WKY Rats Research Articles

Different adaptive NO-dependent Mechanisms in Normal and Hypertensive Conditions.

Myocardial infarction (MI) remains the leading cause of death worldwide. We aimed to investigate the effect of NO deficiency on selective biochemical parameters within discreet myocardial zones after experimentally induced MI. To induce MI, the left descending coronary artery was ligated in two groups of 16-week-old WKY rats. In one group, NO production was inhibited by L-NAME (20 mg/kg/day) administration four weeks prior to ligation. Sham operations were performed on both groups as a control. Seven days after MI, we evaluated levels of nitric oxide synthase (NOS) activity, eNOS, iNOS, NFҡB/p65 and Nrf2 in ischemic, injured and non-ischemic zones of the heart. Levels of circulating TNF-α and IL-6 were evaluated in the plasma. MI led to increased NOS activity in all investigated zones of myocardium as well as circulating levels of TNF-α and IL-6. L-NAME treatment decreased NOS activity in the heart of sham operated animals. eNOS expression was increased in the injured zone and this could be a compensatory mechanism that improves the perfusion of the myocardium and cardiac dysfunction. Conversely, iNOS expression increased in the infarcted zone and may contribute to the inflammatory process and irreversible necrotic changes.

Stabilizing Ryanodine Receptor with Chronic Dantrolene Treatment Does not Affect Hypertension, but Attenuates Atrial Fibrillation Inducibility under Sympathetic Stimulation in Spontaneously Hypertensive Rats

IntroductionRyanodine receptor (RyR) dysfunction in skeletal muscle (RyR1) is linked to malignant hyperthermia, and in cardiac muscle (RyR2) is associated with cardiac arrhythmias and heart failure. We hypothesized that RyR dysfunction in vascular smooth muscle (RyR 1,2 and 3) could increase vascular resistance and blood pressure by increasing intracellular calcium level. RyR dysfunction may also contribute to increased atrial fibrillation (AF) in hypertension. Thus, stabilizing RyR function with chronic dantrolene treatment may attenuate hypertension development and AF inducibility in spontaneously hypertensive rats (SHR).MethodsMale SHR (16 week‐old) were randomized into vehicle‐ (n=10) and dantrolene‐treated (10mg/kg/d, n=10) groups for 4‐weeks. Male Wistar Kyoto (WKY, n=10) rats served as controls. Tail‐cuff blood pressures were recorded before treatment, and at 1‐week, 3‐week during the treatment period. At the end of the 4‐week treatment period, direct blood pressure, echocardiography and hemodynamics were recorded. Atrial fibrillation inducibility tests were performed at baseline and under sympathetic stimulation (SS) with isoproterenol infusion.ResultsCompared with WKY rats, SHR rats had significantly higher blood pressure throughout the experimental period and at the end of 4‐week treatment the systolic blood pressure (SBP) was 212±9mmHg in SHR versus 122±15mmHg in WKY (P<0.001). Dantrolene treatment in SHR had no effect on blood pressure level (208±16 mmHg in dantrolene versus 212±9mmHg in vehicle, P>0.05). AF inducibility was not significantly different between WKY and SHR rats at baseline. However, under SS AF inducibility was significantly increased in SHR compared with WKY rats (58±28% in SHR versus 20±23% in WKY, P<0.01) and dantrolene treatment attenuated AF inducibility under SS in SHR (58±28% in SHR‐vehicle versus 27±21% in SHR‐dantrolene group, P<0.05). Dantrolene treatment also reduced AF duration (0.8±1.4s in dantrolene group versus 10.0±15.0s in vehicle group, P<0.05).Conclusions: Stabilizing RyR with dantrolene treatment did not affect hypertension development in SHR rats, indicating RyR dysfunction does not play an important role in mediating hypertension development in this animal model. SHR rats did not show increased AF inducibility compared with WKY rats at baseline; however, under SS, SHR rats had increased vulnerability to AF induction which was attenuated with dantrolene treatment, indicating that enhanced SS‐induced RyR dysfunction in SHR may be responsible for enhanced AF inducibility in hypertension.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Spontaneously Hypertensive Rats is Susceptible to Bacteria Translocation from Gut to Peripheral Tissues

