Brain Of Spontaneously Hypertensive Rats Research Articles

Gastrodin Improved Baroreflex Sensitivity and Increased Gamma‐Amino Butyric Acid Content in Brains without Decreasing Blood Pressure in Spontaneously Hypertensive Rats

Gastrodin, a bioactive component in Gastrodina elata Bl., has been approved as a drug for the treatment for dizziness and headache in China 1. Combined with antihypertensive drugs, gastrodin was employed to deal with refractory hypertension in the elderly, but the effect of gastrodin alone on hypertension remains unknown. Arterial baroreflex plays a crucial role in the regulation of cardiovascular activities, and baroreflex function impairment should be regarded as an independent risk factor in hypertension. A new strategy, improving the impaired arterial baroreflex function, has been promoted to treat cardiovascular disease 2. Glutamate (GLU) and gamma-amino butyric acid (GABA), the most common neurotransmitters in the central nerve system (CNS), were both involved in baroreflex regulation 3. Our present experiment was designed to investigate the effect of gastrodin on hypertension, baroreflex sensitivity (BRS) and content of GLU and GABA in the CNS in spontaneously hypertensive rat (SHR). Spontaneously hypertensive rat (20–22 weeks of age) received gastrodin (Xi'nan Co., Chongqing, China) via the left femoral vein in doses of 8, 80 and 800 mg/kg (n = 7 per group). Blood pressure, heart period (HP) and BRS were recorded 0.5 h before and 2 h after drug administration in conscious freely moving SHR 4. It was found that BRS was significantly enhanced after administration in 80 and 800 mg/kg (Figure 1D), while no significant difference was found (Figure 1A–C) in systolic blood pressure, diastolic blood pressure and HP. In another experiment, 2 h after bolus injection of gastrodin (80 or 800mg/kg, ip) or vehicle (n = 7–9 per group), content of GLU and GABA in whole brain of SHR was determined 4 using HPLC/MS. Gastrodin increased GABA content significantly in 800 mg/kg group (Figure 2A). No similar results were found about GLU content (Figure 2B). The present study has directly demonstrated for the limited times that, independent of blood pressure reduction, gastrodin can significantly improve baroreflex function. The blood pressure was not the unique but the most common factor that influence arterial baroreflex. When a drug lowers blood pressure, BRS often increased. Gastrodin, similar to low dose of ketanserin 5, was one of the few drugs that can enhance BRS, not attributable to the normalization of BP levels. As baroreflex was an independent factor that causes end-organs damage in hypertension 2, the study of baroreflex mechanism became more and more important. The effect of gastrodin improving baroreflex without reducing blood pressure may help investigating baroreflex mechanism from more directions than blood pressure reduction. Common mechanism of blood pressure and baroreflex in the CNS, especially in the nucleus of the solitary tract (NTS), was well reported 6, 7. GABA regulated both blood pressure and baroreflex in the NTS, and injection of GABA agonists within the NTS produced an inhibition of NTS neurons, which resulted in reduction in the baroreflex bradycardia and an increase in arterial pressure 6. Meanwhile, GLU receptor antagonists block arterial baroreflex as well as regulate blood pressure in the NTS or in the caudal ventrolateral medulla 8. Gastrodin decreased immunoreactivities of GABA shunt enzymes in the hippocampus 9 and ameliorated the cerebral injury in the rats of cerebral ischaemia reperfusion by improving the level of GABA in striatum 10. Our present study indicated gastrodin increased GABA content in the CNS. All these evidence suggested that GABA increased as a secondary effect in the CNS owing to the gastrodin administration. In summary, gastrodin improved BRS without decreasing blood pressure in SHR. The study was supported by Sciences and Technology Developing Plan grant (No. 2008GG2NS02006) of Shandong Province, and the authors appreciate expert technical assistances from the Second Military Medical University and the Academy of Military Medical Sciences. The authors declare no conflict of interest.

