AJP - Regu AJP: Cell Physiology
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Am J Physiol Regul Integr Comp Physiol 289: R1573-R1579, 2005. First published August 18, 2005; doi:10.1152/ajpregu.00525.2005
0363-6119/05 $8.00
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
289/6/R1573    most recent
00525.2005v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Web of Science (13)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Taylor, N. E.
Right arrow Articles by Cowley, A. W.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Taylor, N. E.
Right arrow Articles by Cowley, A. W., Jr.

TRANSLATIONAL PHYSIOLOGY

Effect of renal medullary H2O2 on salt-induced hypertension and renal injury

Norman E. Taylor and Allen W. Cowley, Jr.

Department of Physiology, Medical College of Wisconsin, Milwaukee, Wisconsin

Submitted 19 July 2005 ; accepted in final form 12 August 2005

ABSTRACT

Dahl salt-sensitive (SS) and consomic, salt-resistant SS-13BN rats possess substantial differences in blood pressure salt-sensitivity even with highly similar genetic backgrounds. The present study examined whether increased oxidative stress, particularly H2O2, in the renal medulla of SS rats contributes to these differences. Blood pressure was measured using femoral arterial catheters in three groups of rats: 1) 12-wk-old SS and consomic SS-13BN rats fed a 0.4% NaCl diet, 2) SS rats fed a 4% NaCl diet and chronically infused with saline or catalase (6.9 µg·kg–1·min–1) directly into the renal medulla, and 3) SS-13BN fed high salt (4%) and infused with saline or H2O2 (347 nmol·kg–1·min–1) into the renal medullary interstitium. After chronic blood pressure measurements, renal medullary interstitial H2O2 concentration ([H2O2]) was collected by microdialysis and analyzed with Amplex red. Blood pressure and [H2O2] were both significantly higher in SS (126 ± 3 mmHg and 145 ± 17 nM, respectively) vs. SS-13BN rats (116 ± 2 mmHg and 56 ± 14 nM) fed a 0.4% diet. Renal interstitial catalase infusion significantly decreased [H2O2] (96 ± 41 vs. 297 ± 52 nM) and attenuated the hypertension (146 ± 2 mmHg catalase vs. 163 ± 4 mmHg saline) in SS rats after 5 days of high salt (4%). H2O2 infused into the renal medulla of consomic SS-13BN fed high salt (4%) for 7 days accentuated the salt sensitivity (145 ± 2 mmHg H2O2 vs. 134 ± 1 mmHg saline). [H2O2] was also increased in the treated group (83 ± 1 nM H2O2 vs. 44 ± 9 nM saline). These data show that medullary production of H2O2 may contribute to salt-induced hypertension in SS rats and that chromosome 13 of the Brown Norway contains gene(s) that protect against renal medullary oxidant stress.

SS-13BN rats; salt sensitivity

THERE IS GROWING EVIDENCE of increased arterial vascular oxidative stress in both genetic (32, 33) and induced forms of hypertension (2, 8, 37), suggesting that reactive oxygen species (ROS) may contribute to the pathogenesis of this disease. Although most of these studies have focused on the ability of superoxide (O2–) to scavenge and inactivate nitric oxide (NO), recent evidence suggests that ROS other than O2– produce hypertension by mechanisms independent of NO. The present study examines the role of the ROS hydrogen peroxide (H2O2) in the development of salt-sensitive (SS) hypertension.

Evidence that H2O2 could participate in hypertension was obtained in transgenic mice that overexpressed the human catalase gene (38). These mice showed significantly blunted arterial pressure and aortic H2O2 compared with wild-type mice in response to chronic infusions of norepinephrine or angiotensin II. In addition to catalase, glutathione peroxidase plays a key role in scavenging H2O2 in vivo. When the substrate of this enzyme, glutathione, was depleted in Sprague-Dawley (SD) rats, mean arterial pressure (MAP) increased significantly (9). Conversely, n-acetyl-L-cysteine, a well-known antioxidant that can increase glutathione stores, prevented the salt-induced hypertension of Dahl S rats when it was supplemented in the chow (40).

Studies in our laboratory in SD rats have found that local excess production of H2O2 within the medulla of the kidney would produce hypertension. In one of these studies, chronic renal medullary infusion of the O2– dismutase (SOD) inhibitor, diethyldithiocarbamic acid, into a single remaining kidney produced hypertension (21). Co-administration of the SOD mimetic, tempol, failed to prevent this hypertension and only with the addition of catalase was hypertension attenutated (22). Because tempol was shown to inhibit the diethyldithiocarbamic acid-induced elevations of medullary O2–, as determined by microdialysis, the anti-hypertensive actions of catalase indicated a role for H2O2 in maintaining hypertension in this model. Insight into the mechanisms of these H2O2 responses was provided in a second study when acute interstitial infusion of H2O2 reduced medullary blood flow and sodium excretion, and these dose-dependent responses were reversed by catalase treatment (3). A third study then showed that chronic renal medullary interstitial (ri) infusion of H2O2 (1.0–1.4 µmol·kg–1·min–1) produced chronic hypertension in SD rats. Microdialysis studies conducted after day 5 of infusion found that renal interstitial H2O2 was nearly five times higher than saline-infused rats (22).