Cerebral small vessel disease (CSVD) is an inflammatory disease characterized by blood‐brain barrier (BBB) breakdown and neuroinflammation. We previous observed that spontaneously hypertensive stroke‐prone rats (SHRSPs) exhibit gut dysbiosis. Furthermore, transplanting SHRSPs with control gut microbiota significantly reduced makers associated with inflammation. Since the gut microbiome is a potential source of inflammation, we hypothesize that bacteria translocating from gut to brain induce neuroinflammation in CSVD. To test our hypothesis, we determined the efficiency of fluorescent‐labeled bacteria (GFP‐E. coli JJ1901, chloramphenicol resistant) translocation in WKY and SHRSP rats. A single dose of E. coli JJ1901 [≈109 colony forming units (CFUs)] was administered to WKY and SHRSP rats (11–15 weeks old) through oral gavage. Peripheral tissues, brain, and cecum content were collected 2 hours later, homogenized, and plated on culture plates. The number of CFUs was counted 24 hours later. When compared to WKY rats, SHRSP showed significantly increased bacteria translocation to peripheral tissues (liver, spleen, and abdominal adipose tissue; n=7, p<0.05). In addition, more bacteria translocation incidents were found in SHRSPs compared to WKY rats (5/7 in SHRSP vs. 3/7 in WKY). However, similar level of acute bacteria translocation to brain was observed in both groups. To assess translocation rates of the chronically colonized bacteria, as opposed to the acute effects following gavage, colonization was established in newborn rat pups by administrating E. coli JJ1901 (≈109 CFUs) to pregnant mother rats through oral gavage. Bacteria colonization in the offspring of the gavaged rats was confirmed by plating diluted fecal samples on culture plates. Colonized SHRSPs (15 weeks old) showed significantly increased bacteria translocation to peripheral tissues when compared to WKY controls. Additionally, SHRSPs demonstrated a trend towards increased translocation of E. coli JJ1901 to the brain than WKY rats. We conclude that translocation of gut bacteria can be an underlying source responsible for inflammation in pathological states.Support or Funding InformationNational Heart, Lung, and Blood Institute R01HL134838 (D.J. Durgan), and by National Institute of Neurological Disorders and Stroke R01NS080531 (R.M. Bryan)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Group III Metabotropic Glutamate Receptors Regulate Excitability of Hypothalamic Presympathetic Neurons and Sympathetic Output in Hypertension

Enhanced glutamatergic input to the hypothalamic paraventricular nucleus (PVN) contributes to the development of hypertension. Group III metabotropic glutamate receptors (mGluRs) are expressed in the PVN. In the present study, we determined the role of group III mGluRs in the control of the excitability of spinally projecting PVN neurons and sympathetic outflow in hypertension. We found that stimulation of group III mGluRs with L‐(+)‐2‐amino‐4‐phosphonobutyric acid (L‐AP4) consistently inhibited the firing activity of PVN neurons in normotensive WKY rats. However, L‐AP4 inhibited 37%, but excited 48%, of spinally projecting PVN neurons in spontaneously hypertensive rats (SHRs). Both the inhibitory and excitatory effects of L‐AP4 on the firing activity were abolished by the group III mGluR antagonist CPPG. Inclusion of GDP‐β‐S in the intracellular recording solution eliminated the excitatory effect of L‐AP4 on the firing activity of PVN neurons in SHRs, but it did not significantly change the inhibitory effect of L‐AP4 on PVN neurons in WKY rats or SHRs. L‐AP4 significantly reduced the frequency of spontaneous excitatory postsynaptic currents of PVN neurons in both WKY rats and SHRs. Interestingly, pretreatment of brain slices with the group I mGluR agonist DHPG converted the L‐AP4 effect on PVN neurons from inhibitory to excitatory in WKY rats. Also, L‐AP4 consistently inhibited PVN neurons in SHRs after treatment with the group I mGluR antagonist MPEP. In addition, microinjection of L‐AP4 into the PVN dose‐dependently decreased arterial Bp and lumbar sympathetic nerve activity (LSNA) in WKY rats and SHRs. Microinjection of L‐AP4 in the presence of MPEP produced a more significant reduction in arterial BP and LSNA than did with microinjection of L‐AP4 alone in SHRs. These findings suggest that group III mGluRs in the PVN regulate the synaptic plasticity and excitability of PVN presympathetic neurons and sympathetic vasomotor activity in hypertension. The signaling of postsynaptic group III mGluRs is influenced by increased group I mGluR activity in PVN presympathetic neurons in hypertension. Supported by NIH grants HL131161 and HL142133.Support or Funding InformationThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Hypertensive Neuropathy: A New Clinical Entity?

