Mechanism based evaluation of botanical bioactive compounds enhancing neurological resilience (Project 2)

项目摘要-项目2 涉及植物膳食补充剂的有效临床试验的设计需要试验的最优化 产品中含有足够数量的活性化合物。这些活动的识别 因此，化合物必须是任何严肃的植物学研究的一部分。人们还认识到，总体上 一种植物的活性图谱可能由多个活性化合物介导，这些活性化合物可以独立发挥作用， 相加的，协同的，或相互对抗的。虽然植物学的功能效应可以是 在活体实验中，活性化合物的鉴定及其相互作用的初步研究最多 使用体外、体外或体内中等吞吐量方法进行有效评估。这些也有 以机制为基础的优势。积雪草(Centella asiatica(CA))和睡莲(Withan ia Somnifera(WS)) 影响认知、睡眠和情绪，在这个项目中，我们将使用初级神经元分析、大脑切片和 以果蝇为模型探讨CA和WS的作用机制及活性成分。初级神经元将是 用于通过测量神经元来研究CA和WS及其化合物对神经元健康的影响 树枝形成、抗氧化反应、活性氧物种和线粒体功能。此外，我们还将 测定对小鼠脑片血管张力和血管收缩弹性的影响，以测试 这些萃取物有助于血管健康。果蝇模型将被用来识别能够改善 与年龄相关的运动和反应能力下降。我们还将测试对所描述的类似抑郁症的状态的影响 在果蝇中，以求爱为测试手段，我们将测量神经元亚群中的5-羟色胺水平 被确认为调节这种类似抑郁症的状态。此外，果蝇将被用来确定对 睡眠模式，苍蝇和哺乳动物的睡眠模式也会随着年龄的变化而改变。细胞和分子途径 调节睡眠在果蝇中是众所周知的，因此我们也可以研究CA和/或WS 通过改变神经元的活动和神经递质信号来促进健康的睡眠模式 促进或抑制睡眠的人群。除了5-羟色胺和乙酰胆碱，这还包括GABA。至 将这些研究扩展到小鼠模型，我们将测量小鼠脑片中的神经元活动并解决 CA和/或WS治疗是否影响GABA信号转导(使用GABA抑制剂)。总而言之，这些研究 将识别CA和WS中的活性化合物，并将通过以下方式显示它们是否支持健康和弹性 促进认知、睡眠和/或情绪。此外，它们还将提供对以下机制的见解： 调节这些效应，如改善神经元活动，降低氧化应激和/或促进 线粒体功能。未来的研究可以在活体小鼠模型中证实这些效应，并最终 为在人体内测试活性化合物提供了基础。

PROJECT SUMMARY_PROJECT 2 The design of valid clinical trials involving botanical dietary supplements requires the optimization of the trial product such that it contains sufficient levels of the active compounds. The identification of these active compounds therefore has to be a part of any serious study of botanicals. It is also recognized that the overall activity profile of a botanical may be mediated by multiple active compounds, which can act independently, additively, synergistically or be antagonistic to each other. While functional effects of a botanical can be demonstrated in vivo, initial studies on the identification of active compounds and their interaction is most efficiently evaluated using in vitro, ex vivo, or moderate throughput in vivo approaches. These also have the advantage of being mechanism based. Centella asiatica (CA) and Withania somnifera (WS) have been shown to affect cognition, sleep and mood and in this project, we will use primary neuron assays, brain slices, and Drosophila models to explore mechanisms and active compounds of CA and WS. Primary neurons will be used to address effects of CA and WS and their compounds on neuronal health by measuring neuronal arborization, antioxidant responses, reactive oxygen species, and mitochondrial function. In addition, we will determine effects on vascular tone and resilience to vasoconstriction in mouse brain slices to test whether these extracts promote vascular health. Drosophila models will be used to identify compounds that improve the age-related decline in locomotion and reactivity. We will also test effects on a described depression-like state in Drosophila, using courtship as an assay, and we will measure Serotonin levels in neuronal subpopulations identified as mediating this depression-like state. Furthermore, Drosophila will be used to determine effects on sleep patterns, which are also altered by age in flies and mammals. The cellular and molecular pathways regulating sleep are well known in Drosophila and we can therefore also investigate whether CA and/or WS promotes healthy sleep patterns by altering neuronal activity and neurotransmitter signaling in the neuronal populations that promote or suppress sleep. Besides Serotonin and Aceteylcholine, this includes GABA. To expand these studies to the mouse model, we will measure neuronal activity in mouse brain slices and address whether treatment with CA and/or WS affects GABA signaling (using GABA inhibitors). Together, these studies will identify active compounds in CA and WS and they will show whether they support health and resilience by promoting cognition, sleep and/or mood. In addition, they will provide insights into mechanism that may mediate these effects, like improving neuronal activity, decreasing oxidative stress and/or promoting mitochondrial function. Future studies can then confirm these effects in an in vivo mouse model and eventually provide the basis for testing active compounds in humans.