Evaluating the Effect of Thymoquinone on Keap-1/Nrf-2 Signaling, Oxidative Stress, and Behavior in Rats

Recent developments in pesticide exposure research have increased interest in antioxidant-based protective strategies. Thymoquinone (TQ), a plant-derived bioactive compound, has been reported to exert protective effects against various toxic agents. This study describes a protocol to evaluate the effects of TQ on oxidative stress parameters, the Keap-1/Nrf-2 signaling pathway, and behavioral outcomes in rats exposed to low-dose Deltamethrin (DTM). Here, 24 adult male Wistar Albino rats (250 ± 20 g) were randomly assigned to four groups (n = 6/group): control, TQ, DTM, and DTM+TQ. DTM (1.28 mg/kg) and TQ (10 mg/kg) were administered intragastrically for 30 days. Body weight was monitored throughout the study. Locomotor activity and anxiety-like behavior were assessed using the open field test on day 31, and depression-like behavior was evaluated using the forced swim test on day 32. At the end of the experimental period, plasma and brain tissues were collected for biochemical, histopathological, and molecular analyses. Oxidative stress markers, including malondialdehyde, total nitric oxide, glutathione, and sulfhydryl group levels, were measured in plasma and cerebral cortex samples. Kelch-like ECH-associated protein 1 (Keap-1) and Nuclear factor erythroid 2-related factor 2 (Nrf-2) expression levels were analyzed using Western blotting and immunohistochemistry. This protocol provides a comprehensive and reproducible approach for investigating pesticide-induced neurotoxicity and the modulatory effects of antioxidant compounds.