Hypertension is a main risk factor for stroke and vascular dementia and may cause important changes to the cerebrovascular tree, turning the brain more susceptible to infarcts, microaneurysms and ischemia. In spite of the well documented influence of hypertension on the brain, data on the sensitivity of peripheral nerves in hypertension is scarce. Our laboratory has been working for decades to help understanding how hypertension affects peripheral nerve morphology and function, using the spontaneously hypertensive rats (SHR) model. Spontaneously hypertensive rats (SHR), first inbred from Wistar Kyoto rats (WKY), are considered a good experimental model of human essential hypertension. The purpose of this study was to investigate the alterations on the morphology and morphometric data on sural, phrenic, vagus and recurrent laryngeal nerves of SHR, previous to the development of hypertension and with well‐established hypertension. Male and female SHR and normotensive WKY rats aged 5, 8 and 20 weeks (N = 6 in each group) were investigated. After arterial pressure and heart rate recordings in anesthetized animals, right and left sural, vagus, phrenic and recurrent laryngeal nerves were removed and prepared for epoxy resin embedding and light microscopy. Morphometric analysis was performed with the aid of computer software, and took into consideration the fascicle area and diameter, as well as myelinated fiber number, density, area and diameter. Nerves from young animals did not show differences between sides and genders. Significant differences were observed for the myelinated fiber number and density, comparing different genders of WKY and SHR in all nerves. Also, significant differences for the morphological (thickening of the endoneural blood vessel walls and lumen reduction) and morphometric (myelinated fibers diameter and G ratio) parameters of myelinated fibers were identified in nerves from adult hypertensive animals. Morphological exam of the myelinated fibers suggested the presence of a neuropathy due to hypertension in both SHR genders. These results indicate that hypertension altered important morphometric parameters related to nerve conduction in hypertensive animals. Moreover the comparison between males and females of WKY and SHR showed that the morphological and morphometric alterations due to hypertension are not gender related. The presence of a so called “hypertensive neuropathy” is documented with endoneural vascular lesions, axonal atrophy and loss of small myelinated fibers.Support or Funding InformationSupport: CNPq, FAPESP and FAEPAThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Altered Cardiopulmonary Reflexes Evoked by Irritants in Spontaneously Hypertensive Rats – Effect of Route of Administration and Anesthetics

Inhalation of noxious substances such as allyl isothiocyanate (AITC) evoke reflex bradycardia in conscious, healthy animals following the activation of airway nociceptive afferents. These afferents are also activated by air pollutants including particulate matter and ozone. Nevertheless, in patients with cardiovascular disease such pollutants evoke sympathoexcitation (tachycardia, hypertension). We hypothesize that preexisting cardiovascular disease remodels the pulmonary‐cardiac reflexes stimulated by noxious airway stimuli. Here, we studied the cardiopulmonary responses to two separate administration methods of AITC (inhalation and intravenous (i.v.) injection) in conscious and anesthetized normotensive Wistar‐Kyoto rats (WKY) and Spontaneously Hypertensive rats (SH). Both methods of AITC administration evoked bradycardia with increased RR‐interval (RRi) and atrial‐ventricular (AV) block in conscious WKY rats. AITC i.v. evoked similar bradycardic responses in conscious SH rats, however, AITC inhalation evoked a complex brady‐tachycardia which alternated between increased and decreased RRi and both AV block and premature ventricular contractions (PVCs). AITC inhalation caused bradypnea with sporadic episodes of tachypnea in both strains, while only bradypnea was seen after AITC i.v. administration. These responses were shown to be vagally mediated as they were abolished by bilateral vagotomy in a decerebrate model. Anesthesia was also found to prevent the effect of AITC inhalation on HR in both strains, while the bradycardic response to AITC i.v. persisted in the WKY but not in the SH. Furthermore, anesthesia prevented breathing rate responses to inhaled AITC in the SH, but not in the WKY. Differences were also seen in the effects of anesthesia on breathing rate responses to AITC iv between the two strains as bradypnea persisted in the WKY, while the SH was found to have bouts of tachypnea in addition to the bradypnea seen in the conscious studies. The differential responses evoked by AITC in the SH along with the differences in effects of anesthesia on these responses lead us to conclude that multiple unique circuits work independently to control cardiopulmonary reflexes and that preexisting hypertension remodels nociceptive cardiopulmonary reflexes towards sympathoexcitation.Support or Funding InformationNIH SPARC AwardThis abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Safety and efficacy of global intracoronary administration of cardiosphere-derived cells or conditioned medium immediately after coronary reperfusion in rats