Although these studies demonstrated that elevations of H2O2 in the renal medulla could induce hypertension, the significance of endogenously produced H2O2 in this region of the kidney that may occur in genetic forms of hypertension remains to be established. The present study examined the role H2O2 produced within the renal medulla of SS rats might play in the development of salt-induced hypertension and related renal end-organ damage. SS rats were compared with an inbred consomic control strain, in which chromosome 13 of the Brown Norway rat was introgressed onto the genetic background of the SS rat (SS-13BN). This strain exhibits only a 1.95% allelic difference from the SS/Medical College of Wisconsin (MCW) rat over the whole genome, compared with 30% for the Dahl salt-resistant rat (R), yet exhibits more than a 50% reduction of salt-induced hypertension, proteinuria, and glomerular disease (4).

The aims of the present study were twofold: 1) to determine whether SS rats exhibit greater levels of oxidative stress, as reflected by H2O2 in the renal medulla, relative to consomic SS-13BN rats; and 2) to determine the role that H2O2 within the renal medulla plays in salt-induced hypertension and renal injury in SS rats. Catalase was therefore infused chronically into the ri of SS rats fed a high (4%) NaCl diet to examine its effects on MAP, medullary H2O2, and kidney damage. Conversely, H2O2 was infused ri into SS-13BN rats fed a 4% NaCl diet to determine whether the trait of SS hypertension could be exacerbated in this strain. Microdialysis was used to collect samples of renal medullary interstitial H2O2 in each of these studies.

METHODS

Animal preparation. Experiments were performed on inbred lines of male Dahl SS/JrHsdMcwi (SS) and consomic SS-13BN rats maintained as inbred colonies at the MCW (4). The MCW Institutional Animal Care and Use Committee approved all experimental protocols. Rats were maintained ad libitum on tap water, and a purified AIN-76A rodent diet containing 0.4 or 4% NaCl (Dyets, Bethlehem, PA).

Male SS and SS-13BN rats fed a 0.4% NaCl diet were uninephrectomized at 9 wk of age. After a 7- to 10-day recovery period, medullary interstitial and femoral arterial catheters were implanted as previously described (24). Five to six days later, daily 3-h measurements of MAP were begun using an online data collection and analysis system (5).

After the last day of blood pressure recording, acute surgical preparation for in vivo microdialysis of the left kidney was performed as described previously (21). After a 2-h equilibration period, dialysate effluent was collected over two 30-min intervals and renal medullary interstitial H2O2 concentrations ([H2O2])were determined by fluorescence spectrometry using the Amplex Red H2O2 Assay Kit (Molecular Probes, Eugene, OR).

Group 1. Eleven-week-old SS and SS-13BN rats were prepared only with femoral arterial catheters, recovered 5–6 days, and blood pressure measured for 3 consecutive days before determination of medullary concentrations of H2O2 by microdialysis.

Group 2. SS rats were prepared with a renal interstitial catheter as well as a femoral arterial catheter and allowed to recover. After 3 days of stable baseline MAP measurements, ri catalase (Sigma) infusion was begun at 6.9 µg·kg–1·min–1 (13.3 mU·kg–1·min–1), a dose that was shown to effectively decrease elevated renal medullary H2O2 levels previously (22). Blood pressure was then recorded for 3 days with the rats maintained on a 0.4% salt diet. Control SS rats were continuously infused with saline (8 µl/min). The control and catalase-treated rats were then switched to a 4.0% NaCl diet, and infusions continued for 5 more days. On the last day of chronic infusion, the rats were prepared for in vivo microdialysis measurement of renal interstitial [H2O2] as described above.

Group 3. SS-13BN rats were prepared as in group 2, except that H2O2 (347 nmol·kg–1·min–1) was infused into the interstitium instead of catalase, and the experiment was extended to 7 days of 4.0% NaCl. A previous study demonstrated that ri infusions of three to four times this dose of H2O2 produced hypertension in SD rats (22). As we were trying to determine the role of H2O2 in salt-induced hypertension, a dose was selected that elevated blood pressure only after the change to a 4% NaCl diet.

Histology preparation and measurements. Rats were euthanized, and the left kidney was removed, immersion-fixed in 10% neutral buffered formalin, and paraffin embedded. Sections were prepared and stained with Gomori trichrome stain to highlight the fibrotic tissue and hematoxylin and eosin to quantify cast formation as an index of tubular dysfunction. Interstitial fibrosis was determined by immunostaining with antibodies for {alpha}-smooth muscle actin (SMA) (Dako Cytomation), and detected using an Envision/horseradish peroxidase detection kit (DAKO Cytomation). Cast and SMA staining were quantified in 20 random frames using the Metamorph analysis software, as previously described (29), and reported as the mean % positive area per frame for cast or SMA staining.

