Nitric oxide (NO) is generated in substantial amounts by inducible nitric oxide synthase (iNOS) in renal cortex after renal ischemia and reperfusion (I/R). As there was evidence that the renal organic cation transporters rOct1 (Slc22a1) and rOct2 (Slc22a2) might be regulated by NO, we investigated the effect of iNOS specific NO inhibition by L-NIL on the expression and function of the latter transporters after I/R. Experiments were performed in vivo and in vitro by using established model systems of renal I/R-injury. Expression was determined by qPCR. Function was determined by measuring fractional excretion (FE) of the endogenous substrate serotonin in vivo, whereas in vitro MMP was used as a substrate. In vivo L-NIL completely inhibited NO generation after I/R. Moreover, L-NIL completely abolished the ischemia induced down regulation of rOct1 and rOct2. In agreement with expression of the rate limiting organic cation transporters, FE of serotonin was diminished after ischemia which was totally abolished by L-NIL. In vitro, ischemia induced down regulation of rOct1 and rOct2 is also abolished by L-NIL. The same is true for the MMP uptake. In summary we have shown that renal I/R down regulates rOct1 and rOct2, which is mediated via NO. This may be an autocrine effect of the proximal tubular epithelial cells. We furthermore have shown that rOct1 and rOct2 are rate limiting for the renal excretion of serotonin. As these transporters are also down regulated in diabetes we speculate that this may be involved in serotonin induced vascular damage in diabetes.