Abstract
Purpose :
The astrocytes of the optic nerve head are in a unique position in glaucoma as the predominant glial cell in the unmyelinated portion of the optic nerve. We tested the metabolic changes these cells face in glaucoma by exposing them in vitro to degrees of deformation/stretch that these cells would similarly experience from increased intraocular pressure in order to better understand the role of metabolism in glaucomic degeneration.
Methods :
Primary astrocytes were cultured from the cortices of P1 mouse pups or P7 optic nerve head explants then seeded on collagen-coated FlexCell plates. The astrocytes were then biaxially stretched by 12% for 24 hours using the FX-6000T FlexCell system. Conditions were chosen to mimic the forces applied to optic nerve head astrocytes (ONHAs) during glaucoma. ONHA glycolytic rates and mitochondrial fuel preferences were measured using the Seahorse XFe24 Analyzer, while changes in the proteome were measured using mass spectroscopy.
Results :
Stretched cortical astrocytes showed at least twofold increases in metabolic proteins such as glutamate dehydrogenase 1, isocitrate dehydrogenase 1, and aldolase fructose-bisphosphate c (n=3, p=0.00024, p=0.022, and p=0.00065). There was also a twofold decrease in citrate synthase, a 10-fold decrease in isocitrate dehydrogenase 2, and 2.5-fold decreases in glycogen phosphorylase B and adenylate kinase 1. In the Seahorse Analyzer, stretched ONHAs showed an increased glycolytic extracellular acidification rate (ECAR), maximal ECAR, and baseline oxygen consumption rate (n=18, p=0.0032, p=0.0070, and p=0.0203). Stretched astrocytes showed no difference in their dependence on pyruvate compared to controls, but a significant decrease in their capacity for mitochondrial respiration from pyruvate (n=18, p=0.0828 and p=0.0001).
Conclusions :
Exposing astrocytes, both cortical and ONHAs, to glaucoma-associated deformation altered their metabolism in ways that indicated increased glycolytic activity compared to control astrocytes. The increase in glutamate dehydrogenase 1 combined with a decrease in isocitrate dehydrogenase 1 and citrate synthase suggest a shift away from mitochondrial energy production. The decrease in pyruvate-based mitochondrial respiration capacity is consistent with these findings.
This is a 2021 ARVO Annual Meeting abstract.