Statistical methods. Data are presented as means ± SE. In cases where each animal served as its own control in the pre- and posttreatment periods, the data were analyzed using a one-way ANOVA for repeated measures followed by a Tukey's multiple range test. A P value of <0.05 was considered significant. Between-group comparisons were performed using a two-way ANOVA followed by a Tukey's multiple range test to compare individual points. A Pearson correlation was performed using the data from individual rats in all three experimental groups to assess whether MAP was correlated to medullary H2O2 levels and tubular cast formation.

RESULTS

Group 1: renal medullary interstitial H2O2 in rats fed a 0.4% NaCl diet. SS rats exhibited higher MAP (126 ± 3 mmHg, n = 7) than SS-13BN rats (116 ± 2 mmHg, n = 7), even when these rats were fed a 0.4% NaCl diet from weaning, as shown in Fig. 1. Renal medullary interstitial H2O2 was also found to be significantly higher in SS compared with SS-13BN rats (145 ± 17 nM, n = 7 vs. 56 ± 14 nM, n = 7).



View larger version (10K):
[in this window]
[in a new window]
  		Fig. 1. Mean arterial pressure (MAP; top) and renal medullary H2O2 concentrations (bottom) in 12-wk-old salt-sensitive (SS) and SS-13BN rats maintained on a 0.4% NaCl diet. Values are means ± SE. *P < 0.05 vs. control.

 
Group 2: effect of renal medullary interstitial infusion of catalase on salt-induced hypertension and medullary H2O2 concentration in SS rats. Catalase was infused chronically into the renal medulla of SS rats to determine the contribution of medullary H2O2 production to salt-induced hypertension. As summarized in Fig. 2, MAP of the saline-infused control group increased from 129 ± 2 to 163 ± 4 mmHg after 5 days of high salt (4%), whereas MAP in SS rats infused with catalase increased from 122 ± 1 to only 146 ± 2 mmHg after 5 days of high salt. At the end of 5 days, renal medullary [H2O2] in saline-infused rats averaged 297 ± 52 nM (n = 7) and was significantly reduced (96 ± 41 nM, n = 7) in catalase-infused rats. These data show that medullary production of H2O2 contributes importantly to the salt-induced hypertension in the SS rat.



View larger version (16K):
[in this window]
[in a new window]
  		Fig. 2. Effect of chronic renal interstitial (ri) catalase (6.9 µg·kg–1·day–1) infusion on salt-induced hypertension (top) and medullary interstitial H2O2 (bottom) in SS rats ({circ}) compared with saline-infused controls ({bullet}). Values are means ± SE. * and {dagger}P < 0.05 vs. control.

 
Group 3: effect of renal medullary interstitial infusion of H2O2 on salt-induced hypertension and medullary [H2O2] in SS-13BN rats. To further evaluate the importance of H2O2 in salt-induced hypertension, renal medullary [H2O2] was increased in relatively salt-insensitive SS-13BN rats to determine whether the SS trait of salt sensitivity could be restored in this consomic strain (Fig. 3). H2O2 was infused at a dose that did not change MAP over 3 days on a 0.4% salt diet (347 nmol·kg–1·min–1). When the salt in the diet was increased from 0.4 to 4%, MAP increased 25 mmHg (from 120 ± 2 to 145 ± 2 mmHg). In saline vehicle-infused SS-13BN rats, MAP increased only 16 mmHg (118 ± 2 to 134 ± 1 mmHg) when fed the high-salt diet. As determined by microdialysis at the end of the experiment, saline-infused SS-13BN rats exhibited H2O2 concentrations of 44 ± 9 nM (n = 6) compared with 83 ± 11 nM for H2O2-infused rats (n = 6). Renal medullary [H2O2] levels were highly correlated with MAP in the three experimental groups (r = 0.602, P < 0.001). Of particular note, SS rats infused with catalase exhibited the same level of hypertension as the H2O2-infused SS-13BN rats (146 ± 2 vs. 145 ± 2 mmHg) and also had similar levels of renal medullary H2O2 (96 ± 41 vs. 83 ± 11 nM). Blood pressure salt sensitivity was exacerbated in the consomic SS-13BN simply by elevating medullary H2O2, indicating that excess H2O2 production in this region of the kidney contributes to salt-induced hypertension in SS rats.



View larger version (17K):
[in this window]
[in a new window]
  		Fig. 3. Effect of chronic renal interstitial H2O2 (347 nmol·kg–1·min–1) infusion on salt-induced hypertension (top) and medullary interstitial H2O2 (bottom) in SS-13BN rats ({circ}) compared with saline-infused controls ({bullet}). Values are means ± SE. * and {dagger}P < 0.05 vs. control.

 
Histological quantification of tubular necrosis and interstitial fibrosis. The percentage of the outer medulla exhibiting protein casts was used as an index of tubular necrosis. As shown in Fig. 4, cast formation in SS rats receiving catalase infusions (7.5 ± 1.0%, n = 6) was significantly less than saline-infused controls (16.0 ± 2.0%, n = 6). Despite considerable reduction of tubular damage, catalase-infused SS rats continued to exhibit significantly more tubular casts than the saline-infused SS-13BN rats. There was no significant difference in cast staining between saline and H2O2-infused SS-13BN rats (4.7 ± 0.6%, n = 6 vs. 6.0 ± 1.3%, n = 6). Tubular necrosis was significantly correlated to MAP in these studies (r = 0.83, P < 0.05).



