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Updated: Sep 3, 2025
In Vivo Chronic Two-Photon Imaging of Microglia in the Mouse Hippocampus
Published on: July 6, 2022
In vivo Chronic Two-Photon Imaging of Microglia in the Mouse Hippocampus
Ryosuke Kamei1, Shinji Urata1, Hisato Maruoka1
1Department of Cellular Neurobiology, Graduate School of Medicine, The University of Tokyo.
Abstract:
Microglia, the only immune cells resident in the brain, actively participate in neural circuit maintenance by modifying synapses and neuronal excitability. Recent studies have revealed the differential gene expression and functional heterogeneity of microglia in different brain regions. The unique functions of the hippocampal neural network in learning and memory may be associated with the active roles of microglia in synapse remodeling. However, inflammatory responses induced by surgical procedures have been problematic in the two-photon microscopic analysis of hippocampal microglia. Here, a method is presented that enables the chronic observation of microglia in all layers of the hippocampal CA1 through an imaging window. This method allows the analysis of morphological changes in microglial processes for more than 1 month. Long-term and high-resolution imaging of the resting microglia requires minimally invasive surgical procedures, appropriate objective lens selection, and optimized imaging techniques. The transient inflammatory response of hippocampal microglia may prevent imaging immediately after surgery, but the microglia restore their quiescent morphology within a few weeks. Furthermore, imaging neurons simultaneously with microglia allows us to analyze the interactions of multiple cell types in the hippocampus. This technique may provide essential information about microglial function in the hippocampus.
Insights
This study presents a new chronic imaging method for observing microglia in the hippocampus, enabling long-term analysis of their role in neural circuits and synapse remodeling without immediate post-surgical inflammation interference.
Area of Science:
- Neuroscience
- Immunology
- Cell Biology
Background:
- Microglia are brain-resident immune cells crucial for neural circuit maintenance, synapse modification, and neuronal excitability.
- Microglia exhibit regional heterogeneity in gene expression and function, with significant roles in hippocampal learning and memory.
- Surgical procedures for in vivo imaging can induce inflammatory responses in hippocampal microglia, complicating analysis.
Purpose of the Study:
- To develop and validate a minimally invasive method for chronic, high-resolution imaging of microglia in all hippocampal CA1 layers.
- To overcome the challenge of transient inflammatory responses post-surgery for long-term microglial observation.
- To enable simultaneous imaging of microglia and neurons for analyzing cell-cell interactions in the hippocampus.
Main Methods:
- Implementation of a chronic imaging window for sustained access to the hippocampal CA1 region.
- Application of minimally invasive surgical techniques and optimized imaging parameters for high-resolution microscopy.
- Longitudinal monitoring of microglial morphological dynamics over a period exceeding one month.
- Simultaneous visualization of microglial and neuronal activity.
Main Results:
- The developed method allows for the chronic observation of microglia in all hippocampal CA1 layers for over a month.
- Microglia demonstrate transient inflammatory responses post-surgery, but restore quiescent morphology within weeks.
- The technique facilitates the analysis of microglial process dynamics and morphological changes over time.
- Simultaneous imaging of microglia and neurons reveals potential interactions crucial for hippocampal function.
Conclusions:
- This novel imaging technique provides a robust platform for long-term, high-resolution studies of hippocampal microglia.
- The method overcomes surgical-induced inflammation challenges, allowing for the study of quiescent microglial states.
- It offers essential insights into microglial roles in synaptic remodeling and neural circuit function within the hippocampus.
