Manual versus automatic analysis of microglial density and distribution: a comparison in the hippocampus of healthy and lipopolysaccharide-challenged mature male mice

Abstract

Microglia, the immune resident cells of the central nervous system (CNS), are now recognized as performing crucial roles for maintaining homeostasis and determining the outcomes of various pathological challenges across life. While brightfield microscopy is a powerful and established tool to study microglia-mediated mechanisms underlying neurological diseases, microglial density and distribution are some of the most frequently investigated parameters. Their quantitative assessment provides relevant clues regarding dynamic densitometric changes in the microglial population across various CNS regions. Investigators often rely on a manual identification and analysis of these cells within key regions of interest, which can be time-consuming and introduce an experimenter bias. Automation of this process, which has been gaining popularity in recent years, represents a potential solution to minimize both experimenter's bias and time investment, thus increasing the efficacy of the experiment and uniformity of the collected data. We aimed to compare manual versus automatic analysis methods to determine whether an automatic analysis is efficient and accurate enough to replace a manual analysis in both homeostatic and pathological contexts (i.e., adult healthy and lipopolysaccharide-challenged adolescent male mice, respectively). To do so, we used a script that runs on the ImageJ software to perform microglial density analysis by automatic detection of microglial cells from brightfield…