Involvement of sex hormones, oxidative stress, ACE and ACE2 activity in the impairment of renal function and remodelling in SHR.

AIMS: Hypertension is a relevant sex and sex hormones-dependent risk factor where the cardiovascular and renal health of the population are concerned. Men experience greater losses of renal function (RF) than women, but the mechanisms remain somewhat unclear. Our goal was to evaluate the relationship between oxidative stress (OS), angiotensin-converting enzyme (ACE) and angiotensin-converting enzyme 2 (ACE2) activities and RF in male and female SHR. MAIN METHODS: Twelve-week-old spontaneously hypertensive rats (SHR) were submitted to either castration or SHAM surgery and divided into 4 groups, SHAM or Castrated (CAST) males or females. After 51 days we evaluated RF (inulin and sodium para-aminohippurate), ACE and ACE2 activities (fluorimetry), OS (flow cytometry), collagen deposition (picrosirius red) and protein expression (western blot). KEY FINDINGS: Males presented lower RF than females and castration impaired this parameter in both groups. Sexual dimorphism was not observed regarding OS and inflammation; however, castration increased this parameter more severely in males than in females. SHAM males exhibited higher collagen deposition than females, though castration increased it in both sexes, eliminating the difference. We found sexual dimorphism regarding renal ACE and ACE2 activities, which were lower in males than in females. Although castration did not alter ACE activity, it reduced ACE2 activity in females and increased it in males. SIGNIFICANCE: These results indicate that sex hormones affect RF in SHR. As alterations in the oxidative system were capable of promoting podocyte injury, inflammation, and collagen deposition, we put forward that these effects are differently modulated by ACE and ACE2.

High-intensity intermittent training is as effective as moderate continuous training, and not deleterious, in cardiomyocyte remodeling of hypertensive rats.

Exercise training offers possible nonpharmacological therapy for cardiovascular diseases including hypertension. High-intensity intermittent exercise (HIIE) training has been shown to have as much or even more beneficial cardiovascular effect in patients with cardiovascular diseases than moderate-intensity continuous exercise (CMIE) training. The aim of this study was to investigate the effects of the two types of training on cardiac remodeling of spontaneously hypertensive rats (SHR) induced by hypertension. Eight-week-old male SHR and normotensive Wistar-Kyoto rats (WKY) were divided into four groups: normotensive and hypertensive control (WKY and SHR-C) and hypertensive trained with CMIE (SHR-T CMIE) or HIIE (SHR-T HIIE). After 8 wk of training or inactivity, maximal running speed (MRS), arterial pressure, and heart weight were all assessed. CMIE or HIIE protocols not only increased final MRS and left ventricular weight/body weight ratio but also reduced mean arterial pressure compared with sedentary group. Then, left ventricular tissue was enzymatically dissociated, and isolated cardiomyocytes were used to highlight the changes induced by physical activity at morphological, mechanical, and molecular levels. Both types of training induced restoration of transverse tubule regularity, decrease in spark site density, and reduction in half-relaxation time of calcium transients. HIIE training, in particular, decreased spark amplitude and width, and increased cardiomyocyte contractility and the expression of sarco(endo)plasmic reticulum Ca2+-ATPase and phospholamban phosphorylated on serine 16. NEW & NOTEWORTHY High-intensity intermittent exercise training induces beneficial remodeling of the left ventricular cardiomyocytes of spontaneously hypertensive rats at the morphological, mechanical, and molecular levels. Results also confirm, at the cellular level, that this type of training, as it appears not to be deleterious, could be applied in rehabilitation of hypertensive patients.

Anti-Renal Fibrotic Effect of Exercise Training in Hypertension.

