Tárnok Krisztián, Gulyás Márton, Bencsik Norbert, Ferenc Katalin, Pfizenmaier Klaus, Hausser Angelika, Schlett Katalin
Department of Physiology and Neurobiology, Eötvös Loránd University, Pázmány P. stny. 1/C, H-1117, Budapest, Hungary.
Cytometry A. 2015 Jan;87(1):89-96. doi: 10.1002/cyto.a.22569. Epub 2014 Sep 24.
Dendritic filopodia are tiny and highly motile protrusions formed along the dendrites of neurons. During the search for future presynaptic partners, their shape and size change dynamically, with a direct impact on the formation, stabilization and maintenance of synaptic connections both in vivo and in vitro. In order to reveal molecular players regulating synapse formation, quantitative analysis of dendritic filopodia motility is needed. Defining the length or the tips of these protrusions manually, however, is time consuming, limiting the extent of studies as well as their statistical power. Additionally, area detection based on defining a single intensity threshold can lead to significant errors throughout the image series, as these small structures often have low contrast in fluorescent images. To overcome these problems, the open access Dendritic Filopodia Motility Analyzer, a semi-automated ImageJ/Fiji plugin was created. Our method calculates the displacement of the centre of mass (CoM) within a selected region based on the weighted intensity values of structure forming pixels, selected by upper and lower intensity thresholds. Using synthetic and real biological samples, we prove that the displacement of the weighted CoM reliably characterizes the motility of dendritic protrusions. Additionally, guidelines to define optimal parameters of live cell recordings from dendritic protrusions are provided. © 2014 International Society for Advancement of Cytometry.
树突丝状伪足是沿着神经元树突形成的微小且高度可移动的突起。在寻找未来的突触前伙伴过程中，它们的形状和大小会动态变化，这对体内和体外突触连接的形成、稳定及维持都有直接影响。为了揭示调控突触形成的分子机制，需要对树突丝状伪足的运动进行定量分析。然而，手动定义这些突起的长度或尖端非常耗时，限制了研究的范围及其统计效力。此外，基于定义单个强度阈值的面积检测在整个图像序列中可能会导致显著误差，因为这些小结构在荧光图像中通常对比度较低。为克服这些问题，我们创建了开放获取的树突丝状伪足运动分析仪，这是一个半自动的ImageJ/Fiji插件。我们的方法基于由强度上限和下限阈值选择的构成结构的像素的加权强度值，计算选定区域内质心（CoM）的位移。通过使用合成样本和真实生物样本，我们证明加权质心的位移能够可靠地表征树突突起的运动。此外，还提供了定义从树突突起进行活细胞记录的最佳参数的指南。© 2014国际细胞计量学促进会。
