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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

Time-course imaging of the neurodegeneration of UV-ablated spinal neurons. (A–D) UV-irradiation of a single spinal neuron (cmet:GFP; A; circle) resulted in the soma of the neuron shrinking over time, followed by axonal fragmentation (B; arrowheads). This axonal degeneration radiated anterogradely toward the distal end of the axon (C), until finally the fluorescence in the soma disappears and the entire axon shows “blebbing” (D). Scale bars = 20 μm. Supplementary Video 1 shows the time-lapse video of this process.
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Figure 2: Time-course imaging of the neurodegeneration of UV-ablated spinal neurons. (A–D) UV-irradiation of a single spinal neuron (cmet:GFP; A; circle) resulted in the soma of the neuron shrinking over time, followed by axonal fragmentation (B; arrowheads). This axonal degeneration radiated anterogradely toward the distal end of the axon (C), until finally the fluorescence in the soma disappears and the entire axon shows “blebbing” (D). Scale bars = 20 μm. Supplementary Video 1 shows the time-lapse video of this process.

Mentions: Microglia constantly and efficiently scan for any alterations in their microenvironment, ranging from changes in neuronal activity to signals of damage-associated processes (Nimmerjahn et al., 2005; Ransohoff and Perry, 2009). We applied UV laser ablation (405 nm) to the soma of spinal motor neurons to selectively induce a localized microglial response in the living zebrafish spinal cord. UV ablation of the fluorescent-labeled neurons (cmet:GFP) reproducibly led to selective death of the targeted neuron within minutes to hours (Figure 2). The time-course of neuronal death was dependent upon parameters such as laser-power, ablation size and dwell time. Ablated and dying neurons showed characteristic morphological changes, including the shrinkage of the cell soma with intact membrane structures (Figures 2A,B), and progressive anterograde degeneration (blebbing) commencing at the targeted soma and continuing along the axon over time (Figures 2C–D; Supplementary Video 1).


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)

Time-course imaging of the neurodegeneration of UV-ablated spinal neurons. (A–D) UV-irradiation of a single spinal neuron (cmet:GFP; A; circle) resulted in the soma of the neuron shrinking over time, followed by axonal fragmentation (B; arrowheads). This axonal degeneration radiated anterogradely toward the distal end of the axon (C), until finally the fluorescence in the soma disappears and the entire axon shows “blebbing” (D). Scale bars = 20 μm. Supplementary Video 1 shows the time-lapse video of this process.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4553390&req=5

Figure 2: Time-course imaging of the neurodegeneration of UV-ablated spinal neurons. (A–D) UV-irradiation of a single spinal neuron (cmet:GFP; A; circle) resulted in the soma of the neuron shrinking over time, followed by axonal fragmentation (B; arrowheads). This axonal degeneration radiated anterogradely toward the distal end of the axon (C), until finally the fluorescence in the soma disappears and the entire axon shows “blebbing” (D). Scale bars = 20 μm. Supplementary Video 1 shows the time-lapse video of this process.
Mentions: Microglia constantly and efficiently scan for any alterations in their microenvironment, ranging from changes in neuronal activity to signals of damage-associated processes (Nimmerjahn et al., 2005; Ransohoff and Perry, 2009). We applied UV laser ablation (405 nm) to the soma of spinal motor neurons to selectively induce a localized microglial response in the living zebrafish spinal cord. UV ablation of the fluorescent-labeled neurons (cmet:GFP) reproducibly led to selective death of the targeted neuron within minutes to hours (Figure 2). The time-course of neuronal death was dependent upon parameters such as laser-power, ablation size and dwell time. Ablated and dying neurons showed characteristic morphological changes, including the shrinkage of the cell soma with intact membrane structures (Figures 2A,B), and progressive anterograde degeneration (blebbing) commencing at the targeted soma and continuing along the axon over time (Figures 2C–D; Supplementary Video 1).

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