View larger version (107K):
[in this window]
[in a new window]
  		Fig. 4. Top: Gomori trichrome staining shows that tubular necrosis with protein casts and interstitial fibrosis were more prominent in the outer medulla of SS rats than SS-13BN rats. The % cast-positive region was quantified in 20 randomly chosen frames per outer medulla using morphology software. Cast formation in SS rats receiving catalase infusions (B; n = 6) was significantly (*) lower than saline-infused controls (A; n = 6) but was significantly ({dagger}) elevated compared with saline-infused SS-13BN rats. There was no significant difference in cast staining between saline- (C; n = 6) and H2O2 (D; n = 6)-infused SS-13BN rats. Tubular necrosis was significantly correlated to MAP (r = 0.83). Values are means ± SE.

 
The percentage of tissue that stained positive for {alpha}-SMA in the outer medulla was used as an early index of interstitial fibrosis. SS rats receiving the medullary infusion of catalase exhibited significantly reduced SMA staining (0.6 ± 0.2%, n = 6) compared with saline-infused SS rats (2.3 ± 0.6%, n = 6), as shown in Fig. 5. Conversely, SS-13BN rats that were infused with H2O2 and fed a 4% NaCl diet exhibited an elevated {alpha}-SMA staining compared with saline-infused controls (2.8 ± 0.7%, n = 6 vs. 0.7 ± 0.2%, n = 6). Taken together, these observations indicate that elevations of H2O2 contribute to the development of medullary interstitial fibrosis.



View larger version (98K):
[in this window]
[in a new window]
  		Fig. 5. Tubular interstitial fibrosis, determined by {alpha}-SMA-positive regions immunostained with monoclonal antibody (top), was significantly (*) reduced in SS rats receiving medullary interstitial infusions of catalase (B; n = 6) compared with saline-infused rats (A; n = 6). Conversely, SS-13BN rats that were infused with H2O2 and fed a 4% NaCl diet (D) exhibited elevated ({dagger}) {alpha}-SMA staining than saline-infused rats (C; n = 6). The %SMA-positive region was quantified in 20 randomly chosen frames per outer medulla using morphology software. Values are means ± SE.

 
DISCUSSION

The present study examined whether H2O2 levels are elevated in SS rats and tested the hypothesis that increased renal medullary H2O2 in this strain contributes to salt-induced hypertension and renal injury. The results indicate that H2O2 is elevated in the renal interstitium of SS rats compared with the consomic SS-13BN control strain and that arterial pressures were significantly elevated in the SS, even before the introduction of a high-salt diet. Renal interstitial infusion of catalase in the SS significantly reduced H2O2 levels and greatly attenuated the salt-induced increase in arterial pressure. Conversely, increasing renal medullary H2O2 in SS-13BN rats increased the salt sensitivity. This suggests that the gene(s) protecting the SS-13BN from increased oxidative stress may also be responsible for the protection from salt-induced hypertension.

Reduction of medullary H2O2 in SS rats also significantly reduced renal injury as measured by a reduction in protein casts and {alpha}-SMA staining in the outer medulla. Tubulointerstitial fibrosis and capillary injury occur first in the outer medulla and juxtamedullary glomeruli in many forms of experimental hypertension, including Dahl S rats (16), and is considered a final common pathway of progressive kidney disease. Myofibroblasts, which contain {alpha}-SMA (20), are rarely found in the normal kidney, and when they become detectable in the interstitium, collagen deposition accumulates in the areas in which they reside (1), making {alpha}-SMA staining an important early index of interstitial fibrosis. {alpha}-SMA staining was significantly increased in SS-13BN rats infused with H2O2, suggesting that this ROS could play a significant role in initiating interstitial fibrosis in the kidney. These data support a role for H2O2 in the maintenance of salt-induced arterial pressure and renal injury in the SS rat.

Consomic SS-13BN rats as a control for SS rats. The selection of appropriate controls for the Dahl S and other hypertensive strains has been constrained by the absence of available methods to carry out extensive whole genome scans. Control strains for hypertension have been generally selected on the basis of the ancestral origins of the strains, even though all of the traditional "control" strains (Wistar-Kyoto, Dahl R, etc.) are now known to exhibit considerable genetic divergence from the hypertensive strains. This results in hundreds of phenotypic differences between the control and hypertensive strains, most of which are unrelated to the pathogenesis of hypertension. In support of this argument, there is a 77% allelic difference between Brown-Norway/MCW and SS/MCW rats, a 48% difference between spontaneously hypertensive and Wistar-Kyoto rats, a 52% difference between SD and SS/MCW rats, and a 30% difference between Dahl R and SS/MCW rats as assessed by whole genome scans using 1,541 microsatellite markers (13). In contrast, the consomic SS-13BN rat exhibits only a 1.95% allelic difference from SS/MCW rats over the entire genome, and yet salt-induced hypertension, proteinuria, and glomerular disease is greatly reduced (4). Based on our genetic analysis, we believe that the SS-13BN rat is currently the best available control strain for phenotypic comparison with the SS/MCW rats because it possesses less genetic difference than any other available strain.

