Background and purpose: Methamphetamine (METH) abuse has devastating consequences on the nervous system. There are limited therapy choices in METH abuse with reduced effectiveness and elevated recurrence rates. Thymoquinone (TQ), the most bioactive constituent of Nigella sativa seeds exerts neuroprotective effects mainly via antioxidant properties. This study aimed to evaluate the effect of TQ against METH-induced striatal neurotoxicity and hyperlocomotor activity in mice. Experimental approach: Our groups of animals received METH (10 mg/kg) four times a day with 2 h intervals. Normal saline or TQ (5, 10, or 20 mg/kg) was injected intraperitoneally 30 min before METH administration. Control and sham groups received vehicle or TQ, respectively. The rectal temperature and behavioral tests including the open field for locomotor activity and rotarod for motor coordination were evaluated. The level of superoxide dismutase (SOD), as well as pathological changes, were also assessed in the striatum region. Findings/Results: No significant differences in rectal temperatures were observed among treated groups. Administration of METH increased locomotor activity and did not change motor coordination. TQ co-administration with METH significantly reduced the central and total locomotion and the mean latency to fall off the rotarod in a dose-dependent manner compared with the METH group. TQ also alleviated the METH-induced decrease in the activity of SOD.TQ, especially at the high dose, reduced the METH-induced reactive gliosis level. Conclusion and implications: In conclusion, TQ prevents the enhanced locomotor activity, antioxidant impairment, and morphological striatal damage caused by METH in mice. TQ may be a potential candidate for the treatment of specific METH-induced brain disorders or neurological diseases.