The present study was designed to determine whether arginine vasopressin (AVP) can stimulate nitric oxide (NO) production within the renal medulla and thereby modulate renal medullary blood flow. An in vivo microdialysis/NO trapping technique was used to determine changes in medullary interstitial [NO]. AVP (2 ng/kg per minute) was delivered into the renal medullary interstitium and resulted in a significant increase in renal medullary [NO] of 35%, which was blocked by pretreatment with nitro-L-arginine methyl ester (L-NAME) (1.3 microg/kg per minute) administered into the renal medullary interstitium. The vasopressin V2 receptor agonist 1-desamino-8-D-arginine vasopressin (dDAVP) resulted in a significant increase of 32% in renal medullary interstitial [NO]. No change in renal medullary interstitial [NO] was observed after selective vasopressin V1 receptor stimulation. Laser-Doppler flowmetry with implanted optical fibers was performed to measure cortical and medullary blood flow changes within the kidney. Renal interstitial infusion of dDAVP in rats pretreated with a vasopressin V1 receptor antagonist resulted in a 15% increase (P<0.05) in medullary blood flow, which was completely blocked by pretreatment with L-NAME (1.3 microg/kg per minute). This study demonstrates that AVP increases renal medullary interstitial [NO] through vasopressin V2 receptor stimulation, which in turn elevates blood flow to the renal medulla.