Endogenous Hydrogen Sulfide in Carotid Bodies Correlates with the Initiation of Hypertension in Spontaneously Hypertensive Rats (SHR)

Hydrogen sulfide (H2S) is a new gaseous mediator of carotid chemoreceptor responses to hypoxia. Its production is determined by cystathionine γ lyase (CSE), cystathionine β synthetase (CBS), and 3‐mercaptopyruvate sulfurtransferase (3MST). Since CSE deletion severely impaired mouse carotid body response to hypoxia, we hypothesized that genetic overexpression of the H2S enzymes occurs in carotid bodies of SHR and explains their chemoreceptor hypersensitivity that contributes to hypertension. Carotid bodies were removed from anesthetized male SHR (and their normotensive control WKY rats) when they were young and prehypertensive at 4 weeks, when hypertension was established (at 12 weeks), and when hypertension had been long standing (at 38 weeks). The mRNA expression of CSE, CBS and 3MST was over 100 fold higher (p<0.01) in SHR than in WKY at 4 weeks of age. The expression decreased in SHR to levels that were only slightly higher than WKY at 12 and 38 weeks. The enzyme expression in WKY remained relatively low and stable over time. Expressions in brains of SHR and WKY were comparable. The results indicate a marked overexpression of H2S in carotid bodies of SHR during the critical prehypertensive stage. We suggest that this important molecular determinant of chemoreceptor hypersensitivity causes an increase in sympathetic nerve activity, and induces the initiation and development of hypertension. (HL14388)

NTS (pro)renin receptor (PRR)‐mediated antihypertensive effect involves NF‐KappaB‐cytokine signaling in the spontaneously hypertensive rats (SHR)

Increased PRR expression in the NTS of SHR, coupled with the evidence on role of the brain inflammation in cardiovascular functions, has led us to hypothesize that the NTS PRR may play a part in blood pressure (BP) regulation by modulation of the brain inflammatory pathways. Acute NTS injections of renin (0.02–2 pmol) resulted in a significantly greater dose‐dependent decrease in systolic BP (ΔSBP; −12±5 to −32±5 mmHg) and heart rate (ΔHR; −13±7 to −40±10 bpm) in the SHR compared to the WKY rats (ΔSBP: −3±1 to−8±4 mmHg; ΔHR: −2±4 to −12±7 bpm). These effects were not blocked by the AT1R or AT2R blocker. Consistent with this, chronic PRR knockdown with AAV2‐PRR‐shRNA in the SHR NTS caused an increase in mean arterial pressure (shRNA: 173±5; Control: 151±6 mmHg), and decreases in baroreflex bradycardia (shRNA: 0.26±0.04; Control: 0.44±0.04) and parasympathetic tone (shRNA: 31±4; Control: 49±6). This was accompanied by a decrease in the inflammatory markers TNFa, IL6 and Ccl5, and their transcription factors, NF‐κB and AP‐1. Finally, PRR activation in the SHR brain neuronal cultures activated NF‐κB, and increased the mRNA of IL1β (250 fold), TNFα (32 fold), IL6 (35 fold) and Ccl5 (12 fold) in an NF‐κB‐ and AP1‐dependent, but AT1R/AT2R‐independent manner. Therefore, the NTS PRR mediates Ang II‐independent antihypertensive effects by stimulation of NF‐κB/AP‐1/cytokine signaling (AHA 11SDG7420029)

Aerobic Training (T) Improves Cerebral Circulation of Normotensive (WKY) and Spontaneously Hypertensive Rats (SHR)

We compared the effects of hypertension and T in brain macro and microcirculation. Rats were submitted to treadmill T (55% of maximum capacity, 1h/day, 5 d/week) or kept sedentary (S) for 3 months. After hemodynamic measurements at rest, rats were anesthetized for iv. administration of FITC‐dextran (capillary density) or 4% PFA perfusion (morphometry of cerebral arteries). Brains were removed, post‐fixed and sliced for analysis of the capillary density within the NTS (25 μm) and of the basilar (BA), middle (MCA) and posterior cerebral (PCA) arteries (10 μm) (Image ProPlus). Compared to WKY, SHR exhibited elevated MAP and HR (180±2 mmHg, 359±6 b/min), increased wall/lumen ratio (+40%, +100% and +80% for BA, MCA and PCA, respectively), but no NTS capillary rarefaction. T was equally effective to improve treadmill performance and to cause resting bradycardia (−9%) in both groups; pressure fall (−8%) was only observed in the SHR. T reduced wall/lumen ratio only in the SHR (BA= −40%, MCA= −47%; PCA= −37%) and caused marked increase in capillary density in medial, intermediate and commissural NTS in both groups (on average 7.05 ±0.18 and 7.07±0.88%, corresponding to changes of +55% and +43%, WKY and SHR, respectively). Data showed that T improves brain perfusion by causing eutrophic outward remodeling in SHR brain arteries and capillary angiogenesis in both groups. Financial Support: CAPES, FAPESP

