Roselyn White

Role of central nervous system aldosterone synthase and mineralocorticoid receptors in salt-induced hypertension in Dahl salt-sensitive rats

In Dahl salt-sensitive (S) rats, high salt intake increases cerebrospinal fluid (CSF) Na(+) concentration ([Na(+)]) and blood pressure (BP). Intracerebroventricular (ICV) infusion of a mineralocorticoid receptor (MR) blocker prevents the hypertension. To assess the role of aldosterone locally produced in the brain, we evaluated the effects of chronic central blockade with the aldosterone synthase inhibitor FAD286 and the MR blocker spironolactone on changes in aldosterone and corticosterone content in the hypothalamus and the increase in CSF [Na(+)] and hypertension induced by high salt intake in Dahl S rats. After 4 wk of high salt intake, plasma aldosterone and corticosterone were not changed, but hypothalamic aldosterone increased by approximately 35% and corticosterone tended to increase in Dahl S rats, whereas both steroids decreased by approximately 65% in Dahl salt-resistant rats. In Dahl S rats fed the high-salt diet, ICV infusion of FAD286 or spironolactone did not affect the increase in CSF [Na(+)]. ICV infusion of FAD286 prevented the increase in hypothalamic aldosterone and 30 mmHg of the 50-mmHg BP increase induced by high salt intake. ICV infusion of spironolactone fully prevented the salt-induced hypertension. These results suggest that, in Dahl S rats, high salt intake increases aldosterone synthesis in the hypothalamus and aldosterone acts as the main MR agonist activating central pathways contributing to salt-induced hypertension.

Changes in cardiac ANG II postmyocardial infarction in rats: effects of nephrectomy and ACE inhibitors

We evaluated in rats the time course of changes in cardiac versus plasma ANG I and II postmyocardial infarction (MI) and the effects of nephrectomy and angiotensin-converting enzyme (ACE) inhibitors on the early changes post-MI. Acute coronary artery ligation was induced in conscious rats using the two-stage model, and plasma and cardiac tissue were obtained shortly (6 h, 1 and 3 days) and chronically (1, 4, and 8-9 wk) after MI. In an additional group of rats, bilateral nephrectomy was performed 18 h before the coronary artery ligation, and samples were obtained at 6 h post-MI. Furthermore, in two additional groups of rats, treatment with enalapril and quinapril was started 3 days before the ligation, and samples were obtained at 1 or 3 days post-MI. In these groups of rats, plasma and left ventricular (LV) (infarct and infarct free) ANG I and II were measured by RIA after separation on HPLC. In control rats, plasma ANG I and II showed a clear increase at 6 h post-MI but subsequently only minor increases were observed. In contrast, LV ANG II showed major increases at 6 h and 1 day post-MI, which had returned to normal by 3 days in the infarct-free LV and after 1(-2) wk in the infarct LV. LV ANG I showed a more gradual increase and remained elevated in the infarct up to 8-9 wk. Nephrectomy preceding the MI lowered ANG I and II in plasma but enhanced their increases in the heart at 6 h post-MI. Both ACE inhibitors decreased plasma ANG II associated with large increases in plasma ANG I. They also inhibited the increases in LV ANG II in both the infarct and infarct-free LV at 1 and 3 days post-MI with however no significant increase in LV ANG I. In conclusion, induction of a MI in conscious rats leads to rapid and marked, but only short-lived, increases in cardiac tissue ANG II in both the infarct and infarct-free parts of the LV. Pretreatment with ACE inhibitors, but not nephrectomy, blocks this increase. Local production appears to play a major role in the increases in cardiac ANG II post-MI.We evaluated in rats the time course of changes in cardiac versus plasma ANG I and II postmyocardial infarction (MI) and the effects of nephrectomy and angiotensin-converting enzyme (ACE) inhibitors on the early changes post-MI. Acute coronary artery ligation was induced in conscious rats using the two-stage model, and plasma and cardiac tissue were obtained shortly (6 h, 1 and 3 days) and chronically (1, 4, and 8-9 wk) after MI. In an additional group of rats, bilateral nephrectomy was performed 18 h before the coronary artery ligation, and samples were obtained at 6 h post-MI. Furthermore, in two additional groups of rats, treatment with enalapril and quinapril was started 3 days before the ligation, and samples were obtained at 1 or 3 days post-MI. In these groups of rats, plasma and left ventricular (LV) (infarct and infarct free) ANG I and II were measured by RIA after separation on HPLC. In control rats, plasma ANG I and II showed a clear increase at 6 h post-MI but subsequently only minor increases were observed. In contrast, LV ANG II showed major increases at 6 h and 1 day post-MI, which had returned to normal by 3 days in the infarct-free LV and after 1(-2) wk in the infarct LV. LV ANG I showed a more gradual increase and remained elevated in the infarct up to 8-9 wk. Nephrectomy preceding the MI lowered ANG I and II in plasma but enhanced their increases in the heart at 6 h post-MI. Both ACE inhibitors decreased plasma ANG II associated with large increases in plasma ANG I. They also inhibited the increases in LV ANG II in both the infarct and infarct-free LV at 1 and 3 days post-MI with however no significant increase in LV ANG I. In conclusion, induction of a MI in conscious rats leads to rapid and marked, but only short-lived, increases in cardiac tissue ANG II in both the infarct and infarct-free parts of the LV. Pretreatment with ACE inhibitors, but not nephrectomy, blocks this increase. Local production appears to play a major role in the increases in cardiac ANG II post-MI.

