Previous studies have indicated that angiotensin II (Ang II) concentrations in renal interstitial fluid are much higher than plasma levels. In the present study, we performed experiments to explore renal interstitial fluid concentrations of Ang I and Ang II further and to determine whether these levels are altered by acute arterial infusion of an ACE inhibitor (enalaprilat) or by volume expansion. Microdialysis probes (molecular weight cutoff point: 30 000 Da) were implanted in the renal cortex of anesthetized rats and were perfused at a rate of 2 &mgr;L/min. Using relative equilibrium rates, the basal renal interstitial fluid Ang II concentration averaged 3.07±0.43 nmol/L, a value much higher than the plasma Ang II concentration of 107±8 pmol/L (n=7). Interstitial fluid Ang I concentrations (0.84±0.04 nmol/L) were consistently lower than the Ang II concentrations but higher than the plasma Ang I concentrations (112±14 pmol/L). Intra-arterial infusion of enalaprilat (7.5 &mgr;mol/kg/min, n=5) for 120 minutes resulted in a significant decrease in mean arterial pressure (from 114±4 to 68±4 mm Hg) along with reductions in plasma and renal ACE activity (by −99% and −52%, respectively). Enalaprilat resulted in a significant increase in plasma Ang I from 133±21 to 1167±328 pmol/L and a decrease in plasma Ang II from 110±12 to 67±9 pmol/L. During enalaprilat infusion, interstitial fluid concentration of Ang I was significantly increased from 0.78±0.06 to 0.97±0.08 nmol/L; however, Ang II concentrations were not altered significantly (3.67±0.28 versus 3.67±0.25 nmol/L). Acute volume loading with Ringer’s solution containing 1% bovine serum albumin at a rate of 150 &mgr;L/min for 2 hours (6% to 7% of body weight) lowered plasma concentrations of Ang I from 110±23 to 16±2 pmol/L and Ang II from 100±23 to 36±6 pmol/L; however, renal interstitial fluid concentrations of Ang I and Ang II were not altered significantly during volume expansion (Ang I, from 0.77±0.05 to 0.69±0.03 nmol/L; Ang II, from 3.76±0.43 to 3.59±0.39 nmol/L, n=5). These data indicate that renal interstitial fluid concentrations of Ang I and Ang II are substantially higher than the corresponding plasma concentrations. Furthermore, the fact that the high interstitial fluid concentrations of Ang II are not responsive to acute ACE inhibition or volume expansion suggests the compartmentalization and independent regulation of renal interstitial fluid Ang II.