Abstract
Purpose :
The cystine/glutamate antiporter (system xc-) mediates the exchange of extracellular cystine for intracellular glutamate. An xCT knockout mouse (xCT KO), in which the light chain subunit (xCT) of system xc- is removed, displays age related pathologies in the retina which are exacerbated compared to wild-type (WT) mice. In this study, we have investigated the effects of loss of xCT function on the retina with a particular focus on energy metabolism, the glutamate/glutamine cycle, and antioxidant capacity.
Methods :
C57/BL6J WT and xCT KO (C57/BL6J background) mice at 6 weeks (where retinal pathologies were not apparent in the KO; 6W) and 9 months (where retinal pathologies were apparent; 9M) were used. Retinas for each group were collected (6W: WT n=7; KO n=8; 9M: WT n=12; KO n=12). Lactate dehydrogenase (LDH) activity and ATP concentration were measured via biochemical assays. Mitochondrial activity was measured via high-resolution respirometry using the Oroboros Oxygraph-2k system (WT n=6; KO n=5). Glutamate, glutamine, and glutathione (GSH) were localised in the retina and quantified using silver-intensified immunogold labelling (WT n=6; KO n=6).
Results :
A significant decrease in LDH activity was seen in 6 week-, but not 9 month-old KO retinas compared to WT (p<0.05). However, ATP levels were maintained between WT and KO in both age groups (6W p=0.32; 9M p=0.81). Preliminary analysis of retinal mitochondrial activity reveals increased mitochondrial activation of complex 1 of the electron transport chain in the 6 week KO retina compared to WT (p<0.05). In the 6 week KO group, glutamate immunoreactivity significant increased in the outer plexiform layer (p<0.05) and photoreceptors (p<0.01), and glutamine levels significantly decreased in Müller cells (p=0.01) compared to WT. A significant decrease in GSH immunoreactivity was also observed in the photoreceptors in the KO relative to the WT (p<0.05).
Conclusions :
Lack of xCT function results in biochemical changes in the retina including disruption of lactate metabolism, alterations in glutamate-glutamine balance, increased mitochondrial activity of complex 1, and decreased GSH levels. These effects were evident at 6 weeks in the KO retina which precedes the development of retinal pathologies seen at 9 months, suggesting that early metabolic dysfunction may increase the susceptibility of the KO retina to the early development of age related pathologies.
This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.