Abstract
Purpose :
The function and cellular localization of glycogen in the retina are debated. Our goal is to determine the metabolic sources for glycogen synthesis and catabolism in the mouse retina and RPE-choroid.
Methods :
C57BL/6J retinas or RPE-choroid were cultured for 0, 6, 24, 48, or 72 hours. We supplied tissue with 13C6-glucose, 1-13C1-glucose, and/or 13C3-lactate, then flash-froze tissue and medium samples in liquid N2. We extracted intracellular glucose and glycogen-derived glucose residues from tissue samples. We used enzymatic assays and liquid chromatography-mass spectrometry to quantify glycogen and the extent to which 13C labeled carbons from the added glucose or lactate contribute to glycogen synthesis. Single-cell transcriptomic data was used to generate hypotheses on the cellular specificity of gluconeogenesis.
Results :
At steady state in culture, ~40% of glucose residues in mouse retina glycogen originate from added glucose. Glycogen turnover in the mouse RPE-choroid is slower, and by 48 hours approximately 10% of glucose subunits on glycogen come from glucose in the culture medium. Deletion of the glucose transporter Glut1 from the whole retina reduces total glycogen content, but glycogen levels are unaffected by Glut1 deletion in Müller glia or in photoreceptors. We traced formation of glycogen by gluconeogenesis using 13C lactate. In the retina, gluconeogenesis can produce glucose that is incorporated into glycogen. ~15% of glycogen in the retina is derived from lactate carbons. However, 13C from lactate does not label glycogen in the RPE-choroid. Based on the analysis of transcriptomic data from mouse retinas, Müller glia are primary site of gluconeogenesis and glycogen storage.
Conclusions :
Glucose is a major substrate for glycogen synthesis in the retina and RPE, while lactate is a major substrate for glycogen synthesis solely in the retina. Neither substrate fully accounts for all glucosyl residues on glycogen, which may be due to lower turnover of residues in the core of the glycogen molecule. We are investigating the purpose of retina and RPE-choroid glycogen and the factors that control its synthesis and breakdown.
This abstract was presented at the 2024 ARVO Annual Meeting, held in Seattle, WA, May 5-9, 2024.