ZFIN ID: ZDB-FIG-080604-73
Peri et al., 2008 - Live Imaging of Neuronal Degradation by Microglia Reveals a Role for v0-ATPase a1 in Phagosomal Fusion In Vivo. Cell   133(5):916-927 Full text @ Cell
ADDITIONAL FIGURES
EXPRESSION / LABELING:
Genes:
Fish:
Knockdown Reagent:
Anatomical Terms:
Stage: Protruding-mouth
PHENOTYPE:
Fish:
Knockdown Reagent:
Observed In:
Stage: Protruding-mouth

Fig. 5 v0-ATPase a1 Knocked-Down Microglia Are Unable to Digest Apoptotic Neurons, However Vesicular Acidification Is Not Effected

(A and B) Two time points (0 and 210 min) of confocal time-lapse of one APO-E-GFP wild-type microglial cell containing apoptotic material labeled by AnnexinV-Cy5 binding. Time is in minutes. (A′ and B′) AnnexinV-Cy5 labeling in the same microglial cell.

(C and D) Two time points (0 and 210 min) of confocal time-lapse of one APO-E-GFP v0-ATPase a1 knocked-down microglial cell containing the apoptotic material labeled by AnnexinV-Cy5 binding. Time is in minutes. (C′ and D′) Confocal time-lapse of AnnexinV-Cy5 labeling in the same microglial cell.

(E) Quantification of fluorescent intensity emission values at 670 nm (AnnexinV-Cy5) in wild-type microglia (green bars) and atp6v0a1 knocked-down microglia (purple bars). Error bars indicate standard deviation. Over time, AnnexinV-Cy5 staining disappears only from wild-type embryos. The data represent the average ± SD of four independent experiments.

(F) We have measured fluorescent intensity values at 505nm (LysoSensor) in several wild-type vesicular clusters.

(G–I) Dorsal views of a 3 dpf (days post fertilization) embryonic wild-type brain. (G) LysoSensor. (H) NBT-DsRed. (I) Merge. There is partial colocalization between acidic vesicles (green) and apoptotic cluster (red).

(J–L) Dorsal views of a 3 dpf (days post fertilization) embryonic v0-ATPase-a1 knocked-down brain. (J) LysoSensor. (K) NBT-DsRed. (L) Merge. Acidic vesicles can be detected in v0-ATPase a1 knocked-down and they colocalize with the neuronal apoptotic clusters.

(M–O) Dorsal views of a 3 dpf (days post fertilization) embryonic wild-type brain treated with Bafilomycin A1. (M) LysoSensor. (N) NBT-DsRed. (O) Merge. After Bafilomycin A1 treatment vesicles are no longer acidic and are not labeled by the LysoSensor.

(P) and (Q) Examples of wild-type (P) and atpv0a1-MO (Q) acidic vesicular clusters, (purple circle [P]) and in atpv0a1-MO (yellow circle [Q]). We have also measured background fluorescent emission at 505nm in wild-type (purple square [P]) and in atpv0a1-MO (yellow square [Q]). Values from several different experiments have been plotted. Error bars indicate standard deviation.

Gene Expression Details
Gene Antibody Fish Conditions Stage Anatomy Assay
DsRed zf148Tg standard conditions Protruding-mouth CNS neuron (sensu Vertebrata) IFL
zf148Tg + MO1-atp6v0a1a standard conditions Protruding-mouth CNS neuron (sensu Vertebrata) IFL
EGFP zf147Tg standard conditions Protruding-mouth microglial cell IFL
zf147Tg + MO1-atp6v0a1a standard conditions Protruding-mouth microglial cell IFL
Antibody Labeling Details No data available
Phenotype Details
Fish Conditions Stage Phenotype
zf147Tg + MO1-atp6v0a1a standard conditions Protruding-mouth phagolysosome assembly arrested, abnormal
zf148Tg + MO1-atp6v0a1a standard conditions Protruding-mouth lysosomal lumen acidification process quality, normal
Protruding-mouth phagolysosome assembly arrested, abnormal
Acknowledgments:
ZFIN wishes to thank the journal Cell for permission to reproduce figures from this article. Please note that this material may be protected by copyright.

Reprinted from Cell, 133(5), Peri, F., and Nüsslein-Volhard, C., Live Imaging of Neuronal Degradation by Microglia Reveals a Role for v0-ATPase a1 in Phagosomal Fusion In Vivo, 916-927, Copyright (2008) with permission from Elsevier. Full text @ Cell