An open-source pipeline for analysing changes in microglial morphology.

Changes in microglial morphology are powerful indicators of the inflammatory state of the brain. Here, we provide an open-source microglia morphology analysis pipeline that first cleans and registers images of microglia, before extracting 62 parameters describing microglial morphology. It then compares control and 'inflammation' training data and uses dimensionality reduction to generate a single metric of morphological change (an 'inflammation index'). This index can then be calculated for test data to assess inflammation, as we demonstrate by investigating the effect of short-term high-fat diet consumption in heterozygous Cx3CR1-GFP mice, finding no significant effects of diet. Our pipeline represents the first open-source microglia morphology pipeline combining semi-automated image processing and dimensionality reduction. It uses free software (ImageJ and R) and can be applied to a wide variety of experimental paradigms. We anticipate it will enable others to more easily take advantage of the powerful insights microglial morphology analysis provides.