Brain Of Spontaneously Hypertensive Rats Research Articles

Effects of angiotensin II type 1 receptor blockade on the oxidative stress in spontaneously hypertensive rat tissues

The aim of this work was to investigate the production of oxidative damage in homogenized kidney, liver and brain of spontaneously hypertensive rats (SHR), as well as the involvement of angiotensin (Ang) II in this process. Groups of 12-week-old SHR and Wistar Kyoto rats (WKY) were given 10 mg/kg/day losartan in the drinking water during 14 days. Other groups of WKY and SHR without treatment were used as controls. The production of thiobarbituric acid reactive substances (TBARS), reduced glutathione (GSH) and the activity of the antioxidant enzymes catalase (CAT), superoxide dismutase (SOD) and glutathione peroxidase (Gpx) were determined. No significant difference in TBARS was observed between untreated SHR or WKY rats; GSH content was lower in the liver but higher in the brain of SHR compared to WKY rats. In tissues from the SHR group, SOD and Gpx activities were reduced, whereas CAT activity was slightly increased in kidney. TBARS levels did not change in WKY rats after losartan administration, but were reduced in SHR liver and brain. Losartan treatment decreased GSH content in WKY kidney, but increased GSH in SHR liver. The activity of the antioxidant enzymes was not modified by losartan in WKY rats; however, their activities increased in tissues from treated SHR. The lower activity of antioxidant enzymes in tissues from hypertensive rats compared to those detected in normotensive controls, indicates oxidative stress production. Ang II seems to play no role in this process in normotensive animals, although AT 1 receptor blockade in SHR enhances the enzymatic activity indicating that Ang II is implicated in oxidative stress generation in the hypertensive animals.

Modulation of Nitric Oxide Synthase Activity in Brain, Liver, and Blood Vessels of Spontaneously Hypertensive Rats by Ascorbic Acid: Protection from Free Radical Injury

End organ damage in essential hypertension has been linked to increased oxygen free radical generation, reduced antioxidant defense, and/or attenuation of nitric oxide synthase (NOS) activity. Ascorbic acid (AA), a water-soluble antioxidant, has been reported as a strong defense against free radicals in both aqueous and nonaqueous environment. In this study we examined the hypothesis that antioxidant ascorbic acid may confer protection from increased free radical activity in brain, liver, and blood vessels of spontaneously hypertensive rats (SHR). Male SHRs were divided into groups: SHR + AA (treated with AA, 1 mg/rat/day; for 12 weeks) or SHR (untreated). Wister-Kyoto rats (WKY) served as the control. Mean systolic blood pressure (SBP) in treated and untreated SHR was 145 ± 7 mmHg and 142 ± 8 mmHg, respectively. AA treatment prevented the increase in systolic blood pressure in SHR by 37 ± 1% (p < 0.05). NOS activity in the brain, liver, and blood vessels of WKY rat was 1.82 ± 0.02, 0.14 ± 0.003, and 1.54 ± 0.06 pmol citruline/mg protein, respectively. In SHR, total NOS activity was significantly reduced by 52 ± 1%, 21 ± 3%, and 44 ± 4%, respectively. AA increased NOS activity in brain, liver, and blood vessels of SHR from 0.87 ±.03, 0.11 ±.01, and 0.87 ±.08 pmol citruline/mg protein to 0.93 ± 0.01, 0.13 ± 0.001, and 1.11 ± 0.03 pmol citruline/mg protein (p < 0.05), respectively. Lipid peroxides in the brain, liver, and blood vessels from WKY rats were 0.87 ± 0.06, 0.11 ± 0.005, and 0.47 ± 0.04 nmol MDA equiv/mg protein, respectively. In SHR, lipid peroxides in brain, liver, and blood vessels were significantly increased by 40 ± 3%, 64 ± 3%, and 104 ± 13%, respectively. AA reduced lipid peroxidation in liver and blood vessels by 17 ± 1% and 34 ± 3% but not in brain. Plasma lipid peroxides were almost doubled in SHR (p < 0.01) together with a reduction in total antioxidant status (6 ± 0.1%; p < 0.05), nitrite (53 ± 2%; p < 0.05) and superoxide dismutase (SOD) activity (36 ± 2%; p < 0.05). AA treatment reduced plasma lipid peroxide (p < 0.001), and increased TAS (p < 0.001), nitrite (p < 0.001), and SOD activity (p < 0.001). From this study, we conclude that brain, liver, and blood vessels in SHR are susceptible to free radical injury, which reduces the availability of NO either by scavenging it or by reducing its production via inhibiting NOS. In addition, brain, liver, and blood vessels in SHR; may be protected by antioxidant, which improves total antioxidant status, and SOD thus may prevent high blood pressure and its complications.

