Abstract
Purpose :
The mechanism that precedes the progression of retinal ganglion cell (RGC) death in autosomal dominant optic atrophy (ADOA) remains unclear. OPA1 is a key gene of mitochondrial dynamics and plays a major role in most cases of ADOA. However, the alterations in the movement of mitochondria in RGCs due to OPA1 deficiency remain unknown. We hypothesize that OPA1 deficiency leads to alterations in mitochondrial movement along the neurites of RGCs, which may be an early alteration contributing to the progression of RGC death in ADOA.
Methods :
RGCs were obtained from wild-type and Opa1+/- mouse pups using two-step immunopanning (Winzeler & Wang, 2013). RGCs were stained with MitoTracker®Red CMXRos (Invitrogen) and the movements of mitochondria along neurites were recorded by time-lapse imaging. Images were captured every 5 seconds for 10 minutes. Key parameters of the movement were calculated from kymographs generated by KymoAnalyzer (Neumann et al., 2017) plugin using FIJI software. The mitochondria were classified as stationary (moved < 5 µm) and motile (moved > 5 µm). The density of mitochondria was calculated as the number of stationary or motile mitochondria per length of the neurite in µm. The velocities of movements were calculated for the motile mitochondria (µm/s). Unpaired Student’s t-test was applied for statistical analysis.
Results :
A significant higher proportion of motile mitochondria (22.95 ± 3.10%) was observed in Opa1+/- RGCs compared with the wild-type (increased by 10.53 ± 4.00%, p < 0.05). Moreover, the density of motile mitochondria per length of the neurite performed a marked increase in Opa1+/- RGCs (0.041 ± 0.0035 mitochondria/µm) compared with the wild-type (0.018 ± 0.0034 mitochondria/µm, p < 0.01). Furthermore, the velocities of mitochondrial movement in Opa1+/- RGCs (0.39 ± 0.024 µm/s) showed a remarkable elevation compared to the wild-type (0.24 ± 0.026 µm/s, p < 0.05).
Conclusions :
Our results demonstrated that Opa1 deficiency caused a significant activation in mitochondrial movement along the neurites of RGCs cultured from Opa1+/- mice, indicating the alterations in the transport of mitochondria are likely to be an early change in the progression of RGC loss caused by Opa1 deficiency. Further genetic measurements are required to investigate the precise mechanism underlying mitochondrial mobility altered by the deficit of Opa1.
This is a 2020 ARVO Annual Meeting abstract.