Decreased α4β2 nicotinic receptor number in the absence of mRNA changes suggests post‐transcriptional regulation in the spontaneously hypertensive rat model of ADHD

The spontaneously hypertensive rat (SHR) is widely used as a model of attention-deficit/hyperactivity disorder (ADHD). Deficits in central nicotinic receptors (nAChRs) have been previously observed in SHRs, which is interesting since epidemiological studies have identified an association between smoking and ADHD symptoms in humans. Here, we examine whether nAChR deficits in SHRs compared with Wistar Kyoto rat (WKY) controls are nAChR subtype-specific and whether these deficits correlate with changes at the level of mRNA transcription in specific brain regions. Levels of binding sites (B(max) ) and dissociation constants (K(d)) for nAChRs were determined from saturation curves of high-affinity [³H]epibatidine- and [³H] Methyllycaconitine (MLA) binding to membranes from cortex, striatum, hippocampus and cerebellum. In additional brain regions, nAChRs were examined by autoradiography with [¹²⁵I]A-85380 and [¹²⁵I]α-bungarotoxin. Levels of mRNA encoding nAChR subunits were measured using quantitative real-time PCR (qPCR). We showed that the number of α4β2 nAChR binding sites is lower globally in the SHR brain compared with WKY in the absence of significant differences in mRNA levels, with the exception of lower α4 mRNA in cerebellum of SHR compared with WKY. Furthermore, nAChR deficits were subtype- specific because no strain difference was found in α7 nAChR binding or α7 mRNA levels. Our results suggest that the lower α4β2 nAChR number in SHR compared with WKY may be a consequence of dysfunctional post-transcriptional regulation of nAChRs.

Nuclear Factor Kappa B and Matrix Metalloproteinase Induced Receptor Cleavage in the Spontaneously Hypertensive Rat

Recent evidence suggests that inflammation in the spontaneously hypertensive rat (SHR) is associated with an uncontrolled matrix metalloproteinase (MMP) activity. We hypothesize that the transcription factor nuclear factor kappa B (NFκB) is overexpressed in the SHR, enhancing its MMP activity and enzymatic cleavage of the β2 adrenergic receptor (β₂AR), thereby diminishing catecholamine-mediated arteriolar vasodilation. NFκB expression level and translocation were compared between Wistar Kyoto rat and SHR kidney, heart, and brain. The animals were treated with NFκB inhibitor, pyrrolidine dithiocarbamate, for 10 weeks and correlations between NFκB and MMP activity were determined. Immunohistochemistry showed that NFκB expression is increased in untreated SHR kidney (≈ 14%) and brain hypothalamus (≈ 22%) compared to that in Wistar Kyoto rats (P < 0.05), but not in myocardium and cerebral cortex. After pyrrolidine dithiocarbamate treatment, the SHR systolic blood pressure was reduced to close to Wistar Kyoto rat levels. NFκB expression level in treated SHR was also decreased in kidney and hypothalamus compared to nontreated animals (P < 0.05). Furthermore, MMP-2 and MMP-9 activities in SHR plasma were significantly reduced (≈ 41%) by pyrrolidine dithiocarbamate treatment. Additionally, zymographic analyses and in situ zymography showed decreased MMP-2 activity in kidney homogenates and decreased MMP-1 and MMP-9 activities in brain. The level of the β₂AR extracellular, but not intracellular, domain density was found to be reduced in kidney, showing a receptor cleavage process that can be blocked by pyrrolidine dithiocarbamate treatment. These results suggest NFκB is an important transcription factor in the SHR and may be involved in the enhanced MMP activity and, consequently, receptor cleavage.

Brain nitric oxide production by a proline-rich decapeptide from Bothrops jararaca venom improves baroreflex sensitivity of spontaneously hypertensive rats

