Figure 5

Measurement of axonal and dendritic branch density from primary hippocampal neurons. Neurons were treated with vehicle (DMSO), or nocodazole for three days starting one day after plating. Neurons were stained with the neuronal marker TuJ1. To quantify the average density of neurite branches, the following formula was used: (neurite cell body attachment points - neurite endpoints)/neurite length. a) Representative images and analysis traces of control and treated neurons show decreased neurite length and increased branch density in the presence of nocodazole. b) Quantification of nocodazole titration shows a dose-dependent decrease in total neurite length but an increase in branch density of TuJ1-positive neurites. In our experimental regime, we detected increases in neurite branching at concentrations between 61-185 nM. Measurements from higher concentrations were not included in our analysis, as cell viability and neurite outgrowth decreased drastically, and measurement artefacts from cell debris prevented reliable determination of branch density.