Renovascular Hypertensive Rats Research Articles

ANG II and Aldosterone Acting Centrally Participate in the Enhanced Sodium Intake in Water-Deprived Renovascular Hypertensive Rats.

Renovascular hypertension is a type of secondary hypertension caused by renal artery stenosis, leading to an increase in the renin–angiotensin–aldosterone system (RAAS). Two-kidney, 1-clip (2K1C) is a model of renovascular hypertension in which rats have an increased sodium intake induced by water deprivation (WD), a common situation found in the nature. In addition, a high-sodium diet in 2K1C rats induces glomerular lesion. Therefore, the purpose of this study was to investigate whether angiotensin II (ANG II) and/or aldosterone participates in the increased sodium intake in 2K1C rats under WD. In addition, we also verified if central AT1 and mineralocorticoid receptor blockade would change the high levels of arterial pressure in water-replete (WR) and WD 2K1C rats, because blood pressure changes can facilitate or inhibit water and sodium intake. Finally, possible central areas activated during WD or WD followed by partial rehydration (PR) in 2K1C rats were also investigated. Male Holtzman rats (150–180 g) received a silver clip around the left renal artery to induce renovascular hypertension. Six weeks after renal surgery, a stainless-steel cannula was implanted in the lateral ventricle, followed by 5–7 days of recovery before starting tests. Losartan (AT1 receptor antagonist) injected intracerebroventricularly attenuated water intake during the thirst test. Either icv losartan or RU28318 (mineralocorticoid receptor antagonist) reduced 0.3 M NaCl intake, whereas the combination of losartan and RU28318 icv totally blocked 0.3 M NaCl intake induced by WD in 2K1C rats. Losartan and RU28318 icv did not change hypertension levels of normohydrated 2K1C rats, but reduced the increase in mean arterial pressure (MAP) produced by WD. c-Fos expression increased in the lamina terminalis and in the NTS in WD condition, and increased even more after WD-PR. These results suggest the participation of ANG II and aldosterone acting centrally in the enhanced sodium intake in WD 2K1C rats, and not in the maintenance of hypertension in satiated and fluid-replete 2K1C rats.

Allisartan isoproxil reduces mortality of stroke-prone rats and protects against cerebrovascular, cardiac, and aortic damage.

Stroke is a common cause of death and disability. Allisartan isoproxil (ALL) is a new angiotensin II receptor blocker and a new antihypertensive drug discovered and developed in China. In the present study we investigated the therapeutic effects of ALL in stroke-prone renovascular hypertensive rats (RHR-SP) and the underlying mechanisms. The model rats were generated via two-kidney two-clip (2K2C) surgery, which led to 100% of hypertension, 100% of cerebrovascular damage as well as 100% of mortality 1 year after the surgery. Administration of ALL (30 mg · kg-1 · d-1 in diet, for 55 weeks) significantly decreased stroke-related death and prolonged lifespan in RHR-SP, but the survival ALL-treated RHR-SP remained of hypertension and cardiovascular hypertrophy compared with sham-operated normal controls. In addition to cardiac, and aortic protection, ALL treatment for 10 or 12 weeks significantly reduced cerebrovascular damage incidence and scoring, along with a steady reduction of blood pressure (BP) in RHR-SP. Meanwhile, it significantly decreased serum aldosterone and malondialdehyde levels and cerebral NAD(P)H oxidase expressions in RHR-SP. We conducted 24 h continuous BP recording in conscious freely moving RHR-SP, and found that a single intragastric administration of ALL produced a long hypotensive effect lasting for at least 12 h on systolic BP. Taken together, our results in RHR-SP demonstrate that ALL can be used for stroke prevention via BP reduction and organ protection, with the molecular mechanisms related to inhibition of angiotensin-aldosterone system and oxidative stress. This study also provides a valuable scoring for evaluation of cerebrovascular damage and drug efficacy.

Selective Discovery of GPCR Ligands within DNA-Encoded Chemical Libraries Derived from Natural Products: A Case Study on Antagonists of Angiotensin II Type I Receptor.

Natural products have failed to meet the urgent need for drug discovery in recent decades due to limited resources, necessitating new strategies for re-establishing the key role of natural products in hit screening. This work introduced DNA-encoding techniques into the synthesis of phenolic acid-focused libraries containing 32 000 diverse compounds. Online selection of the library using immobilized angiotensin II type I receptor (AT1R) resulted in seven phenolic acid derivatives. The half-maximal concentration (IC50) of hit 1 for the right shift of the [125I]-Sar1-AngII competition curve was 19.6 nM. Pharmacological examination of renovascular hypertensive rats demonstrated that hit 1 significantly lowered the blood pressure of the animals without changing their heart rates. These results were used to create a general strategy for rapid and unbiased discovery of hits derived from natural products with high throughput and efficiency.