Central infusion of aldosterone synthase inhibitor prevents sympathetic hyperactivity and hypertension by central Na+ in Wistar rats

In Wistar rats, increasing cerebrospinal fluid (CSF) Na+ concentration ([Na+]) by intracerebroventricular (ICV) infusion of hypertonic saline causes sympathetic hyperactivity and hypertension that can be prevented by blockade of brain mineralocorticoid receptors (MR). To assess the role of aldosterone produced locally in the brain in the activation of MR in the central nervous system (CNS), Wistar rats were infused ICV with artificial CSF (aCSF), Na+ -rich (800 mmol/l) aCSF, aCSF plus the aldosterone synthase inhibitor FAD286 (100 microg x kg(-1) x day(-1)), or Na+ -rich aCSF plus FAD286. After 2 wk of infusion, rats treated with Na+ -rich aCSF exhibited significant increases in aldosterone and corticosterone content in the hypothalamus but not in the hippocampus, as well as increases in resting blood pressure (BP) and sympathoexcitatory responses to air stress, and impairment of arterial baroreflex function. Concomitant ICV infusion of FAD286 prevented the Na+ -induced increase in hypothalamic aldosterone but not corticosterone and prevented most of the increases in resting BP and sympathoexcitatory and pressor responses to air stress and the baroreflex impairment. FAD286 had no effects in rats infused with ICV aCSF. In another set of rats, 24-h BP and heart rate were recorded via telemetry before and during a 14-day ICV infusion of Na+ -rich aCSF with or without FAD286. Na+ -rich aCSF without FAD286 caused sustained increases ( approximately 10 mmHg) in resting mean arterial pressure that were absent in the rats treated with FAD286. These data suggest that in Wistar rats, an increase in CSF [Na+] may increase the biosynthesis of corticosterone and aldosterone in the hypothalamus, and mainly aldosterone activates MR in the CNS leading to sympathetic hyperactivity and hypertension.

Cardiac hypertrophy and cardiac renin-angiotensin system in Dahl rats on high salt intake.

