Based on their rearing response to a novel open field, male Wistar rats were divided into two sub-groups with either high or low behavioural activity (high rearings, versus low rearings). These sub-groups were repeatedly exposed to the same open field and tested for behavioural habituation. Since we previously found neurochemical evidence for different cholinergic reactivities in such high rearing and low rearing rats, their behavioural responses to the muscarinic antagonist scopolamine (0.5 mg/kg) were also investigated in the open field. Additionally, they were exposed to the elevated plus-maze to test for possible differences in measures of anxiety. After behavioural testing, tissue concentrations of biogenic amines were determined in the ventral striatum (nucleus accumbens, olfactory tubercle), frontal cortex, striatum, hippocampus and amygdala. The results show that the higher rearing responses of high rearing rats in the novel open field were paralleled by higher locomotor activity. These behavioural differences between groups decreased with repeated open field exposure, an effect which was largely due to between-session habituation in high rearing rats. Thereby, high rearing rats approached the lower levels of low rearing rats, in which locomotor activity and rearings did not habituate between testing. Nevertheless, habituation was also observed in low rearing rats, especially in the measure of thigmotactic scanning, since the levels of scanning declined both between and within test sessions. The anticholinergic challenge with scopolamine induced a general pattern of behavioural activation. Furthermore, scopolamine partly reinstated the behavioural differences between high and low rearing rats that had been observed in the novel open field, since high rearing rats showed more rearing behaviour than low rearing rats under scopolamine. In contrast to the open field, there were no significant differences between high and low rearing rats in the plus-maze. The neurochemical analysis revealed, among others, higher dopamine levels in the ventral striatum of high rearing rats together with lower serotonin levels in the medial frontal cortex. The current findings thus indicate that high and low rearing rats not only differ in their behavioural response to a novel environment, but also in their patterns of behavioural habituation, and with respect to behaviour induced by an anti-cholinergic challenge. These differential behavioural profiles of high and low rearing animals are discussed with respect to the role of dopaminergic mechanisms in the forebrain, and the potential impact of cholinergic mechanisms.