Quantitative neurite outgrowth measurement based on image segmentation with topological dependence

The study of neuronal morphology and neurite outgrowth has been enhanced by the combination of imaging informatics and high content screening, in which thousands of images are acquired using robotic fluorescent microscopy. To understand the process of neurite outgrowth in the context of neuroregeneration, we used mouse neuroblastoma N1E115 as our model neuronal cell. Six‐thousand cellular images of four different culture conditions were acquired with two‐channel widefield fluorescent microscopy. We developed a software package called NeuronCyto. It is a fully automatic solution for neurite length measurement and complexity analysis. A novel approach based on topological analysis is presented to segment cells. The detected nuclei were used as references to initialize the level set function. Merging and splitting of cells segments were prevented using dynamic watershed lines based on the constraint of topological dependence. A tracing algorithm was developed to automatically trace neurites and measure their lengths quantitatively on a cell‐by‐cell basis. NeuronCyto analyzes three important biologically relevant features, which are the length, branching complexity, and number of neurites. The application of NeuronCyto on the experiments of Toca‐1 and serum starvation show that the transfection of Toca‐1 cDNA induces longer neurites with more complexities than serum starvation. © 2008 International Society for Advancement of Cytometry