Abstract
Development of methods for direct and rapid measurement of glucose concentration in the intact organ of animals permits the study of the kinetics of the energy balance in the brain. To monitor the dynamic variation in the concentration of extracellular glucose in rat brain, in this study, we designed two novel glucose monitoring systems, which are comprised of microdialysis sampling and Cu2+-enhanced fluorescence turn-on detection devises. To measure the content of glucose in microdialysate samples, we used glucose oxidase to transform glucose into H2O2, and used 2',7'-dichlorfluorescin-diacetate (DCFH-DA) as fluorescent probe. However, a long incubation time was needed (60 min at room temperature) prior to the fluorescence measurement if only DCFH-DA was treated to react with H2O2. To conquer this embarrassments, we have successfully used a Fenton-like reaction involving the conversion of H2O2 to hydroxyl radical by aqueous Cu2+ ions, which acted as ionic catalyst, to not only shortened the incubation time, but also reached a higher sensitivity. Based on the analytical results obtained, the detection limit of our developed systems was as low as 100 μM, which was low enough for the determination of basal concentration of extracellular glucose in the brain (2 mM). Furthermore, the linear range was ranging from 100μM to 10 mM, which was also wide enough to cover the brain extracellular glucose range of rats. It also indicated that our developed systems can not only be used for the continuous monitoring of brain glucose, but provided necessary evidence for the study of brain energy metabolism. According to the results obtained from the animal experiments, the extracellular glucose concentration would drop simultaneously due to the raised energy consumption rate when the original ionic balance of extracellular environment had been perturbed by treating high doses of K+ ion. However, based on the natural mechanism, the balance of glucose would be rebuilt when the extracellular potassium concentration returned to normal value. Based on the superiority and uniqueness of our developed methods, it is expected that sufficient solid evidences would clarify the mechanism of brain glucose metabolism.