Abstract: 【Objectives】Elevated CO2 has been shown to play a role in enhancing the photosynthesis of plants, and induce stomatal closure of leaf. Stomatal closure significantly decreases plant transpiration, and contributes to enhanced water use efficiency and resistance to water stress. The effect of elevated CO2 on the aperture of stomata, the level of nitric oxide(NO)in guard cells and the role of NO in CO2 elevation-induced stomatal closure in tomato(Solanum lycocarpum L.)were examined. In order to identify the enzymatic source of endogenous NO in guard cells, the role of nitric oxide synthase(NOS)and nitrate reductase(NR)in the CO2 elevation-induced NO accumulation was investigated. 【Methods】 Tomato(Solanum lycocarpum L.)was used as experimental material. In E7/2 growth chambers, CO2 treatments and/or pharmacological experiment were initiated by treating stomata at a concentration of either 350 or 800 μL/L. Then, the stomatal aperture and NO level in guard cells were measured. The levels of NO in guard cells of tomato were determined using the cell NO-specific fluorescent probe. NO levels in guard cells were measured based on the intensity of fluorescence. NOS inhibitor L-NAME and NR inhibitor tungstate were used to assess the role of NOS and NR in the CO2 elevation-induced NO production, respectively. 【Results】 The present study showed that the stomatal aperture decreased to 2.3 μm after 6 hours of elevated CO2 treatment, and decreased by 32% related to ambient CO2 treatment. The intensity of green fluorescence showed that the level of NO in guard cells were 88% higher under elevated CO2 than that under ambient CO2. CO2 elevation-induced stomatal closure was reversed by treatment with NO scavenger cPTIO, the level of NO in guard cells decreased by 35% and the stomatal aperture increased to 3.2 μm, similar to those under ambient CO2. Under elevated CO2, addition of 200 μmol/L L-NAME increased the stomatal aperture by 30%, and decreased NO accumulation in guard cells by 33%; while addition of 100 μmol/L tungstate increased the stomatal aperture by 35% and NO accumulation in guard cells decreased by 40%. 【Conclusions】 Elevated CO2 significantly increased the level of NO in guard cells and decreased aperture of stomata compared to ambient CO2. CO2 elevation-induced stomatal closure was reversed by scavenging NO, indicating that NO plays an important role in the induced stomatal closure of tomato by CO2 elevation. The pharmacological evidences showed that both NOS inhibitor L-NAME and NR inhibitor tungstate significantly decreased NO accumulation in guard cells and inhibited stomatal closure under elevated CO2, suggested that both NOS and NR were involved in CO2 elevation-induced NO accumulation. Therefore, it seems to be concluded that elevated CO2 promotes the production of NO through both NOS and NR, and increases the level of NO in guard cells, and induces stomatal closure in tomato.