The purpose of this study was to evaluate the effects of exercise training on renal fibrosis in hypertensive rats. Masson's trichrome staining and Western blotting were performed on the excised renal cortex from sixteen male spontaneously hypertensive rats (SHR), which were randomly divided into either a sedentary hypertensive group (SHR) or exercise hypertensive group (SHR-EX, running on an exercise treadmill for 60 min/day, 5 sessions/week, for 12 weeks), and from eight male Wistar-Kyoto rats which served as a sedentary normotensive group (WKY). The systolic blood pressure (SBP) and renal fibrosis in hypertensive rats improved after exercise training. The inflammatory-related protein levels of interleukin-6 (IL-6) and cyclooxygenase-2 (COX-2), as well as the fibrotic-related protein levels of transforming growth factor-beta (TGF-ß), phospho-Smad2/3 (p-Smad2/3), connective tissue growth factor (CTGF), matrix metalloproteinase-9 (MMP-9), and matrix metalloproteinase-2 (MMP-2) were decreased in the SHR-EX group when compared with the SHR group. Exercise training suppressed the hypertension-induced renal cortical inflammatory and fibrotic pathways in hypertensive rat models. These findings might indicate a new therapeutic effect for exercise training to prevent renal fibrosis in hypertensive nephropathy.

Dronedarone produces early regression of myocardial remodelling in structural heart disease.

BACKGROUND AND AIMS: Left ventricular hypertrophy (LVH) in hypertension is associated with a greater risk of sustained supraventricular/atrial arrhythmias. Dronedarone is an antiarrhythmic agent that was recently approved for the treatment of atrial fibrillation. However, its effect on early regression of LVH has not been reported. We tested the hypothesis that short-term administration of dronedarone induces early regression of LVH in spontaneously hypertensive rats (SHRs). METHODS: Ten-month-old male SHRs were randomly assigned to an intervention group (SHR-D), where animals received dronedarone treatment (100 mg/kg) for a period of 14 days, or to a control group (SHR) where rats were given vehicle. A third group with normotensive control rats (WKY) was also added. At the end of the treatment with dronedarone we studied the cardiac anatomy and function in all the rats using transthoracic echocardiogram, cardiac metabolism using the PET/CT study (2-deoxy-2[18F]fluoro-D-glucose) and cardiac structure by histological analysis of myocyte size and collagen content. RESULTS: The hypertensive vehicle treated SHR rats developed the classic cardiac pattern of hypertensive cardiomyopathy as expected for the experimental model, with increases in left ventricular wall thickness, a metabolic shift towards an increase in glucose use and increases in myocyte and collagen content. However, the SHR-D rats showed statistically significant lower values in comparison to SHR group for septal wall thickness, posterior wall thickness, ventricular mass, glucose myocardial uptake, size of left ventricular cardiomyocytes and collagen content. All these values obtained in SHR-D rats were similar to the values measured in the normotensive WKY control group. CONCLUSION: The results suggest by three alternative and complementary ways (analysis of anatomy and cardiac function, metabolism and histological structure) that dronedarone has the potential to reverse the LVH induced by arterial hypertension in the SHR model of compensated ventricular hypertrophy.

Cerebral Venous Collagen Remodeling in a Modified White Matter Lesions Animal Model.

To mimic the expected pathological changes of white matter lesions (WMLs) and increase the stability, we applied modified two-vessel occlusion (modified 2VO) (1-week interval bilateral carotid artery occlusion) in stroke-prone renovascular hypertensive rats (RHRSP) and established a modified WMLs model (RHRSP/modified 2VO) that compared their phenotypes with RHRSP and sham-operated rats. In addition, we tried to differentiate small veins from small arteries through the presence of smooth muscle to study the pathological changes of small veins detailed in the model. RHRSP/modified 2VO rats showed higher stability and more extensive white matter damage without an obvious increase in mortality rate at 12 weeks after the modified 2VO operation compared to RHRSP rats. RHRSP/modified 2VO rats showed more severe small venous collagen deposition than RHRSP rats, and the majority of the deposition was collagen I and IV rather than collagen III. In addition, RHRSP/modified 2VO rats possessed cognitive impairment, mild wall thickness and blood-brain barrier disruption. Our findings suggest that the modified WMLs model (RHRSP/modified 2VO) mimics cognitive impairment and small vessel pathological changes similar to WMLs in humans. Differentiating small veins from small arteries through smooth muscle is feasible, and marked small venous deposition may play an important role in the hypertensive white matter lesions.

