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In vivo characterization of microglial engulfment of dying neurons in the zebrafish spinal cord.

Morsch M, Radford R, Lee A, Don EK, Badrock AP, Hall TE, Cole NJ, Chung R - Front Cell Neurosci (2015)

Bottom Line: In vivo imaging confirmed the motile nature of microglia within the uninjured spinal cord.This process of microglial engulfment is highly dynamic, involving the extension of processes toward the lesion site and consequently the ingestion of the dying neuron. 3D rendering analysis of time-lapse recordings revealed the formation of phagosome-like structures in the activated microglia located at the site of neuronal ablation.This real-time representation of microglial phagocytosis in the living zebrafish spinal cord provides novel opportunities to study the mechanisms of microglia-mediated neuronal clearance.

View Article: PubMed Central - PubMed

Affiliation: Motor Neuron Disease Research Group, Faculty of Medicine and Health Sciences, Macquarie University Sydney, NSW, Australia.

ABSTRACT
Microglia are specialized phagocytes in the vertebrate central nervous system (CNS). As the resident immune cells of the CNS they play an important role in the removal of dying neurons during both development and in several neuronal pathologies. Microglia have been shown to prevent the diffusion of damaging degradation products of dying neurons by engulfment and ingestion. Here we describe a live imaging approach that uses UV laser ablation to selectively stress and kill spinal neurons and visualize the clearance of neuronal remnants by microglia in the zebrafish spinal cord. In vivo imaging confirmed the motile nature of microglia within the uninjured spinal cord. However, selective neuronal ablation triggered rapid activation of microglia, leading to phagocytic uptake of neuronal debris by microglia within 20-30 min. This process of microglial engulfment is highly dynamic, involving the extension of processes toward the lesion site and consequently the ingestion of the dying neuron. 3D rendering analysis of time-lapse recordings revealed the formation of phagosome-like structures in the activated microglia located at the site of neuronal ablation. This real-time representation of microglial phagocytosis in the living zebrafish spinal cord provides novel opportunities to study the mechanisms of microglia-mediated neuronal clearance.

No MeSH data available.


Related in: MedlinePlus

Visualization of microglial activity in the spinal cord of 3 day old transgenic zebrafish expressing GFP-positive neurons (islet1:GFP) and mCherry-positive microglia (mpeg1:mCherry). (A) Overview (lateral) of the spinal cord and (B) enlarged lateral view of spinal neurons and a single microglia. (C) Dorsal view of the spinal cord microglia and (D) enlarged dorsal view. Schematic inserts in (A,C) depict orientation of the fish and outline the presented area.
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Figure 1: Visualization of microglial activity in the spinal cord of 3 day old transgenic zebrafish expressing GFP-positive neurons (islet1:GFP) and mCherry-positive microglia (mpeg1:mCherry). (A) Overview (lateral) of the spinal cord and (B) enlarged lateral view of spinal neurons and a single microglia. (C) Dorsal view of the spinal cord microglia and (D) enlarged dorsal view. Schematic inserts in (A,C) depict orientation of the fish and outline the presented area.

Mentions: We firstly characterized the behavior of microglia in the uninjured spinal cord in mpeg1:mCherry zebrafish. This line was chosen as the Gal4 regulatory element causes mosaic expression of mCherry in microglia, allowing us to confidently visualize individual microglia. Accordingly, we observed a non-overlapping distribution of fluorescent microglia throughout the spinal cord (on average 12 microglia ± 1.4; N = 7). The mpeg1-fluorescent cells were on average 240 μm2 (±15.4 μm2; n = 27; N = 21) in size and displayed several features that are characteristic of microglia in the zebrafish and mouse brain (Nimmerjahn et al., 2005; Ellett et al., 2011; Svahn et al., 2013). Hence, spinal microglia exhibited a branched morphology, with filopodia-like processes and bulbous-tipped processes that extended and retracted over minutes (Figure 1). A subset of microglial cells (~30%) showed a highly dynamic behavior, patrolling up and down the spinal cord. This behavior was obvious even in the absence of an “activating” trigger and without characteristic phagocytic activities, such as engulfment of neuronal structures or rapid extension/retraction of their processes. These motile microglia traveled at an average speed of 1.5 μm/min, averaging distances of 98 μm per hour (±7.7 μm; n = 18; N = 7) and a maximum of 241 μm within less than 2 h.


