(Hypertension. 1997;30:409.)
© 1997 American Heart Association, Inc.
Articles |
Nitric Oxide Synthase and Renin-Angiotensin System Gene Expression in Salt-Sensitive and Salt-Resistant Sabra Rats
From the Franz Volhard Clinic and the Max Delbrück Center for Molecular Medicine (A.L., V.G., K.S., W.S., M.B., F.C.L.), Virchow Klinikum, Humboldt University, Berlin, Germany; Department of Neuroscience, Division of Cellular and Molecular Neurochemistry, Karolinska Institute (A.H.), Stockholm, Sweden; INSERM U358 (S.N.), Paris, France; and Barzilai Medical Center (C.Y., Y.Y.), Ashkelon, Ben Gurion University, Israel.
Correspondence to Friedrich C. Luft, Franz Volhard Clinic, Wiltbergstr 50, 13122 Berlin, Germany. E-mail fcluft{at}mdc-berlin.de
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Abstract The molecular mechanisms of salt sensitivity and the contribution of the kidney to salt-induced hypertension in Sabra rats are imperfectly defined. We investigated the expression of the nitric oxide (NO) system (endothelial, inducible, and neural NO synthases) and renin-angiotensin system (renin, angiotensinogen, and angiotensin II type 1A receptor) gene components in the kidneys of SBN/y (salt-resistant) and SBH/y (salt-sensitive) Sabra rat substrains, with and without deoxycorticosterone acetate (DOCA)–salt treatment. We also looked for immunocytochemical evidence of angiotensin II, the effector peptide of the renin-angiotensin system. Inducible and neural NO synthase gene expression values were lower in SBH/y than in SBN/y before and after DOCA-salt treatment. The gene expression level of endothelial NO synthase was not different in SBH/y and SBN/y, either with or without DOCA salt. Renin gene expression was significantly higher in kidneys of SBN/y than in kidneys of SBH/y rats, whereas angiotensinogen gene expression was significantly lower in SBN/y. After DOCA-salt treatment, renin gene expression was strongly suppressed in both strains but more so in SBH/y. Angiotensinogen gene expression, on the other hand, was increased by DOCA salt in SBN/y rats so that the two strains were no longer different. Angiotensin II immunoreactivity was significantly higher in SBN/y than in SBH/y; however, after DOCA salt, immunoreactivity in both strains was no longer detectable. Angiotensin II type 1A receptor gene expression was not different between the two strains, either before or after DOCA-salt administration. We conclude that DOCA salt induced a decrease in the activity of the renin-angiotensin system but did not change NO synthase gene expression in SBH/y and SBN/y. Inducible and neural NO synthase gene expression values were less in SBH/y than in SBN/y, independent of DOCA-salt administration. Thus, the NO system could explain, at least in part, the salt resistance of SBN/y.
Key Words: hypertension, renal • salt • gene expression • renin-angiotensin system • nitric oxide
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Introduction |
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The recently rebred homogeneous SBH/y is especially prone to the development of DOCA-salt hypertension, whereas the SBN/y is not.1 In a recent study,2 we showed that prehypertensive SBH/y were not different from SBN/y with regard to sodium and water excretion, which is at variance with earlier findings in the original Sabra rat strains.3 4 5 However, DOCA-salt treatment decreased the slope and/or shifted the pressure-diuresis and pressure-natriuresis curves rightward in SBH/y.2 In vivo experiments have demonstrated that nonpressor doses of the NOS inhibitor nitro-L-arginine depress the pressure-natriuretic relationship.6 eNOS, iNOS, and nNOS are present in the kidney and may play an important role in control of renal function and the long-term regulation of blood pressure. For instance, Ikeda et al7 found that nNOS activity was decreased in Dahl salt-sensitive rats given a high-salt diet, but eNOS and iNOS remained unchanged. Continuous infusion of the NOS substrate L-arginine in Dahl salt-sensitive rats prevented changes in the pressure-natriuresis relationship as well as the development of salt-induced hypertension.8 In SBH/y, NO production may be decreased compared with SBN/y.9 Furthermore, salt loading regulates the gene expression of NO synthases in Sprague-Dawley rats.10 We examined gene expression of eNOS, iNOS, and nNOS in SBH/y and SBN/y, with or without DOCA-salt treatment. Because the renin-angiotensin system is influenced by salt intake and interacts with the NO system,11 12 13 we examined genes relevant to these systems.