Salt sensitivity and renal medullary oxidative stress. SS rats have been shown to be deficient in NO (34) and 20-HETE (15), while also exhibiting increased sympathetic tone (14) and oxidative stress (25, 26, 33), all of which contribute to the salt sensitivity of this strain. This study provides direct evidence of increased oxidative stress within the renal medulla of the SS rat and supports indirect measurements made by others. Studies where tempol attenuated salt-induced hypertension in SS rats (12, 25) support a role for O2– in maintaining elevated pressure in this strain. However, just as in our diethyldithiocarbamic acid model of hypertension, administration of tempol was not fully protective. Indeed, Williams et al. (36) observed that tempol administered in the drinking water of SD rats was unable to sustain the inhibition of hypertension produced by ETB receptor blockade plus a high-salt diet and that this was accompanied by increased urinary H2O2 excretion. The purpose of this study was to determine whether H2O2 within the renal medulla of SS rats plays a role in salt-induced hypertension and renal injury separate from the direct effects of other ROS.

Increased H2O2 concentrations in the renal medulla of SS rats may be due to increased production or decreased scavenging of this ROS. H2O2 is a relatively stable species that diffuses freely into cells and is thought to be generated predominately from the SOD dismutation of O2–, although it can also be produced directly by certain oxidases, such as glucose oxidase, through a two-electron reduction of O2 (31). Evidence suggests that NADPH oxidase is an important source of ROS in hypertension (17), playing an especially important role in tubular O2– production in the renal medulla (41). Renal cortical protein and mRNA levels of the NADPH oxidase subunit p47phox were shown to be higher in Dahl S than Dahl R rats when fed an 8% NaCl diet, and this increase could be attenuated by either administration of an angiotensin-converting enzyme inhibitor (35), L-arginine (7), or n-acetyl-L-cysteine (40). Studies have begun to assess the role that ROS scavengers play in the kidneys of SS rats. A study by Meng et. al (26) showed that the Dahl S rat has lower renal SOD levels compared with Dahl R rats. This question was also recently addressed in DNA microarray studies to determine differential gene expression in renal outer medullary tissue of SS and SS-13BN rats fed a 0.4 or 4% NaCl diet. Although these data did not resolve whether NADPH oxidase enzyme subunits were differentially expressed, no difference was found in the expression of SOD, catalase, glutathione peroxidase, and synthetase along with several genes related to the biopterin pathway. Glutathione-S-transferase and peroxiredoxin-6 (1-Cys peroxiredoxin) were differentially expressed between SS and SS-13BN rats, although they were expressed lower in the SS-13BN rats. It is now important to follow up these studies by measuring the activities of the enzymes contributing to H2O2 synthesis/degradation given the possibility of posttranscriptional influences on these events.

It is presently unknown what gene or set of genes on chromosome 13 protect the consomic SS-13BN rat from oxidative stress. Cyclooxygenase-2 is located on chromosome 13 and is a potential source of O2–. Administration of the selective cyclooxygenase-2 inhibitor, celecoxib, was found to modestly decrease salt-induced hypertension in the Dahl S rat and significantly reduce plasma 8-isoprostane levels (11), suggesting a possible role for the gene in this model of hypertension. Currently, the genetic basis responsible for the protection afforded by BN chromosome 13 remains to be established.

Role of H2O2 in hypertension. The role that H2O2 plays in vascular function and hypertension is just beginning to be studied. H2O2 has been shown to be both a vasodilator and a vasoconstrictor. It has been implicated as an endothelial-derived dilating factor in both animal and human studies based on observations that catalase inhibited dilation of mesenteric arteries from endothelial NO synthase knockout mice (23) and in human coronary arterial microvessels (27). This response has been attributed to membrane hyperpolarization through the activation of calcium-dependent K+ channels (27). In contrast, studies in several different vascular beds have reported that H2O2 acts as a vasoconstrictor (28, 30, 39). Aortic contraction was found to be Ca2+ dependent with evidence that hydroxyl radicals, cyclooxygenase products, protein kinase C, and products of protein tyrosine phosphorylation may play some role in these responses (39).

Recent studies in mesenteric arteries (10) and rat gracilus arterioles (6) provide some insight into these discrepancies by showing that responses to H2O2 are dose dependent in these vessels. Vasoconstriction occurred at lower doses (10–100 µM in mesenteric arteries) via a PGH2/TxA2 receptor-dependent mechanism, whereas vasodilation predominated at concentrations above 0.3 mM via a K+ channel-mediated hyperpolarization (10). The present study shows that interstitial [H2O2] ranges between 50 and 300 nM. Even after accounting for the 20–30% probe efficiency, levels are still well within the dose range that purportedly constricts vessels in other tissues. Indeed, acute studies in our laboratory have shown that ri H2O2 infusion in SD rats at a dose that increased H2O2 in medullarydialysates from 116 to 211 nM decreased medullary blood flow, urinary sodium excretion, and urine volume, suggesting that a physiological increase of H2O2 produces vasoconstriction in this region of the kidney (3).

