Neurodegenerative processes are exacerbated by changes in the cell's redox potential. Reductive stress, the predominance of reduced forms of coenzymes such as NADH, NADPH, and GSH, disrupts the functions of cellular organelles, such as mitochondria. The aim of the study is to identify the pathways through which reductive stress affects cerebral endothelial cells. For the first time, this study investigated the effect of reductive stress on mitochondrial function and the metabolic profile of cerebral endothelial cells in vitro using fluorescence microscopy with fluorescence lifetime measurement (FLIM). A 24-hour incubation with 500 μM dithiothreitol (DTT) did not alter mitochondrial membrane potential or the total pool of NAD+ and NADH, but increased the concentration of superoxide anion, indicating the onset of oxidative stress. Studies using phase-resolved FLIM revealed an increase in the short-lived NADH component as early as 1 hour after incubation, indicating metabolic shifts. The results suggest an adaptation of cellular metabolism toward enhanced glycolysis, which, with prolonged exposure, may disrupt the permeability of the blood-brain barrier and accelerate neurodegenerative processes. The data reveal early mechanisms of metabolic restructuring in cerebral endothelial cells under oxidative stress and demonstrate the potential of FLIM in identifying therapeutic targets aimed at preserving the integrity of the blood-brain barrier. Keywords: reductive stress, cerebral endothelium, FLIM, microscopy, mitochondrial function, NADH.