Dahl Salt-sensitive Hypertensive Rats Research Articles

Exclusion of the Catechol-O-Methyltransferase Gene from Genes Contributing to Salt-Sensitive Hypertension in Dahl Salt-Sensitive Rats

Catechol-O-methyltransferase (COMT) is an enzyme that inactivates catecholamines. Several studies have suggested that this enzyme may play a role in blood pressure regulation. We previously reported that the expression levels of Comt mRNA in Dahl salt-sensitive (DS) rats were lower than those in Lewis (LEW) rats. However, the physiological significance of this phenomenon has not been investigated. The purpose of the present study was to evaluate the significance of lower expression of Comt in Dahl salt-sensitive hypertension. The Comt gene in DS rats has a palindromic insertion in 3'-untranslated region, which appears to be responsible for reduced mRNA stability. A genome-wide quantitative trait loci (QTL) analysis of blood pressure using 107 F2 rats indicated that a statistically significant QTL for pulse pressure was located at the Comt locus in chromosome 11. Microarray analysis confirmed that Comt was the only gene differentially expressed between DS and LEW rats in this chromosomal region. However, COMT inhibitors had no significant effects on blood pressure in either DS or LEW rats. Thus, Comt was excluded from the candidate genes contributing to salt-sensitive hypertension in DS rats. A true gene responsible for pulse pressure in this chromosome 11 region remains to be determined.

Isohumulones Derived from Hops Ameliorate Renal Injury via an Anti-Oxidative Effect in Dahl Salt-Sensitive Rats

Previous studies have reported that isohumulones, the bitter compounds in beer, improve insulin resistance and hyperlipidemia in several animal models. In this study, we examined whether isohumulones ameliorate renal injury. Dahl salt-sensitive hypertensive rats were fed a low-salt diet (LS), a high-salt diet (HS) or a high-salt diet containing 0.3% isohumulones (HS+IH) for 4 weeks. Urinary nitrite/nitrate (NOx) excretion was measured at 4 weeks along with blood pressure and urinary protein excretion. Renal injury was evaluated histologically and reactive oxygen species (ROS) and nitric oxide (NO) production in the renal cortex was visualized. Oxidative stress and NO synthase (NOS) expression were evaluated by immunohistochemical staining and Western blot analysis. Mean blood pressure was significantly decreased in the HS+IH group compared with the HS group at 4 weeks (158.1+/-8.7 vs. 177.5+/-3.7 mmHg; p<0.05). Isohumulones prevented the development of proteinuria in the HS+IH group compared with the HS group at 2 weeks (61.7+/-26.8 vs. 117.2+/-9.8 mg/day; p<0.05). Glomerulosclerosis and interstitial fibrosis scores were significantly decreased in the HS+IH group compared with the HS group (0.61+/-0.11 vs. 1.55+/-0.23, 23.7+/-6.8 vs. 36.1+/-3.5%; p<0.05 for both). In the HS group, increased ROS and decreased NO were observed in glomeruli in vivo. Isohumulones reduced the ROS production, leading to the restoration of bioavailable NO. Urinary NOx excretion was significantly increased in the HS+IH group compared with the HS group. Furthermore, renal nitrotyrosine was increased in the HS group compared with the LS group, and this effect was prevented by isohumulones. Renal NOS expression did not differ among the three groups. These results suggest that isohumulones may prevent the progression of renal injury caused by hypertension via an anti-oxidative effect.

DIFFERENTIAL EFFECTS OF A HIGH-SALT DIET ON RENAL EXPRESSIONS OF AT1 RECEPTOR, NAD(P)H OXIDASE, COX-1, COX-2, SOD, GPX, MCP-1, AND PAI-1 IN SALT-SENSITIVE AND -RESISTANT RATS.