We conclude that elevation of H2O2 in the renal medulla of SS rats plays an important role in salt-induced hypertension and renal injury, and therapies that reduce both O2– and H2O2 should prove to be more effective in the long-term treatment of hypertension.

GRANTS

This work was supported by National Heart, Lung, and Blood Institute Grants HL-29587 and HL-54998, and predoctoral fellowship AHA-04100437.

ACKNOWLEDGMENTS

The authors thank Meredith M. Skelton for careful review of the manuscript, Glenn Slocum for microscopy assistance, and Carol Bobrowitz for technical assistance with the immunohistochemistry.

FOOTNOTES


Address for reprint requests and other correspondence: N. E. Taylor, Dept. of Physiology, Medical College of Wisconsin, 8701 Watertown Plank Rd., Milwaukee, WI 53226 (e-mail: ntaylor{at}mcw.edu)

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

REFERENCES

  1. Badid C, Vincent M, McGregor B, Melin M, Hadj-Aissa A, Veysseyre C, Hartmann DJ, Desmouliere A, and Laville M. Mycophenolate mofetil reduces myofibroblast infiltration and collagen III deposition in rat remnant kidney. Kidney Int 58: 51–61, 2000.[CrossRef][Web of Science][Medline]
  2. Bouloumie A, Bauersachs J, Linz W, Scholkens BA, Wiemer G, Fleming I, and Busse R. Endothelial dysfunction coincides with an enhanced nitric oxide synthase expression and superoxide anion production. Hypertension 30: 934–941, 1997.[Abstract/Free Full Text]
  3. Chen YF, Cowley AW Jr, and Zou AP. Increased H2O2 counteracts the vasodilator and natriuretic effects of superoxide dismutation by tempol in renal medulla. Am J Physiol Regul Integr Comp Physiol 285: R827–R833, 2003.[Abstract/Free Full Text]
  4. Cowley AW Jr, Roman RJ, Kaldunski ML, Dumas P, Dickhout JG, Greene AS, and Jacob HJ. Brown Norway chromosome 13 confers protection from high salt to consomic Dahl S rat. Hypertension 37: 456–461, 2001.[Abstract/Free Full Text]
  5. Cowley AW Jr, Stoll M, Greene AS, Kaldunski ML, Roman RJ, Tonellato PJ, Schork NJ, Dumas P, and Jacob HJ. Genetically defined risk of salt sensitivity in an intercross of Brown Norway and Dahl S rats. Physiol Genomics 2: 107–115, 2000.[Abstract/Free Full Text]
  6. Cseko C, Gai Z, and Koller A. Biphasic effect of hydrogen peroxide on skeletal muscle arteriolar tone via activation of endothelial and smooth muscle signaling pathways. J Appl Physiol 97: 1130–1137, 2004.[Abstract/Free Full Text]
  7. Fujii S, Zhang L, Igarashi J, and Kosaka H. L-Arginine reverses p47phox and gp91phox expression induced by high salt in Dahl rats. Hypertension 42: 1014–1020, 2003.[Abstract/Free Full Text]
  8. Fukai T, Siegfried MR, Ushio-Fukai M, Cheng Y, Kojda G, and Harrison DG. Regulation of the vascular extracellular superoxide dismutase by nitric oxide and exercise training. J Clin Invest 105: 1631–1639, 2000.[Web of Science][Medline]
  9. Ganafa AA, Socci RR, Eatman D, Silvestrova N, Abukhalaf IK, and Bayorh MA. Acute inhibition of glutathione biosynthesis alters endothelial function and blood pressure in rats. Eur J Pharmacol 454: 217–223, 2002.[CrossRef][Web of Science][Medline]
  10. Gao YJ, Hirota S, Zhang DW, Janssen LJ, and Lee RM. Mechanisms of hydrogen-peroxide-induced biphasic response in rat mesenteric artery. Br J Pharmacol 138: 1085–1092, 2003.[CrossRef][Web of Science][Medline]
  11. Hermann M, Camici G, Fratton A, Hurlimann D, Tanner FC, Hellermann JP, Fidler M, Thiery J, Neidhar M, Gay RE, Gay S, Luscher TF, and Fuschitzka F. Differential effects of selective cyclooxygenase-2 inhibitors on endothelial function in salt-induced hypertension. Circulation 108: 2308–2311, 2003.[Abstract/Free Full Text]
  12. Hoagland KM, Maier KG, and Roman RJ. Contributions of 20-HETE to the antihypertensive effects of Tempol in Dahl salt-sensitive rats. Hypertension 41: 697–702, 2003.[Abstract/Free Full Text]
  13. Huang BS, Wang H, and Leenen FH. Enhanced sympathoexcitatory and pressor responses to central Na+ in Dahl salt-sensitive vs. -resistant rats. Am J Physiol Heart Circ Physiol 281: H1881–H1889, 2001.[Abstract/Free Full Text]
  14. Ito O and Roman RJ. Role of 20-HETE in elevating chloride transport in the thick ascending limb of Dahl SS/Jr rats. Hypertension 33: 419–423, 1999.[Abstract/Free Full Text]
  15. Johnson RJ, Gordon KL, Giachelli C, Kurth T, Skelton MM, and Cowley AW Jr. Tubulointerstitial injury and loss of nitric oxide synthases parallel the development of hypertension in the Dahl-SS rat. J Hypertens 18: 1497–1505, 2000.[CrossRef][Web of Science][Medline]
  16. Lassegue B and Clempus RE. Vascular NAD(P)H oxidases: specific features, expression, and regulation. Am J Physiol Regul Integr Comp Physiol 285: R277–R297, 2003.[Abstract/Free Full Text]
  17. Liang M, Yuan B, Rute E, Greene AS, Zou AP, Soares P, MCQuestion GD, Slocum GR, Jacob HJ, and Cowley AW Jr. Renal medullary genes in salt-sensitive hypertension: a chromosomal substitution and cDNA microarray study. Physiol Genomics 8:139–149, 2002.[Abstract/Free Full Text]