Electroacupuncture at "Taichong"(LR3) improves baroreflex sensitivity and α2-adrenergic receptor activity in nucleus tractus solitarii of hypertensive rats

To investigate the effect of electroacupuncture (EA) at "Taichong"(LR3) on blood pressure, sympathetic nerve activity, baroreflex sensitivity, and α 2-adrenergic receptor (α2AR) expression in nucleus tractus solitarii (NTS) in hypertensive rats, so as to reveal its mechanisms underlying improvement of hypertension. Male Sprague Dawley rats were randomly divided into 4 groups: sham-operation, model, EA, and sham-EA (non-acupoint) groups, with 12 rats in each group. The hypertension model was established by occlusion of the right renal aorta (two-kidney-one clip method). Rats of the sham-operation group received the same surgery but without occlusion of the renal artery. EA (2 Hz/15 Hz, 2 mA) was applied to bilateral LR3 for 30 min, once a day for 28 days, and sham EA was applied to the skin of the rat tail near the buttock on both sides. The mean arterial pressure (MAP) of the abdominal aorta and heart rate (HR) were recorded. The autonomic nerve function was assessed by using frequency domain analysis of heart rate variability (HRV), and the baroreflex sensitivity detected by sequential method. Plasma norepinephrine (NE) level was measured by ELISA, and the α2AR positive neurons and α2AR protein expression in NTS were detected by using fluorescence immunohistochemistry and Western blot, respectively. The functions of α2AR within the NTS in modulating MBP and HR were verified by microinjection of its agonist (clonidine) and antagonist (yohimbine) separately. Compared to the sham operation rats, the hypertension rats displayed significant increases in the MAP (P<0.01), plasma norepinephrine content (P<0.01), ratios of low frequency/total power (LF/TP) and low frequency/high frequency (LF/HF) (P<0.01), and significant reduction in the overall gain, uplink sequence gain and downlink sequence gain of baroreflex (P<0.01), the number α2AR positive neurons and α2AR protein expression level in NTS (P<0.01). The rats in the EA group (rather than in the sham-EA group) showed significant reduction in MAP at the 3rd and 4th week, plasma NE content, LF/TP and LF/HF (P<0.01), and obvious increase in the overall gain, uplink sequence gain and downlink sequence gain of baroreflex (P<0.01), and the number of α2AR positive neurons and α2AR protein expression in comparison (P<0.05) with those of the model group. Microinjection of clonidine into NTS induced an evident decrease in both MAP and HR in the model group relevant to the sham operation group (P<0.01), while the MAP and HR changes of the EA (not sham EA) group were considerably bigger than those of the model group (P<0.05), being similar to those of the sham-operation group (P>0.05), which suggested an elimination of the BP-lowering effect of clonidine after EA. EA at LR3 can reduce MBP, sympathetic activities, improve baroreflex sensitivity in renovascular hypertensive rats, which may be associated with its effects in up-regulating the decreased NTS α2AR expression and functional activities.

Paraventricular Nucleus Infusion of Oligomeric Proantho Cyanidins Improves Renovascular Hypertension.

Oxidative stress plays an important role in the pathogenesis of hypertension. Oligomeric proantho cyanidins (OPC) is the main polyphenol presents in grape seed and is known for its potent antioxidant and anti-inflammatory properties. In the present study, we hypothesize that OPC can attenuate oxidative stress in the paraventricular nucleus of hypothalamus (PVN), ameliorate neurotransmitter imbalance, decrease the blood pressure and sympathetic activity in renovascular hypertensive rats. After induction of renovascular hypertension by the two-kidney one-clip (2K-1C) method, male Sprague-Dawley rats received chronic bilateral PVN infusion of OPC (20 μg/h) or vehicle via osmotic minipump for 4 weeks. We found that hypertension induced by 2K-1C was associated with the production of reactive oxygen species (ROS) in the PVN. Infusion of OPC in the PVN significantly reduced the systolic blood pressure and norepinephrine in plasma of 2K-1C rats. In addition, PVN infusion of OPC decreased the level of ROS and the expression of stress-related nicotinamide adenine dinucleotide phosphate (NADPH) oxidases subunit NOX4, increased the levels of nuclear factor E2-related factor 2 (Nrf2) and antioxidant enzyme, balanced the content of cytokines, increased expression of glutamic acid decarboxylase and decreased the expression of tyrosine hydroxylase in the PVN of 2K-1C rats. Our findings provided strong evidence that PVN infusion of OPC inhibited the progression of renovascular hypertension through its potent anti-oxidative and anti-inflammatory function in the PVN.

