Biochemical methods and X‐ray based imaging strategies to evaluate retinal glucose metabolism

Abstract Purpose Evaluating the coordinated energy metabolism between neurons and glia in situ as a means to evaluate retinal glucose metabolism and function. Methods The imaging of metals conjugated to sugar substrates or metals linked to compounds that affect glycolysis were detected using synchrotron‐based low and high energy x‐ray fluorescence imaging. X‐ray fluorescence maps with <1 micron resolution were placed in a morphological context using simultaneously acquired transmission images of the preparation. Spectrophotometric enzymatic microassays with high selectivity and sensitivity were performed to confirm the intracellular incorporation and metabolism of the delivered substances. Results In the dark‐adapted rat retina, glucose transport and phosphorylation were specifically localized to the Müller glia in situ and an activated glycolysis was not measurable in neurons. Glial glucose metabolism was moreover coordinated with excitatory synaptic transmission in the mid to outer retina. Conclusion Given that oxygen metabolism predominates in neurons and that oxidative metabolism is fuelled by glucose metabolism, fuel transport obligatorily occurs from glia to neurons in intact healthy retina. Combining x‐ray techniques with micron to submicron resolution and biochemical microassays with nM sensitivity offers: (1) a unique experimental strategy to evaluate retinal and cerebral energy metabolism and compartmentation at the cellular level in situ; and (2) is important to the interpretation of images using in vivo functional imaging techniques in the clinic.