Abstract：
Primary neuronal cultures are commonly used to study genetic and exogenous factors influencing neuronal development and maturation. During development, neurons undergo robust morphological changes involving expansion of dendritic arbor, formation of dendritic spines, and expression of synaptic proteins. In this chapter, we will cover methodological approaches allowing quantitative assessment of in vitro cultured neurons. Various quantitative characteristics of dendritic arbor can be derived based on immunostaining against anti-microtubule-associated protein 2 followed by dendrite tracing with the SNT plug-in of the FIJI software package. The number and subtypes of dendritic spines can be assessed by double labeling with DiI and Phalloidin iFluor448 followed by laser scanning confocal microscopy analysis. Finally, expression of presynaptic and postsynaptic proteins can be determined by immunohistochemistry and quantification using several available software packages including FIJI and Imaris, which also allows for 3D rendering and statistical displaying of the expression level of synaptic proteins.
摘要：
原代神经元培养物通常用于研究影响神经元发育和成熟的遗传和外源因素。在开发过程中,神经元经历强烈的形态变化，涉及树突状乔木的扩张，树突棘的形成，和突触蛋白的表达。在这一章中,我们将涵盖方法学方法，允许定量评估体外培养的神经元。基于抗微管相关蛋白2的免疫染色，然后用FIJI软件包的SNT插件进行树突示踪，可以得出树突状乔木的各种定量特征。树突棘的数量和亚型可以通过用DiI和PhalloidiniFluor448双重标记，然后进行激光扫描共聚焦显微镜分析来评估。最后,突触前和突触后蛋白的表达可以通过免疫组织化学和定量使用几个可用的软件包，包括FIJI和Imaris，这也允许3D渲染和统计显示突触蛋白的表达水平。
