Factors that modulate the psychomotor activating effects of amphetamine and cocaine, such as environmental novelty and dose, also regulate the ability of these drugs to induce c-fos mRNA expression in the subthalamic nucleus (STN). We hypothesized therefore that engagement of the STN may be important for stimulant-induced psychomotor activation. To further test this hypothesis we examined whether repeated treatment with cocaine, which enhances its psychomotor activating effects (i.e. produces behavioural sensitization), also enhances its ability to induce c-fos expression in the STN. In addition, given that STN activity is thought to be influenced by preproenkephalin mRNA-containing (ENK+) neurons in the caudate-putamen, we also examined whether repeated cocaine treatment alters c-fos expression in ENK+ cells. We report that: (i) cocaine pretreatment enhances the ability of a cocaine challenge to induce c-fos mRNA expression in the STN, and this effect is most robust at challenge doses where behavioural sensitization is observed; (ii) the ability of cocaine to induce c-fos in the STN is independent of the ability of cocaine to engage ENK+ cells. These results support the idea that the STN is involved in stimulant-induced psychomotor activation and sensitization, but suggest that stimulant-induced engagement of the STN is not dependent on ENK+ cells in the caudate-putamen. These findings may have implications concerning the neurobiological mechanisms underlying the behavioural effects of psychostimulant drugs.