Increased Expression of Glial Fibrillary Acidic Protein in the Brain of Spontaneously Hypertensive Rats

Astrogliosis, consisting in astroglial proliferation and increased expression of the specific cytoskeletal protein glial fibrillary acid protein (GFAP) is common in several situations of brain damage. Arterial hypertension, which induces cerebrovascular changes, can cause also brain damage, neurodegeneration and dementia (vascular dementia). This study was designed to assess astroglial reaction in different brain areas (frontal cortex, occipital cortex, hippocampus and striatum) of spontaneously hypertensive rats (SHR) in the pre‐hypertensive phase (2 months of age), in the developing phase of hypertension (4 months of age) and in established hypertension (6 months of age). SHR were compared to age‐matched normotensive Wistar‐Kyoto (WKY) rats. Analysis included reverse transcription‐polymerase chain reaction (RT‐PCR) of GFAP mRNA, GFAP immunochemistry (Western blot analysis) and immunohistochemistry. A significant increase of GFAP mRNA and an increase of GFAP immunoreactivity were noticeable in different brain areas of SHR compared to normotensive WKY rats at 6, but not at 2 or 4 months of age. Immunohistochemistry revealed a numerical augmentation (hyperplasia) and an increase in size (hypertrophy) of GFAP‐immunoreactive astrocytes in frontal cortex, occipital cortex and striatum of SHR. In the hippocampus of SHR only a numerical increase of GFAP‐immunoreactive astrocytes was found. These finding demonstrating the occurrence of astrogliosis in the brain of SHR with established hypertension suggest that hypertension induces a condition of brain suffering enough to increase biosynthesis and expression of GFAP similarly as reported in several neurodegenerative disorders and in brain ischemia.

Increased brain uptake and CSF clearance of 14C-glutamate in spontaneously hypertensive rats

Blood-to-brain and blood-to-CSF transport kinetics of 14C-glutamate in the spontaneously hypertensive rats (SHR) were studied using the in situ brain perfusion technique. Also, clearance of 14C-glutamate from CSF of SHR was studied using the ventriculo-cisternal (VC) perfusion technique. Blood-to-brain and blood-to-CSF transport kinetics showed greater rate of maximal transport into both brain and CSF of SHR compared to normotensive Wistar Kyoto (WKY) rats ( p>0.05). Uptake into CSF of WKY and uptakes into brains of WKY and SHR did not show any significant diffusion ( K d) of 14C-glutamate ( p<0.05). However, some diffusion of 14C-glutamate only into CSF of SHR was observed, 0.031+0.006 μl min −1 g −1. Clearance of 14C-glutamate from CSF was greater in the SHR (28.33±6.9 μl min −1) compared to that in WKY rats (19.42±4.7 μl min −1). However, 14C-glutamate uptake by brain from CSF side was not significantly different between SHR and WKY rats ( p>0.05). These results suggest that the greater blood-to-brain and blood-to-CSF entry of 14C-glutamate during hypertension may be balanced by greater removal of 14C-glutamate from CSF back to blood.

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Symposium

Effects of simultaneous administration of EPA, DHA, Taurine and peptide from muscle of Pacific mackerel on blood pressure and serum lipids in SHR fed hypercholesterolemic diets