Objective On high salt intake, Dahl salt-sensitive rats develop cardiac hypertrophy disproportionate to the degree of hypertension. In the present studies, we assessed whether the cardiac hypertrophy induced by high salt depends on the development of hypertension per se, and leads to over-activity of the cardiac renin–angiotensin system (RAS). Methods Cardiac angiotensin converting enzyme (ACE) mRNA and activity, cardiac and plasma angiotensin I and II (AngI, II), as well as plasma renin activity (PRA) were assessed in Dahl salt-sensitive (Dahl S) and salt-resistant (Dahl R) rats on high (1370 μmol/g food) or regular salt (120 μmol/g food) diet for 2–5 weeks. Cardiac ACE and hypertrophic response in Dahl S on high salt were also assessed after central blockade of sympathetic hyperactivity and hypertension. Results In Dahl S rats, ACE mRNA and activity of the left ventricle (LV) increased markedly after 4–5 weeks of high salt diet compared with Dahl S on the control diet and Dahl R on either diet. Chronic intra-cerebroventricular treatment with Fab fragments blocking brain ‘ouabain’ prevented the hypertension by high salt in Dahl S rats but did not affect the salt-induced increases in LV weight or in LV ACE mRNA and activity. On regular salt diet, Dahl S rats demonstrated significantly lower cardiac AngI and AngII than Dahl R rats. However, high salt intake did not cause significant changes in cardiac AngI and II in either strain. On regular salt diet, PRA, plasma AngI and II were all significantly lower in Dahl S versus R. In Dahl S rats, high salt did not cause further decreases of the already low PRA or plasma AngI and II. Conclusions These data indicate a low activity of both circulatory and cardiac RAS in Dahl S versus R rats. The marked cardiac hypertrophy and increase in cardiac ACE mRNA and activity induced by high salt in Dahl S do not depend on the increase in blood pressure. High salt intake did not increase cardiac AngII in Dahl S, suggesting that the increase in ACE mRNA and activity may be relevant for non-angiotensinergic mechanisms involved in cardiac hypertrophy.

Central infusion of aldosterone synthase inhibitor attenuates left ventricular dysfunction and remodelling in rats after myocardial infarction

AIMS Blockade of mineralocorticoid receptors in the central nervous system (CNS) prevents sympathetic hyperactivity and improves left ventricle (LV) function in rats post-myocardial infarction (MI). We examined whether aldosterone produced locally in the brain may contribute to the activation of mineralocorticoid receptors in the CNS. METHODS AND RESULTS Two days after coronary artery ligation, Wistar rats received an intra-cerebroventricular (icv) infusion via osmotic mini-pumps of the aldosterone synthase inhibitor FAD286 at 100 microg/kg/day or vehicle for 4 weeks. LV function was assessed by echocardiography at 2 and 4 weeks, and by Millar catheter at 4 weeks. At 4 weeks post-MI, aldosterone in the hippocampus was increased by 70% and tended to increase in the hypothalamus by 20%. These increases were prevented by FAD286. Across groups, aldosterone in the hippocampus and hypothalamus showed a high correlation. There were no differences in brain corticosterone levels. Compared to sham rats, at both 2 and 4 weeks post-MI rats treated with vehicle showed increased LV dimensions and decreased LV ejection fraction. Icv infusion of FAD286 attenuated these changes in LV dimensions and ejection fraction by approximately 30%. At 4 weeks post-MI, LV peak systolic pressure (LVPSP) and dP/dt(max/min) were decreased and LV end-diastolic pressure (LVEDP) was increased. In rats treated with icv FAD286, LVPSP and dP/dt(min) remained normal and LVEDP and dP/dt(max) were markedly improved. Post-MI increases in cardiac fibrosis and cardiomyocyte diameter were substantially attenuated by icv FAD286. CONCLUSION These data suggest that aldosterone produced locally in the brain acts as the main agonist of mineralocorticoid receptors in the CNS and contributes substantially to the progressive heart failure post MI.

