Renal Interstitial Fluid Research Articles

Renal interstitial corticosterone and 11‐dehydrocorticosterone in conscious rats

Genetic or induced abnormalities in the conversion between active and inactive glucocorticoids in the kidney can lead to local glucocorticoid excess and hypertension. It has been difficult, however, to directly assess local glucocorticoid levels in the kidney. We used chronic microdialysis techniques to sample renal cortical and medullary interstitial fluid from conscious rats. The level of corticosterone (compound B, active) and 11-dehydrocorticosterone (compound A, inactive) was analyzed with liquid chromatography-tandem mass spectrometry (LC-MS/MS). Levels of B (ng/ml, n = 13–16) were 0.8 ± 0.1, 1.0 ± 0.1, 66.7 ± 8.1, and 7.9 ± 1.1 in cortical microdialysate, medullary microdialysate, the plasma, and urine, respectively. The B/A ratio, an index of 11 β-hydroxysteroid dehydrogenase activity, was 0.8 ± 0.1, 0.6 ± 0.1, 10.6 ± 1.4, and 1.7 ± 0.1, respectively, in those four types of sample. Microdialysate levels of B were approximately 55% higher in afternoons than in mornings, while plasma levels differed by 2.8 fold. An increase of dietary salt content from 0.4% to 4% did not significantly alter levels of B or B/A ratios in renal microdialysate. The protein abundance (fold) of 11 β-hydroxysteroid dehydrogenase type 2, which inactivates B to A, was 1.0 ± 0.2, 1.6 ± 0.4, and 0.7 ± 0.3 in the cortex, the outer medulla, and the inner medulla, respectively. Type 1 of the enzyme, which likely reactivates A to B, was 1.0 ± 0.1, 1.3 ± 0.2, and 2.0 ± 0.3 in the three regions. Renal interstitial levels of active and inactive glucocorticoids provide an important basis for understanding local metabolism of glucocorticoids in the kidney. (NIH R01HL077263)

Renal interstitial fluid ATP responses to arterial pressure and tubuloglomerular feedback activation during calcium channel blockade

A close relationship between changes in renal interstitial fluid (RIF) ATP concentrations and renal autoregulatory or tubuloglomerular feedback (TGF)-dependent changes in renal vascular resistance (RVR) has been demonstrated, but it has not been determined whether the changes in RIF ATP are a consequence or the cause of the changes in RVR. The present study was performed in anesthetized dogs to assess the changes in RIF ATP following changes in renal arterial pressure (RAP) or stimulation of the TGF mechanism under conditions where changes in RVR were prevented by nifedipine, a calcium channel blocker. RIF ATP levels were measured by using microdialysis probes. Intra-arterial infusion of nifedipine (0.36 microg x kg(-1) x min(-1)) increased renal blood flow (RBF: from 4.49 +/- 0.27 to 5.34 +/- 0.39 ml x min(-1) x g(-1)) and glomerular filtration rate (GFR: from 0.84 +/- 0.07 to 1.09 +/- 0.11 ml x min(-1) x g(-1)). Under conditions of nifedipine infusion, autoregulatory adjustments in RBF, GFR, and RVR were not observed during stepwise reductions in RAP within the autoregulatory range (from 135 +/- 7 to 76 +/- 1 mmHg, n = 7). Furthermore, stimulation of the TGF mechanism with intra-arterial infusion of acetazolamide (100 microg x kg(-1) x min(-1)) did not alter RBF, GFR, and RVR (n = 7). During treatment with nifedipine, RIF ATP levels were significantly decreased in response to reductions in RAP (10.7 +/- 0.7, 5.8 +/- 0.7 and 2.8 +/- 0.3 nmol/l at 135 +/- 7, 101 +/- 4, and 76 +/- 1 mmHg, n = 7) and increased by acetazolamide infusion (from 8.8 +/- 0.8 to 17.0 +/- 1.8 nmol/l, n = 7). These results are similar to those that occurred in dogs not treated with nifedipine and thus demonstrate that the changes in RIF ATP can occur in the absence of autoregulatory or TGF-mediated changes in RVR. The data provide further support to the hypothesis that RIF ATP contributes to adjustments in RVR associated with renal autoregulation and changes in activity of the TGF mechanism.

