Fig. 5 | Cellular and Molecular Life Sciences

Fig. 5

From: Bafilomycin A1 activates respiration of neuronal cells via uncoupling associated with flickering depolarization of mitochondria

Fig. 5

Baf induces flickering depolarization of the ΔΨm in dPC12 cells. a Colocalization of the TMRM and mitoCase12 probes in cells treated with Baf for 45 min decreases from 90–95% to 50–80%. b, c Large fluctuations of TMRM signals in individual mitochondria located in cell neurites and costained with mitoCase12 probe (b inset) reveal a flickering pattern of ΔΨm depolarization. The dashed line (c) shows a cross-section of representative mitochondria selected for analysis. d, e In individual mitochondria localized in neurites (d arrows), TMRM intensity periodically drops by 90 ± 5% and then returns to the basal level, while the mitoCase12 fluorescence shows 15–25% fluctuations. f Inhibition of PTP opening by CsA (2 μM) strongly increases the effect of Baf on ΔΨm depolarization. In contrast, thapsigargin (10 μM, Thaps) does not change the effect of Baf on the TMRM signal. However, the morphology of the mitochondria changes dramatically: they become large round compartments with an average diameter of 1–2 μm. g 15 min after the addition of Baf to the cells preincubated with CsA, TMRM intensity decreases to 15–50%, which is significantly different to the cells treated with Baf alone. Inhibition of F0F1 ATPase with oligomycin (10 μM, Olig) has a similar effect. Antimycin A completely depolarizes the mitochondria. Thapsigargin strongly increases variability of the response to Baf. All preincubations were done for 30 min. DMSO was used as a negative control. Bar 20 μm; asterisks significant differences

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