Contribution of independent and pleiotropic genetic effects in the metabolic syndrome in a hypertensive rat.

Hypertension is a major risk factor for cardiovascular disease, Type 2 diabetes, and end organ failure, and is often found concomitant with disorders characteristic of the Metabolic Syndrome (MetS), including obesity, dyslipidemia, and insulin resistance. While the associated features often occur together, the pathway(s) or mechanism(s) linking hypertension in MetS are not well understood. Previous work determined that genetic variation on rat chromosome 17 (RNO17) contributes to several MetS-defining traits (including hypertension, obesity, and dyslipidemia) in the Lyon Hypertensive (LH) rat, a genetically determined MetS model. We hypothesized that at least some of the traits on RNO17 are controlled by a single gene with pleiotropic effects. To address this hypothesis, consomic and congenic strains were developed, whereby a defined fragment of RNO17 from the LH rat was substituted with the control Lyon Normotensive (LN) rat, and MetS phenotypes were measured in the resultant progeny. Compared to LH rats, LH-17LN consomic rats have significantly reduced body weight, blood pressure, and lipid profiles. A congenic strain (LH-17LNc), with a substituted fragment at the distal end of RNO17 (17q12.3; 74-97 Mb; rn4 assembly), showed differences from the LH rat in blood pressure and serum total cholesterol and triglycerides. Interestingly, there was no difference in body weight between the LH-17LNc and the parental LH rat. These data indicate that blood pressure and serum lipids are regulated by a gene(s) in the distal congenic interval, and could be due to pleiotropy. The data also indicate that body weight is not determined by the same gene(s) at this locus. Interestingly, only two small haplotypes spanning a total of approximately 0.5 Mb differ between the LH and LN genomes in the congenic interval. Genes in these haplotypes are strong candidate genes for causing dyslipidemia in the LH rat. Overall, MetS, even in a simplified genetic model such as the LH-17LN rat, is likely due to both independent and pleiotropic gene effects.

Nonlinearities of heart rate variability in animal models of impaired cardiac control: contribution of different time scales.

Heart rate variability (HRV) has been extensively explored by traditional linear approaches (e.g., spectral analysis); however, several studies have pointed to the presence of nonlinear features in HRV, suggesting that linear tools might fail to account for the complexity of the HRV dynamics. Even though the prevalent notion is that HRV is nonlinear, the actual presence of nonlinear features is rarely verified. In this study, the presence of nonlinear dynamics was checked as a function of time scales in three experimental models of rats with different impairment of the cardiac control: namely, rats with heart failure (HF), spontaneously hypertensive rats (SHRs), and sinoaortic denervated (SAD) rats. Multiscale entropy (MSE) and refined MSE (RMSE) were chosen as the discriminating statistic for the surrogate test utilized to detect nonlinearity. Nonlinear dynamics is less present in HF animals at both short and long time scales compared with controls. A similar finding was found in SHR only at short time scales. SAD increased the presence of nonlinear dynamics exclusively at short time scales. Those findings suggest that a working baroreflex contributes to linearize HRV and to reduce the likelihood to observe nonlinear components of the cardiac control at short time scales. In addition, an increased sympathetic modulation seems to be a source of nonlinear dynamics at long time scales. Testing nonlinear dynamics as a function of the time scales can provide a characterization of the cardiac control complementary to more traditional markers in time, frequency, and information domains.NEW & NOTEWORTHY Although heart rate variability (HRV) dynamics is widely assumed to be nonlinear, nonlinearity tests are rarely used to check this hypothesis. By adopting multiscale entropy (MSE) and refined MSE (RMSE) as the discriminating statistic for the nonlinearity test, we show that nonlinear dynamics varies with time scale and the type of cardiac dysfunction. Moreover, as complexity metrics and nonlinearities provide complementary information, we strongly recommend using the test for nonlinearity as an additional index to characterize HRV.