ObjectiveCardiosphere-derived cells (CDCs) have been shown to reduce infarct size after myocardial infarction (MI). In the present study we investigated the safety and efficacy of global intracoronary administration (GIA) of CDCs or CDC-conditioned medium (CM) immediately after reperfusion in a rat model of ischemia-reperfusion. MethodsCDCs were grown from myocardial biopsies obtained from male Wistar Kyoto rats (WKY). Female WKY rats underwent MI for 45minutes, followed by reperfusion for 1hour. Infarcted rats were randomized to receive GIA of CDCs (CDC group), CM (CM group) or vehicle (control group) immediately after the onset of reperfusion. Cell retention was quantified by PCR for the male specific SRY gene; area at risk (AR) and no reflow area (NR) were measured by histopathology. Cardiac function was evaluated by echocardiography at 1 and 2 months post-MI. ResultsCell retention at 1hour after GIA was 25.1% ±5.1. The myocardial AR and NR (measured at 1 hour post-reperfusion) were similar between groups [AR: 28.8% ±7.4 of LV mass in control vs 27.2% ±8 in CM vs 27% ±7 in CDCs group. NR: 7.0% ±3.3 in control vs 7.3% ±3.8 in CM vs 7.1% ±3.6 in CDCs]. One and 2 months post-MI, systolic function and LV volumes did not differ between control and CM groups. ConclusionIntracoronary administration of CDCs during the acute phase of MI, at the beginning of reperfusion, does not aggravate microvascular obstruction and results in high cell retention. Delivery of CM in the acute phase of MI did not confer long-term cardiac functional benefits.

β‑estradiol alleviates hypertension‑ and concanavalin A‑mediated inflammatory responses via modulation of connexins in peripheral blood lymphocytes

Gap junctions (GJs) formed by connexins (Cxs) in T lymphocytes have been reported to have important roles in the T lymphocyte-driven inflammatory response and hypertension-mediated inflammation. Estrogen has a protective effect on cardiovascular diseases, including hypertension and it attenuates excessive inflammatory responses in certain autoimmune diseases. However, the mechanisms involved in regulating the pro-inflammatory response are complex and poorly understood. The current study investigated whether β-estradiol suppresses hypertension and pro-inflammatory stimuli-mediated inflammatory responses by regulating Cxs and Cx-mediated GJs in peripheral blood lymphocytes. Male, 16-week-old spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats (WKY) rats were randomly divided into the following three groups: WKY rats, vehicle (saline)-treated SHRs, and β-estradiol (20 µg/kg/day)-treated SHRs. β-estradiol was administered subcutaneously for 5 weeks. Hematoxylin and eosin staining was performed to evaluate target organ injury. Flow cytometry and ELISA were used to measure the populations of T lymphocyte subtypes in the peripheral blood, and expression of Cx40/Cx43 in T cell subtypes, and pro-inflammation cytokines levels, respectively. ELISA, a dye transfer technique, immunofluorescence and immunoblotting were used to analyze the effect of β-estradiol on pro-inflammatory cytokine secretion, Cx-mediated GJs and the expression of Cxs in concanavalin A (Con A)-stimulated peripheral blood lymphocytes isolated from WKY rat. β-estradiol significantly decreased blood pressure and inhibited hypertension-induced target organ injury in SHRs. Additionally, β-estradiol treatment significantly improved the immune homeostasis of SHRs, as demonstrated by the decreased percentage of cluster of differentiation (CD)4+/CD8+ T-cell subset ratio, reduced serum levels of pro-inflammatory cytokines and increased the percentage of CD4+CD25+ T cells. β-estradiol also markedly reduced the expression of Cx40/Cx43 in T lymphocytes from SHRs. In vitro, β-estradiol significantly suppressed the production of pro-inflammatory cytokines, reduced communication via Cx-mediated gap junctions and decreased the expression of Cx40/Cx43 in Con A-stimulated lymphocytes. These results indicate that β-estradiol attenuates inflammation and end organ damage in hypertension, which may be partially mediated via downregulated expression of Cxs and reduced function of Cx-mediated GJ.

Metabolomic Study of Obesity and Its Treatment with Palmitoylated Prolactin-Releasing Peptide Analog in Spontaneously Hypertensive and Normotensive Rats.

In this study, the combination of metabolomics and standard biochemical and biometric parameters was used to describe the metabolic effects of diet-induced obesity and its treatment with the novel antiobesity compound palm11-PrRP31 (palmitoylated prolactin-releasing peptide) in spontaneously hypertensive rats (SHR) and normotensive Wistar Kyoto rats (WKY). The results showed that SHR on a high-fat (HF) diet were normoglycemic with obesity and hypertension, while WKY on the HF diet were normotensive and obese with prediabetes. NMR-based metabolomics revealed mainly several microbial cometabolites altered by the HF diet, particularly in urine. The HF diet induced similar changes in both models. However, two groups of genotype-specific metabolites were defined: metabolites specific to the genotype at baseline (e.g., 1-methylnicotinamide, phenylacetylglycine, taurine, methylamine) and metabolites reacting specifically to the HF diet in individual genotypes (2-oxoglutarate, dimethylamine, N-butyrylglycine, p-cresyl sulfate). The palm11-PrRP31 lowered body weight and improved biochemical and biometric parameters in both strains, and it improved glucose tolerance in WKY rats on the HF diet. In urine, the therapy induced significant decrease of formate and 1-methylnicotinamide in SHR and alanine, allantoin, dimethylamine, and N-butyrylglycine in WKY. Altogether, our study confirms the effectiveness of palm11-PrRP31 for antiobesity treatment.

