Abstract: AimTo establish the rat model of morphine-induced conditioned place preference (CPP) and to investigate the effects of morphine psychical dependence on the levels of neurosteroids in rat brain. MethodsRats were ip administered morphine 5 mg·kg^-1for 10 days to induce CPP in morphine group. The concentrations of dehydroepiandrosterone (DHEA), pregnenolone (PREG), allopregnanolone (AP), dehydroepiandrosterone sulfate (DS) and pregnenolone sulfate (PS) in nucleus accumbens (Nac), hypothalamus (Ht), amygdale (A) and plasma of rats were determined with liquid chromatography-negative atmospheric pressure ionization mass spectrometry (LC-MS). ResultsTrained with morphine for 10 days resulted in the acquisition of CPP in morphine group with the time that the rats spent in drug-pairing room was longer than that of control group. Compared with control group, morphine treatment could significantly decrease the contents of DHEA in Nac and plasma, decrease that of PREG in Ht. ConclusionMorphine could induce the CPP in rats and affected the contents of some neurosteroids in rat brain, which suggests that endogenous neurosteroids might be related to the development of morphine dependence.