Role of brain aldosterone and mineralocorticoid receptors in aldosterone-salt hypertension in rats

Central blockade of mineralocorticoid receptors (MRs) or angiotensin II type 1 receptors (AT1Rs) attenuates aldosterone (aldo)-salt induced hypertension. We examined the role of the subfornical organ (SFO), aldo synthesized locally in the brain, and MR and AT1R specifically in the paraventricular nucleus (PVN) in aldo-salt hypertension. Wistar rats were treated with subcutaneous aldo (1 μg/h) plus saline as drinking fluid, and gene expression was assessed by real-time qPCR. Other sets of rats received chronic intra-cerebroventricular (icv) infusion of aldo synthase (AS) inhibitor FAD286, MR blocker eplerenone or vehicle, electrolytic or sham lesions of the SFO, or intra-PVN infusion of AAV-MR-siRNA or AAV-AT1aR-siRNA. Infusion of aldo had no effect on 11βHSD2, MR and AT1R mRNA in different nuclei but increased CYP11B2 mRNA in the SFO, and serum and glucocorticoid-kinase 1 (Sgk1) and epithelial sodium channel (ENaC) γ subunit mRNA in the SFO and supraoptic nucleus (SON). MR-siRNA decreased both MR and AT1R mRNA in the PVN by ∼ 60%, but AT1aR-siRNA only decreased AT1R mRNA. SFO lesion, blockade of brain AS or MR, or knockdown of MR or AT1R in the PVN similarly attenuated aldosterone-induced saline intake by ∼ 50% and hypertension by ∼ 70%. These results suggest that an increase in circulating aldosterone may via MR and AT1R in the SFO increase local aldosterone production in hypothalamic nuclei such as the SON and PVN, and via MR enhance AT1R signaling in the PVN. This central aldosterone-MR-AT1R neuro-modulatory pathway appears to play a major role in the progressive hypertension.

Inhibition of brain angiotensin III attenuates sympathetic hyperactivity and cardiac dysfunction in rats post myocardial infarction

AIMS In rats post-myocardial infarction (MI), activation of angiotensinergic pathways in the brain contributes to sympathetic hyperactivity and progressive left ventricle (LV) dysfunction. The present study examined whether angiotensin III (Ang III) is one of the main effector peptides of the brain renin-angiotensin system controlling these effects. METHODS AND RESULTS After coronary artery ligation, Wistar rats were infused intracerebroventricularly for 4 weeks via minipumps with vehicle, the aminopeptidase A (APA) inhibitor RB150 (0.3 mg/day), which blocks the formation of brain Ang III, or losartan (0.25 mg/day). Blood pressure (BP), heart rate, and renal sympathetic nerve activity in response to air stress and acute changes in BP were measured, and LV function was evaluated by echocardiography and Millar catheter. At 4 weeks post-MI, brain APA activity was increased, sympatho-excitatory and pressor responses to air stress enhanced, and arterial baroreflex function impaired. LV end-diastolic pressure (LVEDP) was increased and ejection fraction (EF) and maximal first derivative of change in pressure over time (dP/dt(max)) were decreased. Central infusion of RB150 during 4 weeks post-MI normalized brain APA activity and responses to stress and baroreflex function, and improved LVEDP, EF, and dP/dt(max). Central infusion of losartan had similar effects but was somewhat less effective, and had no effect on brain APA activity. CONCLUSION These results indicate that brain APA and Ang III appear to play a pivotal role in the sympathetic hyperactivity and LV dysfunction in rats post-MI. RB150 may be a potential candidate for central nervous system-targeted therapy post-MI.

Role of brain corticosterone and aldosterone in central angiotensin II-induced hypertension.

Circulating angiotensin II (Ang II) activates a central aldosterone–mineralocorticoid receptor neuromodulatory pathway, which mediates most of the Ang II–induced hypertension. This study examined whether specific central infusion of Ang II also activates this central aldosterone–mineralocorticoid receptor pathway. Intracerebroventricular infusion of Ang II at 1.0, 2.5, and 12.5 ng/min for 2 weeks caused dose-related increases in water intake, Ang II concentration in the cerebrospinal fluid, and blood pressure. Intracerebroventricular Ang II, at 2.5 and 12.5 ng/min, increased hypothalamic aldosterone and corticosterone, as well as plasma aldosterone and corticosterone without affecting plasma Ang II levels. Intracerebroventricular infusion of the aldosterone synthase inhibitor FAD286—but not the mineralocorticoid receptor blocker eplerenone—inhibited by ≈60% the Ang II–induced increase in hypothalamic aldosterone. Both blockers attenuated by ≈50% the increase in plasma aldosterone and corticosterone with only minimal effects on hypothalamic corticosterone. By telemetry, intracerebroventricular infusion of Ang II maximally increased blood pressure within the first day with no further increase over the next 2 weeks. Intracerebroventricular infusion of FAD286 or eplerenone did not affect the initial pressor responses but similarly prevented 60% to 70% of the chronic pressor responses to intracerebroventricular infusion of Ang II. These results indicate distinctly different patterns of blood pressure increase by circulating versus central Ang II and support the involvement of a brain aldosterone–mineralocorticoid receptor–activated neuromodulatory pathway in the chronic hypertension caused by both circulating and central Ang II.