Sodium Thiosulfate Ameliorates Renovascular Hypertension-Induced Renal Dysfunction and Injury in Rats

Background/Aims: Arterial stenosis activates the renin-angiotensin-aldosterone system subsequently resulting in renovascular hypertension (RVHT) and renal oxidative injury. We explored the effect of sodium thiosulfate (STS, Na2S2O3), a developed antioxidant in clinical trial, on RVHT-induced hypertension and renal oxidative injury in rats. Methods: We induced RVHT in male Wistar rats with bilaterally partial ligation of renal arteries in the 2-kidney 2-clip model. We evaluated the STS effect on RVHT-induced oxidative injury and apoptosis by a chemiluminescence amplification method, Western blot, and immunohistochemistry. Results: We found STS displayed a dose-dependent antioxidant H2O2 activity and adapted the maximal scavenging H2O2 activity of STS at the dosage of 0.1 g/kg intraperitoneally 3 times/week for 4 weeks in RVHT rats. RVHT induced a significant elevation of arterial blood pressure, blood reactive oxygen species amount, neutrophil infiltration, 4-HNE and NADPH oxidase gp91 expression, Bax/Bcl-2/poly(ADP-ribose) polymerase (PARP)-mediated apoptosis formation, blue Masson-stained fibrosis, and urinary protein level. STS treatment significantly reduced hypertension, oxidative stress, neutrophil infiltration, fibrosis, and Bax/Bcl-2/PARP-mediated apoptosis formation and depressed the urinary protein level in the RVHT models. Conclusion: Our results suggest that STS treatment could ameliorate RVHT hypertension and renal oxidative injury through antioxidant, antifibrotic, and antiapoptotic mechanisms.

Renal Sensory Activity Regulates the γ-Aminobutyric Acidergic Inputs to the Paraventricular Nucleus of the Hypothalamus in Goldblatt Hypertension.

Renal sensory activity is centrally integrated within brain nuclei involved in the control of cardiovascular function, suggesting that renal afferents regulate basal and reflex sympathetic vasomotor activity. Evidence has shown that renal deafferentation (DAx) evokes a hypotensive and sympathoinhibitory effect in experimental models of cardiovascular diseases; however, the underlying mechanisms involved in this phenomenon need to be clarified, especially those related to central aspects. We aimed to investigate the role of renal afferents in the control of γ-aminobutyric acid (GABA)ergic inputs to the paraventricular nucleus (PVN) of the hypothalamus in renovascular hypertensive (2K1C) rats and their influence in the regulation of cardiovascular function. Hypertension was induced by clipping the left renal artery. After 4 weeks, renal DAx was performed by exposing the left renal nerve to a 33 mM capsaicin solution for 15 min. After 2 weeks of DAx, microinjection of muscimol into the PVN was performed in order to evaluate the influence of GABAergic activity in the PVN and its contribution to the control of renal sympathetic nerve activity (rSNA) and blood pressure (BP). Muscimol microinjected into the PVN triggered a higher drop in BP and rSNA in the 2K1C rats and renal DAx mitigated these responses. These results suggest that renal afferents are involved in the GABAergic changes found in the PVN of 2K1C rats. Although the functional significance of this phenomenon needs to be clarified, it is reasonable to speculate that GABAergic alterations occur to mitigate microglia activation-induced sympathoexcitation in the PVN of 2K1C rats.

The baroreflex afferent pathway plays a critical role in H2S-mediated autonomic control of blood pressure regulation under physiological and hypertensive conditions.

Hydrogen sulfide (H2S), which is closely related to various cardiovascular disorders, lowers blood pressure (BP), but whether this action is mediated via the modification of baroreflex afferent function has not been elucidated. Therefore, the current study aimed to investigate the role of the baroreflex afferent pathway in H2S-mediated autonomic control of BP regulation. The results showed that baroreflex sensitivity (BRS) was increased by acute intravenous NaHS (a H2S donor) administration to renovascular hypertensive (RVH) and control rats. Molecular expression data also showed that the expression levels of critical enzymes related to H2S were aberrantly downregulated in the nodose ganglion (NG) and nucleus tractus solitarius (NTS) in RVH rats. A clear reduction in BP by the microinjection of NaHS or L-cysteine into the NG was confirmed in both RVH and control rats, and a less dramatic effect was observed in model rats. Furthermore, the beneficial effects of NaHS administered by chronic intraperitoneal infusion on dysregulated systolic blood pressure (SBP), cardiac parameters, and BRS were verified in RVH rats. Moreover, the increase in BRS was attributed to activation and upregulation of the ATP-sensitive potassium (KATP) channels Kir6.2 and SUR1, which are functionally expressed in the NG and NTS. In summary, H2S plays a crucial role in the autonomic control of BP regulation by improving baroreflex afferent function due at least in part to increased KATP channel expression in the baroreflex afferent pathway under physiological and hypertensive conditions.

