article accepted in Cytometry Part A:
A new tool for the quantitative analysis of dendritic filopodial motility.
Krisztián Tárnok 1*, Márton Gulyás 1*, Norbert Bencsik 1, Katalin Ferenc 1, Klaus Pfizenmaier 2, Angelika Hausser 2, Katalin Schlett 1
1 Dept. Physiology and Neurobiology, Eötvös Loránd University, Pázmány P. stny. 1/C, H-1117 Budapest, Hungary;
2 Inst. Cell Biology and Immunology, University of Stuttgart, Allmandring 31, D-70569 Stuttgart, Germany
* Authors contributed equally
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, we created the open access Dendritic Filopodia Motility Analyser, a semi-automated ImageJ/Fiji plugin. Our method calculates the displacement of the centre of mass 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 centre of mass reliably characterizes the motility of dendritic protrusions. Additionally, we provide guidelines to define optimal parameters of live cell recordings from dendritic protrusions.