l‐Theanine Improves Learning and Memory Through Attenuation of NOX4‐Mediated Ferroptosis in Hippocampal Neurons of Sleep‐Deprived Mice

ABSTRACT Sleep deprivation (SD) can negatively affect the central nervous system owing to its detrimental effects on learning and memory. l ‐theanine, known for its antioxidative properties, can confer neuroprotection by enhancing learning and memory. Here, we explored whether l ‐theanine could improve learning and memory by inhibiting the NOX4‐mediated ferroptosis in hippocampal neurons caused by SD. The Morris water maze (MWM) was employed to assess learning and memory ability. MDA, SOD, GSH, LDH, and Fe 2+ levels were measured using their specific assay kits. The degree of damage in hippocampal neurons was examined by Nissl staining. The process of ferroptosis in the hippocampus of SD mice and HT22 cells was analyzed using transmission electron microscopy (TEM), immunohistochemistry, flow cytometry, and Western blotting assays. l ‐Theanine treatment significantly increased the frequency of mice crossing the platform. Moreover, it conferred protection on hippocampal neurons and mitochondria in SD mice. Furthermore, l ‐theanine mitigated Erastin‐induced oxidative stress and ferroptosis in HT22 cells. In addition, it reversed the abnormal expression of proteins including PSD‐95, SYN, NOX4, TFRC, ACSL4, Nrf2, Keap1, SLC7A11, P53, HO‐1, NQO1, FTH1, and GPX4 in the hippocampus of SD mice and in the HT22 cells induced by Erastin. Moreover, the neuroprotection of l ‐theanine was reversed by the GLX351322 agent. l ‐Theanine treatment inhibited hippocampal ferroptosis by attenuating NOX4, suggesting that it can be used to mitigate the adverse effects of SD.