This study investigated the effects of the simultaneous administration of eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA) and taurine (Tau) on the blood pressure, serum lipids, free amino acid and fatty acids of the brain of SHR (spontaneously hypertensive rats) fed hypercholesterolemic diets.In addition, we examined the relationship between the angiotensin- I converting enzyme (ACE) activity and the blood pressure by the simultaneous administration of these components and peptide from the muscle of Pacific mackerel. Moreover, we examined the effects on the blood pressure and serum lipids when replacing casein with defatted sardine meal as the protein source.The results obtained are summarized as follows:1. With the simultaneous administration of EPA, DHA and Tau for SHR, an improvement in the serum lipids and the inhibitive action of blood pressure elevation were recognized.2. Compared with the control, a significant inhibitive action on the elevation of ACE activity and the blood pressure were recognized by the simultaneous administration of EPA, DHA, Tau and peptide.3. When replacing a casein with the defatted serdine meal protein as the protein source, a significant inhibitive action on the elevation was recognized with regard to the serum lipids, but not with regard to ACE activity and the blood pressure compared with the control.4. By simultaneous administration of these components, the Tau content showed a small increase compared to the control. Also, a large difference was not observed with regard to the fatty acid composition in the brain, but the absolute quantity of DHA increased in the I and IV groups, and a remarkable increase was recognized in the II group compared with the control.

Angiotensinogen gene-activating elements regulate blood pressure in the brain.

Although the angiotensinogen gene is a possible candidate as a determinant of hypertension, the molecular mechanisms of tissue angiotensinogen gene regulation have yet to be clarified. We identified essential transcription regulators of angiotensinogen production in the central nervous system using synthetic double-stranded oligodeoxynucleotides (ODNs) as "decoy" cis elements to block the binding of nuclear factors to promoter regions of the targeted gene. Using a gel mobility shift assay, angiotensinogen gene-activating element (AGE) 2 binding protein was detected in the brain nuclear extracts of both spontaneously hypertensive rats (SHRs) and normotensive Wistar Kyoto rats (WKYs). Importantly, the binding activity of AGE 2 and angiotensinogen mRNA level were significantly higher in the brain of SHRs than in that of WKYs. Using the decoy approach, we demonstrated a significant decrease in the blood pressure of SHRs by transfection of AGE 2 decoy, but not mismatched, ODNs into the lateral cerebroventricle, accompanied by a significant decrease in brain angiotensinogen concentration and mRNA, and angiotensin II level. That these effects, demonstrated herein, are due to central effects is confirmed by the fact that no changes in circulating levels of angiotensinogen or angiotensin II concentrations were observed. Notably, AGE 2 decoy ODNs did not decrease the blood pressure of WKYs. We conclude that the abnormal expression of AGE 2 binding protein in the central nervous system plays a crucial role in high blood pressure of a genetically hypertensive rat model.

Role of Phosphatidylinositol 3-Kinase in Angiotensin II Regulation of Norepinephrine Neuromodulation in Brain Neurons of the Spontaneously Hypertensive Rat

Chronic stimulation of norepinephrine (NE) neuromodulation by angiotensin II (Ang II) involves activation of the Ras-Raf-MAP kinase signal transduction pathway in Wistar Kyoto (WKY) rat brain neurons. This pathway is only partially responsible for this heightened action of Ang II in the spontaneously hypertensive rat (SHR) brain neurons. In this study, we demonstrate that the MAP kinase-independent signaling pathway in the SHR neuron involves activation of PI3-kinase and protein kinase B (PKB/Akt). Ang II stimulated PI3-kinase activity in both WKY and SHR brain neurons and was accompanied by its translocation from the cytoplasmic to the nuclear compartment. Although the magnitude of stimulation by Ang II was comparable, the stimulation was more persistent in the SHR neuron compared with the WKY rat neuron. Inhibition of PI3-kinase had no significant effect in the WKY rat neuron. However, it caused a 40-50% attenuation of the Ang II-induced increase in norepinephrine transporter (NET) and tyrosine hydroxylase (TH) mRNAs and [3H]-NE uptake in the SHR neuron. In contrast, inhibition of MAP kinase completely attenuated Ang II stimulation of NET and TH mRNA levels in the WKY rat neuron, whereas it caused only a 45% decrease in the SHR neuron. However, an additive attenuation was observed when both kinases of the SHR neurons were inhibited. Ang II also stimulated PKB/Akt activity in both WKY and SHR neurons. This stimulation was 30% higher and lasted longer in the SHR neuron compared with the WKY rat neuron. In conclusion, these observations demonstrate an exclusive involvement of PI3-kinase-PKB-dependent signaling pathway in a heightened NE neuromodulatory action of Ang II in the SHR neuron. Thus, this study offers an excellent potential for the development of new therapies for the treatment of centrally mediated hypertension.

Angiotensin II-induced phosphorylation of the AT1 receptor from rat brain neurons.