Transient receptor potential vanilloid 4 channels are important regulators of parenchymal arteriole dilation and cognitive function.

Hypertension-associated PA dysfunction reduces cerebral perfusion and impairs cognition. This is associated with impaired TRPV4-mediated PA dilation; therefore, we tested the hypothesis that TRPV4 channels are important regulators of cerebral perfusion, PA structure and dilation, and cognition. Ten- to twelve-month-old male TRPV4 knockout (WKY-Trpv4em4Mcwi ) and age-matched control WKY rats were studied. Cerebral perfusion was measured by MRI with arterial spin labeling. PA structure and function were assessed using pressure myography and cognitive function using the novel object recognition test. Cerebral perfusion was reduced in the WKY-Trpv4em4Mcwi rats. This was not a result of PA remodeling because TRPV4 deletion did not change PA structure. TRPV4 deletion did not change PA myogenic tone development, but PAs from the WKY-Trpv4em4Mcwi rats had severely blunted endothelium-dependent dilation. The WKY-Trpv4em4Mcwi rats had impaired cognitive function and exhibited depressive-like behavior. The WKY-Trpv4em4Mcwi rats also had increased microglia activation, and increased mRNA expression of GFAP and tumor necrosis factor alpha suggesting increased inflammation. Our data indicate that TRPV4 channels play a critical role in cerebral perfusion, PA dilation, cognition, and inflammation. Impaired TRPV4 function in diseases such as hypertension may increase the risk of the development of vascular dementia.

ADHD pathogenesis in the immune, endocrine and nervous systems of juvenile and maturating SHR and WKY rats

RationaleAttention-deficit/hyperactivity disorder (ADHD) is one of the most common neurobehavioural disorders with morphological and functional brain abnormalities. However, there is a growing body of evidence that abnormalities in the immune and endocrine systems may also account for the ADHD pathogenesis.ObjectivesTo test ADHD pathogenesis in neurological, immune and endocrine systems, this study examined the concentrations of cytokines, chemokines, oxidative stress markers, metabolic parameters, steroid hormones and steroidogenic enzymes in the serum and/or tissues of spontaneously hypertensive rats (SHRs, animal model of ADHD) and Wistar Kyoto rats (WKYs, control animals). Moreover, the volume of the medial prefrontal cortex (mPFC) as well as the density of dopamine 2 (D2) receptor-expressing cells and tyrosine hydroxylase (TH)-positive nerve fibres in it was also elucidated.MethodsPeripheral blood, spleen and adrenal gland samples, as well as brain sections collected on day 35 (juvenile) and day 70 (maturating) from SHRs and WKYs, were processed by ELISA and immunohistochemistry, respectively.ResultsThe results show significant increases of serum and/or tissue concentrations of cytokines, chemokines and oxidative stress markers in juvenile SHRs when compared to the age-matched WKYs. These increases were accompanied by a lowered volume of the mPFC and up-regulation of D2 in this brain region. In maturating SHRs, the levels of inflammatory and oxidative stress markers were normalised and accompanied by elevated contents of steroid hormones.ConclusionsSignificant elevations of serum and/or tissue contents of cytokines, chemokines and oxidative stress markers as well as volumetric and neurochemical alterations in the mPFC of juvenile SHRs may suggest the cooperation of neurological and immune systems in the ADHD pathogenesis. Elevated levels of steroid hormones in maturating SHRs may be a compensatory effect involved in reducing inflammation and ADHD symptoms.

Atorvastatin inhibits pro-inflammatory actions of aldosterone in vascular smooth muscle cells by reducing oxidative stress