Myrtus communis improves cognitive impairment in renovascular hypertensive rats.

Myrtus communis has anti-inflammatory, neuroprotective and anticholinesterase activities yet there have been limited studies examining effects of Myrtus communis on cognitive functions. This study investigated the possible effects of Myrtus communis on changes in the cognitive functions of experimental renovascular hypertensive rats. Fifty-six Wistar-Albino rats were equally divided into 4 groups; sham-operated control, renovascular hypertension (RVH), ramipril (RVH + Ram) and Myrtus communis extract (RVH + MC) treatment groups. Goldblatt's 2-kidney 1-clip (2K1C) method was used to induce RVH. At the end of 9 weeks of treatment, after blood pressure recording, the animals underwent new object recognition test and Morris water maze (MWM) task. Following these tests, blood brain barrier (BBB) integrity was examined in 6 animals from each group. In the others after decapitation, osteopontin and interleukin (IL)-10 levels were measured in blood samples; while matrix metalloproteinase (MMP)-13, sodium potassium adenosine triphosphatase (Na+,K+-ATPase), cluster of differentiation (CD) 36, amyloid beta (Aβ), neprilysin levels, and acetylcholinesterase (AChE) activity were investigated in hippocampal tissues. In RVH group, high systolic blood pressure decreased serum IL-10 levels, increased serum osteopontin levels and also impaired BBB permeability. Hippocampal MMP-13, CD36, Aβ, neprilysin levels and AChE activities were elevated, while there were decreases in Na+,K+-ATPase levels. In new objet recognition test, discrimination index (DI) was determined as lower in saline-treated RVH group compared to control animals. In MWM training trail, 4th day performance in finding platform was significantly reduced in saline-treated RVH group compared to control group. RVH also decreased the time spent in target quadrant in probe test of MWM task compared to control group. In both of the treatment groups, all biochemical parameters were restored in parallel with improvement in the behavioral test performances. The results of this study suggest that Myrtus communis extract may improve the cognitive dysfunctions in hypertension through antihypertensive, anti-inflammatory and anticholinesterase activities.

Different reactive species modulate the hypotensive effect triggered by angiotensins at CVLM of 2K1C hypertensive rats

Hypertension is associated with increased central activity of the renin-angiotensin system (RAS) and oxidative stress. Here, we evaluated whether reactive species and neurotransmitters could contribute to the hypotensive effect induced by angiotensin (Ang) II and Ang-(1−7) at the caudal ventrolateral medulla (CVLM) in renovascular hypertensive rats (2K1C). Therefore, we investigated the effect of Ang II, Ang-(1-7), and the Ang-(1-7) antagonist A-779 microinjected before and after CVLM microinjection of the nitric oxide (NO)-synthase inhibitor, (L-NAME), vitamin C (Vit C), bicuculline, or kynurenic acid in 2K1C and SHAM rats. Baseline values of the mean arterial pressure (MAP) in 2K1C rats were higher than in SHAM rats. CVLM microinjection of Ang II, Ang-(1−7), l-NAME, or bicuculline induced decreases in the MAP in SHAM and 2K1C rats. In addition, Vit C and A-779 produced decreases in the MAP only in 2K1C rats. Kynurenic acid increased the MAP in both SHAM and 2K1C rats. Only the Ang-(1−7) effect was increased by l-NAME and reduced by bicuculline in SHAM rats. L-NAME also reduced the A-779 effect in 2K1C rats. Only the Ang II effect was abolished by CVLM Vit C and enhanced by CVLM kynurenic acid in SHAM and 2K1C rats. Overall, the superoxide anion and glutamate participated in the hypotensive effect of Ang II, while NO and GABA participated in the hypotensive effect of Ang-(1−7) in CVLM. The higher hypotensive response of A-779 in the CVLM of 2K1C rats suggests that Ang-(1−7) contributes to renovascular hypertension.

Abstract P029: Aspartate And Potassium Aspartate May Contribute To A Mechanism By Which Saccharina Japonica Extract Attenuates Hypertension In 2-kidney, 1-clip Renovascular Hypertensive Rats.

