ProMoIJ: A new tool for automatic three-dimensional analysis of microglial process motility

Area of Science:

• Neuroscience
• Cell Biology
• Bioimaging

Background:

• Microglia are central nervous system immune cells essential for brain homeostasis.
• Microglial process motility reflects their surveying function in health and disease.
• Current analysis methods are manual, time-consuming, and lack standardization, often losing 3D information.

Purpose of the Study:

• To develop an automated, 3D analysis tool for microglial process motility.
• To overcome limitations of manual and 2D analysis methods.
• To provide an open-source, standardized solution for researchers.

Main Methods:

• Development of ProMoIJ, a pack of ImageJ macros for 3D analysis.
• Automation of cellular process selection and 3D skeleton reconstruction.
• Analysis of process and tip velocity for motility quantification.

Main Results:

• ProMoIJ significantly reduces analysis time and experimenter bias.
• The tool is more robust than manual analysis across varying process numbers.
• 3D analysis with ProMoIJ reveals underestimation of motility by 2D methods.
• ProMoIJ demonstrated that only lesion-adjacent microglia extend processes towards injury and that inflammation reduces motility.

Conclusions:

• ProMoIJ offers a standardized, accelerated, and robust method for 3D microglial process motility analysis.
• The tool enhances the understanding of microglial dynamics in physiological and pathological contexts.
• ProMoIJ is freely available, promoting wider adoption and reproducibility in neuroscience research.