Vascular inflammatory responses play an important role in several cardiovascular diseases. Of the many pro-inflammatory vasoactive factors implicated in this process, is aldosterone, an important mediator of vascular oxidative stress. Statins, such as atorvastatin, are cholesterol-lowering drugs that have pleiotropic actions, including anti-oxidant properties independently of their cholesterol-lowering effect. This study investigated whether atorvastatin prevents aldosterone-induced VSMC inflammation by reducing reactive oxygen species (ROS) production. Vascular smooth muscle cells (VSMC) from WKY rats were treated with 1 μM atorvastatin for 60 min or for 72 h prior to aldosterone (10-7 mol/L) stimulation. Atorvastatin inhibited Rac1/2 and p47phox translocation from the cytosol to the membrane, as well as reduced aldosterone-induced ROS production. Atorvastatin also attenuated aldosterone-induced vascular inflammation and macrophage adhesion to VSMC. Similarly EHT1864, a Rac1/2 inhibitor, and tiron, ROS scavenger, reduced macrophage adhesion. Through its inhibitory effects on Rac1/2 activation and ROS production, atorvastatin reduces vascular ROS generation and inhibits VSMC inflammation. Our data suggest that in conditions associated with aldosterone-induced vascular damage, statins may have vasoprotective effects by inhibiting oxidative stress and inflammation.

Abstract TMP118: The Gut Microbiota Can Protect Against the Onset of Symptoms in an Animal Model of Cerebral Small Vessel Disease

It has become increasingly evident that disruption of the gut microbiota plays an important role in multiple host disease states. We hypothesize that maintaining a healthy gut microbiome protects against the development of cerebral small vessel disease (CSVD), an inflammatory disease accompanied by blood-brain barrier (BBB) breakdown and neuroinflammation. To test our hypothesis, we altered the gut microbiota of spontaneously hypertensive stroke-prone rats (SHRSPs), a model of CSVD associated with hypertension, to resemble that of healthy/asymptomatic WKY control rats. At birth SHRSP pups were removed from their birth moms and nursed by either WKY (SW rats) or SHRSP (SS) foster dams until 21 days of age. At 20 weeks, markers associated with neuroinflammation were significantly decreased in SW rats compared to SS rats (NF-κB1 by 68%, TLR-2 by 58%, and TLR-4 by 54%, P≤ 0.01, n=5-6/ group). When compared to SS rats, SW rats showed a 33% (P=0.045) decrease in IgG extravasation into brain indicating greater BBB integrity. Systolic blood pressure at 20 weeks in SW was 173±6 mm Hg compared to 201±6 mm Hg in SS rats (P=0.003) and these pressures remained stable for until at least 30 weeks. Analysis of microbiota in feces revealed that gut microbiota of SW rats were significantly different from SS rats (P<0.05, principal components analysis of weighted Unifrac distances, n=5-6), resembled the microbiome in WKY rats, and showed > 2-fold increase in the abundance of the genus, Akkermansia, in SW compared to SS rats (6.1% vs 3.0%, P=0.01). Akkermansia can enhance gut integrity by stimulating epithelial cell mucus production and enhancing tight junctions. We conclude that the gut microbiota can influence the onset of CSVD-like symptoms even in the genetic SHRSP model. Maintaining a microbiota similar to WKY control rats, which includes an increased abundance of Akkermansia, appears to protect against the onset of CSVD. Altering the gut microbiota in humans could be an effective prophylactic measure to protect against CSVD.

Regulation of DNA methylation and 2-OG/TET signaling by choline alleviated cardiac hypertrophy in spontaneously hypertensive rats

DNA methylation is a well-defined epigenetic modification that regulates gene transcription. However, the role of DNA methylation in the cardiac hypertrophy seen in hypertension is unclear. This study was performed to investigate genome-wide DNA methylation profiles in spontaneously hypertensive rats (SHRs) and Wistar-Kyoto rats (WKY), and the cardioprotective effect of choline. Eight-week-old male SHRs received intraperitoneal injections of choline (8 mg/kg/day) for 8 weeks. SHRs showed aberrant methylation distribution on chromosomes and genome regions, with decreased methylation levels at CHG and CHH sites. A total of 91,559 differentially methylated regions (DMRs) were detected between SHRs and WKY rats, of which 28,197 were demethylated and 63,362 were methylated. Choline treatment partly restored the DMRs in SHRs, which were related to 131 genes. Gene ontology analysis and Kyoto Encyclopedia of Genes and Genomes analysis of DMRs suggested that choline partly reversed the dysfunctions of biological processes, cellular components and molecular functions in SHRs. Moreover, the inhibition of 2-oxoglutarate accumulation by choline, thereby inhibiting excessive activation of ten-eleven translocation methylcytosine dioxygenase enzymes, may correlate with the beneficial effects of choline on methylation levels, cardiac hypertrophy and cardiac function of SHRs, as indicated by decreased heart rate and blood pressure, and increased ejection fraction and fractional shortening. This study provides the first genome-wide DNA methylation profile of the hypertrophic myocardium of SHRs and suggests a novel role for this epigenetic modification in hypertension. Choline treatment may represent a promising approach for modification of DNA methylation and optimization of the epigenetic profile for antihypertensive therapy.

