Figure 5. Mitofusin activation reverses mitochondrial pathology and stimulates growth of CMT2A dorsal root ganglion neurons in vitro.

(a) Confocal micrographs of CMT2A mouse DRGs cultured for 48 hr with MiM111 or its vehicle. Note greater neuronal process length and branching in MiM111-treated neuron. Exploded insets (right) show neuronal process termini. Mitochondria express mitoDS Red; neuronal processes stained for β-III tubulin are green. (b) Kymographs of mitochondrial motility in neuronal processes of live DRGs from studies shown in (a). Top panel is raw data. Bottom panels emphasize motile mitochondria with red and blue lines transiting left to right or right to left, respectively. (c-f) Quantitative group data demonstrating effect of MiM111 on CMT2A DRG mitochondrial aspect ratio (c), motility (d, e), neuronal process length and branching (e), and proportion of neuronal process termini containing mitochondria (f).

Figure 5—figure supplement 1. Mitofusin activation with Chimera C reverses mitochondrial pathology and stimulates growth of CMT2A dorsal root ganglion neurons in vitro.

(a) CMT2A mouse DRGs cultured for 48 hr with Chimera C (100 nM) or its vehicle (Me₂SO₄). Exploded insets show mitochondria expressing mitoDS Red; neuronal processes stained for β-III tubulin are green. Kymographs (below) show mitochondrial motility in live cell studies; quantitative group data for aspect ratio and motility are to the right. (b) Neuronal process length and branching in CMT2A DRG neurons cultured with vehicle or Chimera C. Quantitative group data are to the right. Note greater length and branching of neuronal processes in MiM111-treated neuron. Mitochondria are expressing mitoDS Red. (c). Proportion of neuronal process termini (encircled) containing mitochondria (red) in vehicle- or Chimera C-treated CMT2A DRGs. P values by t-test.

Figure 5—figure supplement 2. Time course studies of DRG mitochondria responses to mitofusin activation after aspiration axotomy.

Studies were performed as in Figure 5. *P < 0.05 vs. basal (ANOVA). Three independent studies were performed for each endpoint; each symbol represents the mean value for all three biological replicates, each of which is the average of approximately 3 (mitochondrial motility in different neurons), 20 (outgrowth of different cell neuronal processes), or 300 (aspect ratio of individual mitochondria), technical replicates. The increase in mitochondrial motility (red) was rapid, being half-maximal after 2 hr. The increase in neuron outgrowth (green) was significant after 24 hr. Increased mitochondrial aspect ratio (blue), reflecting the morphological consequences of enhanced fusion, was only significant after 48 hr.