Objective To investigate the apoptotic pathway of IL-1β in rat NP cells in vitro. Methods Primary rat NP cells were cocultured with different doeses of IL-1β(0, 10, 20, 50 ng/mL)in 1% FBS medium for 24 h, then the apoptosis incidence was evaluated by flow cytometry with double stain with annexin V-FITC and propidium iodide. The expression levels of Fas, Caspase-8, Caspase-9, Caspase-3, and Bid mRNA were detected by RT-PCR, the enzyme activity of Caspase-8, Caspase-9, and Caspase-3 was evaluated with enzyme activity detection kit, membrane potential change of mitochondria was determined using fluorescence microscope and flow cytometry. Results The apoptosis incidence, the expression levels of Caspase-9 and Caspase-3 mRNA, the enzyme activity of Caspase-9 and Caspase-3, and the membrane potential change of mitochondria of rat NP cells in vitro increased with the dose of IL-1β. The mRNA expression level of Caspase-8 and Bid, and the enzyme activity of Caspase-8 increased by 50 mg/mL IL-1β significantly. Conclusion IL-1β can induce rat NP cells apoptosis through mitochondrion and membrane apoptotic pathway. Mitochondrion apoptotic pathway is the main one, and membrane apoptotic pathway can be activated by a higher dose of IL-1β.