Altered contractile responses of arteries from spontaneously hypertensive rat: The role of endogenous mediators and membrane depolarization.

AIMS: The goal of our study was to reveal the important mechanism(s) responsible for the enhanced contractility of isolated arteries from animals suffering genetic hypertension. MAIN METHODS: Contractile force of endothelium-denuded arteries, modulated by various interventions, was measured by wire myography. KEY FINDINGS: Spontaneously hypertensive rat (SHR) and Wistar-Kyoto rat (WKY) arteries were stimulated by norepinephrine, increased extracellular K+ or tyramine. Strain difference was not observed in the contraction elicited by exogenous norepinephrine but SHR arteries responded more to tyramine (causing endogenous norepinephrine release from neuronal varicosities). K+-induced contraction was enhanced in SHR arteries, with no involvement of endogenous catecholamines. The α-adrenoceptor blockade lowered tyramine-induced contraction more in SHR arteries; similar effect was achieved by guanethidine-induced sympathectomy. Partial depolarization of WKY arteries by 20mM K+ enhanced its contraction to SHR level. The blockade of ß-adrenoceptors by propranolol or selective ß2-antagonist ICI-118,551 induced contraction of SHR endothelium-denuded arteries but was without significant effects on WKY arteries unless they were stimulated with K+. Both tyramine-induced and propranolol-induced contractions were attenuated by flupirtine and abolished by nifedipine. SIGNIFICANCE: The differences of SHR and WKY arteries were not related to vascular expression of α- and ß-adrenoceptors or G-proteins. Enhanced contractility of SHR arteries is related to both increased presence of endogenous norepinephrine in vascular wall and also to altered vascular smooth muscle membrane potential.

The effect of multivitamin-multimineral supplementation on the health status of inbred Wistar and spontaneously hypertensive rat strains.

The nutraceutical industry has proliferated in recent years, with the most popular form of supplementation being the multivitamin-multimineral (MVMM) supplement. In the animal health sector, supplement use has also expanded. The objective of this study was to determine the effects of MVMM supplementation, beneficial or otherwise, on the general health status of the spontaneously hypertensive rat (SHR) strain, an animal model used in hypertension research. A commercially prepared MVMM supplement was given tri-weekly via oral dosing for 8 weeks to two groups of seven adult female SHR and Wistar rats. Their corresponding control groups were dosed with deionised water only. Systolic and diastolic blood pressure, fasting blood glucose, growth rate and food and water intake were measured weekly. At the end of 8 weeks, the animals were euthanased and a full blood profile, urine sodium to potassium ratio, blood urea nitrogen levels and total plasma cholesterol was measured for all groups. The results indicated that growth rate was higher for the SHR supplemented group. Supplementation also decreased diastolic blood pressure in both Wistar and SHR groups and increased red blood cell count and decreased total cholesterol in the SHR group. No adverse effects on the general health status of the animals were observed. MVMM supplementation may therefore be useful in aiding growth and delaying the onset of hypertension and its effects. It may also assist in the longevity of the breeding stock of SHR rats.

Isolation of a Genomic Region Affecting Most Components of Metabolic Syndrome in a Chromosome-16 Congenic Rat Model.