Increased renal production of angiotensin II and thromboxane B 2 in conscious diabetic rats

The mechanisms involved in development of cardiovascular complications associated with diabetes mellitus are not well elucidated. Among the vasoactive factors that may play a role in development of these complications are angiotensin II and thromboxane B2 (TXB2). We hypothesized that diabetes increases renal production of TXB2 through stimulation of angiotensin type-1 receptor. We used a microdialysis technique to monitor changes in renal interstitial fluid (RIF) TXB2 in conscious streptozotocin-induced diabetes rat model. The RIF levels of angiotensin II and TXB2 were monitored before and during 6 weeks after development of diabetes and during treatment with the angiotensin type-1 receptor blocker valsartan at 10 mg/kg. Measurement of the urinary albumin excretion (UAE) was used to monitor the development and progression of diabetic nephropathy. The UAE was 81.62 +/- 1.31 ng/min, 184.75 +/- 9.41 ng/min (P < .01), and 229.84 +/- 4.49 ng/min (P < .0001) at baseline, week 3, and week 6, respectively, after induction of diabetes. Basal levels of RIF angiotensin II were 4.28 +/- 0.02 pg/mL and significantly increased to 6.24 +/- 0.31 pg/mL (P < .001) and 7.66 +/- 0.05 pg/mL (P < .001) at 3 and 6 weeks after development of diabetes. Similarly, basal RIF TXB2 was 197 +/- 27 pg/mL and increased to 488 +/- 80 pg/mL (P < .01) and 703 +/- 130 pg/mL (P < .01) at 3 and 6 weeks after development of diabetes. Valsartan caused further increase in RIF angiotensin II levels. In contrast, valsartan decreased RIF TXB2 levels at baseline to 85 +/- 11 pg/mL (P < .01), at 3 weeks to 141 +/- 17 pg/mL (P < .01), and at 6 weeks to 255 +/- 45 pg/mL (P < .01) after development of diabetes. These results demonstrate that diabetes mellitus is accompanied by increased renal production of angiotensin II and TXB2. The increase in TXB2 is mediated through stimulation of angiotensin type-1 receptor.

MICROFILAMENTS ARE INVOLVED IN RENAL CELL RESPONSES TO SUSTAINED HYDROSTATIC PRESSURE

Increased pressures within renal interstitial fluid, as associated with a number of renal pathologies, could affect cell function and gene expression. The long-term objective of this research is to elucidate kidney cell responses to pathological hydrostatic pressures. In vitro studies were performed in 2 kidney cell lines (cortical tubular and medullary) to determine changes in cell numbers and cytoskeletal (specifically microfilament, microtubule and intermediate filament) arrangement following exposure to pathological (60 cm H2O) pressures. A novel pressure system was used to apply pressure to renal cells for up to 7 days. Cell counts and fluorescent staining were performed to determine alterations in response to pressure. Exposure to pressures of 60 cm H2O resulted in increased renal cell numbers and rearrangement in individual microfilament structures after 7 days. These results prove that hydrostatic pressure alters the function of renal cells. In the future such knowledge of renal cell responses to pressure along with an understanding of the mechanisms involved will aid in the design of novel, targeted drug therapies for treating kidney pathologies.

Blood flow-dependent changes in intrarenal nitric oxide levels during anesthesia with halothane or sevoflurane