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Methods |
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Animals
Experiments were performed in 16 male SBN/y weighing 292±11 g each and 16 male SBH/y weighing 309±8 g each. The animals were bred in the animal facility of the Barzilai Medical Center, Ashkelon, Israel. The animals were allowed free access to standard rat chow (0.3% sodium chloride, SSNIFF Specialitäten GmbH) and drinking water ad libitum. Eight SBN/y and 8 SBH/y were implanted with a 25-mg DOCA tablet (Innovative Research of America). The pellets were placed below the skin at the nape of the neck. The animals were provided with 1% NaCl solution to drink. After 3 weeks (20±1 days) of DOCA-salt treatment, these rats as well as the age-matched controls were killed, and the kidneys were removed.
Tissue Preparation
For RNase protection assay, half of the right kidney was
snap-frozen in isopentane (-35°C) and put to RNA isolation. For
immunohistochemistry, half of the right kidney from each rat was
immersion-fixed in buffered 4% paraformaldehyde/0.2%
picric acid for 24 hours. Thereafter, the kidneys were transferred to
10% sucrose in phosphate buffer, immersed for 48 hours at 4°C, and
snap-frozen in isopentane (-35°C). The kidneys were cut at 10-µm
thickness in a cryostat (Leica Frigocut).
RNA Probe Synthesis
A rat renin cDNA fragment including base pairs 12 to 306 was
subcloned into the transcription vector pGEM 4
(Promega).14 The plasmid was linearized with the use of
Pst I. The probe was transcribed with the use of the T7 RNA
polymerase (Boehringer Mannheim) for antisense RNA
production. The size of the probe was 297 bp.
The angiotensinogen probe was obtained from a 291-bp-long BamHI/Pvu II rat angiotensinogen-cDNA template, representing positions 133 through 424 of the angiotensinogen mRNA,15 subcloned into the BamHI site of the vector pGEM 4 (Promega). For linearization, EcoRI was used. A 291-bp-long antisense RNA was transcribed with the use of T7 RNA polymerase (Boehringer Mannheim).
The AT1A probe was obtained from a 3' PCR fragment representing the positions 1394 through 1622 of rat cDNA16 cloned into the vector pCR II (Clontech). For in vitro transcription, the plasmid was linearized with Xba I and transcribed with SP6 RNA polymerase to get a labeled antisense RNA probe. The protected sequence was 237 bp.
The iNOS probe was obtained from a 426-bp fragment inserted into pGEM-5Zf. This fragment, representing positions 1490 through 1915 of human iNOS mRNA,17 was subcloned into the EcoRV site of the vector. For linearization, Sac I was used. A 519-bp antisense probe was transcribed with the use of T7 RNA polymerase (Boehringer Mannheim).
The eNOS probe was obtained from a 616-bp-long rat cDNA fragment spanning exons 21 to 25. This fragment was cloned into the EcoRV site of the BSKS vector. For linearization, Ava II was used, and T7 RNA polymerase was used to transcribe a 221-bp-long antisense RNA.
The nNOS probe was obtained from a 1-kilobase HindIII fragment (base 3121 to 4119 of rat brain NOS cDNA).18 The fragment was inserted into the HindIII site of the transcription vector pGEM7 (Promega). Before transcription, the vector was cleaved with Pvu II, resulting in a 300-bp transcript.
The probes were labeled with [
-32P]UTP for RNase
protection assay and Northern blot analysis, respectively, by
in vitro transcription19 for 90 minutes at 37°C,
followed by treatment with RNase-free DNase I (Boehringer
Mannheim) for 20 minutes at 37°C. The counts of the radioactively
labeled probes were measured with the use of a liquid scintillation
counter. We checked the labeling quality by separating the probes via a
denaturing formaldehyde gel electrophoresis.
RNase Protection Assay
The tissue was homogenized and the RNA extracted
according to a modified guanidinium isothiocyanate/cesium chloride
centrifugation method. RNA was quantified by
measurement of optical density at 260 nm.20 The RNase
protection assay was performed with an Ambion RPA III kit (ITC
Biotechnology GmbH) according to the instructions of the manufacturers
as described previously.19 The specific activity of the
32P-labeled antisense RNA was in the range of
>3.0x108 cpm/mg RNA. Ten to 25 µg of the isolated RNA
was hybridized with the corresponding labeled probe and subsequently
treated with RNase A/T1. The protected fragments were
electrophoretically separated via a 5% denaturing
polyacrylamide gel. The gel was exposed on an image plate, and
the fragments were visualized in a FUJIX BAS 2000 Phospho-imager and
densitometrically quantified. The variability of the assay was
controlled by cohybridization with a 32P-labeled rat
ß-actin antisense RNA.