The neuronal angiotensin II (Ang II) type 1 (AT1) receptor is coupled to the Ras-Raf-1-mitogen-activated protein (MAP) kinase signal-transduction pathway (Yang H, Lu D, Yu K, Raizada MK. Regulation of neuromodulatory actions of angiotensin II in the brain neurons by the Ras-dependent mitogen-activated protein kinase pathway. J Neurosci. 1996;16:4047-4058). In this study we compared the effects of angiotensin II (Ang II) on AT1 receptor phosphorylation and the ability of the phosphorylated receptor to bind Ang II in neuronal cultures of Wistar-Kyoto rat (WKY) and spontaneously hypertensive rat (SHR) brains to further our understanding of the Ang II signaling mechanism. Ang II caused a time-dependent phosphorylation of AT1 receptors in both WKY and SHR brain neurons. The level of phosphorylation was higher in the SHR brain neurons; this finding was consistent with increased AT1 receptors in these cells. MAP kinase was involved in this phosphorylation, a conclusion supported by the following evidence: (1) exogenous MAP kinase phosphorylated the AT1 receptor; (2) PD98059, a MAP kinase kinase inhibitor, attenuated Ang II-stimulated AT1 receptor phosphorylation; and (3) MAP kinase and AT1 receptors were coimmunoprecipitated in Ang II-stimulated neurons. Finally, MAP kinase phosphorylation was associated with the loss of 125I-[Sar1-Ile8]-Ang II binding ability of the AT1 receptor in both strains of neurons. These observations show that Ang II stimulates phosphorylation of the neuronal AT1 receptor by a mechanism involving MAP kinase and that the phosphorylated neuronal AT1 receptor does not exhibit Ang II binding activity in the brains of either WKY or SHR.

Regulation of preprotachykinin-A mRNA in genetic hypertensive and normotensive rats

It is well-known that central administration of tachykinins (Tks) inhibit salt intake in rats. Recent studies have shown that conditions that arouse salt appetite, such as adrenalectomy and sodium depletion, induce a decrease in preprotachykinin-A (PPT-A) mRNA in discrete regions of the rat brain, suggesting that reduced levels of PPT-A mRNA in the brain may have a permissive role on the expression of salt appetite. It has also been shown that spontaneously hypertensive rats (SHR) show higher avidity for salty solutions than their normotensive control Wistar–Kyoto (WKY) rats. In this regard, the present study tested whether SHR and WKY rats differ in expression of the gene coding for PPT-A, the precursor for Tks peptides. Using semi-quantitative in situ hybridization histochemistry, we examined the level of PPT-A mRNA in discrete rat brain regions of SHR and WKY rats under no treatment, after 1 or 3 days of Na + depletion. Levels of PPT-A mRNA were analysed in the olfactory tubercle (Tu), in the lateral olfactory tubercle (LOT), in the dorsal and ventral caudate putamen (d/v CPu), in the medial preoptic area (mPOA), in the bed nucleus of the stria terminalis (BNST), in the habenula (Hb) and in the postero-dorsal part of the amygdala (MePD). Semi-quantitative analysis of silver grains revealed a 27.5% lower expression of the PPT-A mRNA levels in SHR opposite to WKY rats under no treatment in v-CPu, mPOA, BNST and Hb. 1 day of Na + depletion reduced PPT-A mRNA levels when opposite to Na +-repleted animals in Tu and mPOA in both SHR and WKY rats. On the other hand, when comparing SHR and WKY rats after 1 day of Na + depletion, a 26% lower level of PPT-A mRNA was detected in Tu and d-CPu of SHR opposite to WKYrats whereas a 14% and an 18% lower level was detected in v-CPu and Hb, respectively. A lower expression of PPT-A mRNA in SHR compared to WKY rats was also found in BNST and MePD, although no statistical significance was detected in these two brain areas. In the last experiment, 3 days of Na + depletion reduced PPT-A mRNA levels in mPOA while negligibly increased mRNA levels in d-CPu and v-CPu, in BNST, Hb and MePD, both in SHR and WKY rats. Conversely, when making comparisons between the two strains, a 35% lower level of PPT-A mRNA in SHR with respect to WKY rats was found after 3 days of Na + depletion in d-CPu, v-CPu and mPOA. A lower gene expression, even though not statistically significant, was found in Tu, LOT, MePD. These findings show a consistent difference of PPT-A mRNA levels in discrete regions of the SHR brain opposite to WKY rats and confirm that 1 day of Na + depletion reduces PPT-A mRNA in discrete brain regions. Since SHR are notoriously more salt-avid than WKY rats and Tks are potent inhibitors of sodium intake, the down-regulation of PPT-A mRNA may contribute to the higher natriophilia and, therefore, to the etiology of the hypertensive disease.