We previously demonstrated that intrarenal nitric oxide (NO) levels and renal blood flow are reduced during halothane anesthesia. Studies were performed to determine if volatile anesthetics-induced reductions in renal NO levels are associated with blood flow changes. Halothane and sevoflurane at 0.8 and 2.4 Mac were administered by inhalation to dogs, and cGMP and NOx concentrations in the renal interstitial fluid were measured by a microdialysis method. Neither halothane nor sevoflurane at 0.8 Mac altered renal blood flow and renal interstitial cyclic guanosine monophosphate (cGMP) and NOx levels, but both anesthetics significantly decreased these values at 2.4 Mac. Using an adjustable aortic clamp, renal perfusion pressure was reduced in 2 steps without halothane and sevoflurane anesthesia. Renal blood flow as well as cGMP and NOx concentrations in the renal interstitial fluid were unchanged within the autoregulatory range, but significantly decreased below the autoregulatory range. Changes in cGMP and NOx concentrations in the renal interstitial fluid were highly correlated with renal blood flow changes during halothane or sevoflurane anesthesia, and during stepwise reductions in renal perfusion pressure. The results suggested that halothane- and sevoflurane-induced decreases in intrarenal NO levels result from reductions in blood flow.

Renal nitric oxide production is decreased in diabetic rats and improved by AT1 receptor blockade.

Diabetes mellitus is associated with increased incidence of cardiovascular complications. Lack of nitric oxide production may exacerbate these complications. We hypothesized that diabetes decreases renal nitric oxide (NO) production, an effect that is reversed via inhibition of angiotensin subtype-1 receptor. We monitored changes in renal interstitial fluid nitric oxide by a microdialysis technique in the renal cortex of conscious Sprague-Dawley rats. Rats (n = 8 each group) were given streptozotocin 30 mg/kg intravenously to induce diabetes. Changes in renal interstitial fluid angiotensin II and NO were evaluated at baseline before and over 12 weeks during the development of diabetes and at 4 and 8 h after oral administration of the angiotensin subtype-1 (AT1) receptor blockers, losartan (30 mg/kg) or valsartan (10 mg/kg). Renal interstitial fluid angiotensin II significantly increased after development of diabetes. In contrast, basal renal interstitial fluid nitric oxide decreased significantly over 12 weeks after development of diabetes. Both losartan and valsartan caused a further increase in renal angiotensin II levels. Some 4 h after administration, there was significantly greater increase in renal nitric oxide after administration of valsartan than of losartan. At 8 h post- treatment, only valsartan caused a significant increase in renal nitric oxide levels. These results demonstrate that diabetes mellitus is associated with an increase in renal production of angiotensin II, while renal production of nitric oxide is reduced. The decrease in renal NO is reversed by AT1 receptor blockade.

Why are angiotensin concentrations so high in the kidney?

Recent studies have reported that intrarenal angiotensin II content and angiotensin II concentrations in the proximal tubular fluid and renal interstitial fluid are much greater than the circulating angiotensin II levels. These high intrarenal angiotensin II levels are responsible for regulating renal hemodynamics and tubular transport. Intrarenal angiotensin II levels have been assessed from total tissue contents as well as renal interstitial fluid and proximal tubular fluid concentrations. Total tissue contents expressed per gram of tissue weight are greater than plasma angiotensin II concentrations; tubular fluid concentrations and renal interstitial fluid concentrations are even greater in the range of 3-10 pmoles/ml. In hypertensive states, there is also an increased intracellular accumulation of angiotensin II mediated by angiotensin type 1 receptor-dependent endocytosis. The high intrarenal angiotensin II levels are also caused by the presence of angiotensinogen messenger RNA and protein in the proximal tubule cells. Furthermore, there is positive amplification by which increases in circulating angiotensin II stimulate increased production and secretion of angiotensinogen, which is also manifested as an increased urinary excretion rate. The ability of the kidney to generate high intratubular and interstitial concentrations allows the kidney to regulate intrarenal levels in accord with the homeostatic needs for the regulation of renal hemodynamics and tubular reabsorption and the regulation of sodium balance. When inappropriately stimulated, high intrarenal angiotensin II levels contribute to excessive salt and water retention, the development of hypertension, and long-term proliferative effects leading to renal injury.

Angiotensin II type 1 receptor-mediated augmentation of renal interstitial fluid angiotensin II in angiotensin II-induced hypertension.