  18. Liang M, Yuan B, Rute E, Greene AS, Olivier M, and Cowley AW Jr. Insights into Dahl salt-sensitive hypertension revealed by temporal patterns of renal medullary gene expression. Physiol Genomics 12: 229–237, 2003.[Abstract/Free Full Text]
  19. MacPherson BR, Leslie KO, Lizaso KV, and Schwarz JE. Contractile cells of the kidney in primary glomerular disorders: an immunohistochemical study using an anti-alpha-smooth muscle actin monoclonal antibody. Hum Pathol 24: 710–716, 1993.[CrossRef][Web of Science][Medline]
  20. Makino A, Skelton MM, Zou AP, Roman RJ, and Cowley AW Jr. Increased renal medullary oxidative stress produces hypertension. Hypertension 39: 667–672, 2002.[Abstract/Free Full Text]
  21. Makino A, Skelton MM, Zou AP, and Cowley AW Jr. Increased renal medullary H2O2 leads to hypertension. Hypertension 42: 25–30, 2003.[Abstract/Free Full Text]
  22. Matoba T, Shimokawa H, Kubota H, Morikawa K, Fujiki T, Kunihiro I, Mukai Y, Hiakawa Y, and Takeshita A. Hydrogen peroxide is an endothelium-derived hyperpolarizing factor in mice. J Clin Invest 106: 1521–1530, 2000.[Web of Science][Medline]
  23. Mattson DL, Lu S, and Cowley AW Jr. Role of nitric oxide in the control of the renal medullary circulation. Clin Exp Pharmacol Physiol 24: 587–590, 1997.[Web of Science][Medline]
  24. Meng S, Cason GW, Gannon AW, Racusen LC, and Manning RD Jr. Oxidative stress in Dahl salt-sensitive hypertension. Hypertension 41: 1346–1352, 2003.[Abstract/Free Full Text]
  25. Meng S, Roberts 2nd LJ, Cason GW, Curry TS, and Manning RD Jr. Superoxide dismutase and oxidative stress in Dahl salt-sensitive and -resistant rats. Am J Physiol Regul Integr Comp Physiol 283: R732–R738, 2002.[Abstract/Free Full Text]
  26. Miura H, Bosnjak JJ, Nong G, Saito T, Miura M, and Guttermann DD. Role for hydrogen peroxide in flow-induced dilation of human coronary arterioles. Circ Res 92: 31–40, 2003.
  27. Mori T and Cowley AW Jr. Role of pressure in angiotensin II-induced renal injury—chronic servo-control of renal perfusion pressure in rats. Hypertension 43: 752–759, 2004.[Abstract/Free Full Text]
  28. Prabhakar R, Siegbahn PE, and Minaev BF. A theoretical study of the dioxygen activation by glucose oxidase and copper amine oxidase. Biochim Biophys Acta 1647: 173–178, 2003.[Medline]
  29. Rhoades RA, Packer CS, Poepke DA, Jin N, and Meiss RA. Reactive oxygen species alter contractile properties of pulmonary arterial smooth muscle. Can J Physiol Pharmacol 68: 1581–1589, 1990.[Web of Science][Medline]
  30. Rubanyi GM and Vanhoutte PM. Oxygen derived free radicals, endothelium and responsiveness of vascular smooth muscle. Am J Physiol Heart Circ Physiol 250: H815–H821, 1986.[Abstract/Free Full Text]
  31. Suzuki H, Swei A, Zweifach BW, and Schmid-Schonbein GW. In vivo evidence for microvascular oxidative stress in spontaneously hypertensive rats. Hydroethidine microfluorography. Hypertension 25: 1083–1089, 1995.[Abstract/Free Full Text]
  32. Swei A, Lacy F, DeLano FA, and Schmid-Schonbein GW. Oxidative stress in the Dahl hypertensive rat. Hypertension 30: 1628–1633, 1997.[Abstract/Free Full Text]
  33. Szentivanyi M Jr, Zou AP, Mattson DL, Soares P, Moreno C, Roman RJ, and Cowley AW Jr. Renal medullary nitric oxide deficit of Dahl S rats enhances hypertensive actions of angiotensin II. Am J Physiol Regul Integr Comp Physiol 283: R266–R272, 2002.[Abstract/Free Full Text]
  34. Tojo A, Onozato L, Kobayashi N, Goto A, Matsuoka H, and Fujita T. Angiotensin II and oxidative stress in Dahl salt-sensitive rat with heart failure. Hypertension 40: 834–839, 2002.[Abstract/Free Full Text]
  35. Williams JM, Pollock JS, and Pollock DM. Arterial pressure response to the antioxidant tempol and ETB receptor blockade in rats on a high-salt diet. Hypertension 44: 770–775, 2004.[Abstract/Free Full Text]
  36. Wu R, Millette E, Wu L, and de Champlain J. Enhanced superoxide anion formation in vascular tissues from spontaneously hypertensive and deoxycorticosterone acetate-salt hypertensive rats. J Hypertens 19: 741–748, 2001.[CrossRef][Web of Science][Medline]
  37. Yang H, Shi M, VanRemmen H, Chen X, Vijg J, Richardson A, and Guo Z. Reduction of pressor response to vasoconstrictor agents by overexpression of catalase in mice. Am J Hypertens 16: 1–5, 2003.[Web of Science][Medline]
  38. Yang ZW, Zheng T, Zhang A, Altura BT, and Altura BM. Mechanisms of hydrogen peroxide-induced contraction of rat aorta. Eur J Pharmacol 344: 169–181, 1998.[CrossRef][Web of Science][Medline]
  39. Zhang L, Fujii S, Igarashi J, and Kosaka H. Effects of thiol antioxidant on reduced nicotinamide adenine dinucleotide phosphate oxidase in hypertensive Dahl salt-sensitive rats. Free Radic Biol Med 37: 1813–1820, 2004.[CrossRef][Web of Science][Medline]
  40. Zou AP, Li N, and Cowley AW Jr. Production and actions of superoxide in the renal medulla. Hypertension 37: 547–553, 2001.[Abstract/Free Full Text]



