Abstract
Purpose: :
To understand the metabolic requirements for photoreceptor viability. Photoreceptors do not survive without glucose. We investigated the basis for this by evaluating how photoreceptor viability is influenced by:a. Inadequate concentrations of intracellular ATP.b. Damage to mitochondria or inadequate mitochondrial activity.c. Inadequate concentrations of building blocks for anabolic metabolism.In this study we focused on the importance of intracellular ATP.
Methods: :
Intact mouse retinas were isolated and incubated in bicarbonate buffered physiological solutions with or without glucose, pyruvate, glutamine, leucine or malate. Oxygen consumption was measured using a perfusion apparatus, ATP was measured with a luciferase assay and citric acid cycle and other metabolites were measured by GC-MS. Cell viability was evaluated by propidium iodide uptake in the photoreceptor layer measured by confocal microscopy.
Results: :
When mouse retinas are deprived of glucose oxygen consumption wanes, and ATP levels gradually reach a steady state at ~ 1/3 their normal level. Supplying retinas with mitochondria-specific fuels like pyruvate, glutamine, leucine and malate restores oxygen consumption, mitochondrial membrane potential and citric acid cycle intermediates to their normal levels but it does not restore ATP. Incubating retinas with 2 deoxyglucose further depletes ATP to less than 5% of its normal level. Remarkably, when mitochondria-specific fuels are added in the presence of 2 deoxyglucose retinas consume oxygen, survive more than 90 minutes and do not take up propidium iodide despite still being severely depleted of ATP. Longer term incubations with 2 deoxyglucose and mitochondrial fuels eventually result in propidium iodide uptake and irreversible cell damage.
Conclusions: :
Photoreceptors can survive 90 minutes with less than 5% of their normal ATP levels as long as mitochondria are supplied with fuel so that they are polarized and loaded with citric acid cycle intermediates.
Keywords: metabolism • photoreceptors • retina