Semi-Automated Quantitative Evaluation of Neuron Developmental Morphology In Vitro Using the Change-Point Test

Neuron morphology gives rise to distinct axons and dendrites and plays an essential role in neuronal functionality and circuit dynamics. In rat hippocampal neurons, morphological development occurs over roughly one week in vitro. This development has been qualitatively described as occurring in 5 stages. Still, there is a need to quantify cell growth to monitor cell culture health, understand cell responses to sensory cues, and compare experimental results and computational growth model predictions. To address this need, embryonic rat hippocampal neurons were observed in vitro over six days, and their processes were quantified using both standard morphometrics (degree, number of neurites, total length, and tortuosity) and new metrics (distance between change points, relative turning angle, and the number of change points) based on the Change-Point Test to track changes in path trajectories. Of the standard morphometrics, the total length of neurites per cell and the number of endpoints were significantly different between 0.5, 1.5, and 4 days in vitro, which are typically associated with Stages 2-4. Using the Change-Point Test, the number of change points and the average distance between change points per cell were also significantly different between those key time points. This work highlights key quantitative characteristics, both among common and novel morphometrics, that can describe neuron development in vitro and provides a foundation for analyzing directional changes in neurite growth for future studies.