High-fat diet (HFD) is a major problem causing neuronal damage. However, the mechanisms and potential therapeutic targets have not been evaluated yet. Thymoquinone (TQ) could regulate oxidative stress and inflammatory process. Hence, the present study elucidated the significant role of TQ on oxidative stress, inflammation as well as morphological changes in the cerebellum of rats with HFD. The present study establishes the role of TQ on oxidative stress, anti-inflammatory process and morphological changes in the cerebellum following high-fat diet supplementation in rats. Rats were divided into three groups as (1) Control, (2) HFD (6 mg/day) for eight weeks and (3) HFD supplementation (4 weeks) followed TQ 300 mg/kg/day treated (4 weeks). After treatment, body weight and serum cholesterol profile were measured. Also, blood samples were collected to measure oxidative stress markers glutathione (GSH), superoxide dismutase (SOD) and inflammatory cytokines. Furthermore, neuronal morphological changes were also observed in the cerebellum of the rats. HFD rats show higher in body weight as compared with the control group. TQ treatment significantly (p<0.05) lowers the body weight with significant (p<0.05) reduction in cholesterol, triglycerides, high-density lipoprotein (HDL) and low-density lipoprotein (LDL). The anti-oxidative enzymes significantly reduced in HFD rats as compared with the control group. Surprisingly, treatment with TQ could improve the level of GSH and SOD. TQ treatment significantly (p<0.05) reduced the inflammatory markers as compared with HFD alone. The histological study revealed that neuronal damage was prevented in the cerebellum following treatment of TQ. TQ treatment minimizes neuronal damage as well as reduces inflammation and improves anti-oxidant enzymes.