Name: Video: Confocal Imaging and Tracking Analysis of Microglial Dynamics in Zebrafish Embryos
Uploaded: 2024-05-17T14:30:00
Duration: 6 min 47 s
Description: 105 Views. Confocal Imaging and Tracking Analysis of Microglial Dynamics in Zebrafish Embryos

confocal imaging and tracking analysis of microglial dynamics in zebrafish embryos

Elucidating Microglia Behavior in Zebrafish Optic Tectum Development

As resident macrophages in the central nervous system (CNS), microglia represent a distinct non-neuronal population that accounts for up to 15% of all glial cells in the adult brain. Studying microglia biology has gained increasing attention in recent years due to their established importance in development, physiology, and disease1. Under physiological conditions, microglial cells are highly dynamic, continuously surveying the brain parenchyma2,3. This behavior allows microglia to colonize the brain and play pivotal roles in its development such as shaping neuronal circuitry4, synaptic pruning5, and vasculogenesis6. Furthermore, this inherent dynamic nature allows microglia to constantly monitor the CNS for signs of infection, injury, or any deviations from homeostasis7. To dissect these intricate cell dynamics, live imaging of microglia across space and time is indispensable. Fortunately, the optical transparency of externally developing zebrafish embryos, coupled with the availability of well-characterized transgenic reporter lines that fluorescently label microglia, positions zebrafish as an ideal vertebrate model for such investigations. Live imaging in zebrafish embryos offers a non-invasive approach that does not require surgery or extensive tissue manipulation, minimizing potential perturbations to the CNS status. This is a critical consideration when studying microglial cells, as they are highly sensitive to even subtle changes in the extracellular environment8.
Here, we provide a guideline to successfully track 3D microglial cell movements in the zebrafish embryo, allowing for an unprecedented view of microglia behavior within the intact architecture of the developing brain parenchyma (see Figure 1&#160;for a graphical overview of the protocol). This step-by-step protocol details how to set up and image zebrafish microglia at different developmental stages and how to extract high-resolution data on microglia cell motility to provide valuable insights into their migratory patterns and responses to environmental cues. We also demonstrate this protocol can be adapted to perform live multi-color imaging, thus extending its applicability to studying microglia in combination with transgenic lines that mark neighboring cells, including neurons3, oligodendrocytes9, and endothelial cells10&#160;(as shown in Figure 2). By adding to the toolbox that allows one to directly observe and characterize the dynamics of microglia behavior&#160;in real time and in their natural environment, this protocol will likely contribute to better elucidating microglia functionality during early development, both in physiology and disease.

Author Spotlight: Using Zebrafish to Explore Microglia Migration During Brain Development

Live Imaging and Characterization of Microglia Dynamics in the Zebrafish Embryo

Confocal Imaging and Tracking Analysis of Microglial Dynamics in Zebrafish Embryos

confocal-imaging-tracking-analysis-microglial-dynamics-zebrafish

Research

JoVE Journal

Immunology and Infection

105 Views. Confocal Imaging and Tracking Analysis of Microglial Dynamics in Zebrafish Embryos

Video: Confocal Imaging and Tracking Analysis of Microglial Dynamics in Zebrafish Embryos

Microglia are highly dynamic cells and their migration and colonization of the brain parenchyma is a crucial step for proper brain development and function. Externally developing zebrafish embryos possess optical transparency, which along with well-characterized transgenic reporter lines that fluorescently label microglia, make zebrafish an ideal vertebrate model for such studies. In this paper, we take advantage of the unique features of the zebrafish model to visualize the dynamics of microglia cells in vivo and under physiological conditions. We use confocal microscopy to record a timelapse of microglia cells in the optic tectum of the zebrafish embryo and then, extract tracking data using the IMARIS 10.0 software to obtain the cells' migration path, mean speed, and distribution in the optic tectum at different developmental stages. This protocol can be a useful tool to elucidate the physiological significance of microglia behavior in various contexts, contributing to a deeper characterization of these highly motile cells.

We demonstrate a method that takes advantage of fast scanning confocal microscopy to perform live imaging of microglia cells in the developing zebrafish optic tectum, allowing for the analysis of the dynamics of these cells in vivo.