用免疫印迹方法检测。神经活动和抑制性微环路采用膜片钳电生理记录。γ氨基丁酸（GABA）A型受体激动剂蝇蕈素和地西泮用以研究上调GABA抑制性微环路对嗅觉功能、嗅球病理生理以及嗅球Aβ的作用。结果如下：3月龄APP/PS1转基因小鼠嗅觉阈值和分辨障碍伴有嗅球Aβ三聚体的产生、僧帽细胞自发、诱发放电频率增高以及“信噪比”降低；这种过度兴奋可被GABAA受体阻断剂阻塞；进一步发现自发和微小抑制性突触后电流频率降低，幅度不变，表明AD早期嗅球抑制性微环路的功能障碍是由于突触前机制；更重要的是蝇蕈素挽救了3月龄APP/PS1转基因小鼠僧帽细胞的输出、回返/侧向抑制，改善了嗅觉探测和分辨障碍并降低了嗅球Aβ三聚体。地西泮也恢复了嗅球输出，改善了嗅觉探测和分辨障碍。此外，在体打孔膜片钳记录幼年斑马鱼僧帽细胞发现突触前自然气味刺激后关键时间窗内给予重复突触后放电可诱导谷氨酸能、GABA能突触传递、GABA能侧向抑制和回返抑制不同方向的长期变化。这种长期改变依赖于不同部位的NMDA受体。以上研究结果表明嗅球抑制性微环路受损是AD早期嗅觉障碍和嗅球Aβ产生的重要原因，并且提示上调GABAA受体相应信号通路是AD潜在的治疗干预措施。