Genetic analysis of renin gene expression in the central nervous system of spontaneously hypertensive rats

Renin expression in the central nervous system in spontaneously hypertensive rats (SHR) has been reported to be higher than that in age-matched Wistar–Kyoto (WKY) rats. In the present study, we examined the mechanisms of enhancement of renin mRNA concentration in the brainstem of SHR using F 2 hybrid rats derived from cross-breeding SHR with WKY. The SHR allele of the renin gene was significantly associated with higher brain renin mRNA concentration in both 10- and 24-week-old F 2 populations. However, neither the genotype of the renin gene nor the brain renin mRNA concentration was associated with blood pressure or urinary volume. Thus, the higher renin mRNA concentration in the brainstem of SHR was indicated to be genetically determined by the genotype of the renin gene. Further study is required to clarify the pathophysiological meaning of this increased renin expression in the brain of SHR.

Age-related changes in the DNA and RNA content of the brain in spontaneously hypertensive rats.

To elucidate the effects of aging accompanied with hypertension on brain nucleic acid, we measured both the DNA and RNA contents of six specific brain regions in adult (5-6 months old) and aged (18-22 months old) female spontaneously hypertensive rats (SHRs). Although no statistical difference was observed in the RNA content, the DNA content did tend to increase in the hippocampal CA1 of aged SHR (4.24 +/- 0.55 ng/microgram protein, mean +/- SD, n = 6) in comparison to that of adult SHR (3.21 +/- 0.71 ng/microgram protein, n = 4). Hence, aged SHRs showed a significant decrease in the RNA to DNA ratio in the CA1 subfield of the hippocampus (3.79 +/- 0.61) compared to adult SHR (5.27 +/- 0.81). On the other hand, no other regions, except for the dorsolateral region of the striatum, showed any difference in the RNA/DNA ratio between aged and adult SHR. We therefore conclude that subtle changes in the nucleic acid occur in vulnerable regions of the brain in aged SHRs.

Angiotensin II regulation of tyrosine hydroxylase gene expression in the neuronal cultures of normotensive and spontaneously hypertensive rats.

In the present study we investigated the regulation of tyrosine hydroxylase (TH) by angiotensin II (Ang II) in an attempt to provide cellular and molecular evidence that this hormone has increased neuromodulatory actions in the spontaneously hypertensive (SH) rat brain. Neuronal cells in primary culture from the hypothalamus-brain stem of both normotensive [Wistar-Kyoto (WKY)] and SH rats have been used. These cultures mimic in vivo situations. Ang II caused a time-dependent increase in TH activity in WKY rat brain neurons. A maximal increase of 2.5-fold was observed with 100 nM Ang II in an actinomycin- and cycloheximide-dependent process. In addition, Ang II caused a parallel increase in TH messenger RNA (mRNA) levels, with a maximal stimulation of 5-fold in 4 h by 100 nM Ang II in WKY rat brain neurons. The stimulation of TH mRNA was mediated by the AT1 receptor subtype, resulted from an increase in its transcription, and involved activation of phospholipase C and protein kinase C. Antisense oligonucleotide for c-fos attenuated Ang II stimulation of TH mRNA in a time- and dose-dependent fashion, indicating an involvement of c-fos as a putative third messenger in Ang II stimulation of TH. Ang II also caused stimulation of TH activity and its mRNA levels in neuronal cultures of SH rat brain by a mechanism similar to that observed for neuronal cultures of WKY rat brain, involving AT1 receptors, protein kinase C, and c-fos. However, the stimulation of TH activity and that of TH mRNA were approximately 30% and 80% higher, respectively, in the SH rat brain neurons than those in the WKY rat brain neurons. In vivo experiments have been carried out to validate the elevated response of TH gene expression to Ang II in SH rat brain neuronal cultures. Ang II stimulated both TH activity and TH mRNA levels in the hypothalami and brain stems of adult WKY and SH rats. The level of stimulation in the brain of the SH rat was significantly higher than that in the WKY rat. These observations are consistent with an increase in AT1, receptor gene expression and suggest that increased TH gene expression could be the cellular/molecular basis for the greater neuromodulatory action of Ang II in the SH rat brain.