Metabolic syndrome is a highly prevalent human disease with substantial genomic and environmental components. Previous studies indicate the presence of significant genetic determinants of several features of metabolic syndrome on rat chromosome 16 (RNO16) and the syntenic regions of human genome. We derived the SHR.BN16 congenic strain by introgression of a limited RNO16 region from the Brown Norway congenic strain (BN-Lx) into the genomic background of the spontaneously hypertensive rat (SHR) strain. We compared the morphometric, metabolic, and hemodynamic profiles of adult male SHR and SHR.BN16 rats. We also compared in silico the DNA sequences for the differential segment in the BN-Lx and SHR parental strains. SHR.BN16 congenic rats had significantly lower weight, decreased concentrations of total triglycerides and cholesterol, and improved glucose tolerance compared with SHR rats. The concentrations of insulin, free fatty acids, and adiponectin were comparable between the two strains. SHR.BN16 rats had significantly lower systolic (18-28 mmHg difference) and diastolic (10-15 mmHg difference) blood pressure throughout the experiment (repeated-measures ANOVA, P < 0.001). The differential segment spans approximately 22 Mb of the telomeric part of the short arm of RNO16. The in silico analyses revealed over 1200 DNA variants between the BN-Lx and SHR genomes in the SHR.BN16 differential segment, 44 of which lead to missense mutations, and only eight of which (in Asb14, Il17rd, Itih1, Syt15, Ercc6, RGD1564958, Tmem161a, and Gatad2a genes) are predicted to be damaging to the protein product. Furthermore, a number of genes within the RNO16 differential segment associated with metabolic syndrome components in human studies showed polymorphisms between SHR and BN-Lx (including Lpl, Nrg3, Pbx4, Cilp2, and Stab1). Our novel congenic rat model demonstrates that a limited genomic region on RNO16 in the SHR significantly affects many of the features of metabolic syndrome.

Blood pressure, sex, and female sex hormones influence renal inner medullary nitric oxide synthase activity and expression in spontaneously hypertensive rats.

BACKGROUND: We previously reported that sexually mature female spontaneously hypertensive rats (SHRs) have greater nitric oxide (NO) synthase (NOS) enzymatic activity in the renal inner medulla (IM), compared to age-matched males. However, the mechanisms responsible for this sexual dimorphism are unknown. The current study tested the hypothesis that sex differences in renal IM NOS activity and NOS1 expression in adult SHRs develop with sexual maturation and increases in blood pressure (BP) in a female sex hormone-dependent manner. METHODS AND RESULTS: Renal IM were isolated from sexually immature 5-week-old and sexually mature 13-week-old male and female SHRs. Whereas NOS activity and NOS1 expression were comparable in 5- and 13-week-old male SHRs and 5-week-old female SHRs, 13-week-old females had greater NOS activity and NOS1 expression, compared to 5-week-old female SHRs and age-matched males. NOS3 expression was greater in 5-week-old than 13-week-old SHRs regardless of sex. Treatment with antihypertensive therapy (hydrochlorothiazide and reserpine) from 6 to 12 weeks of age to attenuate age-related increases in BP abolished the sex difference in NOS activity and NOS1 expression between sexually mature SHR males and females. To assess the role of female sex hormones in age-related increases in NOS, additional females were ovariectomized (OVX), and NOS activity was studied 8 weeks post-OVX. OVX decreased NOS activity and NOS1 expression. CONCLUSIONS: The sex difference in renal IM NOS in SHR is mediated by a sex hormone- and BP-dependent increase in NOS1 expression and NOS activity exclusively in females.

Cardiac hypertrophy reduction in SHR by specific silencing of myocardial Na(+)/H(+) exchanger.

We examined the effect of specific and local silencing of sodium/hydrogen exchanger isoform 1 (NHE1) with a small hairpin RNA delivered by lentivirus (L-shNHE1) in the cardiac left ventricle (LV) wall of spontaneously hypertensive rats, to reduce cardiac hypertrophy. Thirty days after the lentivirus was injected, NHE1 protein expression was reduced 53.3 ± 3% in the LV of the L-shNHE1 compared with the control group injected with L-shSCR (NHE1 scrambled sequence), without affecting its expression in other organs, such as liver and lung. Hypertrophic parameters as LV weight-to-body weight and LV weight-to-tibia length ratio were significantly reduced in animals injected with L-shNHE1 (2.32 ± 0.5 and 19.30 ± 0.42 mg/mm, respectively) compared with L-shSCR-injected rats (2.68 ± 0.06 and 21.53 ± 0.64 mg/mm, respectively). Histochemical analysis demonstrated a reduction of cardiomyocytes cross-sectional area in animals treated with L-shNHE1 compared with L-shSCR (309,81 ± 20,86 vs. 424,52 ± 21 µm(2), P < 0.05). Echocardiography at the beginning and at the end of the treatment showed that shNHE1 expression for 30 days induced 9% reduction of LV mass. Also, animals treated with L-shNHE1 exhibited a reduced LV wall thickness without changing LV diastolic dimension and arterial pressure, indicating an increased parietal stress. In addition, midwall shortening was not modified, despite the increased wall tension, suggesting an improvement of cardiac function. Chronic shNHE1 expression in the heart emerges as a possible methodology to reduce pathological cardiac hypertrophy, avoiding potentially undesired effects caused from a body-wide inhibition of NHE1.

