The anxiolytic potential of melatonin and agomelatine, a potent MT(1/2) receptor agonist, and their combined effects with diazepam, were investigated in rats using the punished drinking test, the safety signal withdrawal operant paradigm, the elevated-plus-maze and hypophagia-induced novelty. In the punished drinking test, evening injections of melatonin (80 mg/kg, IP, but not 20 and 40 mg/kg) and agomelatine (40 mg/kg, IP) increased the number of foot shocks received. However, neither melatonin (40-80 mg/kg) nor agomelatine (20-40 mg/kg) released response suppression during the period associated with the safety signal withdrawal and affected rats' behaviour in the elevated-plus-maze. Furthermore, agomelatine (40 mg/kg) did not enhance food consumption in unfamiliar environment. However, the co-administration of melatonin (80 mg/kg) or agomelatine (20-40 mg/kg) with diazepam, at a dose (0.25 mg/kg) inactive on its own, induced an anxiolytic-like effect in the punished drinking test and the elevated plus-maze. These results indicate that, although mostly devoid of anxiolytic-like action per se, melatonin and agomelatine can potentiate the anxiolytic effects of diazepam.