Angiotensin II (Ang II)-dependent hypertension is associated with augmented intrarenal concentrations of Ang II; however, the distribution of the increased intrarenal Ang II has not been fully established. To determine the changes in renal interstitial fluid Ang II concentrations in Ang II-induced hypertension and the consequences of treatment with an angiotensin II type 1 (AT1) receptor blocker. Rats were selected to receive vehicle (5% acetic acid subcutaneously; n = 6), Ang II (80 ng/min subcutaneously, via osmotic minipump; n = 7) or Ang II plus an AT1 receptor antagonist, candesartan cilexetil (10 mg/kg per day, in drinking water; n = 6) for 13-14 days, at which time, experiments were performed on anesthetized rats. Microdialysis probes were implanted in the renal cortex and were perfused at 2 microl/min. The effluent dialysate concentrations of Ang I and Ang II were measured by radioimmunoassay and reported values were corrected for the equilibrium rates at this perfusion rate. Ang II-infused rats developed greater mean arterial pressures (155 +/- 7 mmHg) than vehicle-infused rats (108 +/- 3 mmHg). Ang II-infused rats showed greater plasma (181 +/- 30 fmol/ml) and kidney (330 +/- 38 fmol/g) Ang II concentrations than vehicle-infused rats (98 +/- 14 fmol/ml and 157 +/- 22 fmol/g, respectively). Renal interstitial fluid Ang II concentrations were much greater than plasma concentrations, averaging 5.74 +/- 0.26 pmol/ml in Ang II-infused rats - significantly greater than those in vehicle-infused rats (2.86 +/- 0.23 pmol/ml). Candesartan treatment prevented the hypertension (87 +/- 3 mmHg) and led to increased plasma Ang II concentrations (441 +/- 27 fmol/ml), but prevented increases in kidney (120 +/- 15 fmol/g) and renal interstitial fluid (2.15 +/- 0.12 pmol/ml) Ang II concentrations. These data indicate that Ang II-infused rats develop increased renal interstitial fluid concentrations of Ang II, which may contribute to the increased vascular resistance and reduced sodium excretion. Furthermore, the augmentation of renal interstitial fluid Ang II is the result of an AT1 receptor-mediated process and can be dissociated from the plasma concentrations.

Urinary and renal interstitial concentrations of TNF-α increase prior to the rise in albuminuria in diabetic rats

The development of diabetic nephropathy has been linked to the release of vasoactive hormones and growth factors. Currently the role of inflammatory cytokines in this pathogenic process is not clear. We utilized the microdialysis technique to monitor early changes in tumor necrosis-alpha (TNF-alpha) levels in the renal interstitial fluid and urine of conscious Sprague-Dawley rats (N = 8) before and after induction of diabetes with streptozotocin (STZ). Measurement of the urinary albumin excretion (UAE) was utilized to monitor the development and progression of diabetic nephropathy. UAE increased from 0.56 +/- 0.20 microg/min to 8.14 +/- 2.98 microg/min 17 days after induction of diabetes (P = 0.01). Renal interstitial fluid TNF-alpha increased from 11.96 +/- 5.32 pg/mL at baseline to 45.02 +/- 11.69 pg/mL 5 days after induction of diabetes (P = 0.03). Renal interstitial fluid TNF-alpha levels remained elevated throughout the remainder of the study period. Urinary TNF-alpha also increased significantly compared to baseline 3 days after induction of diabetes (294.18 +/- 36.94 pg/mL vs. 16.05 +/- 6.07 pg/mL, P < 0.002). There was a second significant rise in urinary TNF-alpha concentration to 638.16 +/- 36.94 pg/mL 21 days after induction of diabetes (P < 0.001). Serum TNF-alpha levels were undetectable before STZ injection and remained undetectable by the end of the study. Urinary and renal interstitial fluid TNF-alpha in the control rats (N = 5) did not change throughout the study. We found an early rise in renal TNF-alpha levels after induction of diabetes with STZ, which precedes the rise in UAE by about 2 weeks. These findings suggest a possible contribution of TNF-alpha in the complicated pathogenic process resulting in microalbuminuria in diabetes.