Comparison of the validity of the use of the spontaneously hypertensive rat as a model of attention deficit hyperactivity disorder in males and females.

The spontaneously hypertensive rat (SHR) is a commonly used and well-studied rodent model of attention deficit hyperactivity disorder (ADHD). Sex differences in the cognitive symptoms of ADHD are reported. However, the female SHR rat is much less studied than its male counterpart. The goal of the current study was to assess the validity of the SHR rodent model of ADHD by examining attentional performance, inhibitory control, and hyperactivity in both male and female SHR rats. Adult SHR and control Wistar-Kyoto rats were trained on the 5-choice serial reaction time task, a self-paced test of attention and inhibitory control. This task requires animals to identify the location of a brief light stimulus among five possible locations under several challenging conditions. Analyses of percent correct revealed that attentional performance in SHR females was not significantly different from control females, whereas attentional performance in SHR males was significantly different from control males. Analyses of the number of premature responses revealed that SHR rats made more inhibitory control errors than did control rats and that this decrease in inhibitory control was present in both SHR males and females. Analyses of activity in the open field revealed that SHR rats were more hyperactive than were control rats and that this increased hyperactivity was present in both SHR males and females. The current findings have implications for the study of sex differences in ADHD and for the use of SHR rats as a model of ADHD in females.

Heterogeneity in arterial remodeling among sublines of spontaneously hypertensive rats.

OBJECTIVES: Spontaneously hypertensive rats (SHR) have been used frequently as a model for human essential hypertension. However, both the SHR and its normotensive control, the Wistar Kyoto rat (WKY), consist of genetically different sublines. We tested the hypothesis that the pathophysiology of vascular remodeling in hypertension differs among rat sublines. METHODS AND RESULTS: We studied mesenteric resistance arteries of WKY and SHR from three different sources, at 6 weeks and 5 months of age. Sublines of WKY and SHR showed differences in blood pressure, body weight, vascular remodeling, endothelial function, and vessel ultrastructure. Common features in small mesenteric arteries from SHR were an increase in wall thickness, wall-to-lumen ratio, and internal elastic lamina thickness. CONCLUSIONS: Endothelial dysfunction, vascular stiffening, and inward remodeling of small mesenteric arteries are not common features of hypertension, but are subline-dependent. Differences in genetic background associate with different types of vascular remodeling in hypertensive rats.

Cytochrome P450 1B1 contributes to increased blood pressure and cardiovascular and renal dysfunction in spontaneously hypertensive rats.

