We investigated the interplay between the intracellular energy sensor AMP-activated protein kinase (AMPK), prosurvival kinase Akt, oxidative stress, and autophagy in the cytotoxicity of parkinsonian neurotoxin 1-methyl-4-phenyl piridinium (MPP+) towards SH-SY5Y human neuroblastoma cells. MPP+-mediated oxidative stress, mitochondrial depolarization, and apoptotic cell death were associated with rapid (within 2 h) activation of AMPK, its target Raptor, and prosurvival kinase Akt. Antioxidants N-acetylcysteine and butylated hydroxyanisole suppressed MPP+-induced cytotoxicity, AMPK, and Akt activation. A genetic or pharmacological inhibition of AMPK increased MPP+-triggered production of reactive oxygen species and cell death, and diminished Akt phosphorylation, while AMPK activation protected SH-SY5Y cells from MPP+. On the other hand, genetic or pharmacological inactivation of Akt stimulated MPP+-triggered oxidative stress and neurotoxicity, but did not affect AMPK activation. At later time-points (16–24 h), MPP+ inhibited the main autophagy repressor mammalian target of rapamycin, which coincided with the increase in the levels of autophagy marker microtubule-associated protein 1 light-chain 3B. MPP+ also increased the concentration of a selective autophagic target sequestosome-1/p62 and reduced the levels of lysosomal-associated membrane protein 1 and cytoplasmic acidification, suggesting that MPP+-induced autophagy was coupled with a decrease in autophagic flux. Nevertheless, further pharmacological inhibition of autophagy sensitized SH-SY5Y cells to MPP+-induced death. Antioxidants and AMPK knockdown reduced, whereas genetic inactivation of Akt potentiated neurotoxin-triggered autophagy. These results suggest that MPP+-induced oxidative stress stimulates AMPK, which protects SH-SY5Y cells through early activation of antioxidative Akt and late induction of cytoprotective autophagy.
Strategische Forschungsbereiche und Zentren
- Querschnittsbereich: Medizinische Genetik