Baroreflex sensitivity is disturbed in many people with cardiovascular diseases such as hypertension. Brain deficiency of nitric oxide (NO), which is synthesized by NO synthase (NOS) in the citrulline-NO cycle (with argininosuccinate synthase (ASS) activity being the rate-limiting step), contributes to impaired baroreflex. We recently showed that a decapeptide isolated from Bothrops jararaca snake venom, denoted Bj-PRO-10c, exerts powerful and sustained antihypertensive activity. Bj-PRO-10c promoted vasodilatation dependent on the positive modulation of ASS activity and NO production in the endothelium, and also acted on the central nervous system, inducing the release of GABA and glutamate, two important neurotransmitters in the regulation of autonomic systems. We evaluated baroreflex function using the regression line obtained by the best-fit points of measured heart rate (HR) and mean arterial pressure (MAP) data from spontaneously hypertensive rats (SHRs) treated with Bj-PRO-10c. We also investigated molecular mechanisms involved in this effect, both in vitro and in vivo. Bj-PRO-10c mediated an increase in baroreflex sensitivity and a decrease in MAP and HR. The effects exerted by the peptide include an increase in the gene expression of endothelial NOS and ASS. Bj-PRO-10c-induced NO production depended on intracellular calcium fluxes and the activation of a G(i/o)-protein-coupled metabotropic receptor. Bj-PRO-10c induced NO production and the gene expression of ASS and endothelial NOS in the brains of SHRs, thereby improving baroreflex sensitivity. Bj-PRO-10c may reveal novel approaches for treating diseases with impaired baroreflex function.

A Novel Function of MicroRNA Let‐7d in Regulation of Galectin‐3 Expression in Attention Deficit Hyperactivity Disorder Rat Brain

In this study we investigated the locomotor activity and non-selective attention in spontaneously hypertensive rats (SHR) with control Wistar-Kyoto (WKY) rats, which were employed as an attention deficit hyperactivity disorder (ADHD) model. In open-field test and làt maze, SHR rats were found to be much more spontaneously active than WKY rats. As compared with WKY rats, a lower level of galectin-3 was observed in SHR brain prefrontal cortex (PFC), which was the major affected brain area of ADHD. Through miRNA microarray screening, rno-let-7d was noted to be solely upregulated in SHR PFC. Interestingly, rno-let-7d had a binding site at galectin-3 mRNA and was shown to regulate galectin-3 3' untranslated region (UTR) directly. Mutation of galectin-3 3'UTR by one nucleotide of the seed sequence prevented rno-let-7d regulation of the 3' UTR completely. Although rno-let-7d did not directly regulate tyrosine hydroxylase (TH) 3'UTR, the level of galectin-3 was important for cAMP response element binding protein, the major transcript factor for TH gene. Either overexpression or downexpression of galectin-3 could result in modulation of TH expression in both PC12H and PC12L cells. In conclusion, our data suggested a novel function of rno-let-7d in regulation of galectin-3 and in ADHD development. Rno-let-7d, which is increased in the PFC of SHR brain, negatively regulated galectin-3, which is coupled with TH expression regulation.

Nuclear factor‐ kappa B expression and matrix metalloproteinase activity in hypertension

Our recent evidences suggest that inflammation of the spontaneously hypertensive rat (SHR) is associated with an uncontrolled matrix metalloproteinase (MMP) activity. We hypothesize that the transcription factor NF‐κB is overexpressed in the SHR and involved in the overexpression of MMPs.NF‐κB expression level and translocation were compared between Wistar Kyoto rat and SHR kidney, heart, and brain. In addition, the animals were treated with a NF‐κB inhibitor, pyrrolidine dithiocarbamate (PDTC), for ten weeks. To determine the correlation between NF‐κB and MMP, MMP‐2 and −9 activities were examined after treatment.Immunohistochemistry showed that NF‐κB expression is significantly increased in SHR in renal glomerular and tubulointerstitial areas and brain hypothalamus compared to that in WKY (p&lt;0.05), but not in myocardium and cerebral cortex. Significant NF‐κB translocation into the nucleus was found in renal tubular cells, cardiac muscle cells, and arterioles of kidney and brain in SHR.After PDTC treatment, the systolic blood pressure in SHR was suppressed by 36 % as compared to no significant difference in WKY rats. NF‐κB expression level in treated‐SHR was also decreased in renal glomerular and tubulointerstitial areas and hypothalamus compared to non‐treated animals (p &lt;0.05). Furthermore, MMP‐2 activity was significantly reduced by PDTC treatment in both WKY and SHR. The enhanced MMP‐9 activity in SHR was also significantly decreased by PDTC (p &lt;0.01). These results suggest NF‐κB is an important transcription factor in the SHR and may be responsible for its enhanced MMP activity.Supported by HL 10881