Chronic consumption of a high-salt diet causes hypertension (HTN) and renal injury in Dahl salt-sensitive (DSS), but not salt-resistant (DSR) rats. These events are, in part, mediated by oxidative stress and inflammation in the kidney and cardiovascular tissues. Local activation of AT1 receptor plays an important role in the pathogenesis of oxidative stress and inflammation in hypertensive disorders. However, systemic renin-angiotensin system is typically depressed in salt-sensitive HTN. It is of note that circulating and tissue renin-angiotensin systems operate independently. Since renal tissue angiotensinogen is reportedly elevated in hypertensive DSS rats, we hypothesized that differential response to high-salt diet in DSS versus DSR rats may be related to dysregulations of intrarenal AT1 receptor and factors involved in inflammation and reactive oxygen species production. Subgroups of DSS and DSR rats were fed a high-salt (8% NaCl) or low-salt (0.03% NaCl) diet for 2 weeks. DSS rats consuming a regular diet exhibited significant increases in AT1 receptor and COX-2 abundance in the kidney. High-salt intake resulted in marked HTN, proteinuria, and up-regulations of P22phox, MCP-1, and PAI-1, as well as further rises in AT1 receptor and COX-2 abundance in the DSS but not the DSR rat kidneys. Among animals fed a low-salt diet, glutathione peroxidase (GPX) was significantly lower in the DSS compared with DSR kidneys. Consumption of a high-salt diet raised kidney GPX in DSR but not DSS rats and lowered kidney CuZn SOD in DSS rats. Thus, high-salt intake results in up-regulations of AT1 receptor, COX-2, NAD(P)H oxidase, MCP-1, and PAI-1 and down-regulations of key antioxidant enzymes, GPX and CuZn SOD, in DSS rats. These events can contribute to oxidative stress, inflammation, and tissue damage in the DSS kidney.

High‐Throughput Production and Phenotyping of Rat Knockout Models for Hypertension

The inability to generate knock-out (KO) rats has been a major limitation for studying gene function in rats. PhysGen-MCWuses a strategy that combines ENU mutagenesis to induce spermatic gene mutations, with TILLING, a high-throughput assay which identifies mutations between wild-type and mutated alleles to confirm and identify KO rats. The goal is to identify the induced point mutations in 112 selected genes in three different rat strains Dahl Salt-Sensitive, Brown Norway, and Fawn-Hooded Hypertensive rat with the broad goal of developing 99 total strains of KO rats (33 genes on 3 different genome backgrounds), which are then characterized physiologically for of cardiovascular, pulmonary and behavioral traits. We have currently developed 26 gene-mutation strains, of which six are KO for different genes. Males are injected with a variable dose of ENU and then bred to wild-type females to produce the KO rats, which is then brought to homozygosis. An ultrasound protocol assesses cardiac function by echocardiography in basal and stressed (dobutamine administration) conditions. Vascular function is studied in aortic rings with responsiveness to vasoconstrictors and vasodilators. Plasma levels 29 hormones, electrolytes and blood indicators are also measured. Blood pressure is determined and kidney function is assessed by protein excretion. KO rats availability and data is posted in the PGA web site periodically (pga.mcw.edu).

Hyperhomocysteinemia – association with renal transsulfuration and redox signaling in rats

Despite substantial evidence indicating the association of hyperhomocysteinemia (hHcys) and end-stage renal disease (ESRD), the pathogenic role of increased plasma homocysteine (Hcys) levels in the progression of ESRD remains unclear. This review will briefly summarize recent findings regarding the role of hHcys in the development of glomerulosclerosis, the association of hHcys with reduced renal transsulfuration and Hcys-induced changes of redox signaling in the development of glomerulosclerosis in rat kidneys. Based on these results, it is concluded that hHcys is implicated in glomerular sclerosis in hypertension, elevated plasma Hcys in Dahl salt-sensitive (SS) hypertensive rats is due to downregulation of cystathionine beta-synthase (CBS) expression and consequent abnormality of transsulfuration in the kidney compared with normotensive rats. Hcys-induced superoxide (O(2)(*-)) production by activation of NADPH oxidase as a triggering mechanism contributes to the effects of Hcys on the homeostasis of extracellular matrix and consequent sclerosis in the glomeruli, and NADPH oxidase activation by Hcys is associated with enhanced Rac GTPase activity.

