Fig. 6

The Phosphorylation of p38 mediated the neuronal uptake of astrocytic mitochondria. a Representative Western blot plots showing protein levels of MRIO1, TNFaIP2, total and phosphorylated-p38 (p-P38) and GAPDH in control DA neurons and after rotenone exposure. b An inhibitor of p-38, SB203580, inhibited the rotenone-induced increase in the p-p38 protein levels in neurons. c The neuroprotective role of ACM was abolished by the pre-treatment with SB203580 which inhibits p38 mitogen-activated protein kinase phosphorylation. d Flow cytometry data plot shows the percentage of recipient DA neurons that took up mitochondria from astrocytes with or without SB203580 exposure. The first plot (top left) shows MitoTracker Green labeled donor astrocytes in the first quadrant. The second plot (top right) shows CellTrace Red labeled recipient DA neurons in the third quadrant. The third plot (bottom left) shows in the co-culture setup in which the double-positive (green and red) DA neurons are in the second quadrant. The fourth plot (bottom right) also shows the double-positive DA neurons in the second quadrant. The uptake of mitochondria was decreased when neurons were pre-treated with SB203580, and the mitochondrial transfer rate was reduced from 29.3% ± 1.3% to 14.6% ± 0.6% (e). f Immunostaining was done to detect astrocytic derived mitochondria in TH labeled (red) DA neurons. N = 3, results were presented as mean + SEM. ** P < 0.01, *** P < 0.001 and ^###P < 0.001 versus controls. Scale bars in all panels represent 20 μm