Roles of cocaine- and amphetamine-regulated transcript peptide in the rostral ventrolateral medulla in cardiovascular regulation in rats

Cocaine- and amphetamine-regulated transcript peptide (CARTp) is present in neurons and varicose fibers in the rostral ventrolateral medulla (RVLM) that is crucial in the control of cardiovascular function. Prior research indicated that intracisternal administration of CARTp evokes hypertension and accumulation of Fos in the RVLM. Despite the interaction among CARTp, cardiovascular effect, and the RVLM, no studies have directly examined whether CARTp participates in cardiovascular regulation in the RVLM. The current study directly examined the modulation of blood pressure and baroreflex sensitivity by CARTp in the RVLM in the different strain of rats. Immunohistochemical study showed that CARTp immunoreactive (CART-IR) cell bodies and varicose CART-IR fibers were observed throughout the RVLM in the SD, WKY, and SHRs. Varicose CART-IR nerve fibers were particularly abundant in the WKY and SHRs. Bilateral microinjection of CARTp (30 pmol) into the RVLM caused a significant increase in mean arterial pressure (MAP) in WKY and SHRs. Bilateral microinjection of CARTp antibody (1:5000) into the RVLM displayed a fall in the basal level of the MAP in SHRs but had no effects in WKY rats. In SD rats, bilateral microinjection of CARTp (6, 30 or 60 pmol) into the RVLM did not change the MAP but attenuated phenylephrine-induced bradycardia in a dose-dependent manner. We propose that CARTp acting in the RVLM may involvement in the cardiovascular regulation either by increases in the blood pressure or by decreases in the baroreflex sensitivity in rats. Moreover, endogenous CARTp in the RVLM is associated with the maintenance of basal blood pressure of SHRs.

Assessing the role of extracellular vesicles in renin-angiotensin system signalling in cardiomyocyte hypertrophy

Assessing the role of extracellular vesicles in renin-angiotensin system signalling in cardiomyocyte hypertrophy

Superoxide anions modulate the performance of apelin in the paraventricular nucleus on sympathetic activity and blood pressure in spontaneously hypertensive rats

The present study was designed to determine how apelin in paraventricular nucleus (PVN) modulates the renal sympathetic nerve activity (RSNA), arterial blood pressure (ABP), mean arterial pressure (MAP), and heart rate (HR), and whether superoxide anions regulate the performance of PVN apelin in spontaneously hypertensive rats (SHRs). Acute experiment was carried out with 13-week-old male Wistar-Kyoto rats (WKY) and SHRs under anaesthesia. RSNA, ABP, MAP and HR after PVN microinjection were measured. Apelin microinjection into PVN increased RSNA, ABP, MAP and HR in WKY rats and SHRs, more obviously in SHRs. APJ antagonist F13A decreased the RSNA, ABP, MAP and HR in SHRs, and inhibited the effects of apelin. Apelin and APJ mRNA levels were higher in the PVN in SHRs. PVN microinjection of superoxide anion scavengers tempol and tiron, or NAD(P)H oxidase inhibitor apocynin, decreased the RSNA, ABP, MAP and HR in SHRs, and inhibited the effects of apelin, but the superoxide dismutase (SOD) inhibitor diethyldithiocarbamic acid (DETC) potentiated the effects of apelin. NAD(P)H oxidase activity and superoxide anion levels in PVN were increased by apelin, but decreased by APJ antagonist F13A. The apelin-induced increases in NAD(P)H oxidase activity and superoxide anion level were abolished by pre-treatment with F13A. These results indicate that apelin in PVN increases the sympathetic outflow and blood pressure via activating APJ receptor. The enhanced activity of endogenous apelin and APJ receptor in PVN contributes to sympathetic activation in hypertension, and the superoxide anion is involved in these apelin–mediated processes in PVN.

Fatty acid amide hydrolase inhibitor (URB597) as a regulator of myocardial lipid metabolism in spontaneously hypertensive rats.