Pravastatin Enhances Beneficial Effects of Olmesartan on Vascular Injury of Salt-Sensitive Hypertensive Rats, via Pleiotropic Effects

This work was undertaken to investigate comparative effect of AT1 receptor blocker (ARB), 3-hydroxy-3-methylglutaryl (HMG) coenzymeA (CoA) reductase inhibitor (statin), and their combination on vascular injury of salt-sensitive hypertension. Salt-loaded Dahl salt-sensitive hypertensive rats (DS rats) were treated with (1) vehicle, (2) hydralazine (5 mg/kg/d), (3) olmesartan (0.5 mg/kg/d), (4) pravastatin (100 mg/kg/d), and (5) combined olmesartan and pravastatin for 4 weeks. Olmesartan or pravastatin significantly and comparably improved vascular endothelium-dependent relaxation to acetylcholine, coronary arterial remodeling, and eNOS activity of DS rats. Olmesartan prevented vascular eNOS dimer disruption or the downregulation of dihydrofolate reductase (DHFR) more than pravastatin, whereas Akt phosphorylation was enhanced by pravastatin but not olmesartan, indicating differential pleiotropic effects between olmesartan and pravastatin. Add-on pravastatin significantly enhanced the improvement of vascular endothelial dysfunction and remodeling by olmesartan in DS rats. Moreover, pravastatin enhanced the increase in eNOS activity by olmesartan, being associated with additive effects of pravastatin on phosphorylation of Akt and eNOS. Olmesartan and pravastatin exerted beneficial vascular effects in salt-sensitive hypertension, via differential pleiotropic effects. Pravastatin enhanced vascular protective effects of olmesartan. Thus, the combination of ARB with statin may be the potential therapeutic strategy for vascular diseases of salt-sensitive hypertension.

Individual QTLs controlling quantitative variation in blood pressure inherited in a Mendelian mode

We studied three possible genotypes at 10 well-defined blood pressure (BP) QTLs using congenic rat lines. The central question was whether the hypertensive or normotensive allele is dominant, or whether there is partial dominance. The congenic strains were employed to investigate the BP effects of alleles originating from normotensive rats in the background of hypertensive Dahl salt-sensitive (DSS) rats. The normotensive alleles at eight QTLs were fully dominant over DSS alleles, which we tentatively interpreted as indicating that DSS rats incurred a loss of function at these loci and that the QTLs produced BP-reducing agents. In contrast, the normotensive allele of only one QTL was recessive over its DSS counterpart, implying a gain of function at this QTL or a null allele involved in generating a BP-elevating agent. Only one locus, C17QTL, had alleles exhibiting partial dominance. These estimates of dominance differ considerably from those obtained by QTL analysis in a F2 cross. This disagreement demonstrates the importance of establishing a cause-effect relationship between a QTL and its phenotypic effect via congenic strains. The dominance relationships suggest pertinent strategies for gene identification and pharmaceutical intervention.

NO Synthase Uncoupling in the Kidney of Dahl S Rats

NO synthase (NOS) can paradoxically contribute to the production of reactive oxygen species when l-arginine or the cofactor R-tetrahydrobiopterin (BH(4)) becomes limited. The present study examined whether NOS contributes to superoxide production in kidneys of hypertensive Dahl salt-sensitive (SS) rats compared with an inbred consomic control strain (SS-13(BN)) and tested the hypothesis that elevated dihydrobiopterin (BH(2)) levels are importantly involved in this process. This was assessed by determining the effects of l-nitroarginine methyl ester (l-NAME) inhibition of NOS on superoxide production and by comparing tissue concentrations of BH(4) and BH(2). A reverse-phase high-performance liquid chromatography method was applied for direct measurements of BH(4) and BH(2) using (S)-tetrahydrobiopterin as an internal standard. Superoxide concentrations were measured in vivo from medullary microdialysis fluid using dihydroethidine and in vitro using lucigenin. The results indicate the following: (1) that superoxide levels were elevated in the outer medulla of SS rats fed a 4% salt diet and could be inhibited by l-NAME. In contrast, l-NAME resulted in elevated superoxide production in consomic SS-13(BN) rats because of higher NOS activity; (2) SS rats showed a reduced ratio of BH(4)/BH(2) in the outer medulla that was driven by increased concentrations of BH(2); and (3) lower superoxide dismutase and catalase activities contributed to elevated reactive oxygen species in SS samples. Based on the shift of BH(4) to BH(2) and the observation of l-NAME inhibitable superoxide production, we conclude that NOS uncoupling occurs in the renal medulla of hypertensive SS rats fed a high-salt diet.

