Tracking individual mitochondria over time reveals that fusion and fission are paired. (A) An INS1 cell expressing mtPA-GFP and co-labeled with TMRE. The two mitochondria fused 20 s after photoactivation (arrow in ‘20 s'). Then, a voltage difference was established between the two poles (60–120 s) and they were later physically separated (120 s). The pattern is qualitatively similar to that shown in . Scale bar, 2 μm. (B) Fusion and fission time chart for individual mitochondria. Individual mitochondria tagged with mtPA-GFP were monitored for a period of 1 h and the occurrence of fusion and fission was recorded. Mitochondria in INS1 and COS7 cells spend 77s (±71 s) and 87 s (±78 s), respectively, in the post-fusion, connected state, and an average of 1434s and 1172 s, respectively, in the solitary, post-fission state. (C) Fusion triggers fission. Cumulative probability of the occurrence of fission events at different intervals after fusion events occur (fraction of all fission events occurring after the given time interval). The data are fit to a hyperbolic function representing linkage between the two events (INS1, R2=0.98; COS7, R2=0.96). The straight line shows the predicted linear growth rate in fission event probability over time that would be expected if the fission had occurred independently from the fusion events (dashed lines INS1, R2=0.77; COS7, R2=0.84; see ). (D) Fission does not trigger fusion. Cumulative probability of the occurrence of fusion events at different intervals after the fission event occurred. The data points were fitted to a linear relationship (R2=0.92), as predicted if fusion occurred independently of fission (0.05 versus 0.06%/second, respectively; see ).