PURPOSE: We investigated the contribution of cytochrome P450 (CYP) 1B1 to hypertension and its pathogenesis by examining the effect of its selective inhibitor, 2,4,3',5'-tetramethoxystilbene (TMS), in spontaneously hypertensive rats (SHR). METHODS: Blood pressure (BP) was measured bi-weekly. Starting at 8 weeks, TMS (600 µg/kg, i.p.) or its vehicle was injected daily. At 14 weeks, samples were collected for measurement. RESULTS: TMS reversed increased BP in SHR (207 ± 7 vs. 129 ± 2 mmHg) without altering BP in Wistar-Kyoto rats. Increased CYP1B1 activity in SHR was inhibited by TMS (RLU: aorta, 5.4 ± 0.7 vs. 3.7 ± 0.7; heart, 6.0 ± 0.8 vs. 3.4 ± 0.4; kidney, 411 ± 45 vs. 246 ± 10). Increased vascular reactivity, cardiovascular hypertrophy, endothelial and renal dysfunction, cardiac and renal fibrosis in SHR were minimized by TMS. Increased production of reactive oxygen species and NADPH oxidase activity in SHR, were diminished by TMS. In SHR, TMS reduced increased plasma levels of nitrite/nitrate (46.4 ± 5.0 vs. 28.1 ± 4.1 µM), hydrogen-peroxide (36.0 ± 3.7 vs. 14.1 ± 3.8 µM), and thiobarbituric acid reactive substances (6.9 ± 1.0 vs. 3.4 ± 1.5 µM). Increased plasma levels of pro-inflammatory cytokines and catecholamines, and cardiac activity of extracellular signal-regulated kinase, p38 mitogen-activated protein kinase, c-Src tyrosine kinase, and protein kinase B in SHR were also inhibited by TMS. CONCLUSIONS: These data suggests that increased oxidative stress generated by CYP1B1 contributes to hypertension, increased cytokine production and sympathetic activity, and associated pathophysiological changes in SHR. CYP1B1 could be a novel target for developing drugs to treat hypertension and its pathogenesis.

Chronic depression of hypothalamic paraventricular neuronal activity produces sustained hypotension in hypertensive rats.

Changes in the sympathetic nervous system are responsible for the initiation, development and maintenance of hypertension. An important central sympathoexcitatory region is the paraventricular nucleus (PVN) of the hypothalamus, which may become more active in hypertensive conditions, as shown in acute studies previously. Our objective was to depress PVN neuronal activity chronically by the overexpression of an inwardly rectifying potassium channel (hKir2.1), while evaluating the consequences on blood pressure (BP) and its reflex regulation. In spontaneously hypertensive rats (SHRs) and Wistar rats (WKY) lentiviral vectors (LVV-hKir2.1; LV-TREtight-Kir-cIRES-GFP5 4 × 10(9) IU and LV-Syn-Eff-G4BS-Syn-Tetoff 6.2 × 10(9) IU in a ratio 1:4) were stereotaxically microinjected bilaterally into the PVN. Sham-treated SHRs and WKY received bilateral PVN microinjections of LVV-eGFP (LV-Syn-Eff-G4BS-Syn-Tetoff 6.2 × 10(9) IU and LV-TREtight-GFP 5.7 × 10(9) IU in a ratio 1:4). Blood pressure was monitored continuously by radio-telemetry and evaluated over 75 days. Baroreflex gain was evaluated using phenylephrine (25 µg ml(-1), i.v.), whereas lobeline (25 µg ml(-1), i.v.) was used to stimulate peripheral chemoreceptors. In SHRs but not normotensive WKY rats, LVV-hKir2.1 expression in the PVN produced time-dependent and significant decreases in systolic (from 158 ± 3 to 132 ± 6 mmHg; P < 0.05) and diastolic BP (from 135 ± 4 to 113 ± 5 mmHg; P < 0.05). The systolic BP low-frequency band was reduced (from 0.79 ± 0.13 to 0.42 ± 0.09 mmHg(2); P < 0.05), suggesting reduced sympathetic vasomotor tone. Baroreflex gain was increased and peripheral chemoreflex depressed after PVN microinjection of LVV-hKir2.1. We conclude that the PVN plays a major role in long-term control of BP and sympathetic nervous system activity in SHRs. This is associated with reductions in both peripheral chemosensitivity and respiratory-induced sympathetic modulation and an improvement in baroreflex sensitivity. Our results support the PVN as a powerful site to control BP in neurogenic hypertension.