Northern Blotting
Tissue RNA was extracted as described for the RNase protection
assay. Fifteen micrograms of total tissue RNA was separated via a
denaturing 1% agarose/1 mol/L formaldehyde gel at 100 V for 3
to 4 hours. A mixture of 20 mmol/L
3-[N-morpholine]-propanesulfonic acid, 5 mmol/L sodium
acetate, and 1 mmol/L EDTA, pH 7.0, was used as an
electrophoresis buffer. The buffer was continuously mixed during
electrophoresis. After separation, the RNA was transferred to nylon
membranes by capillary blot in 5x standard saline citrate (1x SSC;
0.15 mol/L sodium chloride, 0.15 mol/L sodium citrate, pH
7.0). After the transfer, the membranes were baked at 80°C for 2
hours in a vacuum oven and subsequently prehybridized at 42°C for 4
hours. The prehybridization mixture contained 5x Denhardt’s solution
(0.05% Ficoll, 0.05% polyvinylpyrrolidone, and 0.05% bovine serum
albumin) and 15x SSC, 1% SDS, and 200 µg of herring sperm
DNA. After prehybridization, the blots were hybridized with the same
buffer containing the radioactively labeled probe at 42°C for 24 to
36 hours. The membranes were washed in 2x SSC for 5 minutes at room
temperature and subsequently twice for 30 minutes with 1x SSC and
0.1x SSC at 45°C and 56°C, respectively. The membranes were
exposed on an image plate, and the fragments were visualized in a FUJIX
BAS 2000 Phospho-imager and densitometrically quantified. The
variability of the assay was controlled by cohybridization with a
32P-labeled rat ß-actin antisense RNA.
Immunocytochemistry
Immunocytochemistry was performed by use of the
avidin-biotin-peroxidase technique (Vectastain Kit, Vector
Laboratories) with DAB used as chromogen. Ang II was detected with a
rabbit polyclonal Ang II antiserum termed Denise.21 This
antiserum cross-reacts with Ang(2-8), Ang(3-8), and Ang(4-8) but only
1% with Ang(1-10). It was used in a dilution of 1:1000 in a PBS buffer
containing 0.04% Triton X-100 and 0.4% dimethyl sulfoxide as well as
1% bovine serum albumin. The staining procedure was done as
described earlier.19 Briefly, the slides were brought to
room temperature, washed in PBS, incubated with
methanol/H2O2 to suppress
endogenous peroxidase activity, blocked with 5% bovine
serum albumin, and subsequently incubated with the diluted
antiserum (see above) overnight at 4°C. For negative controls, the
antiserum was preabsorbed with 15 µg of Ang II per milliliter of
diluted antiserum. After incubation, the slides were washed and the
primary antiserum was detected with the appropriate secondary antibody,
by use of the ABC-Kit (Vectastain, Vector) and DAB. The reaction was
stopped in tap water, glycerin-gelatin was used to coverslip the
slides, and the slides were examined for brownish precipitate under a
microscope (AHBT3, Olympus Optical Co Ltd).
Statistical Analysis
Ang II immunoreactivity was evaluated by counting the signals at
the afferent arterioles versus the glomeruli of the entire kidney
section. The ratio between the number of signals at the afferent
arteriole and the number of glomeruli in the section was used to
determine the percentage of Ang II immunoreactivity. Densitometric mRNA
quantification was done with software supplied by the FUJIX BAS 2000
Phospho-imager system. Data are presented as mean±SE.
Statistical significance of differences in mean values was tested by
two-way ANOVA for repeated measures and the Duncan multiple-range test.
A value of P<.05 was considered statistically
significant.
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Results |
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Tail-cuff blood pressure measurements were slightly higher (P<.05) in SBH/y than in SBN/y, averaging 135±1 and 105±2 mm Hg in the untreated groups and 123±4 and 99±3 mm Hg before DOCA-salt treatment, respectively. After 3 weeks of DOCA-salt treatment, blood pressure increased to 165±5 mm Hg in SBH/y whereas it remained at 106±4 mm Hg in SBN/y. The kidneys of the Sabra rat substrains were morphologically normal and were not affected by DOCA-salt administration (data not shown).