Angiotensin II regulation of plasminogen activator inhibitor-1 gene expression in neurons of normotensive and spontaneously hypertensive rat brains.

Neuronal cells in primary culture from the hypothalamus-brain stem areas of normotensive [Wistar-Kyoto (WKY)] and spontaneously hypertensive (SH) rat brains have been used in the present study to investigate an interaction between the brain renin-angiotensin II system and the plasminogen activator system. This is an attempt to further our understanding of the role of brain Ang II in the control of neuronal development and differentiation through its regulation of the extracellular matrix. Ang II caused a 10-fold stimulation of plasminogen activator inhibitor-1 (PAI-1) messenger RNA (mRNA) in WKY rat brain neuronal cultures. The stimulation was mediated by the AT1 receptor subtype and was accompanied by an increase in PAI-1 gene transcription and the synthesis of cellular PAI-1 protein. The stimulation involved activation of protein kinase C, and alterations in the intracellular Ca2+ pool caused a significant inhibition of Ang II stimulation of PAI mRNA. Ang II stimulation of PAI-1 mRNA succeeded its action on c-fos mRNA and was attenuated by c-fos antisense oligonucleotide. Although PAI-1 gene expression was also stimulated by Ang II in neuronal cultures of SH rat brain, two differences between WKY and SH rat brain neurons were observed: 1) the level of Ang II stimulation in SH rat neurons was 50% of that in WKY rat neurons; and 2) Ang II stimulation of c-fos was 2.4-fold higher in SH neurons than in WKY neurons, but c-fos antisense oligonucleotide did not attenuate the stimulatory action of Ang II on PAI-1 mRNA in SH neurons. These observations suggest that the changes in the Ang II-mediated signaling pathways and/or the regulatory region(s) of the PAI-1 gene may contribute to the differential actions of Ang II in WKY and SH rat brain neurons.

Regulation of norepinephrine transport system by angiotensin II in neuronal cultures of normotensive and spontaneously hypertensive rat brains.

Brain angiotensin II (Ang II) plays a key role in blood pressure control in part by interacting with catecholamines (CA) and by stimulation of sympathetic pathways. The significance of Ang-CA interaction is further heightened by the presence of a hyperactive brain Ang II system in spontaneously hypertensive (SH) rat, a genetic model for essential hypertension. Neuronal cells in primary culture from the hypothalamus-brainstem that mimic in vivo situations in so far as many cellular actions of Ang II are concerned, have been used in the present study to elucidate Ang II regulation of CA by determining its cellular action on the norepinephrine transporter (NET) system. Ang II causes both acute and chronic stimulation of [3H]-norepinephrine (NE) uptake in neuronal cultures of Wistar Kyoto (WKY) rat brain. Acute stimulation begins as early as 5 min, reaches maximal levels in about 30 min in the presence of 100 nM Ang II, and is blocked by losartan, a specific antagonist for AT1 receptor subtype. In addition, this acute stimulation appears to be a posttranscriptional event and does not involve protein kinase C (PKC) or NET gene transcription. Chronic stimulation of [3H]-NE uptake by Ang II persists throughout the duration of Ang II incubation (24 h), is dose dependent, and is also mediated by AT1 receptor subtype. However, chronic stimulation of [3H]-NE uptake involves PKC, cfos, and NET gene transcription. Ang II also stimulates [3H]-NE uptake in neuronal cultures of SH rat brain, both acutely and chronically, by mechanisms similar to those observed in neuronal cultures of WKY rat brain. The stimulation of NET by Ang II is 2-fold higher than that seen in WKY and is consistent with increased AT1 receptor gene transcription and increased functional AT1 receptors in SH rat brain neurons compared with WKY rat brain neurons. The Ang II stimulation of the NET system is also higher in adult SH compared with WKY rats in vivo. These observations show that 1) Ang II stimulates the NET system both acutely and chronically, the former involving activation of preexisting transporters and the latter involving NET gene transcription and translation; and 2) Ang II stimulation of the NET system is elevated in SH rat brain neurons.

