Simultaneous Imaging of Microglial Dynamics and Neuronal Activity in an Awake Mouse

Abstract

Source: Maruoka, H., et. al. Simultaneous Imaging of Microglial Dynamics and Neuronal Activity in Awake Mice. J. Vis. Exp. (2022)

This video shows a two-photon microscopy procedure for imaging neuronal activity and microglial dynamics in an awake mouse during visual stimulation, highlighting calcium transients and cellular interactions.

Protocol

All procedures involving animal models have been reviewed by the local institutional animal care committee and the JoVE veterinary review board.

In vivo two-photon imaging of microglia and neuronal calcium dynamics

1. Habituation of mice to the microscope apparatus

1. Three weeks after the surgery, induce anesthesia in the mouse with 3% isoflurane, and set the mouse under the 25x objective lens of a two-photon microscope using a custom-made stereotaxic instrument (Figure 1C). For the setup, the mouse is set on the top of a treadmill and allowed to run freely.
2. Approximately 10 min later, remove the mouse from th....

Results

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
10-inch LCD monitor	EIZO	DuraVision FDX1003	For presenting grating visual stimuli
Black aluminum foil	THORLABS	BKF12
CX3CR1-EGFP mouse	Jackson laboratory	IMSR_JAX: 005582	CX3CR-1EGFP/+ knock-in/knock-out mice expressing EGFP in microglia in the brain under the control of the endogenous Cx3cr1 locus
DENT SILICONE-V	Shofu Inc	N/A	For the attachment of a shading device to a head-plate
Head-plate	Customized	N/A	Material: SUS304, thickness: 0.5mm
Isoflurane	Pfizer	N/A
Multi-photon excitation microscope	NIKON	N/A	The commercial name is "A1MP+"
Objective lens	NIKON	N/A	The commercial name is "CFI75 apochromat 25xC W".
Psychtoolbox		Free software	For presenting grating visual stimuli
Shading device	Customized	N/A
Stereotaxic instrument	Customized	N/A	For fixing mice under the two-photon microscope
Treadmill	Customized	N/A