University Presentation Showcase: Undergraduate Division
A Semi-Automated Method for Quantifying Neurite Degeneration
Abstract
Neurite degeneration is a cellular dysfunction commonly associated with neurodegenerative pathologies such as Alzheimer’s and Parkinson’s disease (PD). Specifically, in PD patients, neurites of dopaminergic neurons localized in the striatum brain region begin to fragment, leading to motor impairment. While methods for quantifying neurite degeneration have been previously described, there has not been published work with detailed description of the method. Here we set out to provide an in-depth description of a method for quantifying neurite degeneration. We also discuss how to troubleshoot issues that may be commonly encountered when performing the technique. Moreover, we demonstrate how this method may be applied to measure neurite degeneration in an in vitro model of PD. Oxidative stress, a cellular condition caused by an imbalance in reactive oxygen species and antioxidants, contributes to neurite fragmentation associated with PD. To model neurite degeneration associated with PD, we treated dopaminergic neurons with two drugs that induce oxidative stress, HNE or 6-OHDA. These cells were then immunostained and visualized by phase bright and fluorescence microscopy. The resulting images were then processed using ImageJ software to obtain a degeneration index (DI). To automate quantification, we have developed a macro to analyze multiple images simultaneously, and we are currently developing a way to validate the use of fluorescent images in order to reduce the background noise that may occur during image capture. These automated methods may be used by others to investigate neurite fragmentation associated with PD, possibly leading to the development of novel therapies.
A Semi-Automated Method for Quantifying Neurite Degeneration
Neurite degeneration is a cellular dysfunction commonly associated with neurodegenerative pathologies such as Alzheimer’s and Parkinson’s disease (PD). Specifically, in PD patients, neurites of dopaminergic neurons localized in the striatum brain region begin to fragment, leading to motor impairment. While methods for quantifying neurite degeneration have been previously described, there has not been published work with detailed description of the method. Here we set out to provide an in-depth description of a method for quantifying neurite degeneration. We also discuss how to troubleshoot issues that may be commonly encountered when performing the technique. Moreover, we demonstrate how this method may be applied to measure neurite degeneration in an in vitro model of PD. Oxidative stress, a cellular condition caused by an imbalance in reactive oxygen species and antioxidants, contributes to neurite fragmentation associated with PD. To model neurite degeneration associated with PD, we treated dopaminergic neurons with two drugs that induce oxidative stress, HNE or 6-OHDA. These cells were then immunostained and visualized by phase bright and fluorescence microscopy. The resulting images were then processed using ImageJ software to obtain a degeneration index (DI). To automate quantification, we have developed a macro to analyze multiple images simultaneously, and we are currently developing a way to validate the use of fluorescent images in order to reduce the background noise that may occur during image capture. These automated methods may be used by others to investigate neurite fragmentation associated with PD, possibly leading to the development of novel therapies.