Angiotensin II, oxidative stress, and extracellular matrix degradation during transition to LV failure in rats with hypertension

Angiotensin II (Ang-II) plays pivotal roles in the progression of left ventricular (LV) remodeling in diseased hearts; it remains to be elucidated how Ang-II links to degradation of the extracellular matrix (ECM). Using hypertensive Dahl salt-sensitive rats that show the distinctive transition from concentric LV hypertrophy to LV remodeling, we chronically treated them with an angiotensin type-1 receptor blocker (telmisartan 5 mg/kg/day, ARB group) or vehicle (0.5% CMC, CHF group). During the process of LV remodeling, we assessed, (1) in-vivo LV shape and function; (2) animal survival; (3) amounts of ECM in LV using a scanning electron microscope (SEM); (4) mRNA (by real time RT-PCR) and protein (by immunoblotting) levels in LV of NADPH oxidase, glutathione peroxidase-1 (GPX-1), and matrix metalloproteinase (MMP)-2, -9, and -13; (5) immunohistochemical staining of myocardial 4-hydroxy-2-nonenal and 8-hydroxy-2′-deoxyguanosine; (6) nuclear factor kappa-B (NFκB) protein levels in the nuclear extract; and (7) endogenous activities of MMP-2 and -9 by an antibody capture method. Compared with CHF, ARB group showed an improvement of survival and preserved LV shape and function, and ECM density in SEM that was accompanied by decreases in oxidative stress-mediated protein degenerations, activities of GPX-1, NADPH oxidase, NFκB, and MMP-2, -9, and -13. Local activation of Ang-II in hypertrophic LV triggers MMP-mediated ECM degradation, namely LV remodeling, at least in part, through NADPH oxidase-induced oxidative stress and the subsequent NFκB activation.

Distinct rapid and slow phases of salt-induced hypertension in Dahl salt-sensitive rats

To test the hypothesis that Dahl salt-sensitive (Dahl-S) rats exhibit distinct and separable phases of salt sensitivity. Blood pressure (BP) telemetry was used to describe the detailed time course of salt-induced hypertension in Dahl-S rats and in hybrid rats derived from Dahl-S and Dahl salt-resistant strains. Switching to a high salt (4% NaCl) diet led to a biphasic increase in BP. Phase-1 reached a plateau in 4 days whereas phase-2 progressed slowly over the subsequent 5 weeks. In hybrid rats, phase-1 was present in each rat whereas phase-2 was absent in many individuals. A correlation of the amplitude of the first and second phases was of borderline significance in Dahl-S rats (P = 0.053, R2 = 0.44, n = 9) but was clearly significant in hybrid rats (P < 0.0001, R2 = 0.78, n = 22). Increases in BP were reversible following 1 week of high salt but progressively less so after 4 and 7 weeks. Estimation of the chronic pressure-natriuresis relationship suggests that phase-1 is attributable to a reduced slope of this relationship. In contrast, phase-2 corresponds with a further reduction in slope and a progressive and irreversible resetting of the relationship to higher BP levels. Two phases of salt sensitivity coexist and provide distinct contributions to salt-induced hypertension in Dahl-S rats. Our data also suggest that short-term measures of salt-sensitivity may be predictive of the effect of salt on the eventual progression of salt-induced hypertension.