Angiotensin AT 2 Receptors Directly Stimulate Renal Nitric Oxide in Bradykinin B 2 -Receptor–Null Mice

Both bradykinin B2 and angiotensin II type 2 (AT2) receptors are known to stimulate renal production of nitric oxide (NO). To evaluate the individual contributions of AT2 and B2 receptors to renal NO production, we monitored renal interstitial, stable NO metabolites and cGMP by a microdialysis technique in conscious, bradykinin B2-null and wild-type mice (n=8 in each group) during low sodium intake alone or with the angiotensin AT1 or AT2 receptor blockers, valsartan (0.5 microg/min) or PD123319 (0.15 microg/min), or both. During normal salt intake, renal interstitial fluid NO and cGMP levels in B2-null mice were not different from those of wild-type mice. Low sodium intake increased NO and cGMP in wild-type mice but not in B2-null mice. Valsartan increased NO and cGMP in both wild-type and B2-null mice but to a significantly greater degree in the wild-type than in B2-null mice. PD123319 decreased NO and cGMP in both wild-type and B2-null mice. Combined valsartan and PD123319 decreased NO and cGMP in both wild-type and B2-null mice, but there was no significant difference during combined treatment from their levels after administration of PD123319 alone. Our results indicate that during ingestion of a low-salt diet, production of NO is mediated mainly via the AT2-B2 receptor cascade. Blockade of the AT1 receptor enhances the production of NO via the AT2 receptor in both wild-type and B2-null mice. We conclude that NO can be produced by 2 alternative pathways: directly through the AT2 receptor or indirectly from AT2 receptor stimulation of bradykinin via the B2 receptor.

The angiotensin II type 1 receptor mediates renal interstitial content of tumor necrosis factor-alpha in diabetic rats.

A unique microdialysis technique was used to demonstrate that increased levels of angiotensin II (Ang II) and consequent stimulation of the Ang II type 1 (AT(1)) receptor increase the renal content of TNF-alpha in diabetes. Recovery of Ang II and TNF-alpha in renal interstitial fluid (RIF) was measured in conscious rats before and weekly for 12 wk after induction of diabetes with streptozocin and in response to oral valsartan (10 mg/kg.d). Recovery of Ang II in RIF was significantly higher in diabetic rats than in nondiabetic rats. In diabetic rats, RIF recovery of TNF-alpha increased by approximately 67% over baseline, whereas it was unchanged in nondiabetic rats. AT(1) receptor blockade with valsartan prevented the increase in TNF-alpha in the diabetic group. This study shows that diabetes is associated with an increase in the vasoconstrictive hormone Ang II and the inflammatory cytokine TNF-alpha, both of which play a role in accelerating renal function decline in diabetic nephropathy. The study also confirms that valsartan reduces intrarenal level of TNF-alpha by acting on Ang II at the AT(1) receptor level. This finding of a potential antiinflammatory effect for valsartan is new and in addition to its known antihypertensive effects.

Renal Interstitial Fluid Angiotensin I and Angiotensin II Concentrations during Local Angiotensin-Converting Enzyme Inhibition

It was recently demonstrated that angiotensin II (AngII) concentrations in the renal interstitial fluid (RIF) of anesthetized rats were in the nanomolar range and were not reduced by intra-arterial infusion of an angiotensin-converting enzyme (ACE) inhibitor (enalaprilat). This study was performed to determine changes in RIF AngI and AngII concentrations during interstitial administration of ACE inhibitors (enalaprilat and perindoprilat). Studies were also performed to determine the effects of enalaprilat on the de novo formation of RIF AngII elicited by interstitial infusion of AngI. Microdialysis probes (cut-off point, 30,000 D) were implanted in the renal cortex of anesthetized rats and were perfused at 2 micro l/min. The effluent dialysate concentrations of AngI and AngII were measured by RIA, and reported values were corrected for the equilibrium rates at this perfusion rate. Basal RIF AngI (0.74 +/- 0.05 nM) and AngII (3.30 +/- 0.17 nM) concentrations were much higher than plasma AngI and AngII concentrations (0.15 +/- 0.01 and 0.14 +/- 0.01 nM, respectively; n = 27). Interstitial infusion of enalaprilat through the microdialysis probe (1 or 10 mM in the perfusate; n = 5 and 8, respectively) significantly increased RIF AngI concentrations but did not significantly alter AngII concentrations. However, perindoprilat (10 mM in the perfusate, n = 7) significantly decreased RIF AngII concentrations by 22 +/- 4% and increased RIF AngI concentrations. Interstitial infusion of AngI (100 nM in the perfusate, n = 7) significantly increased the RIF AngII concentration to 8.26 +/- 0.75 nM, whereas plasma AngI and AngII levels were not affected (0.15 +/- 0.02 and 0.14 +/- 0.02 nM, respectively). Addition of enalaprilat to the perfusate (10 mM) prevented the conversion of exogenously added AngI. These results indicate that addition of AngI in the interstitial compartment leads to low but significant conversion to AngII via ACE activity (blocked by enalaprilat). However, the addition of ACE inhibitors directly into the renal interstitium, via the microdialysis probe, either did not reduce RIF AngII levels or reduced levels by a small fraction of the total basal level, suggesting that much of the RIF AngII is formed at sites not readily accessible to ACE inhibition or is formed via non-ACE-dependent pathways.