Converting enzyme inhibition and its withdrawal in spontaneously hypertensive rats.

When spontaneously hypertensive rats (SHR) treated at a young age with an inhibitor of angiotensin-converting enzyme (ACE) are withdrawn from treatment, their blood pressure (BP) remains below that of untreated rats. We examined the effects of ACE inhibitor treatment and its withdrawal on angiotensin-(1-7) [Ang-(1-7)], angiotensin II (Ang II) and angiotensin I (Ang I) in plasma, kidney, adrenal, heart, aorta, brown adipose tissue, lung, and brain of male SHR and normotensive Donryu rats. Rats were administered either vehicle or perindopril (3 mg/kg/day) from 6 to 10 weeks, from 6 to 20 weeks, and from 6 to 10 weeks, followed by perindopril withdrawal from 10 to 20 weeks. Angiotensin peptides and plasma levels of renin, angiotensinogen, ACE, and aldosterone were measured at 10 and 20 weeks of age. Perindopril reduced BP of both SHR and Donryu rats, although only SHR showed a reduction of BP of 19 mm Hg after perindopril withdrawal, associated with a reduction of 5% in heart weight/body weight ratio. Perindopril reduced the angiotensin II/angiotensin I ratio in all tissues by > 50%, with strain- and tissue-specific differences in the effects of perindopril on the levels of individual angiotensin peptides. None of the changes in Ang II levels persisted after perindopril withdrawal. In contrast to those of Donryu rats, plasma angiotensinogen levels of perindopril-withdrawn SHR were 14% lower than those of vehicle-treated SHR (p = 0.0356). Although the lower BP of perindopril-withdrawn SHR was not associated with an alteration in Ang II levels, the suppressed plasma angiotensinogen levels may have contributed to the lower BP of these rats. Alternatively, another action of perindopril, such as a change in cardiovascular structure, may have been responsible for the reduced BP of perindopril-withdrawn SHR.

Angiotensin peptides in spontaneously hypertensive and normotensive Donryu rats.

The renin-angiotensin system has been implicated in the pathogenesis of hypertension in spontaneously hypertensive rats (SHR). Given that SHR may have normal or suppressed plasma levels of renin and angiotensin peptides, we examined whether the tissue levels of angiotensin peptides are elevated in these rats. We measured angiotensin-(1-7) [Ang-(1-7)], Ang II, and Ang I in plasma, kidney, adrenal, heart, aorta, brown adipose tissue, lung, and brain of male SHR and normotensive Donryu rats at 6, 10, and 20 weeks of age. SHR had higher blood pressures and ratios of heart weight to body weight at all ages. Plasma renin levels of SHR were 13% to 32% of the levels of Donryu rats. Although plasma angiotensin-converting enzyme activity was lower in SHR than in Donryu rats, lung was the only SHR tissue with a reduced Ang II-Ang I ratio. Ang II levels in SHR adrenal were 24% to 42% of the levels of Donryu adrenal, and for SHR plasma, aorta, brown adipose tissue, and lung, Ang II levels were 38% to 93% of the levels of Donryu rats. For kidney and heart, Ang II levels were similar in SHR and Donryu rats at 6 weeks of age although suppressed in SHR at 10 and 20 weeks. Moreover, brain Ang II levels were higher in SHR than Donryu rats at 6 weeks of age and similar at 10 and 20 weeks of age.(ABSTRACT TRUNCATED AT 250 WORDS)

Diminished brain synaptic plasma membrane ca 2+-atpase activity in spontaneously hypertensive rats: Association with reduced anesthetic requirements

