To explore the mechanisms of the impairment of dopaminergic neurons by alpha-synuclein ( α-Syn) oligomers in the Parkinson’s disease (PD) mouse model established by gavage with constant low-dose rotenone. Methods: A total of 48 elderly male C57 mice were randomly divided into the rotenone group and control group with 24 mice per group. Mice in the rotenone group were administered 0.01 mL/g rotenone by gavage while mice in the control group were administered 0.01 mL/g chloroform. Mice were sacrificed after 12 weeks of continuous treatment. In mice brains harvested after treatment, western blotting and immunohistochemical staining were carried out to detect α-Syn expression levels; electron microscopy was used to observe the ultrastructure of substantia nigra neurons in the midbrain. In mice brains harvested before and after treatment, immunohistochemical staining of tyrosine hydroxylase (TH) was performed to examine the density of TH-positive neurons in the substantia nigra, and levels of superoxide dismutase (SOD), glutathione peroxidase (GSH-PX), and malondialdehyde (MDA) in the brain tissue were determined. Results: After the 12-week treatment, western blotting demonstrated that α-Syn expression in the midbrain of the rotenone group mice was significantly higher than that of the control group mice (P<0.05). Immunohistochemistry showed that there was a significant decrease in TH-positive neurons in the midbrain of the rotenone group mice compared with that of the control group mice (P<0.05). Electron microscopy showed swollen mitochondria and Golgi in the substantia nigra of the rotenone group mice. Compared to the control group, the rotenone group showed significantly decreased levels of SOD and GSH-PX (P<0.05) and significantly increased levels of MDA (P< 0.05). Conclusion: α-Syn oligomers can form in the brain after continuous intragastric administration of low-dose rotenone. α-Syn oligomers in the midbrain might impair dopaminergic neurons via oxidative stress.