Potential roles of interstitial fluid adenosine in the regulation of renal hemodynamics

Adenosine exerts membrane receptor-mediated effects on renal hemodynamics and function. Endogenously produced adenosine is proposed to be formed primarily by renal tubular epithelial cells and approaches the vascular smooth cells of the renal vasculature from the extracellular space, i.e. the interstitium. Hence, it is important to determine the dynamics of adenosine levels in renal interstitial fluid. Indeed, many studies have indicated that the effects of adenosine on renal hemodynamics depend on its concentration in the interstitium. Furthermore, in vitro studies reveal that adenosine in the renal interstitial fluid binds to membrane receptors from the adventitial side and initiates renal microvascular responses. Recently, we developed a renal microdialysis method to monitor the concentration of adenosine in renal interstitial fluid. This method is well suited for in vivo studies investigating alterations in renal interstitial adenosine levels. In this review, we summarize the literature pertaining to alterations in renal interstitial fluid concentrations of adenosine in some pathophysiological conditions. Potential roles of interstitial adenosine for regulating renal hemodynamics were also discussed.

Regulation of intrarenal angiotensin II in hypertension.

Intrarenal angiotensin II (Ang II) is regulated by several complex processes involving formation from both systemically delivered and intrarenally formed substrate, as well as receptor-mediated internalization. There is substantial compartmentalization of intrarenal Ang II, with levels in the renal interstitial fluid and in proximal tubule fluid being much greater than can be explained from the circulating levels. In Ang II--dependent hypertension, elevated intrarenal Ang II levels occur even when intrarenal renin expression and content are suppressed. Studies in Ang II--infused rats have demonstrated that augmentation of intrarenal Ang II is due, in part, to uptake of circulating Ang II via an Ang II type 1 (AT(1)) receptor mechanism and also to sustained endogenous production of Ang II. Some of the internalized Ang II accumulates in the light and heavy endosomes and is therefore potentially available for intracellular actions. The enhanced intrarenal Ang II also exerts a positive feedback action to augment intrarenal levels of angiotensinogen (AGT) mRNA and protein, which contribute further to the increased intrarenal Ang II in hypertensive states. In addition, renal AT(1) receptor protein and mRNA levels are maintained, allowing increased Ang II levels to elicit progressive effects. The increased intrarenal Ang II activity and AGT production are associated with increased urinary AGT excretion rates. The urinary AGT excretion rates show a clear relationship to kidney Ang II content, suggesting that urinary AGT may serve as an index of Ang II--dependent hypertension. Collectively, the data support a powerful role for intrarenal Ang II in the pathogenesis of hypertension.

Renal interstitial fluid concentrations of angiotensins I and II in anesthetized rats.

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 microL/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 micromol/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 microL/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.