Introduction: Saccharina japonica (SJ) is one of brown algae. The extract of SJ (SJE) is often used for soup stock in Japanese cuisine. We recently observed the decreases in blood pressure (BP) by intake of SJ or SJE diet in 2-kidney, 1-clip renovascular hypertensive (2K1C) rats. SJ is rich in aspartate (Asp), which is the umami taste of amino acid. The intake of Asp was reported to reduce BP in Dahl salt sensitive rats. As well, the potassium Asp (AspK) intake was reported to improve BP in hypertensive patients. 70% (70 of 100) of amino acids such as Asp are eluted into the SJE from SJ. Therefore, we hypothesized that Asp in SJE contributes to the mechanism of BP reduced by dietary SJE in 2K1C rats. In this study, we observed BP in 2K1C rats fed a diet containing Asp or AspK and investigated whether those diets alleviate hypertension to the same extent of the diet containing SJE. Methods: Male Sprague-Dawley rats (6 wks) were treated with sham operation (SHAM) or clipping the left renal artery (2K1C). After surgery, the rats fed a control diet (C), a diet containing Asp, AspK or SJE for 6 weeks. The content of SJE was set to be the same as that in our previous study. The amount of Asp in diet was set to be the same as that in our previous study which showed the antihypertensive effect of SJ. The amount of AspK in diet was set to be equimolar to Asp. The systolic BP (SBP) was measured by a tail-cuff method every week. At the end of the procedure, mean arterial BP (MAP) was measured in each rat under anesthesia. Results: Six weeks after the surgery, 2K1C-C significantly increased in SBP compared with SHAM-C (126±2 vs 159±4 mmHg, p&lt;0.05). However, 2K1C-Asp (138±3), -AspK (142±4), and -SJE (135±4) decreased compared with 2K1C-C (p&lt;0.05) respectively. MAP was significantly higher in 2K1C-C than in SHAM-C (162±3 vs 131±5, p&lt;0.05). It showed a significant decrease in 2K1C-Asp (142±3), -AspK (148±3) and -SJE (144±4), compared with 2K1C-C (p&lt;0.05), respectively. There were no significant differences among 2K1C-Asp, -AspK and -SJE, in either SBP or MAP. Conclusions: Asp and AspK may attenuate hypertension in 2K1C renovascular hypertension as much as SJE which contains similar quantities of Asp. It is possible that Asp and/or AspK eluted into SJE contributes to the mechanism of alleviating hypertension by SJE in 2K1C.

Reversion of cardiovascular remodelling in renovascular hypertensive 2K-1C rats by renin-angiotensin system inhibitors.

Evaluate whether the RAS dual blockade would induce additional beneficial effects on cardiovascular remodelling when compared to monotherapy in renal hypertensive two kidneys-one clip (2K-1C) rats. Hypertensive 2K-1C and normotensive (2K) rats were treated for 14days with submaximal doses of losartan (LOS), enalapril (ENA), losartan plus enalapril (LOS+ENA) or vehicle (water). Blood pressure and some parameters of cardiovascular remodelling were evaluated. Systolic blood pressure (SBP) was higher in 2K-1C (209±3mmHg, P<.05) than in 2K (113±1mmHg) rats. There was an additional effect in 2K-1C treated with LOS+ENA (153±9mmHg) on lowering SBP when compared to LOS (184±12mmHg) or ENA (177±9mmHg). None of the treatments had effect on SBP in 2K rats. In 2K-1C, cardiomyocyte hypertrophy was reduced by all treatments, although the cardiac hypertrophy indexes remained unchanged. 2K-1C aortas presented medial thickening that was partially reduced by the treatments. Intimal hyperplasia observed in 2K-1C (15.56±0.89µm vs 8.24±0.80µm) was reversed by ENA (9.52±0.45µm) or LOS+ENA (8.17±0.53µm). Collagen deposition was increased in 2K-1C hearts (1.77±0.16 vs 1.28±0.09) and aortas (8.1±0.6 vs 5.2±0.2). Treatment with LOS reduced (1.12±0.14%) and ENA (0.81±0.11%) or LOS+ENA (0.86±0.11%) additionally diminished collagen only in 2K-1C hearts. Submaximal doses of ACEi and/or ARB have inhibitory actions on cardiac remodelling and vascular hypertrophy not entirely dependent on their effects on blood pressure normalization in renovascular hypertensive rats. Combined therapy produced additional reduction in blood pressure than monotherapy despite a similar inhibition on cardiovascular remodelling.

Rosuvastatin Improves Cognitive Function of Chronic Hypertensive Rats by Attenuating White Matter Lesions and Beta-Amyloid Deposits.

