Abstract
Microglia are highly motile cells that act as the main form of active immune defense in the central nervous system. Attracted by factors released from damaged cells, microglia are recruited towards the damaged or infected site, where they are involved in degenerative and regenerative responses and phagocytotic clearance of cell debris. ATP release from damaged neural tissues has been suggested to mediate the rapid extension of microglial process towards the site of injury. However, the mechanisms of the long-range migration of microglia remain to be clarified. Here, we found that lysosomes in microglia contain abundant ATP and exhibit Ca2+-dependent exocytosis in response to various stimuli. By establishing an efficient in vitro chemotaxis assay, we demonstrated that endogenously-released ATP from microglia triggered by local microinjection of ATPĪ³S is critical for the long-range chemotaxis of microglia, a response that was significantly inhibited in microglia treated with an agent inducing lysosome osmodialysis or in cells derived from mice deficient in Rab 27a (ashen mice), a small GTPase required for the trafficking and exocytosis of secretory lysosomes. These results suggest that microglia respond to extracellular ATP by releasing ATP themselves through lysosomal exocytosis, thereby providing a positive feedback mechanism to generate a long-range extracellular signal for attracting distant microglia to migrate towards and accumulate at the site of injury.
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Acknowledgements
We thank Dr IC Bruce (Zhejiang University, China) for critical comments on the manuscript and Dr XB Yuan (Institute of Neuroscience, SIBS, CAS, China) for valuable discussion. This work was supported by grants from the Major State Basic Research Program of China (2011CB504400, 2011CBA00400) and the National Natural Science Foundation of China (30730037, 30800321).
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Supplementary information, Movie S1
ATP-induced oriented migration in cultured microglia. Example of directional migration of cultured microglia toward the pipette tip at the center of the field during 20Ā min exposure to an ATP gradient created by pulsatile application of ATP-containing solution (1 mM in the pipette). (MOV 352 kb)
Supplementary information, Movie S2
ATPĪ³S-induced microglial migration. Similar directional migration of cultured microglia during 20Ā min exposure to a gradient of ATPĪ³S (1 mM in the pipette). (MOV 322 kb)
Supplementary information, Movie S3
Inhibition of ATPĪ³S-induced microglial migration by the P2Y receptor antagonist RB-2 (2Ā Ī¼M in the medium). (MOV 273 kb)
Supplementary information, Movie S4
Blockade of ATPĪ³S-induced microglial migration in the presence of the ATP degradation enzyme apyrase (30 U/ml in the medium). (MOV 328 kb)
Supplementary information, Movie S5
Directional migration of microglia from wild-type mouse during 20Ā min exposure to a gradient of ATPĪ³S (0.5 mM in the pipette). (MOV 963 kb)
Supplementary information, Movie S6
Decreased migration of microglia from Rab27a-deficient mouse (ashen) during 20Ā min exposure to a gradient of ATPĪ³S (0.5 mM in the pipette). (MOV 868 kb)
Supplementary information, Figure S1
ATP released from purified microglia visualized by NADPH-based fluorescence Microscope. (PDF 121 kb)
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Dou, Y., Wu, Hj., Li, Hq. et al. Microglial migration mediated by ATP-induced ATP release from lysosomes. Cell Res 22, 1022ā1033 (2012). https://doi.org/10.1038/cr.2012.10
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DOI: https://doi.org/10.1038/cr.2012.10
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