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Dhani Tracey-White, Maria Toms, Dhakshi Muhundhakumar, Matthew Smart, Thomas Burgoyne, Glen Jeffery, Clare Futter, Andrew Webster, Mariya Moosajee; KCNJ13-related energy crisis: Mitochondrial dynamics in the retinal degeneration zebrafish model.. Invest. Ophthalmol. Vis. Sci. 2016;57(12):2743.
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© ARVO (1962-2015); The Authors (2016-present)
Mutations in KCNJ13 cause Leber’s Congenital Amaurosis, encoding a defective inward rectifying K+ channel (Kir7.1) expressed in the apical membrane of the RPE, which is essential for the maintenance of photoreceptor excitability. Our previous work has characterised the retinal phenotype of the kcnj13-/- zebrafish, which displays an early onset retinal degeneration. This study investigates the pathophysiology arising from kir7.1 dysfunction, in particular the changes in mitochondrial activity within the retina.
Using kcnj13-/- zebrafish, electron microscopy (EM) of the photoreceptor outer segments (PR-OS) and RPE was undertaken at 3, 6 and 12 months. Mitochondrial number, fission and fusion were assessed with quantitative RT-PCR of polg and nd1, drp1 and opa1, respectively. TUNEL assays were performed to determine levels of cell death. Mitochondrial function was investigated using the JC1 mitochondrial membrane potential assay and measuring levels of ATP production. N=5 for each experiment and timepoint.
The kcnj13-/- retina displayed an increase in mitochondria within the RPE and PR-OS at 6 months on EM, confirmed by qRT-PCR showing a 2.8 and 1.7 fold increase in polg2 and nd1, respectively. Increased levels of apoptosis and a 2-fold decrease in ATP levels were found compared to age-matched wildtype (wt) fish. At 1 year, the kcnj13-/- retina maintained a 1.7-fold increase in nd1, but a 2-fold decrease in mitochondrial fusion opa1, but no significant difference in polg2 and drp1 compared to normal wt levels. Apoptosis was reduced, with a 3.0-fold difference in membrane potential and a 1.5-fold reduction in ATP production compared to wildtypes.
Dysfunction of the kir7.1 channel results in significant mitochondrial biogenesis within the PR-OS and RPE. This is likely to be a compensatory response to metabolic demand and cellular stress. Mitochondrial dynamics are adversely affected with reduced ATP production, increased cell death and aberrant membrane potential resulting in an early onset retinal degeneration. Similar processes have been seen in the ageing retina and models of age-related macular degeneration, however more work is required to understand the triggers in the kcnj13-/- mutant zebrafish.
This is an abstract that was submitted for the 2016 ARVO Annual Meeting, held in Seattle, Wash., May 1-5, 2016.
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