We examined the gene expression values of eNOS, iNOS, and nNOS, before and after DOCA-salt treatment, by means of RNase protection assay. Fig 1 shows a corresponding representative autoradiogram for eNOS, iNOS, and nNOS gene expression. The double bands for eNOS are related to overdigestion of RNA.
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Fig 2 shows the values obtained by RNase protection assay for five rats in each of the four groups. The top left panel demonstrates the gene expression of nNOS in both strains, with and without DOCA salt. In untreated SBN/y and SBH/y, the values leveled off to 114.5±6.9 and 79.4±7.6 AU (P<.05), respectively. After 3 weeks of DOCA-salt administration, the values did not significantly decrease in SBN/y; they leveled off to 114.2±13.8 AU. In SBH/y treated with DOCA salt, nNOS mRNA decreased slightly to 54.5±9.3 AU. Again, there was a significant (P<.05) decreased expression after administration of DOCA salt in SBH/y compared with DOCA-salt–treated SBN/y. The gene expression of eNOS is shown in the top right panel of Fig 2. mRNA for the eNOS shows similar expression levels between the strains and no change after DOCA-salt loading. eNOS mRNA leveled off to 0.07±0.01 and 0.08±0.01 AU in SBN/y and SBH/y, respectively, and was 0.08±0.02 and 0.08±0.003 AU in SBN/y and SBH/y, respectively, after DOCA-salt treatment. In contrast, iNOS mRNA (Fig 2, bottom) was significantly different between the untreated and DOCA-salt–treated Sabra rat strains. In the untreated strains, iNOS leveled off to 101.04±6.8 AU in SBN/y and 72.2±2.6 AU in SBH/y (P<.05). After 3 weeks of DOCA-salt treatment, mRNA levels in the kidneys of SBN/y were 100.8±7.4 AU and leveled off to 68.03±3.9 AU in SBH/y (P<.05). The levels of nNOS mRNA were lower in SBH/y than in SBN/y both before and after DOCA salt. iNOS gene expression was not influenced by DOCA salt in either SBN/y or SBH/y and showed the same pattern as iNOS. eNOS was not different between the strains and was not influenced by DOCA salt.
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Figs 3 and 4 show the expression levels of the renin-angiotensin system genes in SBH/y and SBN/y kidneys before and after DOCA-salt treatment. In Fig 3, the corresponding representative autoradiograms for renin, angiotensinogen, and AT1A receptor mRNA are depicted. Fig 4 demonstrates mRNA values for renin, angiotensinogen, and AT1A receptor in the kidneys of SBH/y and SBN/y, with and without DOCA-salt treatment. Renin mRNA (Fig 4, top left) was significantly (P<.05) increased in untreated SBN/y (0.16±0.05 AU) compared with SBH/y (0.06±0.01 AU). After 3 weeks of DOCA-salt treatment, renin gene expression decreased significantly in SBN/y and SBH/y. In SBH/y, renin mRNA was nearly undetectable after DOCA-salt treatment and was significantly (P<.05) less than in SBN/y treated with DOCA salt. Angiotensinogen gene expression was decreased in untreated SBN/y compared with SBH/y (P<.05) and increased in SBN/y after DOCA-salt treatment (P<.05). In SBH/y, this value remained unchanged. AT1A receptor mRNA (Fig 4, bottom) was also detected in the kidneys of both Sabra rat substrains and was not different between the untreated strains. DOCA-salt treatment exerted no effect.
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We also looked for Ang II immunoreactivity in the kidneys of both Sabra rat strains before and after DOCA-salt treatment. Ang II immunoreactivity was significantly higher (P<.05) in SBN/y (n=5) than in SBH/y (n=5) and leveled off to 19.4±2.4% and 7.7±0.4% (number of glomeruli divided by stained afferent arterioles), respectively. Ang II could not be detected by immunohistochemistry after DOCA-salt treatment in either strain. An example of Ang II immunoreactivity before (left) and after (right) DOCA-salt treatment in SBN/y is demonstrated in Fig 5. With DOCA salt, the signal disappeared.
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Discussion |
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The important findings in this study were that iNOS and nNOS gene expression values in the kidneys of SBN/y were significantly higher than in SBH/y, whereas eNOS gene expression was not different. These values were not influenced by DOCA salt. SBN/y had greater renin gene expression than SBH/y, although its angiotensinogen expression was somewhat lower. Ang II staining in the kidneys of SBN/y was also more prominent than in SBH/y. DOCA salt sharply decreased renin gene expression, and Ang II was no longer detectable in the kidneys. We were not able to find differences in AT1A receptor gene expression and detected no effect of DOCA salt on this gene. These findings support the notion that the reduced NO generation of SBH/y may be related to genetic differences between the rat substrains but is not a consequence of hypertension. Therefore, the higher NO generation in SBN/y may play a role in their resistance to DOCA salt.
