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Updated: Jun 23, 2025
Author Spotlight: Advancing Understanding Through Technological Innovations in Psychoneuroimmunology
Published on: May 31, 2024
Quantification of Microglial Engulfment of Synaptic Material Using Flow Cytometry
Bilge Ugursu1, Susanne A Wolf2
1Psychoneuroimmunology, Max-Delbrück-Center for Molecular Medicine in the Helmholtz Association.
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.
Insights
Researchers developed new flow cytometry assays for rapid, high-throughput quantification of microglial engulfment of synapses. These methods offer a faster alternative to traditional imaging techniques for studying brain synaptic refinement.
Area of Science:
- Neuroscience
- Cell Biology
- Immunology
Background:
- Microglia are key players in synaptic refinement, a critical process for brain development and function.
- Quantifying microglial engulfment of synapses is vital for understanding this role.
- Current methods like immunohistochemistry and imaging are laborious and time-consuming.
Purpose of the Study:
- To develop and validate fast, high-throughput in vitro and in vivo assays for quantifying microglial synapse engulfment.
- To provide efficient alternatives to labor-intensive image analysis-based methods.
Main Methods:
- Development of in vivo flow cytometry assay using intracellular vGLUT1 staining on freshly isolated adult mouse brain cells to quantify microglial engulfment of vGLUT1+ synapses.
- Development of in vitro flow cytometry assay using pHrodo Red-labeled synaptosomes and freshly isolated adult mouse brain cells to quantify microglial synaptosome engulfment.
Main Results:
- The developed assays enable rapid and high-throughput quantification of microglial synapse engulfment.
- These flow cytometry-based methods are significantly more time-efficient than traditional imaging techniques.
- The assays successfully quantify microglial engulfment of labeled synapses in both in vivo and in vitro settings.
Conclusions:
- The novel flow cytometry assays provide a streamlined and efficient approach to measuring microglial synapse engulfment.
- These methods can accelerate research into the role of microglia in synaptic refinement across various physiological and pathological conditions.
- The assays represent promising alternatives for researchers studying microglial function in neuroscience and disease models.
