Supplemental Materials

This article contains the following supporting material:

• Supplemental Figure 1 - Reversible manipulation of mitochondrial ΨΔ_m and SV clustering by creatine and FCCP. The cultured neurons were treated with 100 μM creatine (Cr) for 16 hr or 0.5 μM FCCP for 4 hr. The treated neurons were then incubated in normal medium for another 4 hr before live imaging for JC-1 or fixation for synapsin-1 immunostaining. (A) Quantification of the ratiometric analyses of JC-1 fluorescence intensity. Creatine and FCCP cultures were not significantly different from the untreated cultures after washout, mean ąSEM from 3 independent experiments, n>90 morphologically distinct mitochondria. (B) The ratio of mitochondria with high potential to low potential in creatine- and FCCP-washout cultures was similar to that in untreated cultures. Data are from 3 independent experiments, n>90. (C-D) Quantification of the area (C) and fluorescence intensity (D) of SV clusters showed the reversible effects of creatine and FCCP on SV clustering, mean ąSEM from 3 independent experiments, n>30 neurites.
• Supplemental Video 1 - Time-lapse video of mitochondrial translocation along a control neurite (5s per frame, 100s duration) labeled with MitoTracker. Mitochondrial movement was played back at a rate of 1 frame per second. The first frame is shown in Figure 3B.
• Supplemental Video 2 - Time-lapse video of mitochondrial translocation along a creatine-treated neurite. The culture was treated with creatine for 16 hr and followed by MitoTracker labeling before recording (5s per frame, 100s duration). Playback speed was 1 frame/sec. The first frame is shown in Figure 3E.
• Supplemental Video 3 - Time-lapse video of mitochondrial translocation along a FCCP-treated neurite. The culture was treated with FCCP for 4 hr and followed by MitoTracker labeling before recording (5s per frame, 100s duration). Since FCCP caused a decrease in the MitoTracker loading due to a reduction in ΨΔ_m, the exposure time for each frame was increased in order to visualize mitochondria clearly. Playback speed was 1 frame/sec. The first frame is shown in Figure 3H.
• Supplemental Video 4 - Time-lapse video of mitochondrial translocation along an oligomycin-treated neurite. The culture was treated with oligomycin for 4 hr and followed by MitoTracker labeling before recording (5s per frame, 100s duration). Playback speed was 1 frame/sec.