Objective To investigate the combined effect of CdCl₂ and nicotine on the cell cycle and apoptosis of mouse spermatogonium (GC-1). Methods A CCK-8 assay was used to detect the inhibition of GC-1 cell proliferation in the CdCl₂ groups, nicotine groups, and combined groups at 24 hours. Flow cytometry was used to detect the effects of CdCl₂, nicotine, and the combined treatment on the cell cycle and apoptosis of GC-1 cells at 24 hours. Results The inhibitory effect on the proliferation of GC-1 cells increased with the rise in CdCl₂ and nicotine concentrations. The IC₅₀ values of CdCl₂ and nicotine alone were 5.409 μmol/L and 2814 μmol/L, respectively. The IC₅₀ values of CdCl₂ and nicotine combined for GC-1 cells were 4.422, 4.532, 3.309, and 2.532 μmol/L at nicotine concentrations of 0.175, 0.350, 0.700, and 1.400 mmol/L, respectively, thus there was an obvious decrease with nicotine concentration. The IC₅₀ values were lower than those of the group administered CdCl₂ alone, and the combined action of CdCl₂ and nicotine had synergistic effects. Cell cycle result showed that CdCl₂ concentration of 2.5 μmol/L increased the percentage of G2 /M phase cells in the combination groups. Cells were arrested in the G2 /M phase in the combined groups (P<0.01). When combined, CdCl₂ and nicotine had a synergistic effect on the cell cycle, and the synergistic effect of CdCl₂ was stronger. The result of apoptosis showed the proportion of cells in apoptosis increased in the 2.5 μmol/L CdCl₂ group. Compared with the CdCl₂ and nicotine single-treatment groups, the CdCl₂ and nicotine combined group’s percentage of cells in apoptosis increased significantly(P<0.01), and CdCl₂ had a stronger synergistic effect. Conclusions Combinations of CdCl₂ and nicotine had a synergistic effect on mouse spermatogonium(GC-1)and led to enhanced cytotoxicity, G2 /M cell cycle arrest, and cell apoptosis. CdCl₂ was the main effect factor in the synergistic effect.