Expression of aquaporins 1 and 4 in the brain of spontaneously hypertensive rats

Aquaporins (AQP) 1 and 4 are water channel proteins localized respectively at the level of the blood–cerebrospinal fluids (CSF) and blood brain (BBB) barriers. These barriers represent the sites of exchange between blood and nervous tissue and between blood, choroid plexus and CSF in brain ventricles respectively. Damage of these barriers may alter transfer of substances between blood and nervous tissue. In spontaneously hypertensive rats (SHR) chronic hypertension may induce BBB dysfunction and pronounced defects in the integrity of the blood–CSF barrier. AQP1 is expressed in the apical membrane of choroid plexus epithelium. AQP4 is expressed by astrocyte foot processes near blood vessels. The present study has assessed the expression of AQP1 and AQP4 in the brain of SHR in pre-hypertensive (2 months of age), developing hypertension (4 months of age) and established hypertension (6 months of age) stages. Age-matched Wistar–Kyoto (WKY) rats were used as normotensive reference group. AQP1 expression is increased in choroid plexus epithelium of 6-month-old SHR. An increased expression of AQP4 was found in frontal cortex, striatum, and hippocampus of 4- and 6-month-old SHR compared to younger cohorts and age-matched WKY rats. These findings suggest that the increase in AQP expression may alter fluid exchange in BBB and/or in blood–CSF barrier. This situation in case of an acute or excessively elevated rise of blood pressure can promote BBB changes causing the brain damage occurring in this animal model of hypertension.

Effects after multiple administration of alcohol on some biochemical parameters in liver and brain in spontaneously hypertensive rats (SHR)

Event Abstract Back to Event Effects after multiple administration of alcohol on some biochemical parameters in liver and brain in spontaneously hypertensive rats (SHR) Rumyana Simeonova1, Vessela Vitcheva1* and Mitka Mitcheva1 1 Medical University-Sofia, Departement of Pharmacology, Pharmacotherapy and Toxicology, Bulgaria Introduction. Spontaneously hypertensive rats (SHR) are a genetic model of essential hypertension in human. They are used from many laboratories to characterize some aspects of this pathology, including an influence of some toxic compounds, including alcohol. The aim of the following study was to investigate the effects of multiple alcohol administration on some biochemical brain and hepatic parameters in SHRs, compared to normotensive Wistar-Kyoto rats (WKY). Methods. The parameters: malondialdehyde (MDA), reduced glutathione (GSH) and neuronal nitric oxide synthase (nNOS) were determined in livers and brains from both strains spectrometrically. For the experiment male SHR (strain Okamoto-Aoki, n=12) and WKY (n=12), were used. The animals of each strain were divided into 2 groups: control group and group treated with alcohol (3g/ kg, p.o., 14 days). Results: GSH level was lower in the liver (29%) and in the brain (35%) of SHR, compared to the NTRs, and ethanol administration decreased it in both strains in investigated organs. MDA level was increased in SHR significantly in liver by 38% and in brain by 27% and ethanol administration enhance this parameter in both strains, in both organs, but more pronounced in NTR. nNOS activity was significantly increased in SHR by 31%, compared to NTR and alcohol didn’t affect this parameter in SHR. Conclusions. The difference in the effect of ethanol between the strains might be due to the changed reactivity and pathophysiological characteristics of the hypertensive model. Keywords: alcohol, Ethanol, Malondialdehyde, Glutathione, nNOS, hypertensive models Conference: 8th Southeast European Congress on Xenobiotic Metabolism and Toxicity - XEMET 2010, Thessaloniki, Greece, 1 Oct - 5 Oct, 2010. Presentation Type: Poster Topic: Xenobiotic toxicity Citation: Simeonova R, Vitcheva V and Mitcheva M (2010). Effects after multiple administration of alcohol on some biochemical parameters in liver and brain in spontaneously hypertensive rats (SHR). Front. Pharmacol. Conference Abstract: 8th Southeast European Congress on Xenobiotic Metabolism and Toxicity - XEMET 2010. doi: 10.3389/conf.fphar.2010.60.00197 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 28 Oct 2010; Published Online: 04 Nov 2010. * Correspondence: Dr. Vessela Vitcheva, Medical University-Sofia, Departement of Pharmacology, Pharmacotherapy and Toxicology, Sofia, Bulgaria, vesselavitcheva@yahoo.com Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Rumyana Simeonova Vessela Vitcheva Mitka Mitcheva Google Rumyana Simeonova Vessela Vitcheva Mitka Mitcheva Google Scholar Rumyana Simeonova Vessela Vitcheva Mitka Mitcheva PubMed Rumyana Simeonova Vessela Vitcheva Mitka Mitcheva Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Cortical and Putamen Age-Related Changes in the Microvessel Density and Astrocyte Deficiency in Spontaneously Hypertensive and Stroke-Prone Spontaneously Hypertensive Rats