Renal NF-κB activation and TNF-α upregulation correlate with salt-sensitive hypertension in Dahl salt-sensitive rats

Molecular mechanisms of salt-sensitive (SS) hypertension related to renal inflammation have not been defined. We seek to determine whether a high-salt (HS) diet induces renal activation of NF-kappaB and upregulation of TNF-alpha related to the development of hypertension in Dahl SS rats. Six 8-wk-old male Dahl SS rats received a HS diet (4%), and six Dahl SS rats received a low-sodium diet (LS, 0.3%) for 5 wk. In the end, mean arterial pressure was determined in conscious rats by continuous monitoring through a catheter placed in the carotid artery. Mean arterial pressure was significantly higher in the HS than the LS group (177.9 +/- 3.7 vs. 109.4 +/- 2.9 mmHg, P < 0.001). There was a significant increase in urinary albumin secretion in the HS group compared with the LS group (22.3 +/- 2.6 vs. 6.1 +/- 0.7 mg/day; P < 0.001). Electrophoretic mobility shift assay demonstrated that the binding activity of NF-kappaB p65 proteins in the kidneys of Dahl SS rats was significantly increased by 53% in the HS group compared with the LS group (P = 0.007). ELISA indicated that renal protein levels of TNF-alpha, but not IL-6, interferon-gamma, and CCL28, were significantly higher in the HS than the LS group (2.3 +/- 0.8 vs. 0.7 +/- 0.2 pg/mg; P = 0.036). We demonstrated that plasma levels of TNF-alpha were significantly increased by fivefold in Dahl SS rats on a HS diet compared with a LS diet. Also, we found that increased physiologically relevant sodium concentration (10 mmol/l) directly stimulated NF-kappaB activation in cultured human renal proximal tubular epithelial cells. These findings support the hypothesis that activation of NF-kappaB and upregulation of TNF-alpha are the important renal mechanisms linking proinflammatory response to SS hypertension.

Addition of spironolactone to an angiotensin-converting enzyme inhibitor decreases lung congestion and edema in Dahl hypertensive rats

We investigated the effect of adding spironolactone to treatment with an angiotensin-converting enzyme (ACE) inhibitor, imidapril, in Dahl salt-sensitive (DS) hypertensive heart failure rats with preserved systolic function. Male DS rats were fed laboratory chow containing 8% NaCl from 7 weeks of age. Rats were divided into four groups and treated for 9 weeks with vehicle alone (water; n = 23), the ACE inhibitor imidapril (1 mg kg(-1) day(-1); n = 16), spironolactone (2 mg kg(-1) day(-1); n = 15), or a combination of imidapril and spironolactone at the doses above (n = 16). The left ventricular weight to body weight (BW) ratio was significantly lower in the imidapril group (3.28 +/- 0.30 mg g(-1)) and the combination group (3.34 +/- 0.38 mg g(-1)) than in the vehicle group (3.71 +/- 0.46 mg g(-1)). Adding spironolactone to imidapril inhibited an increase in the ratio of lung weight to BW (4.38 +/- 0.50 mg g(-1)) related to high salt intake, while monotherapy (imidapril group, 4.61 +/- 0.90 mg g(-1); spironolactone group, 5.40 +/- 2.50) did not significantly change the ratio from that seen with vehicle treatment (6.32 +/- 3.62 mg g(-1)). All active treatments (imidapril, 0.66% +/- 0.67%; spironolactone, 0.51% +/- 0.55%; both together, 0.31% +/- 0.26%) inhibited a salt-intake related increase in the percentage area representing fibrous tissue compared with vehicle treatment alone (1.81% +/- 1.51%). These findings suggest that adding spironolactone to an ACE inhibitor is more effective in improving pulmonary congestion and edema in hypertensive heart failure with preserved systolic function than an ACE inhibitor alone.

Benazepril Combined with Either Amlodipine or Hydrochlorothiazide Is More Effective than Monotherapy for Blood Pressure Control and Prevention of End-organ Injury in Hypertensive Dahl Rats