In vivo characterization of microglial engulfment of dying neurons in the zebrafish spinal cord.

Morsch M, Radford R, Lee A, Don EK, Badrock AP, Hall TE, Cole NJ, Chung R - Front Cell Neurosci (2015)

Visualization of microglial activity in the spinal cord of 3 day old transgenic zebrafish expressing GFP-positive neurons (islet1:GFP) and mCherry-positive microglia (mpeg1:mCherry). (A) Overview (lateral) of the spinal cord and (B) enlarged lateral view of spinal neurons and a single microglia. (C) Dorsal view of the spinal cord microglia and (D) enlarged dorsal view. Schematic inserts in (A,C) depict orientation of the fish and outline the presented area.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4553390&req=5

Figure 1: Visualization of microglial activity in the spinal cord of 3 day old transgenic zebrafish expressing GFP-positive neurons (islet1:GFP) and mCherry-positive microglia (mpeg1:mCherry). (A) Overview (lateral) of the spinal cord and (B) enlarged lateral view of spinal neurons and a single microglia. (C) Dorsal view of the spinal cord microglia and (D) enlarged dorsal view. Schematic inserts in (A,C) depict orientation of the fish and outline the presented area.
Mentions: We firstly characterized the behavior of microglia in the uninjured spinal cord in mpeg1:mCherry zebrafish. This line was chosen as the Gal4 regulatory element causes mosaic expression of mCherry in microglia, allowing us to confidently visualize individual microglia. Accordingly, we observed a non-overlapping distribution of fluorescent microglia throughout the spinal cord (on average 12 microglia ± 1.4; N = 7). The mpeg1-fluorescent cells were on average 240 μm2 (±15.4 μm2; n = 27; N = 21) in size and displayed several features that are characteristic of microglia in the zebrafish and mouse brain (Nimmerjahn et al., 2005; Ellett et al., 2011; Svahn et al., 2013). Hence, spinal microglia exhibited a branched morphology, with filopodia-like processes and bulbous-tipped processes that extended and retracted over minutes (Figure 1). A subset of microglial cells (~30%) showed a highly dynamic behavior, patrolling up and down the spinal cord. This behavior was obvious even in the absence of an “activating” trigger and without characteristic phagocytic activities, such as engulfment of neuronal structures or rapid extension/retraction of their processes. These motile microglia traveled at an average speed of 1.5 μm/min, averaging distances of 98 μm per hour (±7.7 μm; n = 18; N = 7) and a maximum of 241 μm within less than 2 h.

Bottom Line: In vivo imaging confirmed the motile nature of microglia within the uninjured spinal cord.This process of microglial engulfment is highly dynamic, involving the extension of processes toward the lesion site and consequently the ingestion of the dying neuron. 3D rendering analysis of time-lapse recordings revealed the formation of phagosome-like structures in the activated microglia located at the site of neuronal ablation.This real-time representation of microglial phagocytosis in the living zebrafish spinal cord provides novel opportunities to study the mechanisms of microglia-mediated neuronal clearance.

View Article: PubMed Central - PubMed

Affiliation: Motor Neuron Disease Research Group, Faculty of Medicine and Health Sciences, Macquarie University Sydney, NSW, Australia.

ABSTRACT
Microglia are specialized phagocytes in the vertebrate central nervous system (CNS). As the resident immune cells of the CNS they play an important role in the removal of dying neurons during both development and in several neuronal pathologies. Microglia have been shown to prevent the diffusion of damaging degradation products of dying neurons by engulfment and ingestion. Here we describe a live imaging approach that uses UV laser ablation to selectively stress and kill spinal neurons and visualize the clearance of neuronal remnants by microglia in the zebrafish spinal cord. In vivo imaging confirmed the motile nature of microglia within the uninjured spinal cord. However, selective neuronal ablation triggered rapid activation of microglia, leading to phagocytic uptake of neuronal debris by microglia within 20-30 min. This process of microglial engulfment is highly dynamic, involving the extension of processes toward the lesion site and consequently the ingestion of the dying neuron. 3D rendering analysis of time-lapse recordings revealed the formation of phagosome-like structures in the activated microglia located at the site of neuronal ablation. This real-time representation of microglial phagocytosis in the living zebrafish spinal cord provides novel opportunities to study the mechanisms of microglia-mediated neuronal clearance.

No MeSH data available.


Related in: MedlinePlus