Hypertensive white matter lesion (WML) is one of common causes of vascular cognitive impairment. In this study, we aimed to investigate the effect of rosuvastatin on cognitive impairment and its underlying mechanisms in chronic hypertensive rats. From the 8th week after establishment of stroke-prone renovascular hypertensive rats (RHRSPs), rosuvastatin (10 mg/kg) or saline as a control was administrated once daily for consecutive 12 weeks by gastric gavage. Cognitive function was assessed with the Morris water maze test and novel object recognition test. WML was observed by Luxol fast blue staining. Aβ deposits, Claudin-5, Occludin, and ZO-1 were determined by immunofluorescence. After rosuvastatin treatment, the escape latencies were decreased and the time of crossing the hidden platform was increased in the Morris water maze, compared with the vehicle-treated RHRSP group. In a novel object recognition test, the recognition index in the rosuvastatin-treated RHRSP group was significantly larger than that in the vehicle-treated RHRSP group. Rosuvastatin treatment presented with the effects of lower WML grades, higher expression of tight junction proteins Claudin-5, Occludin, and ZO-1 in the corpus callosum, and less Aβ deposits in the cortex and hippocampus. The data suggested that rosuvastatin improved the cognitive function of chronic hypertensive rats partly by attenuating WML and reducing Aβ burden.

Autophagy inhibition by chloroquine prevents increase in blood pressure and preserves endothelial functions

Purpose: To determine the effects of lysosomal inhibition of autophagy by chloroquine (CHQ) onhypertension-associated changes in the endothelial functions.&#x0D; Method: Angiotensin II (Ang II)-treated human endothelial cell line EA.hy926 and renovascularhypertensive rats were subjected to CHQ treatment (in vitro: 0.5, 1, and 2.5 μM; in vivo: 50 mg/kg/dayfor three weeks). Changes in the protein expressions of LC3b II (autophagosome formation marker) andp62 (autophagy flux marker) were assessed using immunoblotting. Cell migration assay, tubuleformation assay (in vitro), and organ bath studies (in vivo) were performed to evaluate the endothelialfunctions. Hemodynamic parameters were measured as well.&#x0D; Results: A higher expression of LC3b II and a reduced expression of p62 observed in the Ang II-treatedendothelial cells, as well as in the aorta of the hypertensive rats, indicated enhanced autophagy.Treatment with CHQ resulted in reduced autophagy flux (in vitro as well as in vivo) and suppressed AngII-induced endothelial cell migration and angiogenesis (in vitro). The treatment with CHQ was alsoobserved to prevent increase in blood pressure in hypertensive rats and preserved acetylcholineinducedrelaxation in phenylephrine-contracted aorta from the hypertensive rats. In addition, chloroquineattenuated Ang II-induced contractions in the aorta of normotensive as well as hypertensive rats.&#x0D; Conclusion: These observations indicated that CHQ lowers the blood pressure and preserves thevascular endothelial function during hypertension.&#x0D; Keywords: Angiotensin II, Autophagy, Chloroquine, Endothelial function, Hypertension, Vasculardysfunction

Intermittent Theta Burst Stimulation Improves the Spatial Cognitive Function of Rats with Chronic Hypertension-induced Cerebral Small Vessel Disease

We investigated whether intermittent theta burst stimulation (iTBS) can improve the spatial cognitive function of rats with hypertension-induced cerebral small vessel disease. To prove our hypothesis, stroke-prone renovascular hypertensive rats (RHRSPs) were treated with iTBS beginning at postoperative week 22. The Morris water maze was performed to assess spatial cognitive function. The expression of the N-methyl-d-aspartate receptor (NMDAR) subunits NR1, NR2A and NR2B, calcium/calmodulin-dependent protein kinase IIα (CaMKIIα), p-CaMKIIα and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor subunit 1 (GluR1) in the hippocampus were evaluated by western blot analysis. The distribution of GluR1, glial fibrillary acidic protein (GFAP) and ionized calcium-binding adaptor molecule-1 (IBa-1) in the CA1 and CA3 regions and dentate gyrus (DG) of the hippocampus were evaluated by immunofluorescence analysis. Treatment with iTBS significantly improved the spatial cognitive function of RHRSPs, increased the expression of NR2B, p-CaMKIIα and GluR1 in the hippocampus, and decreased the proliferation of astrocytes and microglia. Our results showed that iTBS treatment had a beneficial effect on the cognitive impairments induced by cerebral small vessel disease, potentially through the activation of the NR2B-CaMKII pathway, an increase in GluR1 expression and the suppression of astrocyte and microglial activation.

