Nitric oxide (NO) is a potent regulator of both vascular tone and oxygen utilization. Diabetes is commonly associated with both NO deficiency and reduced renomedullary oxygen availability. Arginine availability as regulator of NO production has gained growing interest. We hypothesized that arginine limitation causes diabetes-induced renomedullary NO deficiency, which directly influences renomedullary oxygen tension (pO₂). Medullary NO, pO₂and blood flow were measured in control and streptozotocin-induced diabetic rats, which were treated or not treated with -tocopherol, and administered L-arginine followed by L-NAME. Major components of the arginine metabolism were also investigated. Diabetic rats had reduced renomedullary NO levels compared to controls. Arginine selectively increased NO levels in diabetic rats, and totally restored NO levels in -tocopherol-treated animals. -ocopherol prevented the reduction in medullary pO₂in the diabetic animals. Although blood flow increased equally in all groups, arginine increased pO₂exclusively in the diabetic groups. Diabetes decreased plasma arginine and ADMA concentrations, but increased hepatic CAT-2A and plasma ornithine independently of -tocopherol treatment. In conclusion, diabetic rats had reduced renomedullary NO due to decreased plasma arginine following increased hepatic arginine uptake and degradation. This was unrelated to oxidative stress. The diabetes-induced reduction in renomedullary pO₂was restored by either acute arginine administration, which also restored NO levels, or long-term antioxidant treatment. Arginine increased medullary NO and pO₂independently of altered hemodynamics in the diabetics. This reveals a direct regulatory function of NO for renomedullary pO₂especially during situations of elevated oxidative stress.