Cerebral small vessel disease (SVD) is a major contributor to dementia in the elderly, and hypertension represents a major cause for developing the disease. However, little is known about its development and progression. Modifications of large cerebral arteries due hypertension are thought to participate to the development of small ischemic infarcts, but the status of the small vessels before the establishment of hypertension is not well defined. Using spontaneously hypertensive rats (SHR) and stroke-prone SHR (SP-SHR) as a models for SVD, we analysed the effect of hypertension on the microvasculature in the cortex and putamen, and on its relationship with astrocytes in animals aged 2 to 9 months. Compared with the normotensive Wistar-Kyoto rats (WKY), the densities of the collagen type IV-positive capillaries were significantly higher in both brain areas of young SHR and SP-SHR. In contrast, the expression of the astrocytic marker GFAP was significantly lower in these animals, whereas astrogliosis was observed after 6 months in their cortex only. To investigate if chronic hypoxia occurs due to the lower number of astrocytes in young SHR and SP-SHR, we evaluated the levels of HIF-1alpha in both brain regions. The accumulation of HIF-1alpha was not observed at the youngest ages, but was apparent in neurons of 9-month-old SHR and SP-SHR. Our results indicate that the brains of young SHR and SP-SHR rats show evidence of cellular imbalance between microvessels and astrocytes at the neurovascular unit that may lead to their higher vulnerability to hypoxic events at older ages.

Assessment of the effect of fermented papaya preparation on oxidative damage in spontaneously hypertensive rat brain using electron spin resonance (ESR) imaging and L-band ESR spectroscopy

Oxidative and nitrosative stress mechanisms are widely implicated in the biological and pathological processes involved in aging, cardiovascular and neurodegenerative diseases. Although this has continued to fuel suggestions of the benefits of antioxidant functional foods, in vivo methods for assessing the integrity of this remain limited. A novel electron spin resonance (ESR) technique for evaluating oxidative stress and location of its damage in the brain of spontaneously hypertensive rats (SHR) has been described [Lee, M.-C., et al. (2004). Assessment of oxidative stress in the SHR brain using electron spin resonance (ESR) imaging and in vivo L-Band ESR. Hypertension Research, 27, 485–492]. The reconstructed 2D ESR images of the distribution of a blood brain barrier-permeable nitroxyl spin probe, 3-methoxycarbonyl-2,2,5,5-tetramethylpyrrolidine-1-oxyl (MC-PROXYL) was used to investigate the ability of fermented papaya preparation (FPP, a product of yeast fermentation of Carica papaya Linn.) to modulate oxidative stress of SHR brain. Supplementation (5–7 months) with FPP (50 mg/rat/day) significantly increased the decay of the ESR images of the MC-PROXYL, suggesting that FPP may have up-regulated the redox defense activity in the SHR brain. Herein is an in vivo noninvasive technique for the study of oxidative stress and its modulation by dietary factors (that may be intended for applications as neuroprotectants in chronic degenerative disease involving loss of brain function).

The cerebrovascular permeability to protein after bicuculline and amphetamine administration in spontaneously hypertensive rats. Evidence for increased resistance of pressure-induced blood-brain barrier dysfunction.

Hemodynamic and morphometric studies on spontaneously hypertensive rats (SHR) have suggested that the lumen to wall ratio of the resistance vessels is decreased due to hypertrophy of the media. As the tension in the vessel wall increases with the pressure and the radius and decreases with the vessel wall thickness, such vessels would be expected to withstand higher pressure than vessels of normotensive rats (NR) unless local wall defects were present. Bicuculline-induced epileptic activity and amphetamine intoxication, two conditions with cerebral vasodilatation and high blood pressure, give rise to extensive protein leakage in the brain of NR. Consistent with the hypothesis presented above, less protein leakage occurred in brains of SHR than in brains of NR after amphetamine and bicuculline administration.