In our functional study,2 we examined pressure-diuresis and -natriuresis relationships (renal function curves) in SBH/y and SBN/y under two different circumstances. We studied the renal function curves with all extrinsic systems controlled (clamped) by simultaneously infusing aldosterone, 17-OH-corticosterone, norepinephrine, and vasopressin after renal denervation. We then repeated these studies without controlling these extrinsic systems. With this approach, we were able to separate the extrinsic regulatory systems from intrinsic effects emanating from within the kidneys themselves. We found that the renal function curves of SBH/y had a more rightward shift when the extrinsic systems were operative than when they were clamped. This finding suggested that extrinsic factors regulating sodium and water excretion were more important than intrinsic influences in the kidney in terms of shifting the pressure-diuresis and pressure-natriuresis relationships rightward. Nevertheless, a degree of rightward shift remained, indicating that intrinsic renal mechanisms were also operative. We suggest that both the extrinsic and intrinsic renal influences we identified are possibly related to a decrease in iNOS and nNOS gene expression in SBH/y compared with SBN/y. We additionally propose that these two genes are important candidate genes in this form of genetic rat hypertension.
Numerous studies have demonstrated that the different isoforms of NOS are localized in the kidney. Immunohistochemical and PCR data demonstrated that nNOS is present in the macula densa, inner and outer medullas, the glomerulus, vasa recta, and arcuate artery. In contrast, eNOS mRNA is mostly present in high levels in the glomeruli and the preglomerular vasculature. The two separate iNOS isoforms have also been identified in renal vascular and tubular segments of normal rats.10 22 Both short-term and long-term NOS inhibition have a profound effect on renal function and blood pressure and shift the pressure-natriuretic and -diuretic relationship rightward to higher levels of perfusion pressure. In salt-sensitive Dahl rats, renal NO release is decreased, which may contribute to the development of salt-sensitive hypertension.23 DOCA-salt–treated Wistar rats also showed a decreased NO release.24 Furthermore, in Dahl salt-resistant rats, the activity of the NO system is increased compared with Dahl salt-sensitive rats.25 A role of the NO system in the development of hypertension after DOCA-salt treatment has also been suggested for SBH.9 Possibly, the remarkable resistance of SBN/y to DOCA-salt hypertension is related to a greater NO generation. Data from salt-loaded Sprague-Dawley rats suggest that increased NO synthesis in response to salt intake may facilitate sodium excretion and allow maintenance of normal blood pressure.26 In Dahl salt-sensitive rats, high salt intake led to decreased nNOS activity, which may be responsible for their salt sensitivity.7 In the present study, the DOCA salt had no additional effect on nNOS and iNOS mRNAs in either SBH/y or SBN/y. Rats appear to be quite variable in terms of their NOS responses after high salt intake. The reasons for these differences are not clear. They may be genetically determined in part.
DOCA-salt loading per se increases sympathetic nerve activity.27 Because the development of hypertension induced by NG-nitro-L-arginine methyl ester is attenuated in renally denervated rats, Matsuoka et al28 suggested that the integrity of the sympathetic nervous system is a requirement for the development of hypertension after NO inhibition. Thus, the decreased nNOS and iNOS in the kidney of SBH/y may be relevant to the genesis of salt-sensitive hypertension through increased renal sympathetic nerve activity, an extrinsic renal modulator. Such an effect could lead to changes in sodium and water excretion, as well as to a blood pressure increase. This possibility would also be consistent with our finding that extrinsic mechanisms were more important than intrinsic renal regulators. This interpretation implies that the renin-angiotensin system and the NO system operating in the kidney locally are not the key factors responsible for DOCA-salt–induced hypertension in SBH/y.