Pressure overload, which is typical of hypertension, is known to evoke alterations not only in the morphology of the heart but also in the preference of myocardial energetic substrates usage. Nowadays, the endocannabinoid system (ECS) serves as a potential therapeutic target for cardiovascular disorders and, simultaneously, affects whole body metabolism homeostasis. Therefore, an open question is whether ECS, apart from decreasing blood pressure, also affects cardiac muscle metabolism in hypertensive conditions. All experiments were conducted on a genetic model of primary hypertension i.e. spontaneously hypertensive rats (SHRs) and Wistar Kyoto rats (WKY) served as a normotensive control. ECS was chronically activated by 2-weeks intraperitoneal injections of fatty acid amide hydrolase (FAAH) inhibitor - URB597. Lipid analyses in the left ventricle and serum were based on ex vivo heart perfusion in Langendorff perfusion system, thin layer chromatography, and gas liquid chromatography. The total expression of selected proteins was determined using Western blot as well as immunohistochemical techniques. As expected, URB597 markedly reduced systolic as well as mean blood pressures in SHRs. Moreover, prolonged FAAH inhibition resulted in stimulation of 3H-palmitate uptake and incorporation into different lipid fractions in cardiomyocytes in the hypertensive as well as normotensive conditions. An increase in fatty acid oxidation caused by URB597 treatment was observed only in WKY rats, but not SHRs, and was accompanied by an elevation in peroxisome proliferator-activated receptor alpha (PPARα) and β-hydroxyacyl-CoA dehydrogenase (β-HAD) expressions. Chronic activation of ECS significantly upregulates palmitate uptake and its esterification but not oxidation in the SHR's myocardium.

Exposure to methylphenidate in adolescence and adulthood modulates cross-sensitization to amphetamine in adulthood in three genetically variant female rat strains

Attention Deficit Hyperactivity Disorder (ADHD) is a developmental, behavioral disorder that is characterized by patterns of impulsivity and limited attention. Stimulants, such as methylphenidate (MPD) and amphetamine (Amph), are utilized as first-line agents in the treatment of ADHD. While Amph is known to elicit dependence, MPD is not. Interdependence between MPD and Amph is a growing concern. Therefore, it is important to explore this interaction in animal models to gain insight on its mechanisms. In experimental studies, drugs that elicit behavioral sensitization and cross-sensitization in animals are considered to have the potential to elicit dependence. This study evaluated whether chronic repetitive MPD exposure in adolescence and/or adulthood in various genetic strains of female rats elicits behavioral sensitization as well as cross-sensitization with Amph. We used three strains: SHR (ADHD model), WKY, and SD rats. The three rat strains were exposed to chronic MPD in their adolescence and adulthood or exposed to chronic MPD only in adulthood. When the female rats were treated with MPD only in adulthood, the SHR strain exhibited cross-sensitization to Amph but the WKY and SD strains did not. Whereas, when the female rats were pretreated with MPD in adolescence and adulthood, the SHR and WKY strains exhibited cross-sensitization response to Amph, but the SD strain did not. This study showed that the genetic strain and age of exposure to MPD plays a crucial role in cross-sensitization to Amph in female rodents. Furthermore, we showed that genetics associated with ADHD pre-dispose animals to dependence between drugs even if the exposure starts in adulthood compared to control strains that did not show dependence with adult only MPD exposures. Genetic variability, age of initial drug exposure, and sex of the subject are key variables that should be accounted in studies that explore effects of psychostimulants.

Silencing salusin β ameliorates heart failure in aged spontaneously hypertensive rats by ROS-relative MAPK/NF-κB pathways in the paraventricular nucleus

ObjectiveSustained hypertension is a major cause of heart failure in aging hypertensive patients. Salusin β, a novel bioactive peptide of 20 amino acids, has been reported to participate in various cardiovascular diseases, including hypertension. We therefore hypothesized that central knockdown of salusin β might be effective for hypertension-induced heart failure treatment. Methods and resultsEighteen-month-old male aged spontaneously hypertensive rats (SHR) with heart failure and WKY rats were microinjected with either a specific adenoviral vector encoding salusin β shRNA (Ad-Sal-shRNA) or a scramble shRNA (Ad-Scr-shRNA) in the hypothalamic paraventricular nucleus (PVN) for 4 weeks. Radiotelemetry and echocardiography were used for measuring blood pressure and cardiac function, respectively. Blood samples and heart were harvested for evaluating plasma norepinephrine, tyrosine hydroxylase, and cardiac morphology, respectively. The mesenteric arteries were separated for measurement of vascular responses. The PVN was analyzed for salusin β, proinflammatory cytokines (PICs), mitogen-activated protein kinase (MAPK), NF-κB, and reactive oxygen species (ROS) levels. Compared with normotensive rats, aging SHR with heart failure had dramatically increased salusin β expression. Silencing salusin β with Ad-Sal-shRNA attenuated arterial pressure and improved autonomic function, cardiac and vascular dysfunction in aging SHR with heart failure, but not in aging WKY rats. Knockdown of salusin β significantly reduced paraventricular nucleus PICs levels, MAPK and NF-κB activity, and ROS levels in aging SHR with heart failure. ConclusionThese data demonstrate that in aging SHR, the heart failure that was developed during the end stage of hypertension could be ameliorated by silencing salusin β.