Effects of Angiotensin II on the Renal Interstitial Concentrations of NO2 /NO3 and Cyclic GMP in Anesthetized Rats

The present study was conducted to determine whether exogenous angiotensin II (Ang II) may increase the renal interstitial fluid concentrations of NO2/NO3 (NOx) and cyclic guanosine monophosphate (cGMP) concomitantly and which Ang II receptor subtypes may induce these changes in anesthetized rats, using a microdialysis method. Ang II (50 ng/kg per min, i.v.) significantly increased mean blood pressure (MBP), extraction rates of renal interstitial NOx from 23.9+/-1.0 to 31.2+/-1.9 pmol/min, and cGMP from 4.1+/-0.3 to 6.4+/-0.5 fmol/min, and decreased renal blood flow (RBF). The AT1-receptor antagonist CV11974 alone significantly increased RBF, but did not alter MBP, renal interstitial concentrations of NOx and cGMP. A superimposition of Ang II on CV11974 did not affect MBP and RBF, but significantly increased renal interstitial concentrations of NOx and cGMP. The AT2-receptor antagonist PD123319 alone did not change any of the parameters. However, superimposition of Ang II on PD123319 increased MBP and decreased RBF without any effects on renal interstitial concentrations of NOx and cGMP. These results suggest that Ang II stimulates NO production via the AT2-receptor in the kidney.

Obesity and the kidney connection

The importance of the functional and structural changes that occur in the kidneys in association with obesity was discussed previously in the medical literature.1-4 These studies suggested that the kidneys are involved in the development of obesity-induced hypertension. This involvement is explained by an abnormal tubular sodium handling and by deposition of cells or matrix in the renal medulla that causes an increase in the renal interstitial fluid hydrostatic pressure.3,5

Angiotensin-converting enzyme inhibition potentiates angiotensin II type 1 receptor effects on renal bradykinin and cGMP.

Angiotensin (Ang) receptor blockers (ARBs) increase bradykinin (BK) by antagonizing Ang II at its type 1 (AT(1)) receptors and diverting Ang II to its counterregulatory type 2 (AT(2)) receptors. Because the effect of ARBs on BK is constrained by the short half-life of BK and because ACE inhibitors block the degradation of BK, this study was designed to test the hypothesis that an ACE inhibitor can potentiate ARB-induced increases in renal interstitial fluid (RIF) BK levels. We used a microdialysis technique to recover BK and cGMP in vivo from the RIF of sodium-depleted, conscious Sprague-Dawley rats infused for 60 minutes with the AT(1) receptor blocker valsartan (0.17 mg/kg per minute), with the active metabolite of the ACE inhibitor benazepril (benazeprilate, 0.05 mg/kg per minute), or with the specific AT(2) receptor blocker PD 123,319 (50 microg/kg per minute) alone or combined. Each animal served as its own control. RIF BK and cGMP levels increased significantly over 1 hour in response to valsartan, benazeprilate, or both but not to a vehicle control (P<0.01). The combined benazeprilate-valsartan effect was greater than the sum of their individual effects, suggesting potentiation rather than addition, and was abolished by PD 123,319. We demonstrate for the first time that an ACE inhibitor (benazepril) and an ARB (valsartan) potentiate each other, and we postulate that such combinations may be beneficial in clinical states marked by Ang II elevation, such as chronic heart failure, postinfarction left ventricular dysfunction, and hypertension.

Aldosterone as a determinant of cardiovascular and renal dysfunction.

