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The use of animal models (particularly rats) in research for developing drugs for central nervous system diseases is well validated. However a range of strains are often utilised in these models. The Lister-Hooded (LH) strain is beginning to be increasingly used in preclinical investigations. Thus, the objective of the present study was to investigate the comparative behavioural pharmacology of this strain, with the two most widely used rat strains, namely the Sprague–Dawley (SD), and Wistar (W) strains. The tests used were the forced swim test (FST) for antidepressants, the amphetamine-locomotor activity test for antipsychotics, the elevated plus maze (EPM) for anxiolytics, as well as tests of general locomotor activity using home cage monitoring (HCM) and the open field test. Continuous HCM revealed a significantly higher daily activity and lower nocturnal activity for LH compared to the other strains; there were no strain-related differences in the open field test. In the FST, there were no strain differences in immobility time and a similar magnitude of desipramine-induced reduction in immobility across strains. In the locomotor activity test, control LH rats showed significantly higher activity whilst significant amphetamine-induced hyperactivity was seen only with the LH and W strains. In the EPM, control LH rats had a significantly larger percentage of open arm entries, whilst only the SD strain displayed a significant diazepam-induced increase in this parameter. These findings suggest that strain variation can cause markedly different results in behavioural pharmacological tests where locomotor activity plays a significant role, and should be taken into account when selecting a strain for evaluating the behavioural effects of psychotropic drugs. Such differences in locomotor activity in the LH strain could be accounted for by an altered diurnal pattern in this strain.