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| Item Type: | Article |
|---|---|
| Title: | Quantification of microglial engulfment of synaptic material using flow cytometry |
| Creators Name: | Ugursu, B. and Wolf, S.A. |
| Abstract: | Microglia play a pivotal role in synaptic refinement in the brain. Analysis of microglial engulfment of synapses is essential for comprehending this process; however, currently available methods for identifying microglial engulfment of synapses, such as immunohistochemistry (IHC) and imaging, are laborious and time-intensive. To address this challenge, herein we present in vitro and in vivo* assays that allow fast and high-throughput quantification of microglial engulfment of synapses using flow cytometry. In the in vivo* approach, we performed intracellular vGLUT1 staining following fresh cell isolation from adult mouse brains to quantify engulfment of vGLUT1(+) synapses by microglia. In the in vitro synaptosome engulfment assay, we used freshly isolated cells from the adult mouse brain to quantify the engulfment of pHrodo Red-labeled synaptosomes by microglia. These protocols together provide a time-efficient approach to quantifying microglial engulfment of synapses and represent promising alternatives to labor-intensive image analysis-based methods. By streamlining the analysis, these assays can contribute to a better understanding of the role of microglia in synaptic refinement in different disease models. |
| Keywords: | Brain, Flow Cytometry, Microglia, Synapses, Synaptosomes, Vesicular Glutamate Transport Protein 1, Animals, Mice |
| Source: | Journal of Visualized Experiments |
| ISSN: | 1940-087X |
| Publisher: | JoVE |
| Number: | 207 |
| Date: | 31 May 2024 |
| Official Publication: | https://doi.org/10.3791/66639 |
| PubMed: | View item in PubMed |
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