Metabolic changes in optic nerve head astrocytes following glaucoma-associated deformation

Purpose: The astrocytes of the optic nerve head (ONHAs) are the predominant glial cell in the unmyelinated portion of the optic nerve. We tested the metabolic changes these cells undergo in glaucoma by exposing them in vitro to degrees of deformation similarly experienced as a result of increased intraocular pressure in primary open angle glaucoma. 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. ONHA extracellular acidification rate (ECAR) and oxygen consumption rate (OCR) were measured using the Seahorse XFe24 Analyzer, while changes in the proteome were measured using mass spectrometry. 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. There was also at least a twofold decrease in citrate synthase, isocitrate dehydrogenase 2, glycogen phosphorylase B, and adenylate kinase 1. In the Seahorse Analyzer, stretched ONHAs showed increased glycolytic ECAR, maximal ECAR, and maximal OCR. 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. Conclusions: Exposing astrocytes to glaucoma-associated deformation altered their metabolism in ways that indicated stronger commitment to glycolysis compared to control astrocytes, such as increases in glycolytic proteins, decreases in mitochondrial proteins, and increases in glycolytic ECAR.