Immunohistochemical evaluation of microvascular rarefaction in hypertensive humans and in spontaneously hypertensive rats

No data are presently available about changes in capillary density in the skeletal muscle and in the brain of spontaneously hypertensive rats (SHR) in relation to the development of hypertension. We have investigated 4 week-old and 12 week-old SHR and age-matched normotensive Wistar-Kyoto controls (WKY). Microvessel density (MVD) in the cerebral cortex and in a skeletal muscle were evaluated in sections stained for CD31. We also evaluated MVD in the dermal tissue of normotensive subjects and essential hypertensive patients. Subcutaneous small resistance arteries were dissected and mounted in a micromyograph and the media to lumen ratio (M/L) was measured. A significant reduction in MVD in the skeletal muscle and in the brain of SHR was clearly observed at 12 weeks of age, after the development of hypertension, but not at 4 weeks of age (pre-hypertensive condition). In hypertensive patients a significant reduction in the dermal MVD and an inverse correlation between M/L and MVD was observed. Our results suggest that, in the brain and skeletal muscle of adult SHR after the development of hypertension, and in the derma of adult essential hypertensive patients microvascular rarefaction may occur.

Vesicular Acetylcholine Transporter (VAChT) in the Brain of Spontaneously Hypertensive Rats (SHR): Effect of Treatment with an Acetylcholinesterase Inhibitor

The cholinergic marker vesicular acetylcholine transporter (VAChT) was investigated in different cerebral areas of spontaneously hypertensive rats (SHR) by immunochemistry (Western blot analysis) and by immunohistochemistry. SHR were used as an animal model of hypertensive brain damage. The sensitivity of manipulation of cholinergic system on VAChT was assessed in rats treated for four weeks with the acetylcholinesterase (AChE) inhibitor galantamine (3 mg/Kg/day). VAChT concentrations were increased in the brain of control SHR compared to age-matched normotensive Wistar-Kyoto rats. This increase probably represents an up-regulation of VAChT to oppose cholinergic deficits reported in SHR and is countered by galantamine administration. The possibility that cholinergic neurotransmission enhancement may represent a therapeutic strategy in cerebrovascular disease is discussed.

INFLUENCE OF GENDER ON OXIDATIVE STRESS, LIPID PEROXIDATION, PROTEIN DAMAGE AND APOPTOSIS IN HEARTS AND BRAINS FROM SPONTANEOUSLY HYPERTENSIVE RATS

1. Hypertension leads to oxidative stress, lipid and protein damage, apoptosis and impaired cardiac contractile function. However, impact of gender on these hypertension-associated abnormalities has not been elucidated. 2. The present study evaluated the oxidative stress, lipid/protein damage, apoptosis in heart and brain tissues as well as cardiomyocyte contractile function in Wistar Kyoto (WKY) and spontaneously hypertensive rats (SHR) of both genders. Oxidative stress, lipid peroxidation, protein damage and apoptosis were assessed by glutathione (GSH) : reduced glutathione (GSSG) ratio, malondialdehyde (MDA) levels, protein carbonyl levels and caspase-3 activity, respectively. Cardiomyocyte contractile function was examined including peak shortening (PS), time-to-PS (TPS), time-to-90% relengthening (TR90) and maximal velocity of shortening/relengthening (+/-dL/dt). The SHR cardiomyocytes displayed reduced PS and +/-dL/dt compared with gender-matched WKY counterparts. Male but not female SHR cardiomyocytes possessed longer resting cell length, normal TPS and prolonged TR90. All mechanical parameters were comparable between male and female WKY rats with the exception of a higher TR90 in females. Hypertension did not significantly affect the GSH : GSSG ratio in the heart and brain tissues of either gender. Brain from female WKY rats displayed a reduced GSH : GSSG ratio. The MDA levels were unchanged and elevated, respectively, in SHR heart and SHR brain tissues from both genders. Protein carbonyl formation and caspase-3 activity were elevated in male but not female SHR hearts. Nonetheless, brain protein carbonyl level and caspase-3 activity were unaffected by hypertension or gender. 3. In summary, these results suggest that gender affects hypertension-associated oxidative stress, lipid and protein damage, apoptosis in heart and brain tissues and cardiomyocyte contractile function.

(Pro)renin receptor (PRR) expression in the spontaneously hypertensive rats (SHR) brain

The Brain renin-angiotensin system (RAS) stimulates neuronal AT1 receptors (AT1R) exerting cardiovascular effects. Although the cellular and physiological mechanism of neuronal AngII effects is well understood, how AngII is generated and delivered to the AT1R responsive neurons remains unknown. The recent discovery of PRR led us to hypothesize that PRR is a part of the tissue RAS responsible for generation of AngII in an autocrine manner in the brain. Thus, our objectives in this study were two-fold: determine if PRR is expressed in the brain and if its expression localized to neurons or glial cells. Real-time PCR with PRR specific primers and probe was used to quantity mRNA levels. Although, no significant difference in the PRR mRNAs were observed in the RVLM and cortex of SHR, its levels increased 2 fold (P<0.01) in the PVN of SHR. Neuronal and astroglial cells in primary culture from Wistar Kyoto rats (WKY) and SHR were used to compare expression level. PRR mRNA levels were 2.6 fold higher in WKY neuronal culture compared with astroglial culture. In addition, neuronal cells from the SHR brain showed a 50% increase in the PRR level compared to WKY controls. In contrast, no significant difference in the PRR level were observed in astroglial culture of WKY and SHR, these observations demonstrate that PRR is primary expressed in the neurons and its levels are increased in the SHR neuron. (supported by NIH grants HL33610 and HL76312)

