NEW METHODS FOR EVALUATING CHOLINERGIC TOXICITY

描述（来自申请人摘要）突触前胆碱能降解 与诸如阿尔茨海默氏症之类的胆碱中毒性疾病有关。 神经化学标志物浓度变化的直接定量 化合物对选择性胆碱毒性事件的影响是一项具有挑战性的任务。 在这里， 申请人将开发特别敏感的方法， 测量这些神经化学标记物的变化， 胆碱毒性事件。 他们已经制备了6-羟基儿茶酚， 6-羟基多巴胺的拟胆碱类似物，以及许多相关的 一系列胆碱毒性剂，目的是确定 在大脑中引入选择性胆碱能突触前损伤。 选择性胆碱毒性事件将在体外进行研究， 突触体制备，以建立选择性和相对 每种神经毒剂的神经毒性。 随后的体内研究将 通过在插管大鼠脑中进行微透析取样来监测 在生命系统中发生的近实时变化。 条件 选择性神经毒素的应用将寻求最大限度地改变 胆碱和乙酰胆碱的水平，同时尽量减少 儿茶酚胺和色氨酸神经递质的变化 对照 基于毛细管电泳的分析方法 超微量电化学检测将被用来选择性地和 灵敏地监测微量神经递质及其 代谢物。 毛细管电泳技术将允许高效的 基于电泳分离uL至nL的物种样品 流动性，而电化学检测与专门设计的 超微电极装置将允许额外的选择性和 灵敏的检测到痕量级。 这一战略将允许连续 和时间分辨监测改变的代谢产物和毒素摄取-排出 当与特定脑周围组织的微透析结合时， 受到化学毒素威胁的地区。 该方法将定义 时间和浓度依赖性急性和慢性选择性 胆碱能突触前位点被6-羟基儿茶酚中毒， 相关的季铵儿茶酚基氧化还原亲和试剂。 因此，在本发明中， 追踪胆碱中毒状态下化学事件的新方法可能 为将来研究阿尔茨海默病相关的动物模型而开发 和研究潜在的胆碱能贡献， 控制意识和认知的机制。

DESCRIPTION (from applicant's abstract) Presynaptic cholinergic degradation has been linked to cholinotoxic disorders such as Alzheimer's dementia. Direct quantitation of the changes in concentrations of neurochemical marker compounds upon selective cholinotoxic events is a challenging task. Here, the applicants will develop especially sensitive methodology to directly measure the changes in theses neurochemical markers in response to selective cholinotoxic events. They have prepared 6-hydroxycatecholine, a cholinomimetic analogue of 6-hydroxydopamine, and a number of related cholinotoxic agents in a series for the purpose of defining conditions for the introduction of selective cholinergic presynaptic lesions in the brain. Selective cholinotoxic events will be investigated in vitro with synaptosomal preparations to establish the selectivity and relative neurotoxicity of each neurotoxic agent. Subsequent in vivo studies will be conducted by microdialysis sampling in cannulated rat brain to monitor the near real-time changes which occur in a living system. Conditions for the application of selective neurotoxins will be sought to maximize the changes in levels of choline and acetylcholine, while at the same time minimize the changes in catecholamine and tryptophan neurotransmitters relative to controls. Analytical methods based on capillary electrophoresis with ultramicroelectrochemcial detection will be utilized to selectively and sensitively monitor changes of trace levels of neurotransmitters and their metabolites. Capillary electrophoresis techniques will permit efficient separation of uL to nL samples of species based on their electrophoretic mobility, while electrochemical detection with specifically designed ultramicroelectrode devices will allow for additional selectively and sensitive detection a the trace level. This strategy will allow sequential and time-resolved monitoring of altered metabolite an toxin uptake-efflux when coupled with microdialysis of tissues surrounding specific brain regions challenged with chemical toxins. The methodology will define the temporal and concentration-dependent acute and chronic selective intoxication of cholinergic presynaptic sites by 6-hydroxycatecholine and related quaternary ammonium catechol-based redox-affinity reagents. Thus, new methodologies to follow the chemical events in cholinotoxic states may be developed for future studies of animal models relevant to Alzheimer's dementia and for the study of underlying cholinergic contributions to mechanisms that govern consciousness and cognition.