Effects of age on expression of BKca channel in vascular smooth muscle cells from mesenteric arteries of spontaneously hypertensive rats

Abnormally functioning K+ channels during aging might contribute to increased arterial tone of vascular smooth muscle cells. However, changes in large conductance calcium-activated potassium (BKca) channel current density and channel protein composition during aging are not fully understood and are currently investigated here. Spontaneously hypertensive rats (SHR) and Wistar–Kyoto (WKY) rats were grouped by age into very young (4–6 weeks), young (16–18 weeks), and adult (40–42 weeks) rats. BKca current was measured using a whole cell patch clamp in vascular smooth muscle cells from rat mesenteric arteries. BKca channel and subunits were measured using confocal imaging and real-time reverse transcriptase polymerase chain reaction techniques. Systolic and diastolic blood pressure increased significantly with age in both SHR and WKY rats. The expression of BKca á subunits increased in plasma membrane and cytoplasm in both SHR and WKY rats during aging, but had a larger increase in the cytoplasm in SHR than in WKY rats. BKca current density increased with aging in SHR but not in WKY rats. The relative abundance of BKca áand â1 mRNA transcripts changed minimally or not at all with aging in both SHR and WKY rats. In conclusion, plasma membrane BKca protein increased with aging in both SHR and WKY rats. Increases in cytoplasmic BKca protein and in BKca current were greater in SHR than WKY rats (AU)

Entrainment of spontaneously hypertensive rat fibroblasts by temperature cycles.

The functional state of the circadian system of spontaneously hypertensive rats (SHR) differs in several characteristics from the functional state of normotensive Wistar rats. Some of these changes might be due to the compromised ability of the central pacemaker to entrain the peripheral clocks. Daily body temperature cycles represent one of the important cues responsible for the integrity of the circadian system, because these cycles are driven by the central pacemaker and are able to entrain the peripheral clocks. This study tested the hypothesis that the aberrant peripheral clock entrainment of SHR results from a compromised peripheral clock sensitivity to the daily temperature cycle resetting. Using cultured Wistar rat and SHR fibroblasts transfected with the circadian luminescence reporter Bmal1-dLuc, we demonstrated that two consecutive square-wave temperature cycles with amplitudes of 2.5 °C are necessary and sufficient to restart the dampened oscillations and entrain the circadian clocks in both Wistar rat and SHR fibroblasts. We also generated a phase response curve to temperature cycles for fibroblasts of both rat strains. Although some of the data suggested a slight resistance of SHR fibroblasts to temperature entrainment, we concluded that the overall effect it too weak to be responsible for the differences between the SHR and Wistar in vivo circadian phenotype.

Vascular reactivity of arteria femoralis in adult and aged spontaneously hypertensive and Wistar-Kyoto rats.

AIM: The relationship of age and hypertension on endothelial dysfunction and increased responses to vasoconstrictor stimuli. BACKGROUND: Hypertension is a disease accompanied by endothelial dysfunction and is characterized by an impaired vascular reactivity and enhanced activity of sympathetic nervous system. MATERIALS AND METHODS: In our experiment, we used spontaneously hypertensive rats representing model of essential hypertension and the Wistar-Kyoto rats as normotensive strain. Femoral arteries of adult and aged rats were put into the chamber of Mulvany-Halpern isometric myograph. As the nutrient solution, the modified Krebs-Henseleit solution having temperature 37 °C and bubbled with O2 was used. After 30 minutes stabilization of blood vessels, a dose-dependent curve of norepinephrine response was recorded (concentrations 3x10-8 M, 10-7 M, 3x10-7 M, 10-6 M, 3x10-6 M, 10-5 M, 3x10-5 M, 10-4 M), followed by a dose-dependent curve of acetylcholine response (concentrations 3x10-8 M, 10-7 M, 3x10-7 M, 10-6 M, 3x10-6 M). RESULTS: Our experiments recorded an increased reactivity to contraction stimuli in spontaneously hypertensive animals. Vascular reactivity to norepinephrine at 5 month and 12 month old rats from the same group was not significantly affected. Our experiments on the other hand, did not record a reduced endothelium-dependent relaxation in hypertensive compared to normotensive animals, neither in different age groups. CONCLUSIONS: Increased norepinephrine-induced contraction occurs even before development of reduced acetylcholine-induced relaxation in SHR rats. We predict that in our experiment hypertension plays a bigger role in the development of endothelial dysfunction than aging (Fig. 2, Ref. 22).