Department of Pathology, Stanford University Medical Center, Stanford, CA, United States.
Department of Pathology, University of Washington, Seattle, WA, United States.
J Neurosci Methods. 2019 Jan 15;312:73-83. doi: 10.1016/j.jneumeth.2018.11.008. Epub 2018 Nov 20.
Synaptic alterations, especially presynaptic changes, are cardinal features of neurodegenerative diseases and strongly correlate with cognitive decline.
We report "Mass Synaptometry" for the high-dimensional analysis of individual human synaptosomes, enriched nerve terminals from brain. This method was adapted from cytometry by time-of-flight mass spectrometry (CyTOF), which is commonly used for single-cell analysis of immune and blood cells.
Here we overcome challenges for single synapse analysis by optimizing synaptosome preparations, generating a 'SynTOF panel,' recalibrating acquisition settings, and applying computational analyses. Through the analysis of 390,000 individual synaptosomes, we also provide proof-of principle validation by characterizing changes in synaptic diversity in Lewy Body Disease (LBD), Alzheimer's disease and normal brain.
COMPARISON WITH EXISTING METHOD(S): Current imaging methods to study synapses in humans are capable of analyzing a limited number of synapses, and conventional flow cytometric techniques are typically restricted to fewer than 6 parameters. Our method allows for the simultaneous detection of 34 parameters from tens of thousands of individual synapses.
We applied Mass Synaptometry to analyze 34 parameters simultaneously on more than 390,000 synaptosomes from 13 human brain samples. This new approach revealed regional and disease-specific changes in synaptic phenotypes, including validation of this method with the expected changes in the molecular composition of striatal dopaminergic synapses in Lewy body disease and Alzheimer's disease. Mass synaptometry enables highly parallel molecular profiling of individual synaptic terminals.
突触改变，尤其是突触前改变，是神经退行性疾病的主要特征，与认知能力下降密切相关。
我们报告了“大规模突触计量学”，用于对人类突触体（富含大脑神经末梢的分离神经小泡）进行高维分析。该方法源自飞行时间质谱流式细胞术（CyTOF），常用于免疫细胞和血细胞的单细胞分析。
在这里，我们通过优化突触体制备、生成“SynTOF 面板”、重新校准采集设置以及应用计算分析，克服了单个突触分析的挑战。通过对 390,000 个单个突触体的分析，我们还通过对路易体病（LBD）、阿尔茨海默病和正常大脑中突触多样性变化的特征分析，提供了原理验证。
目前用于研究人类突触的成像方法能够分析有限数量的突触，而传统的流式细胞技术通常仅限于少于 6 个参数。我们的方法允许从数万个体突触中同时检测 34 个参数。
我们应用大规模突触计量学同时分析了来自 13 个人脑样本的超过 390,000 个突触体的 34 个参数。这种新方法揭示了突触表型的区域和疾病特异性变化，包括用预期的纹状体多巴胺能突触在路易体病和阿尔茨海默病中的分子组成变化验证该方法。大规模突触计量学能够对个体突触末梢进行高度平行的分子分析。
