Lysosome exocytosis is involved in astrocyte ATP release after oxidative stress induced by H₂O₂

Background and purpose: Studies demonstrated that oxidative damage decreased intracellular ATP level in astrocytes. However, the pathway mediated ATP level decrease is obscure. Our previous study found intracellular ATP could be released via lysosome exocytosis in astrocytes. Here, we explored whether lysosome exocytosis was involved in ATP release during oxidative stress induced by H₂O₂ in astrocytes.

Methods: Astrocytes were isolated from the cortex of neonatal rats. Intracellular lysosomes and calcium signals were stained in astrocytes before and after H₂O₂ stimulation. ATP molecules location and ATP level were detected by immunostaining and bioluminescence method, respectively. Extracellular β-Hexosaminidase and LDH were examined by colorimetric method.

Results: We found that ATP located in lysosome of astrocytes. H₂O₂ stimulation resulted in the decrease of lysosomes staining and the increase of extracellular ATP, compared to the control (p < 0.05). At the same time, intracellular Fluo4 signals and β-Hexosaminidase level were also increased (p < 0.05). Extracellular LDH level did not show an increase, suggesting that there is no cell membrane damage after H₂O₂ stimulation. Glycyl-phenylalanine 2-naphthylamide blocked lysosome exocytosis and inhibited ATP release in astrocytes after H₂O₂-treatment (p < 0.05).

Conclusion: Our results indicated that H₂O₂ induced ATP release from intracellular to extracellular via lysosome exocytosis. The increase of intracellular Ca²⁺ was necessary for lysosome release under oxidative stress induced by H₂O₂.