We confirmed the presence of an intact renin-angiotensin system in the kidneys of the Sabra rat strains. However, we could not detect the increased AT1A receptor mRNA in kidneys of prehypertensive SBH reported by others.29 We measured AT1A mRNA in whole kidney, whereas Bouby et al29 studied the localization and distribution of AT1A receptor mRNA in four different zones of the kidney. They found an increase only in the inner stripe of the outer medulla of SBH rats. In their study, Bouby et al29 detected AT1A mRNA by reverse-transcription PCR. We used RNase protection assay and Northern blotting for quantification of the mRNA levels and also used the new, rebred Sabra rat substrains termed SBH/y and SBN/y. Differences in rat strains and in the experimental design may be responsible for the divergent observations. The finding that the renin content of the afferent arterioles was reduced after DOCA-salt loading both in SBN/y and SBH/y is in agreement with the observation that the renin activity of glomeruli is extremely reduced in DOCA-salt–treated rats.30 This finding fits with the observation that we could not detect Ang II in the afferent arterioles in either rat strain after DOCA-salt treatment. Moreover, Dahl salt-sensitive and salt-resistant rats also react with suppressed renal renin immunoreactivity after salt loading.31
We can only speculate about the mechanism(s) responsible for the difference in renin gene expression we observed between the two strains. The patterns of potassium and sodium plasma levels detected in our earlier study2 were not different between the two strains and therefore cannot be the reason for this difference. The 10- to 20-mm Hg higher blood pressure of SBH/y compared with SBN/y should be too small to induce changes in renin gene expression. After DOCA salt, SBH/y had a significantly lower diuretic and natriuretic response than SBN/y.2 This finding suggests that SBH/y may have a tendency to retain salt and water and therefore may have an increased extracellular fluid volume, which in turn could be responsible for the decreased renin gene expression in SBH/y.
Interestingly, Dahl salt-sensitive rats have also been reported to have lower renal renin activity than the Dahl salt-resistant rat.31 Therefore, the reduced renin gene expression in SBH/y rats may be common in salt-sensitive rat strains and may reflect an altered regulation of the renin gene expression in kidneys of these rats. In this context, it may be interesting that changes in renin mRNA are primarily obtained by changing the number of cells that express the renin gene rather than by changing the mRNA level per cell.31 Our histological examination gave no evidence of an altered morphology of the juxtaglomerular apparatus. Furthermore, whether or not the decreased renin mRNA in SBH/y is connected with the NO content in the kidney is not clear. There are reports suggesting a direct influence of macula densa–derived NO on renin release.32 33 34 35 Conceivably, the lower NO content in the kidneys of SBH/y could have led to decreased renin gene expression because NO may function as a paracrine stimulus for the local regulation of renin release.12 13 Additional support for NO-related genes in salt-sensitive hypertension stems from the recent report by Deng and Rapp,36 who found that a locus for iNOS but not cNOS cosegregates with blood pressure in the Dahl salt-sensitive rat.
The components of the renin-angiotensin system are believed to be candidates for development of nephrosclerosis in DOCA-salt hypertension.37 We found that DOCA salt did not induce nephrosclerosis in SBH/y. Therefore, the observed morphological protection of the kidneys from DOCA-salt influence may be connected with altered regulation of the renin gene expression in SBH/y. Because we did not detect differences in the renin-angiotensin system after DOCA-salt treatment between the two substrains, it seems unlikely that the renin-angiotensin system is responsible for the differences in renal blood flow, glomerular filtration rate, and sodium and water excretion observed between SBN/y and SBH/y after DOCA-salt administration, as reported in our earlier study.2
In summary, whereas renin gene expression is less in SBH/y than in SBN/y, nNOS and iNOS are decidedly less as well. Renin gene expression and immunocytochemical documentation of Ang II are markedly suppressed by DOCA salt. The lower renin gene expression values of SBH/y persisted, and perhaps low renin is a hallmark of salt sensitivity. The lower nNOS and iNOS gene expressions in SBH/y compared with SBN/y suggest lower NO production in SBH/y. The inability to increase local NO production with DOCA salt may contribute to DOCA-salt hypertension in SBH/y.
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Acknowledgments |
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This study was supported by a grant-in-aid from the Deutsche Forschungsgemeinschaft to Volkmar Gross and Friedrich C. Luft. We are grateful to Anita Müller, Edith Richter, Regina Uhlmann, and Christel Lipka for technical assistance.
Received January 30, 1997; first decision February 14, 1997; accepted February 14, 1997.
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References |
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1. Yagil C, Gurion K, Sperenza R, Stolpe C, Kreutz R, Lindpaintner K, Ganten D, Ben-Ishay D, Yagil Y. Development, genotype and phenotype of a new colony of the Sabra hypertension prone (SBH/y) and resistant (SBN/y) rat model of salt sensitivity and resistance. J Hypertens. 1996;14:1175-1182.[Medline] [Order article via Infotrieve]
2. Gross V, Lippoldt A, Yagil C, Yagil Y, Luft FC.
Pressure natriuresis in salt-sensitive and salt-resistant Sabra
rats. Hypertension. 1997;29:1252-1259.