Angiotensin-converting enzyme inhibitors, inhibition of brain and peripheral angiotensin-converting enzymes, and left ventricular dysfunction in rats after myocardial infarction.

The brain renin-angiotensin system contributes significantly to progressive left ventricular (LV) dysfunction in rats after myocardial infarction (MI). The present study evaluated the effects of central versus peripheral plus central angiotensin-converting enzyme (ACE) blockade on sympathetic activity, and LV anatomy and function after MI. Methods: Wistar rats were treated for 4 weeks after MI with the lipophilic ACE inhibitor trandolapril at 5 mg/kg/day or the hydrophilic blocker lisinopril at 50 mg/kg/day by once daily subcutaneous injection, or with a central infusion of lisinopril at 0.1 mg/kg/day. Results: At 24 hours after the last dose, subcutaneous trandolapril caused 70% to 80% ACE inhibition in both brain and kidneys; lisinopril caused 10% to 20% less. Central infusion of lisinopril caused 70% inhibition of brain ACE and minimal (6%) inhibition in the kidneys. All three treatments similarly improved sympathetic reactivity and arterial baroreflex function. All three treatments lowered cardiac Ang I and II, and similarly attenuated the increases in LV end diastolic pressure, circumference, and fibrosis. Both subcutaneous treatments further decreased LV peak systolic pressure and dP/dtmax, whereas icv lisinopril caused no change. Conclusion: Despite marked differences in the extent of peripheral blockade, all three treatments similarly affected sympathetic activity and decreased cardiac Ang II, preload and remodeling after MI. One may speculate that central and peripheral ACE-mediated mechanisms are sequential and therefore only minor additional effects of peripheral ACE blockade are noted.

Cardiac Macrophages and Apoptosis after Myocardial Infarction: Effects of Central MR Blockade

After myocardial infarction (post-MI), inflammation and apoptosis contribute to progressive cardiac remodeling and dysfunction. Cardiac mineralocorticoid receptor (MR) and β-adrenergic signaling promote apoptosis and inflammation. Post-MI, MR activation in the brain contributes to sympathetic hyperactivity and an increase in cardiac aldosterone. In the present study, we assessed the time course of macrophage infiltration and apoptosis in the heart as detected by both terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) and active caspase-3 immunostaining in both myocytes and nonmyocytes, as well as the effects of central MR blockade by intracerebroventricular infusion of eplerenone at 5 μg/day on peak changes in macrophage infiltration and apoptosis post-MI. Macrophage numbers were markedly increased in the infarct and peri-infarct zones and to a minor extent in the noninfarct part of the left ventricle at 10 days post-MI and decreased over the 3-mo study period. Apoptosis of both myocytes and nonmyocytes was clearly apparent in the infarct and peri-infarct areas at 10 days post-MI. For TUNEL, the increases persisted at 4 and 12 wk, but the number of active caspase-3-positive cells markedly decreased. Central MR blockade significantly decreased CD80-positive proinflammatory M1 macrophages and increased CD163-positive anti-inflammatory M2 macrophages in the infarct. Central MR blockade also reduced apoptosis of myocytes by 40-50% in the peri-infarct and to a lesser extent of nonmyocytes in the peri-infarct and infarct zones. These findings indicate that MR activation in the brain enhances apoptosis both in myocytes and nonmyocytes in the peri-infarct and infarct area post-MI and contributes to the inflammatory response.