Fluorescent Biosensors for Neuronal Metabolism and the Challenges of Quantitation

Abstract

Over the past decade, genetically encoded fluorescent biosensors that report metabolic changes have become valuable tools for understanding brain metabolism. These sensors have been targeted to specific brain regions and cell types in different organisms to track multiple metabolic processes at single cell (and subcellular) resolution. Here, we review genetically encoded biosensors used to study metabolism in the brain. We particularly focus on the principles needed to use these sensors quantitatively while avoiding false inferences from variations in sensor fluorescence that arise from differences in expression level or environmental influences such as pH or temperature.

Figure 1.. Metabolic targets of fluorescent biosensors in neurons

Key substrates, intermediates and cofactors in brain energy metabolism are presented in this diagram of neuronal metabolism. Solid green boxes indicate analytes that have been successfully measured using genetically encoded fluorescent biosensors expressed in the cytosol and mitochondrial matrix of neurons. Dashed gray boxes indicate target analytes of existing sensors that have not yet been utilized in neurons. Cytosolic measurements of pyruvate, lactate and glucose have been essential in studying fuel preference in neurons at rest and during stimulation, while NADH:NAD⁺ and ATP sensors have provided insight into the relationship between excitability and energy demand. Targeting sensors to intracellular organelles like the mitochondria have enabled study of the compartmentation of energy production. In the future, biosensors for analytes like NADPH, inorganic phosphate, citrate, α-ketoglutarate, mitochondrial glutamate and O₂ could further inform on the cellular energy state and the overall redox state in neurons.