3. Yagil Y, Mekler J, Wald H, Popovtzer MM, Ben-Ishay D. Sodium handling by the Sabra hypertension prone (SBH) and resistant (SBN) rats. Pflugers Arch. 1986;407:547-551.[Medline] [Order article via Infotrieve]
4. Yagil Y, Ben-Ishay D, Wald H, Popovtzer MM. Water handling by the Sabra hypertension prone (SBH) and resistant (SBN) rats. Pflugers Arch. 1985;404:61-66.[Medline] [Order article via Infotrieve]
5. Ben-Ishay D. The Sabra hypertension-prone and -resistant strains. In: DeJong W, ed. Handbook of Hypertension, Vol 4: Experimental and Genetic Models of Hypertension. Amsterdam, Netherlands: Elsevier Scientific Publishers; 1984:296-313.
6. Majid DS, Williams A, Navar LG. Inhibition of nitric oxide synthesis attenuates pressure-induced natriuretic responses in anesthetized dogs. Am J Physiol. 1993;264:F79-F87.[Medline] [Order article via Infotrieve]
7. Ikeda Y, Saito K, Kin JI, Yokoyama M. Nitric oxide synthase isoform activities in kidney of Dahl salt-sensitive rats. Hypertension. 1995;26(pt 2):1030-1034.
8. Hu L, Manning RD Jr. Role of nitric oxide in regulation of long-term pressure-natriuresis relationship in Dahl rats. Am J Physiol. 1995;268:H2375-H2383.[Medline] [Order article via Infotrieve]
9. Rees D, Ben-Ishay D, Moncada S. Nitric oxide
and the regulation of blood pressure in the hypertension-prone and
hypertension-resistant Sabra rat.
Hypertension. 1996;28:367-371.
10. Mattson DL, Higgins DJ. Influence of dietary sodium intake on renal medullary nitric oxide synthase. Hypertension. 1996;27(pt 2):688-692.
11. Pohlova I, Ziche J. The influence of high salt intake and/or chronic blood volume expansion on renin-angiotensin system in Brattleboro rats. Acta Physiol Scand. 1992;145:115-120.[Medline] [Order article via Infotrieve]
12. Johnson RA, Freeman RH. Renin release in rats during blockade of nitric oxide synthesis. Am J Physiol. 1994;266:R1723-R1729.[Medline] [Order article via Infotrieve]
13. Schricker K, Heggyi I, Hamann M, Kaissling B, Kurtz
A. Tonic stimulation of renin gene expression by nitric oxide is
counteracted by tonic inhibition through angiotensin
II. Proc Natl Acad Sci U S A. 1995;92:8006-8010.
14. Burnham CE, Hawelu-Johnson CL, Frank BM, Lynch
KR. Molecular cloning of rat renin cDNA and its gene.
Proc Natl Acad Sci U S A. 1987;84:5605-5609.
15. Okhubo H, Kageyama R, Ujihara M, Hirose T, Inayama S,
Nakanishi S. Cloning and sequence analysis of cDNA for
rat angiotensinogen. Proc Natl Acad Sci
U S A. 1983;80:2196-2200.
16. Murphy TJ, Alexander WR, Griendling KK, Runge MS, Bernstein KE. Isolation of a cDNA encoding the vascular type 1 angiotensin II receptor. Nature. 1991;351:233-236.[Medline] [Order article via Infotrieve]
17. Geller DA, Lowenstein CJ, Shapiro RA, Nussler AK, Di
Silvio M, Wang SC, Nakayama DK, Simmons RL, Snyder SH, Billiar
TR. Molecular cloning and expression of inducible nitric oxide
synthase from human hepatocytes. Proc Natl
Acad Sci U S A. 1993;90:3491-3495.