We have recently reported that plasma membrane Ca 2+-ATPase ( PMCA) pumping activity in rat brain synaptic plasma membranes (SPM) was reduced by in vitro or prior in vivo exposure to inhalation anesthetics (IA). In addition, rats with streptozocin-induced diabetes were found to have diminished brain synaptic PMCA pumping and a decrease in the partial pressures of several IA required to prevent movement in response to stimulation, defined as the minimum effective dose or MED. Diminished PMCA activity in erythrocytes of spontaneously hypertensive rats (SHR) has been noted. Because PMCA is ubiquitous, it seemed possible that PMCA pumping might be decreased in the brain of SHR and perhaps associated with decreased IA requirement. Eighteen SHR and 18 control, normotensive Wistar-Kyoto rats (WKY) were studied. PMCA activity was assessed by measurement of Ca 2+ uptake into synaptic plasma membrane vesicles prepared from cerebrum and diencephalon-mesencephalon (D-M) in WKY and SHR. Ca 2+ pumping was significantly less in SHR than in WKY, 85% of control in the cerebrum and 90% in the D-M (p < 0.01). The MEDs for halothane, isoflurane and desflurane were also lower in SHR than in WKY, 91%, 90% and 89%, respectively, of control (p < 0.05). Thus, an animal model of primary hypertension (SHR) manifested diminished brain synaptic PMCA activity and reduced MED for several volatile anesthetics. These findings provide further evidence for a role for PMCA in anesthetic action.

Delivery of angiotensin II type 1 receptor antisense inhibits angiotensin action in neurons from hypertensive rat brain.

Increased brain angiotensin II (AII) type 1 receptor (AT1R) expression has been implicated in the hyperactive brain angiotensin system and the development and maintenance of hypertension in the genetically spontaneously hypertensive (SH) rat. Neuronal cells in primary culture from the cardioregulatory-relevant brain areas (hypothalamus/brainstem) mimic increased brain AT1R gene expression and AT1R function of the adult SH rat. They have been utilized in the present study to determine whether cellular actions of AII could be regulated by the transfer of AT1R antisense (AT1R-AS) with the use of a retroviral-mediated gene delivery system developed for the central nervous system cultures. AII stimulates norepinephrine (NE) uptake in neuronal cultures of both normotensive (Wistar Kyoto) and SH rat brains. This neuromodulatory action is mediated by the AT1R subtype, is significantly higher in SH neurons, and is associated with a parallel stimulation of mRNAs for c-fos and NE transporter. Infection of neuronal cultures with a retrovirus vector that contains AT1R-AS (LNSV-AT1R-AS) results in an inhibition of AT1R-mediated stimulation of both c-fos and NE transporter mRNA, as well as NE uptake in both strains of rats; however, the inhibition is more pronounced in SH neurons compared with Wistar Kyoto rat brain neurons. The higher sensitivity of the SH rat brain neurons is further supported by our observation that a certain dose of LNSV-AT1R-AS that fails to induce inhibition of cellular actions of AII in WKY neurons causes a significant inhibition of AII actions in SH neurons. These observations show that retrovirally mediated delivery of AT1R-AS could be used to selectively control the actions of AII in primary neuronal cultures from SH rat brain.

Regulation of angiotensin II type 1 receptor mRNA in neuronal cultures of normotensive and spontaneously hypertensive rat brains by phorbol esters and forskolin.

Neuronal cells in primary culture from the brains of normotensive, Wistar-Kyoto (WKY) rats and spontaneously hypertensive (SH) rats express angiotensin II type 1 (AT1) receptors. Treatment of WKY rat brain cultures with a phorbol ester, phorbol 12-myristate 13-acetate (PMA), causes a time- and dose-dependent increase in the levels of an approximately 2.3-kb AT1 receptor mRNA transcript. A maximal stimulation of 4.5-fold in the AT1 receptor mRNA transcript level is observed with 200 nM PMA in 4 h and is blocked by 1 microM staurosporine. Forskolin also increases the AT1 receptor mRNA levels in WKY rat brain neurons in a time- and dose-dependent manner, and a 4.5-fold stimulation is achieved with 50 microM forskolin in 4 h. The stimulatory effects of both PMA and forskolin are completely abolished by coincubation of neuronal cultures with 1 microM actinomycin D. In addition, nuclear run-on assay indicated an increase in the transcription of AT1 receptor mRNA in WKY rat brain neurons treated with either PMA or forskolin. Both PMA and forskolin also stimulate levels of AT1 receptor mRNA in neuronal cultures from brain of the SH rat. The degree of stimulation in these cultures is comparable to that in WKY rat brain neurons. These observations show that although the basal AT1 receptor gene expression is significantly higher in SH rat brain neurons compared with WKY rat brain neurons, the protein kinase C- and protein kinase A-responsive stimulation is not altered. These data suggest a possible involvement of protein kinase C and protein kinase A response elements in AT1 receptor gene expression.