We studied the effect of hydrochlorothiazide (HCTZ), the angiotensin-converting enzyme inhibitor benazepril, the calcium channel blocker amlodipine, or a combination of benazepril/amlodipine or benazepril/HCTZ on systolic blood pressure (BP) and end-organ injury (left ventricular hypertrophy, proteinuria, and endothelium-dependent relaxation to acetylcholine) in hypertensive Dahl salt-sensitive rats fed either a normal-salt (0.5% NaCl) or high-salt (4% NaCl) diet for 6 weeks. Rats fed a high-salt diet developed hypertension and significant end-organ injury. Monotherapy with HCTZ (75 mg/L in drinking water) or amlodipine (10 mg/kg/day by gavage) reduced systolic BP and proteinuria; benazepril (40 mg/kg/day by gavage) decreased proteinuria without significantly lowering systolic BP. In rats receiving a high-salt diet, only HCTZ reduced left ventricular hypertrophy, whereas endothelium-dependent relaxation was improved by amlodipine and benazepril but not by HCTZ. Combining benazepril with either amlodipine or HCTZ dramatically reduced systolic BP and end-organ injury. These data clearly support clinical studies suggesting that combination therapy is more effective than monotherapy for systolic BP control and prevention of end-organ injury. Complementary mechanisms of action of agents from different antihypertensive classes appear to facilitate the greater benefit on BP and end-organ injury.

Contribution of reactive oxygen species to the pathogenesis of left ventricular failure in Dahl salt-sensitive hypertensive rats: effects of angiotensin II blockade

We investigated the contribution of reduced nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase-dependent reactive oxygen species (ROS) generation to the pathogenesis of diastolic heart failure (DHF) in Dahl salt-sensitive (DS) hypertensive rats, with the aim of testing our hypothesis that the cardioprotective effects of angiotensin II (Ang II) blockade are provided by the suppression of this pathway. DS rats were maintained on high (H: 8.0% NaCl) or low (L: 0.3% NaCl) salt diets from age 7 to 17 weeks. DS/H rats were also treated with candesartan cilexetil (10 mg/kg per day, orally) or a superoxide dismutase mimetic, tempol (3 mmol/l in drinking water) from age 7 to 17 weeks. DS/H rats represented hypertension, left ventricular (LV) relaxation abnormality and myocardial stiffening with preserved systolic heart function. As compared with DS/L rats, DS/H rats showed higher levels of transforming growth factor-beta (TGF-beta), connective tissue growth factor (CTGF), p22phox and gp91phox mRNA expression, NADPH oxidase activity and thiobarbituric acid-reactive substance (TBARS) contents in LV tissues. Gene expression of uncoupling protein-2 (UCP-2), an inner mitochondrial membrane proton transporter, was also 2.8 +/- 0.5-fold higher. In DS/H rats, treatment with candesartan did not alter blood pressure, but resulted in a marked improvement of the hemodynamic deterioration; these therapeutic effects were accompanied by decreases in myocardial NADPH oxidase activity, TBARS contents and the expression of TGF-beta, CTGF, p22phox, gp91phox and UCP-2. Similar therapeutic effects were provided by treatment with tempol in DS/H rats. Our data suggest that NADPH oxidase-mediated ROS production contributes to the pathogenesis of DHF in DS hypertensive rats, and that the cardioprotective effects of AngII blockade are, at least partially, mediated through the suppression of this pathway.

A26. Increased LV hypertrophy and mortality in hypertensive Dahl salt sensitive rats fed high fructose diets

A26. Increased LV hypertrophy and mortality in hypertensive Dahl salt sensitive rats fed high fructose diets

Nicorandil but not ISDN Upregulates Endothelial Nitric Oxide Synthase Expression, Preventing Left Ventricular Remodeling and Degradation of Cardiac Function in Dahl Salt-sensitive Hypertensive Rats With Congestive Heart Failure

