Visualization of microglia in living tissues using Iba1-EGFP transgenic mice
1Department of Neurochemistry, National Institute of Neuroscience, Kodaira, Tokyo, Japan.
Abstract:
Microglia are thought to play important roles not only in repairing injured tissue but in regulating neuronal activity, and visualizing the cells is very useful as a means of further investigating the function of microglia in vivo. We previously cloned the ionized calcium-binding adaptor molecule 1 (Iba1) gene, which is expressed selectively in microglia/microphages. To generate new transgenic mice to visualize microglia with enhanced green fluorescent protein (EGFP), we here constructed a plasmid carrying EGFP cDNA under control of the Iba1 promoter. This construct was injected into C57B/6 mouse zygotes, and the Iba1-EGFP transgenic line was developed. Fluorescent in-situ hybridization analysis revealed that the Iba1-EGFP transgene was located on chromosome 11D. No obvious defects were observed during development or in adulthood, and the EGFP fluorescence remained invariant over the course of at least four generations. Judging from the immunoreactivity with anti-Iba1 antibody, all EGFP-positive cells in the adult brain were ramified microglia. In the developing transgenic embryos, EGFP signals were detected as early as embryonic Day 10.5. The most prominent EGFP signals were found in forebrain, spinal cord, eye, foreleg, yolk sac, liver, and vessel walls. At postnatal Day 6, clear EGFP signals were observed in the supraventricular corpus callosum, known as "fountain of microglia", where ameboid microglia migrate into the brain parenchyma and mature into ramified microglia. Iba1-EGFP transgenic mice thus permit observation of living microglia under a fluorescence microscope and provide a useful tool for studying the function of microglia in vivo.
Insights
Researchers developed new Iba1-EGFP transgenic mice to visualize microglia, essential immune cells in the brain. These mice allow real-time observation of microglia in living animals, aiding in the study of their functions in tissue repair and neuronal activity.
Area of Science:
- Neuroscience
- Immunology
- Genetics
Background:
- Microglia are crucial for brain health, involved in tissue repair and neuronal regulation.
- Visualizing microglia in vivo is essential for understanding their complex functions.
- The ionized calcium-binding adaptor molecule 1 (Iba1) gene is selectively expressed in microglia.
Purpose of the Study:
- To create a novel transgenic mouse model for visualizing microglia using enhanced green fluorescent protein (EGFP).
- To establish a reliable tool for in vivo studies of microglial behavior and function.
Main Methods:
- Constructed a plasmid with EGFP cDNA under the Iba1 promoter.
- Injected the construct into C57B/6 mouse zygotes to generate transgenic offspring.
- Utilized fluorescent in-situ hybridization and anti-Iba1 antibody staining for validation.
Main Results:
- Successfully generated a stable Iba1-EGFP transgenic mouse line with consistent fluorescence across generations.
- Confirmed that EGFP expression accurately labels ramified microglia in adult brains.
- Detected EGFP signals in developing embryos (from E10.5) and specific tissues, including the developing brain and spinal cord.
- Observed clear EGFP signals in the 'fountain of microglia' region at postnatal day 6, illustrating microglial migration and maturation.
Conclusions:
- The Iba1-EGFP transgenic mice provide a powerful, non-invasive method for observing living microglia.
- This new tool facilitates in vivo research into microglial roles in development, disease, and repair.
- The model enables detailed studies of microglial dynamics and interactions within the central nervous system.
