AIM: To study caspase-3 gene expression and [Ca2+]i homeostasis in verapamil (Ver)-induced human retinal pigment epithelium (RPE) cells apoptosis. METHODS: Ver 80mg/L was applied in cultured human RPE cells for 12, 24 and 48 hours to induce RPE cells apoptosis. The expression of apoptotic effector gene caspase-3 was assessed by reverse transcription polymerase chain reaction (RT-PCR). Single cell was measured using fluorescence indicator Fura-3/AM with MetaFluo4.5/coolsnapfx/IX70 intracellular Ca2+ fluorescence imaging system. RESULTS: High levels of expression of caspase-3 mRNA were observed in normal RPE cells and it significantly increased after co-cultured with Ver. The fluorescence in resting RPE cells was strong and distributed throughout the cells. The nucleus appeared more fluorescent than the cytoplasm. Calcium fluorescence of RPE cells attenuated after co-cultured with Ver. CONCLUSION: Up-regulation of caspase-3 gene expression and disturbance of [Ca2+]i homeostasis might play pivotal roles in Ver-induced RPE cells apoptosis.