Cardiac endothelial nitric oxide synthase (ecNOS) was suppressed and inducible NOS (iNOS) enhanced at the decompensated heart failure stage in 18-week-old Dahl salt-sensitive (DS) hypertensive rats to which a high-salt diet had been administered from the age of 6 weeks. Nicorandil (NIC) enhanced ecNOS by activating Adenosine triphosphate-sensitive potassium channels (K-ATP channels) in the normal rat left ventricle. In this study, left ventricular hypertrophy, remodeling, function, cardiac ecNOS, and iNOS were compared between NIC and isosorbide dinitrate (ISDN) treatments in DS hypertensive rats with congestive heart failure. We examined DS hypertensive rats of 18 weeks of age to which 8% NaCl had been administered from the age of 6 weeks, and to which subdepressor doses of NIC (6 mg/kg/d), ISDN (6 mg/kg/d), and vehicle (CON) were administered from the age of 11 weeks. Contractility (Ees), stiffness (Eed), left ventricular end-diastolic volume, and left ventricular end-systolic volume were measured by conductance catheter and micromanometer on the basis of the pressure-volume relationship, and mRNA and protein levels of ecNOS and iNOS in the left ventricle were measured by reverse transcription-polymerase chain reaction and Western blot analysis at 18 weeks. LV mass index and LV dimensions were smaller in the NIC and ISDN groups than in the CON group (P < 0.01), and the first parameter was lower in the NIC than in the ISDN group (P < 0.01). Ees was also better maintained in the NIC and ISDN groups than in the CON group (NIC: 3349 +/- 649; ISDN: 2950 +/- 577, P < 0.05 vs. NIC; CON: 1424 +/- 375 mL/mmHg, P < 0.01 vs. treatments). Eed was exacerbated only in the ISDN group. NIC enhanced whereas ISDN suppressed ecNOS mRNA and protein levels (NIC 2.0-fold and 1.8-fold, ISDN 0.70-fold and 0.8-fold vs. CON; P < 0.01, respectively). However, no intragroup differences in iNOS mRNA or protein levels were observed for the 3 groups. More significant improvements in cardiac function and LV hypertrophy regression were observed in an NIC group than in an ISDN group of DS hypertensive rats. Activation of the K-ATP channel seems to induce this beneficial effect, which may be mediated in part by enhanced ecNOS expression in the heart in DS hypertensive congestive heart failure rat model.

Hyperhomocysteinemia Associated With Decreased Renal Transsulfuration Activity in Dahl S Rats

Elevated plasma homocysteine (Hcys) has been reported to participate in the development of arterial and glomerular sclerosis in Dahl salt-sensitive hypertensive (SS) rats. The mechanism resulting in hyperhomocysteinemia in these animals remains unknown. Disposal of Hcys in the kidneys plays an important role in regulating the plasma Hcys level. We, therefore, examined the activities and expressions of the enzymes involved in the metabolism of Hcys in the kidneys of SS rats, compared with that of Brown Norway rats and SSBN13 rats, a consomic subcolony of SS rats that carries a substituted chromosome 13 from Brown Norway rats. High-performance liquid chromatography analysis demonstrated that plasma Hcys levels were significantly higher in SS rats. The conversion of S-adenosylhomocysteine into Hcys via S-adenosylhomocysteine hydrolase by renal tissue was not different among these 3 rat strains. However, the metabolic rate of Hcys into cysteine was markedly reduced in the SS rat kidneys. The mRNA and protein levels of cystathionine beta-synthase (CBS), one of the key enzymes in the transsulfuration pathway in the kidneys, were significantly lower in SS rats. In microdissected nephron segments, CBS mRNA was shown to be mainly present in renal proximal tubules (PTs). The mRNA levels of CBS in the PTs were also significantly decreased in SS rats, accompanied by a reduced CBS activity in PTs. We conclude that hyperhomocysteinemia is associated with a decreased activity and expression of CBS in renal PTs because of the defect of chromosome 13 in SS rats.

Angiotensin Type 1A Receptors on Glial Cells in Rostral Ventrolateral Medulla and Hypertension

During the 1970s and early 1980s, all the components of the renin–angiotensin system (RAS) were identified within the brain.1 Subsequent physiological studies have shown that angiotensin II (Ang II) can act at various sites within the brain stem and hypothalamus to regulate cardiovascular function and fluid and electrolyte balance. Further, there is increasing evidence that brain Ang II can contribute to increased sympathetic activity in hypertension and heart failure.2 In recent years, a number of investigators have studied the physiological effects of alteration of the expression of genes encoding various components of the RAS. As reviewed recently by Davisson,2 early studies by Ganten and colleagues showed that transgenic rats containing the mouse renin ( Ren -2) gene were hypertensive. Although there is an increased expression of Ang II in the hypothalamus and medulla oblongata of such animals, the extent to which the hypertension was caused by increased levels of central or peripheral Ang II was not clear. Later, with the development of new methods that enabled brain-selective expression of particular genes, transgenic mice in which the angiotensin type 1A (AT1A) receptors were overexpressed in the brain but not in peripheral tissues were produced.3 These animals exhibit exaggerated pressor responses to intracerebroventricular administration of Ang II, but have a normal resting arterial blood pressure.3 Thus, this indicates that an increased density of AT1A receptors alone is not sufficient to produce hypertension. Other genetic models, however, have demonstrated that overactivity of the brain RAS can produce hypertension. In transgenic mice containing both the human angiotensinogen …