18. Bredt DS, Hwang PM, Glatt CE, Lowenstein C, Reed RR, Snyder SH. Cloned and expressed nitric oxide synthase structurally resembles cytochrome P-450 reductase. Nature. 1991;351:714-718.[Medline] [Order article via Infotrieve]
19. Lippoldt A, Gross V, Ganten U, Luft FC. Life-long angiotensin converting enzyme inhibition, pressure natriuresis, and renin angiotensin system gene expression in transgenic (mRen-2)27 rats. J Am Soc Nephrol. 1996;7:2119-2129.[Abstract]
20. Auffray C, Rougeon F. Purification of mouse immunoglobulin heavy chain messenger RNAs from total myeloma tumor RNA. Eur J Biochem. 1980;107:303-314.[Medline] [Order article via Infotrieve]
21. Brownfield MS, Reid IA, Ganten D, Ganong WF. Differential distribution of immunoreactive angiotensin and angiotensin converting enzyme in the rat brain. Neuroscience. 1982;7:1759-1769.[Medline] [Order article via Infotrieve]
22. Mattson DL, Bellehumeur TG. Neural nitric oxide
synthase in the renal medulla and blood pressure regulation.
Hypertension. 1996;28:297-303.
23. Chen PY, Sanders PW. Role of nitric oxide
synthesis in salt-sensitive hypertension in Dahl/Rapp rats.
Hypertension. 1993;22:812-818.
24. Hayakawa H, Hirata Y, Suzuki E, Kimura K, Kikuchi K, Nagano T, Hirobe M, Omata M. Long-term administration of L-arginine improves nitric oxide release from kidney in deoxycorticosterone acetate-salt hypertensive rats. Hypertension. 1994;23(pt 1):752-756.
25. Chen PY, Sanders PW. L-Arginine abrogates salt-sensitive hypertension in Dahl/Rapp rats. J Clin Invest. 1991;88:1559-1567.[Medline] [Order article via Infotrieve]
26. Shultz PJ, Tolins JP. Adaptation to increased dietary salt intake in the rat: role of endogenous nitric oxide. J Clin Invest. 1993;91:642-650.[Medline] [Order article via Infotrieve]
27. Katholi RE, Naftilan AJ, Oparil S. Importance of
renal sympathetic tone in the development of DOCA-salt hypertension in
the rat. Hypertension. 1980;2:266-273.
28. Matsuoka H, Nishida M, Nomura G, Van Vliet BN, Toshima
M. Hypertension induced by nitric oxide synthesis inhibition is
renal nerve dependent. Hypertension. 1994;23:971-975.
29. Bouby N, Bankir L, Llorens-Cortes C. Type 1 angiotensin II receptor subtypes in kidney of normal and salt-sensitive hypertensive rats. Hypertension. 1996;27(pt 1):392-398.
30. Schnermann J, Hermle M, Schmidmeier E, Dahlheim H. Impaired potency for feedback regulation of glomerular filtration rate in DOCA escaped rats. Pflugers Arch. 1975;358:339-348.[Medline] [Order article via Infotrieve]
31. Campbell WG Jr, Gahnem F, Catanzaro DF, James GD,
Camargo MJF, Laragh JH, Sealey JE. Plasma and renal
prorenin/renin, renin mRNA, and blood pressure in Dahl salt-sensitive
and salt-resistant rats. Hypertension. 1996;27:1121-1133.
32. Fischer E, Schnermann J, Briggs JP, Kriz W, Ronco PM, Bachmann S. Ontogeny of NO synthase and renin in juxtaglomerular apparatus of rat kidney. Am J Physiol. 1995;268:F1164-F1176.[Medline] [Order article via Infotrieve]
33. Kurtz A. Cellular control of renin secretion. Rev Physiol Biochem Pharmacol. 1989;113:1-40.[Medline] [Order article via Infotrieve]
34. Takagi M, Franco-Saenz R, Mulrow PJ. Effect of
atrial natriuretic peptide on renin release in a
superfusion system of kidney slices and dispersed
juxtaglomerular cells. Endocrinology. 1988;122:1437-1442.
35. Schricker K, Kurtz A. Liberators of NO exert dual effect on renin secretion from isolated mouse renal juxtaglomerular cells. Am J Physiol. 1993;265:F180-F186.[Medline] [Order article via Infotrieve]
36. Deng AY, Rapp JP. Locus for the inducible, but not a constitutive, nitric oxide synthase cosegregates with blood pressure in the Dahl salt-sensitive rat. J Clin Invest. 1995;95:2170-2177.[Medline] [Order article via Infotrieve]
37. Kim S, Ohta K, Hamaguchi A, Omura T, Yukimura T, Miura
K, Inada Y, Wada T, Ishimura Y, Chatani F, Iwao H. Role of
angiotensin II in renal injury of deoxycorticosterone
acetate-salt hypertensive rats. Hypertension. 1994;24:195-204.
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