Real-time, seconds-resolved measurements of amino acid homeostasis and its regulation - Summary. The plasma concentrations of many metabolites are strictly controlled. Glucose, of course, is a well- known and obviously important example. But because of the critical roles they play in protein synthesis, intermediary metabolism, and neurochemistry, homeostatic control over the concentrations of many amino acids is similarly tight and similarly vital, with their dysregulation similarly leading to disease. The acute (i.e., minutes- scale) mechanisms underlying this homeostasis, however, are far less well characterized than those underlying blood sugar control. In no small part this discrepancy is due to the cumbersome, costly, and poorly-time-resolved nature of current methods for measuring any metabolite other than glucose, all of which rely on sampling (e.g., blood draws, microdialysis) and benchtop analysis. Against this background, the premise of our work is 3-fold: First, that improved understanding of metabolic homeostasis is required to reach a complete, systems- level understanding of physiology. Second, that achieving this will require orders of magnitude improve- ment in the time-resolution of metabolic measurements. Third, that, when coupled with the powerful aptamer discovery strategies we have developed, electrochemical aptamer-based (EAB) sensors are the only technology on the horizon that can enable this paradigm shift. EAB sensors, a platform technology we invented, have already been shown to support seconds-resolved, real-time, multi-hour measurements of more than 2 dozen different molecules in the veins, brains, and solid peripheral tissues of living subjects. Building on this, here we propose to adapt this now well-established (e.g., independently reproduced by others) technology to an important and novel problem: the study of metabolic homeostasis, placing particular emphasis on the amino acids involved in nitrogen and energy metabolism (glutamine, alanine, arginine), neurotransmitter synthesis (the aromatic amino acids), and metabolic syndrome/diabetes (the branched-chain amino acids). To achieve this goal, our established, multi-disciplinary team, whose expertise spans aptamer selection, device design and optimization, and animal studies, employs a rigorous discovery pipeline and multiple layers of independent internal replication to ensure our findings are robust and reproducible. By providing a window into metabolite dynamics at physiologically-most-relevant, seconds-to-minutes time scales, the proposed technology will provide unique opportunities to test and expand models of acute metabolic homeostasis throughout the body.