PPARα agonist protects against salt‐mediated increases in endogenous carbon monoxide production and blood pressure in Dahl salt‐sensitive rats

Heme oxygenase (HO) metabolizes heme to form carbon monoxide (CO). Increased heme-derived CO inhibits nitric oxide synthase and promotes hypertension in Dahl salt-sensitive rats (DS). Peroxisome proliferator-activated receptor α (PPARα) agonists confer cardiovascular protection during hypertension and lower blood pressure in DS. The current study tests the hypothesis that the PPARα agonist fenofibrate prevents the high salt (HS)-diet induced increase in endogenous CO production and blood pressure in DS. Male DS were placed on low (0.3% NaCl, LS) or HS (8% NaCl) diets and some received 90mg/kg/day fenofibrate in the drinking water. Respiratory CO excretion and blood pressure were measured weekly via solid-phase gas chromatography and tail-cuff plethysmography. CO excretion (HS: Δ +0.57±0.07 μmol/hr per kg) and systolic blood pressure (HS: 132±4 to 251±5 mmHg) increased markedly in HS rats. Fenofibrate treatment attenuated the HS diet induced increase in CO excretion (Δ+0.07±0.01 μmol/hr per kg) and blood pressure (145±4 to 172±8 mmHg). These data show that a PPARα agonist protects from HS diet-induced increases in CO production and blood pressure in DS. These results suggest that PPARα agonists might exert their blood pressure lowering effect by “normalizing” endogenous CO production in DS. Supported by NIH/NCRR grant P20 RR017659 (FKJ), NIH/NHLBI RO1 grants HL76187 (RAJ), HL59976 (WD) and HL74966 (WD).

Polymorphisms in 5′UTR of ENaC α, β and γgenes in Dahl and Wistar rats

The activity of the epithelial sodium channel (ENaC) in the kidney is elevated in salt sensitive (SS) versus salt resistant (SR) Dahl rats. Blockade of ENaC in the brain by benzamil prevents the increase in BP in SS Dahl rats on high salt diet. Previously, we did a comprehensive screening of the complete coding regions of ENaC α, β and γ genes and no differences were identified in SS versus SR Dahl rats. In the presentstudy, we assessed whether the promotor regions of ENaC genes might harbor genetic variation(s) that enhance ENaC expression and contribute to salt sensitivity in Dahl SS, but not SR rats. Methods Screening 5′UTR of ENaC genes in Dahl SS, SR and control Wistar rats by Denaturing High Performance Liquid Chromatography (DHPLC) and automatic sequencing. Results Screening 2.2, 1.5, and 2.2 kb of 5′UTR of ENaCα, β and γ respectively showed no differences between Dahl SS and SR rats. A total of 18 SNPs; 4 SNPs in ENaCα 5′UTR, 2 in ENaCβ5′UTR, and 12 in ENaCγ 5′UTR were identified in Dahl SS and SR rats. 4 of these 18 SNPs were previously identified in the genome database of Brown Norway rats. The remaining 14 SNPs are novel polymorphisms. Of these 14 SNPs, 13 were also identified in Wistar rats. None of the identified SNPs in the 5′UTR of ENaC genes were located within the previously reported regulatory element DNA consensus sequences. Conclusions The first comprehensive screening of ENaC genes 5′UTR did not reveal any differences between SS and SR. Therefore, we could not link differences in activity of ENaC in SS versus SR to differences in ENaC 5′UTR sequences. The identified variations in ENaC genes 5′UTR on their own can not explain salt-sensitive hypertension in Dahl SS rats.