Introduction: The cases of contrast induced acute kidney injury (CI-AKI) are increasing. A toxic effect of contrast media on the tubular cells is generally accepted as the key feature in CI-AKI(1,2). We investigate the hypothesis that contrast media induce tubular cell death by generating oxidative stress and reducing nitric oxide. Methods: Male Sprague-Dawley rats were killed using isoflurane and isolated thick ascending limbs (TALs) were perfused with either vehicle solution or iodixanol. Nitric oxide bioavailability and superoxide concentration were quantificated by the fluorescent dyes DAF-FM and dihydroethidium (DHE), respectively. Propidium iodide was used to estimate the cell death rate. The expression of oxidative stress related genes and the activity of superoxide dismutase (SOD) in tubuli were investigated using a microarry and the OxiSelect kit in an ex vivo setup. DHE, DAF-FM and Propidium iodide data were compared using Brunner’s test. RNA microarray and the activity of SOD data were compared by the Student’s t-test. Results: Iodixanol increased DHE fluorescence ratio (9.6±1.4%, n=7) compared to the control group (1.7±1.0%, n=8) in 4min, whereas tempol, a superoxide dismutase mimic, inhibited the increase by iodixanol. DAF-FM fluorescence intensity decreased during iodixanol treatment (-0.6±0.6%, n=7) and by nitric oxide synthase inhibitor L-NAME, whilst it increased by 1.6±0.3% in the control group within 12min (n=7). Propidium iodide intensity was significantly enhanced by iodixanol (n=8) compared to the control group (n=6) within 20min. Moreover, during total 4-hour perfusion, iodixanol increased propidium iodide intensity to 17.7±3.1%, while in control group was 7.0±1.0%. Only two of 84 investigated genes related to oxidative stress were differentially expressed in the tubuli. The SOD activity did not differ between iodixanol perfused rats and control animals. Conclusion: The study shows that contrast media increase superoxide concentration, implying oxidative stress in TALs. Further, an impaired nitric oxide bioavailability happens due to contrast media. TAL cells are damaged by contrast media. This damage is not because of changes in the expression of oxidative stress related genes or SOD activity.