ROLE OF SPINAL AT1, IONOTROPIC GLUTAMATERGIC AND GABAERGIC RECEPTORS ON CONTROL OF RENAL SYMPATHETIC NERVE ACTIVITY IN 2K‐1C RATS

Previous studies performed in our laboratory showed a higher glutamate and angiotensin receptors activation, as well as an alteration in GABAergic pathways in central nuclei involved in the cardiovascular control that may lead to sympathoexcitation in renovascular hypertensive rats (2K‐1C). However, the role of the spinal cord neurons on the renal sympathoexcitation remains unclear in this model. Thus, we aimed to assess the influence of spinal glutamatergic and GABAergic (ionotropic receptors), as well as angiotensin II type 1 receptors (AT1) receptors on the renal sympathetic nerve activity (rSNA) in the 2K‐1C rats. Male Wistar rats (250–300 gr) were used to induce renovascular hypertension by clipping the renal artery with a silver clip (0.2 mm). After six weeks, a polyethylene catheter (PE‐10) was inserted into the subarachnoid space and advanced to the T10–11 vertebrae level in urethane‐anaesthetized rats (1.2 g/Kg, iv). All experiments were approved by the Institutional Ethical Committee of UNIFESP. The effects of intrathecal (i.t.) injection of kynurenic acid (KYN) (160 nmol in 2 μl), losartan (Los) (2nmol in 2 μl) or bicuculline (Bic) (8 mM in 2 μl) on blood pressure (BP) and rSNA were investigated. KYN i.t. injection induced a larger and significant fall on rSNA in the 2K‐1C compared to control rats. Intrathecally Los evoked a larger and significant fall in rSNA in the 2K‐1C compared to control group. KYN decreased BP similarly in both, control and 2K‐1C groups, however, Los significantly decreased BP in the 2K‐1C group only. Intrathecally Bic triggered a more intense BP and rSNA reponses in 2K‐1C compared to control rats. Gene expression analysis by qRT‐PCR showed a spinal (T11–12) AT1 mRNA upregulation in the 2K‐1C rats (1.71‐fold change), while no changes were observed in AT2, ionotropic glutamate and GABA receptors subunits. Therefore, our data show that ionotropic spinal cord excitatory amino acid, GABAergic and AT1 receptors play a significant role in the control of rSNA in the 2K‐1C model that may contribute to the maintenance of hypertension. The participation of spinal AT1 receptors seems to be more important in the establishment of sympathoexcitation in this model. The origin of those projections remains unclear.Support or Funding InformationCNPq, FAPESP (2018/01898‐4) and CAPES

Systemic administration of pentoxifylline attenuates the development of hypertension in renovascular hypertensive rats

There is evidence to suggest that hypertension involves a chronic low-grade systemic inflammatory response; however, the underlying mechanisms are unclear. To further understand the role of inflammation in hypertension, we used a rat renovascular model of hypertension in which we administered the TNF-α synthesis inhibitor pentoxifylline (PTX, 30 mg/kg/day) in the drinking water for 60 days. In conscious rats, PTX administration significantly attenuated the development of hypertension (systolic blood pressure, PTX: 145 ± 8 vs. vehicle (Veh): 235 ± 11 mmHg, after 38 days of treatment, P < 0.05, N = 5/group). This attenuation in hypertension was coupled with a decrease in the low-frequency spectra of systolic blood pressure variability (PTX: 1.23 ± 0.2 vs Veh: 3.05 ± 0.8 arbitrary units, P < 0.05, N = 5/group). Furthermore, systemic PTX administration decreased c-Fos expression within the hypothalamic paraventricular nucleus (PTX: 17 ± 4 vs. Veh: 70 ± 13 cells, P < 0.01, N = 5, PVN) and increased the total number of microglial branches (PTX: 2129 ± 242 vs. Veh: 1415 ± 227 branches, P < 0.05, N = 4/group). Acute central injection of PTX (20 μg) under urethane anesthesia caused a small transient decrease in blood pressure but did not change renal sympathetic nerve activity. Surprisingly, we found no detectable basal levels of plasma TNF-α in either PTX- or vehicle-treated animals. These results suggest that inflammation plays a role in renovascular hypertension and that PTX might act both peripherally and centrally to prevent hypertension.

Renovascular hypertension elevates pulmonary ventilation in rats by carotid body-dependent mechanisms.