Aldosterone is a major regulator of extracellular fluid volume and the principal determinant of potassium metabolism 1,2,3,4,5. These effects are mediated by the binding of aldosterone to the mineralocorticoid receptor in target tissues, primarily the kidney. Volume is regulated through a direct effect on the collecting duct, where aldosterone promotes sodium retention and potassium excretion. The reabsorption of sodium ions produces a fall in the transmembrane potential, thus enhancing the flow of positive ions (such as potassium) out of the cell into the lumen. The reabsorbed sodium ions are transported out of the tubular epithellium into the renal interstitial fluid and from there into the renal capillary circulation. Three primary mechanisms control aldosterone release—the renin-angiotensin system, potassium, and adrenocorticotropic hormone. The renin-angiotensin system controls extracellular fluid volume via regulation of aldosterone secretion. In effect, the renin-angiotensin system keeps the circulating blood volume constant by causing aldosterone-induced sodium retention during volume deficiency and by decreasing aldosterone-dependent sodium retention when volume is ample. In recent years there has been a radical shift in our view of aldosterone's effects on the heart, the vasculature and the kidney6,7,8,9. Aldosterone's endocrine properties have taken on a broader perspective, involving non-classic actions in non-epithelial cells found in non-classic target tissues6, 10,11,12,13,14,15. The traditional concept, that aldosterone is synthesized only in the adrenal glomerulosa cell and acts almost exclusively on the kidney to modify sodium and potassium homoeostasis, needs to be expanded. There is increasing evidence that aldosterone can have an effect on vascular remodelling and collagen formation, and a non-genomic action to modify endothelial function. Among the most intriguing effects of aldosterone are its impact on fibrosis and activity associated with a cell surface receptor in certain target tissues, including endothelial cells 6, 7, 16,17,18,19. These actions contribute substantially to the pathophysiology of congestive heart failure (CHF), as well as progressive renal dysfunction. This new information has increased interest in the development of an antagonist to block aldosterone's effect, not just because of its diuretic action but primarily because of its potential cardiovascular and renal protective effects. In this review I consider the broad spectrum of non-genomic effects of aldosterone. It is becoming increasingly evident that these effects, occurring independently of haemodynamic factors, contribute to enhanced cardiovascular risk manifested by congestive heart failure and progressive renal disease. I also discuss the clinical trials with selective aldosterone receptor antagonists that are currently underway in patients with left ventricular hypertrophy, essential hypertension and systolic hypertension, and in those who have experienced myocardial infarction. Such trials will enhance our understanding of the role of aldosterone in the pathophysiology of cardiovascular disease. Also, selective aldosterone receptor antagonism holds promise for a reduction in cardiovascular and renal disease morbidity and mortality, and for enhancement of patient wellbeing.

Selective inhibition of the renal angiotensin type 2 receptor increases blood pressure in conscious rats.

The angiotensin II type 2 (AT(2)) receptor is present in rat kidney; however, its function is not well understood. The purpose of this study was to evaluate the role of the AT(2) receptor in blood pressure (BP) regulation. The effects of selective inhibition of the renal AT(2) receptor with phosphorothioated antisense oligodeoxynucleotide (AS-ODN) were examined in conscious uninephrectomized rats. Oligodeoxynucleotides (AS-ODN or scrambled [S-ODN]) were infused directly into the renal interstitial space by using an osmotic pump at 1 microL/h for 7 days. Texas red-labeled AS-ODN was distributed in renal tubules in the infused but not the contralateral kidney of normal rats. Continuous renal interstitial infusion of the AS-ODN, but not S-ODN, caused a significant (P<0.01) increase in BP 1 to 5 days after the initiation of the infusion. AS-ODN-treated rats experienced an increase in systolic BP from 109+/-4 to 130+/-4 mm Hg (n=8, P<0.01), whereas S-ODN-treated (n=8) and vehicle-treated (n=8) rats did not show any significant change in BP. On day 5 of the oligodeoxynucleotide infusion, AS-ODN-treated rats exhibited a greater pressor response to systemic angiotensin II infusion (30 ng/kg per hour) than did S-ODN-treated rats (P<0.01). Renal interstitial fluid cGMP decreased from 11.9+/-0.8 to 3.6+/-0.5 pmol/mL (P<0.001), and bradykinin decreased from 0.05+/-0.05 to 0.18+/-0.03 ng/mL (P<0.001) in response to AS-ODN, but they were not significantly changed in response to S-ODN. To evaluate the effects of AS-ODN and S-ODN on AT(2) receptor expression, Western Blot analysis was performed on treated kidneys. Kidneys treated with AS-ODN had approximately 40% less expression of AT(2) receptor than did kidneys treated with S-ODN or vehicle (P<0.05). These results suggest that AS-ODN directed selectively against the renal AT(2) receptor decreased receptor expression and caused an increase in BP. We conclude that the renal AT(2) receptor plays an important role in the regulation of BP via a bradykinin/cGMP vasodilator signaling cascade.