Using ImageJ to Assess Neurite Outgrowth in Mammalian Cell Cultures: Research Data Quantification Exercises in Undergraduate Neuroscience Lab

Brain functions rely critically upon the proper development of neuronal processes (axons and dendrites) and the formation of functional networks. Any genetic factors or environmental compounds that alter the morphological features of neurons may render the nervous system dysfunctional and result in neuronal disorders. In vitro cell culture is an important technique in assessing the effect of chemicals on neurite formation and growth of individual neurons in desired brain regions and has been fundamental in advancing our understanding of the nervous system development and functioning. Despite others offering excellent techniques in cell cultures (Catlin et al., 2016), there is a lack of available resources for teaching students how to analyze neurite outgrowth and run proper statistics on their data. Here, we first briefly discuss culturing cryopreserved mammalian neurons. We then give detailed options to aid upper level undergraduate neurobiology students to quantify neurite outgrowth using NeuronJ, a plugin in the free ImageJ package, Fiji, on both phase contrast and immunofluorescent images. This laboratory exercise provides students the opportunity to culture live neurons, quantify neuronal growth, experiment with the effects of common chemicals on neural development, and conduct statistical data analysis. Previous students expressed their great appreciation for the opportunity to work with live neurons and conduct data quantification and analysis like a true scientist. The ability to accurately measure and calculate the overall growth of neurons using the software ImageJ greatly enhanced students' confidence in presenting their results both in oral and written format.