High dietary alpha tocopherol improves attention deficit/hyperactivity (ADHD)‐like behavior in juvenile spontaneously hypertensive rats (SHR)

The juvenile SHR, a widely used animal model for ADHD, was previously shown to have low levels of alpha tocopherol (AT) in plasma and brain compared to Sprague-Dawley (SD) rats on the same diet. In this study we asked if AT levels are related to the behavior abnormalities associated with ADHD (inattention, hyperactivity, and impulsivity), using measures previously shown to distinguish these behaviors. Because AT might affect behavior by protection of important brain PUFAs such as arachidonic and docosahexaenoic acids (AA and DHA), fatty acid analysis was also done. Three groups of SHR (n=8 each) were fed synthetic AIN93 diets containing, respectively, 0, 50 and 250 U/kg AT acetate, and 8 SD rats were given the 50 U/kg AT diet. The diets were fed ad libitum from the age of 4 weeks. Behavior testing was done from 8 weeks to 12 weeks in conditioning cages equipped with infrared monitors. The animals were killed at 12 weeks. AT concentrations in plasma of SHRs increased with increasing dietary AT (1.47 ± 0.64, 56.3 ± 6.39, 69.4 ± 12.8 μM), approaching that of SDs (71.0 ± 12.1 μM). AT concentrations in brain followed a similar pattern (5.49 ± 0.54, 14.5 ± 0.69, 17.2 ± 1.06 μg/g compared to 18.6 ± 2.00 μg/g). Total n-3 PUFA in SHR plasma and the AA + DHA sum in SHR brain also increased in response to dietary AT. SHR behavior showed significant effects of dietary treatment. Measures of learning (attention), inhibition, and impulsivity improved with increasing AT. Total activity was not reduced by AT. Replenishment of AT in the SHR can improve behavior. Supported by the Purdue Research Foundation.

Role of protein kinase Cβ in phorbol ester-induced c- fos gene expression in neurons of normotensive and spontaneously hypertensive rat brains

We have previously demonstrated that pressure application of the protein kinase C (PKC) activator phorbol 12-myristate 13-acetate (PMA) onto some neurons in the anterior hypothalamic area of rats increases neural activity in vivo and that this PKC activation-induced increase of neural activity is enhanced in spontaneously hypertensive rats (SHR), an animal model for genetic hypertension. Activation of PKC increases expression of the c- fos gene, an important transcription factor and proto-oncogene thought to be a marker of neural activity. To evaluate PKC isoforms responsible for neural activation, we examined which isoforms of PKC are involved in the PKC activation-induced c- fos gene expression in neuronal cultures of Wistar rat and spontaneously hypertensive rat (SHR) brains. PMA increased c- fos gene expression in neuronal cultures of Wistar rat brain and the PMA-induced c- fos gene expression was inhibited by the PKC inhibitor 1-(5-isoquinolinesulfonyl)-2-methylpiperazine dihydrochloride (H-7). The PKCα,β,γ activator thymeleatoxin also increased c- fos gene expression, while the PKCδ,ε activator ingenol did not affect it. In addition, the PMA-induced c- fos gene expression was inhibited by PKCβantisense oligonucleotides (AON) but not by PKCα and PKCγAONs. In SHR brain neuronal cultures, the PMA-induced c- fos gene expression was enhanced as compared with that of Wistar Kyoto rats (WKY), while basal c- fos gene expression was almost the same in both neuronal cultures. The enhancement of PMA-induced c- fos gene expression in SHR brain cultures was abolished by PKCβAON. These findings suggest that in rat brain neuronal cultures, PMA increases c- fos gene expression via activation of PKC and that PKCβisoforms are partly involved in the PMA-induced c- fos gene expression. In neuronal cultures of SHR brain, it appears that the PMA-induced c- fos gene expression is also enhanced via PKCβ.