The two kidney-one clip (2K1C) renovascular hypertension depends on the renin-angiotensin system and sympathetic overactivity. The maintenance of 2K1C hypertension also depends on inputs from the carotid bodies (CB), which when activated stimulate the respiratory activity. In the present study, we investigated the importance of CB afferent activity for the ventilatory responses in 2K1C hypertensive rats and for phrenic and hypoglossal activities in in situ preparations of normotensive rats treated with angiotensin II. Silver clips were implanted around the left renal artery of male Holtzman rats (150 g) to induce renovascular hypertension. Six weeks after clipping, hypertensive 2K1C rats showed, in conscious state, elevated resting tidal volume and minute ventilation compared with the normotensive group. 2K1C rats also presented arterial alkalosis, urinary acidification, and amplified hypoxic ventilatory response. Carotid body removal (CBR), 2 wk before the experiments (4th week after clipping), significantly reduced arterial pressure and pulmonary ventilation in 2K1C rats but not in normotensive rats. Intra-arterial administration of angiotensin II in the in situ preparation of normotensive rats increased phrenic and hypoglossal activities, responses that were also reduced after CBR. Results show that renovascular hypertensive rats exhibit increased resting ventilation that depends on CB inputs. Similarly, angiotensin II increases phrenic and hypoglossal activities in in situ preparations of normotensive rats, responses that also depend on CB inputs. Results suggest that mechanisms that depend on CB inputs in renovascular hypertensive rats or during angiotensin II administration in normotensive animals increase respiratory drive.

Afferent innervation of the ischemic kidney contributes to renal dysfunction in renovascular hypertensive rats.

The ablation of renal nerves, by destroying both the sympathetic and afferent fibers, has been shown to be effective in lowering blood pressure in resistant hypertensive patients. However, experimental studies have reported that the removal of sympathetic fibers may lead to side effects, such as the impairment of compensatory cardiorenal responses during a hemodynamic challenge. In the present study, we evaluated the effects of the selective removal of renal afferent fibers on arterial hypertension, renal sympathetic nerve activity, and renal changes in a model of renovascular hypertension. After 4weeks of clipping the left renal artery, afferent renal denervation (ARD) was performed by exposing the left renal nerve to a 33mM capsaicin solution for 15min. After 2weeks of ARD, we found reduced MAP (~ 18%) and sympathoexcitation to both the ischemic and contralateral kidneys in the hypertensive group. Moreover, a reduction in reactive oxygen species was observed in the ischemic (76%) and contralateral (27%) kidneys in the 2K1C group. In addition, ARD normalized renal function markers and proteinuria and podocin in the contralateral kidney. Taken altogether, we show that the selective removal of afferent fibers is an effective method to reduce MAP and improve renal changes without compromising the function of renal sympathetic fibers in the 2K1C model. Renal afferent nerves may be a new target in neurogenic hypertension and renal dysfunction.

Liposomal and Non-Liposomal Formulations of Vitamin C: Comparison of the Antihypertensive and Vascular Modifying Activity in Renovascular Hypertensive Rats.

Background: Liposomes constitute a promising drug delivery vehicle, and are believed to improve drugs’ effectiveness. This study was aimed to compare antihypertensive and vascular modifying activities of liposomal and non-liposomal forms of ascorbic acid.Methods: Forty-nine male Sprague-Dawley rats were randomly divided into seven groups (n=7): A sham vehicle-receiving (Sham-veh), hypertensive (HTN), vehicle-receiving hypertensive (HTN-Veh), two liposomal Ascorbic acid-treated hypertensive at 50 or 100 mg/kg/day (LVC-50 and LVC-100), and two non-liposomal Ascorbic acid-treated hypertensive at 50 or 100 mg/kg/day (VC-50 and VC-100). Systolic blood pressure (SBP) and heart rate (HR) were measured weekly; after 4 weeks, dose-responses to phenylephrine (PE) in the absence and presence of nitro-L-arginine methyl ester (L-NAME), acetylcholine (Ach), and sodium nitroprusside (SNP) were obtained on aortic rings. Data were analyzed with one-way ANOVA and Duncan’s multiple range test at a P value of <0.05 using Sigmastat statistical softwareResults: Compared to the non-liposomal form, the liposomal one was associated with more prominent effects on the final SBP. Both forms of Ascorbic acid decreased SBP dose-dependently. The basal and stimulated release of Nitric Oxide (NO) was significantly recovered by both forms of Ascorbic acid. The PE maximal responses were not significantly different between the liposomal and non-liposomal groups (P=0.08). Although the Emax of Ach-relaxation response was not different in two preparation forms, Ach-relaxation response induced a lower concentration of the liposomal form of Ascorbic acid (P=0.03Conclusion: The liposomal Ascorbic acid exhibited relaxation activity in significantly lower concentrations. The observed effects were partly mediated by the increased basal release of NO.