This article has been cited by other articles:


Home page
 Am. J. Physiol. Regul. Integr. Comp. Physiol.Home page
C. S. Escano, I. Armando, X. Wang, L. D. Asico, A. Pascua, Y. Yang, Z. Wang, Y.-S. Lau, and P. A. Jose
Renal dopaminergic defect in C57Bl/6J mice
Am J Physiol Regulatory Integrative Comp Physiol, December 1, 2009; 297(6): R1660 - R1669.
[Abstract] [Full Text] [PDF]

Home page
 HypertensionHome page
Z. Tian, Y. Liu, K. Usa, D. Mladinov, Y. Fang, X. Ding, A. S. Greene, A. W. Cowley Jr, and M. Liang
Novel Role of Fumarate Metabolism in Dahl-Salt Sensitive Hypertension
Hypertension, August 1, 2009; 54(2): 255 - 260.
[Abstract] [Full Text] [PDF]

Home page
 HypertensionHome page
R. Pedrosa, N. Goncalves, U. Hopfer, P. A. Jose, and P. Soares-da-Silva
Activity and Regulation of Na+-HCO3- Cotransporter in Immortalized Spontaneously Hypertensive Rat and Wistar-Kyoto Rat Proximal Tubular Epithelial Cells
Hypertension, May 1, 2007; 49(5): 1186 - 1193.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Regul. Integr. Comp. Physiol.Home page
J. C. Sullivan, J. M. Sasser, and J. S. Pollock
Sexual dimorphism in oxidant status in spontaneously hypertensive rats
Am J Physiol Regulatory Integrative Comp Physiol, February 1, 2007; 292(2): R764 - R768.
[Abstract] [Full Text] [PDF]

Home page
 HypertensionHome page
N. E. Taylor, K. G. Maier, R. J. Roman, and A. W. Cowley Jr
NO Synthase Uncoupling in the Kidney of Dahl S Rats: Role of Dihydrobiopterin
Hypertension, December 1, 2006; 48(6): 1066 - 1071.
[Abstract] [Full Text] [PDF]

Home page
 HypertensionHome page
N. E. Taylor, P. Glocka, M. Liang, and A. W. Cowley Jr
NADPH Oxidase in the Renal Medulla Causes Oxidative Stress and Contributes to Salt-Sensitive Hypertension in Dahl S Rats
Hypertension, April 1, 2006; 47(4): 692 - 698.
[Abstract] [Full Text] [PDF]

This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow All Versions of this Article:
289/6/R1573    most recent
00525.2005v1
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via Web of Science (13)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Taylor, N. E.
Right arrow Articles by Cowley, A. W.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Taylor, N. E.
Right arrow Articles by Cowley, A. W., Jr.

HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
Visit Other APS Journals Online
Copyright © 2005 by the American Physiological Society.