Cholinergic neurons and memory in GFP-AD mouse: a novel neurotrophic therapy

描述（由申请人提供）： 阿尔茨海默病（AD）是一种年龄依赖性神经退行性疾病，是影响老年人的最常见的痴呆形式。AD的神经病理学特征包括海马和新皮质中含有β-淀粉样蛋白的细胞外斑块和由磷酸化tau蛋白组成的细胞内神经元缠结，以及基底前脑胆碱能神经元（BFCN）的变性和损失。BFCN投射到海马和大脑皮层，在那里释放它们的神经递质乙酰胆碱（ACh）是认知的中心。在AD患者和AD动物模型中，BFCN功能和胆碱能标记物的下降是明显的。因此，据推测，BFCN的功能障碍和/或变性有助于与AD相关的记忆缺陷。许多可用的AD治疗方法旨在抑制ACh的分解，尽管也探索了支持胆碱能神经元功能和存活的营养因子的管理。一种这样的神经营养因子是通过TrkB受体发出信号的脑源性神经营养因子（BDNF）。从AD患者纯化的BFCN中编码TrkB的mRNA的表达减少，表明BDNF治疗可以预防或恢复AD中这些神经元的功能障碍。这种方法受到BDNF不穿过血脑屏障的不良药代动力学的阻碍。最近，发现了一种有效的和选择性的TrkB激动剂，当外周给药时容易进入大脑，7，8-二羟基黄酮（7，8-DHF）。在帕金森病、AD和创伤后应激障碍模型中已经报道了7，8-DHF的有益作用，我们正在展示DHF治疗AD和肌萎缩侧索硬化小鼠模型的初步数据，这些数据强烈支持这些报道。我们假设，7，8-DHF，一种TrkB激动剂，将通过其对海马和基底前脑胆碱能神经元的神经营养作用延迟或减轻记忆丧失并减少淀粉样蛋白病理。为了检验我们的假设，APP.PS1/CHGFP小鼠将用7，8-DHF处理1至6个月（早期病理学）或12个月（晚期病理学）。我们将通过Morris水迷宫实验检测7，8-DHF处理对动物记忆和学习的影响，通过芯片和FACS纯化的BFCN的RT-PCR检测BFCN基因表达模式，通过高效液相色谱法（HPLC）检测海马ACh含量和释放，淀粉样蛋白病理学（免疫组织化学法）和ELISA以及通过IHC技术评估的海马和大脑皮质功能指数，包括胆碱能神经元和胆碱能纤维、神经发生和突触以及树突棘。我们还将使用磁共振波谱（MRS）测量神经化学特征。我们提出的研究将合理的药理学原理与最先进的神经病理学，神经化学，基因分析和MR技术与认知评估，以全面评估一个独特的阿尔茨海默氏症小鼠模型，表达绿色荧光蛋白特异性胆碱能细胞的大脑。这些研究将有助于 进一步定义了一种新的神经营养化合物的治疗益处，该化合物可以穿过血脑屏障，但也将提供对BFCN的基础生物学及其在AD中的功能的全新理解水平，并表征一种新的模型，用于未来测试与AD患者相关的治疗方法。

DESCRIPTION (provided by applicant): Alzheimer's disease (AD) is an age dependent neurodegenerative disorIer and the most common form of dementia affecting the elderly. The neuropathological hallmarks of AD include extracellular ss-amyloid- containing plaques and intracellular neurofibrillary tangles composed of phosphorylated tau protein in the hippocampus and neocortex, and degeneration and loss of basal forebrain cholinergic neurons (BFCN). BFCN project to the hippocampus and cerebral corte where the release of their neurotransmitter, acetylcholine (ACh), is central to cognition. A decline in BFCN function and cholinergic marker is apparent in AD patients and in animal models of AD. Thus, it has been postulated that the dysfunction and/or degeneration of BFCN contributes to the memory deficits associated with AD. Many of the available treatments for AD are designed to inhibit the breakdown of ACh, though the administration of trophic factors that support the function and survival of cholinergic neurons has also been explored. One such neurotrophin is brain derived neurotrophic factor (BDNF) that signals via the TrkB receptor. The reduced expression of mRNAs encoding TrkB in purified BFCN from AD patients, suggests that BDNF therapy could prevent or restore the functional impairments of these neurons in AD. This approach has been hampered by to the poor pharmacokinetics of BDNF that does not cross the blood-brain barrier. Recently a potent and selective TrkB agonist that readily enters the brain when administered peripherally was discovered 7,8-dihydroxyflavone (7,8-DHF). Beneficial effects of 7,8-DHF have been reported in models of Parkinson's disease, AD and post-traumatic stress disorder and we are presenting preliminary data of DHF treatment in mouse models of AD and amyotrophic lateral sclerosis that strongly support these reports. We hypothesize that 7,8-DHF, a TrkB agonist, will delay or lessen memory loss and reduce amyloid pathology through its neurotrophic effects on hippocampal and basal forebrain cholinergic neurons. To test our hypothesis APP.PS1/CHGFP mice will be treated with 7,8-DHF from 1 to 6 months (early pathology) or 12 months (late pathology). We will measure the effects of 7,8-DHF treatment on animals' memory and learning using Morris water maze test, BFCN gene expression pattern by microarray and RT-PCR of FACS purified BFCN, hippocampal ACh content and release by high-performance liquid chromatography (HPLC), amyloid pathology by immunohistochemistry (IHC) and ELISA and indices of hippocampal and cerebral cortical function assessed by IHC techniques including cholinergic neurons and cholinergic fibers, neurogenesis and synapses and dendritic spines. We will also measure neurochemical profile using magnetic resonance spectroscopy (MRS). Our proposed studies incorporate principals of rational pharmacology with state-of-the-art neuropathological, neurochemical, gene-analysis and MR techniques with cognitive evaluation to comprehensively assess the brain of a unique Alzheimer mouse model that expresses green fluorescent protein specifically in cholinergic cells. These studies will help further define the therapeutic benefits of a novel neurotrophic compound that crosses the blood-brain barrier but also will provide an entirely new level of understanding of the basic biology of BFCN and their function in AD and characterize a novel model for future use to test therapeutics relevant to patients with AD.