Analyzing neurite outgrowth from explants by fitting ellipses

Abstract

The establishment of appropriate connectivity in the developing nervous system depends on many molecular guidance cues. A key method for studying the response of nerve fibers to such guidance cues is to culture explants of neural tissue in three-dimensional collagen gels. However, most previous analyses of the neurite outgrowth patterns from these explants have been very simple, often measuring only one or two parameters. Here we introduce a more sophisticated method for characterizing neurite outgrowth from explants, based on fitting an ellipse to the pattern of outgrowth. This provides 5 parameters describing the outgrowth: x and y position of the center of the ellipse, the elongation, the area and the tilt. We then apply this method to a large dataset of dorsal root ganglion explants grown in the presence of precisely controlled gradients of nerve growth factor. This analysis reveals a number of new features of these data. For instance, we find that it is the position of the center of the ellipse rather than the shape of the ellipse that is correlated with the strength of the gradient. Together these results show that ellipse-fitting of explant data can give new insights into the biological processes underlying neurite guidance by molecular cues.