Alamandine Induces Neuroprotection in Ischemic Stroke Models.

BACKGROUND AND OBJECTIVE: Stroke, a leading cause of mortality and disability, characterized by neuronal death, can be induced by a reduction or interruption of blood flow. In this study, the role of Alamandine, a new peptide of the renin-angiotensin system, was evaluated in in-vitro and in-vivo brain ischemia models. METHODS: In the in-vitro model, hippocampal slices from male C57/Bl6 mice were placed in a glucose-free aCSF solution and bubbled with 95% N2 and 5% CO2 to mimic brain ischemia. An Alamandine concentration-response curve was generated to evaluate cell damage, glutamatergic excitotoxicity, and cell death. In the in-vivo model, cerebral ischemia/ reperfusion was induced by bilateral occlusion of common carotid arteries (BCCAo-untreated) in SD rats. An intracerebroventricular injection of Alamandine was given 20-30 min before BCCAo. Animals were subjected to neurological tests 24 h and 72 h after BCCAo. Cytokine levels, oxidative stress markers, and immunofluorescence were assessed in the brain 72 h after BCCAo. RESULTS: Alamandine was able to protect brain slices from cellular damage, excitotoxicity and cell death. When the Alamandine receptor was blocked, protective effects were lost. ICV injection of Alamandine attenuated neurological deficits of animals subjected to BCCAo and reduced the number of apoptotic neurons/cells. Furthermore, Alamandine induced anti-inflammatory effects in BCCAo animals as shown by reductions in TNFα, IL- 1ß, IL-6, and antioxidant effects through attenuation of the decreased SOD, catalase, and GSH activities in the brain. CONCLUSION: This study showed, for the first time, a neuroprotective role for Alamandine in different ischemic stroke models.

Angiotensin-(1-7) prevents T3-induced cardiomyocyte hypertrophy by upregulating FOXO3/SOD1/catalase and downregulating NF-ĸB.

Clinical studies have shown a correlation between thyroid disorders and cardiac diseases. High levels of triiodothyronine (T3) induce cardiac hypertrophy, a risk factor for cardiac complications and heart failure. Previous results have demonstrated that angiotensin-(1-7) is able to block T3-induced cardiac hypertrophy; however, the molecular mechanisms involved in this event have not been fully elucidated. Here, we evidenced the contribution of FOXO3 signaling to angiotensin-(1-7) effects. Angiotensin-(1-7) treatment increased nuclear FOXO3 levels and reduced p-FOXO3 levels (inactive form) in isolated cardiomyocytes. Knockdown of FOXO3 by RNA silencing abrogated the antihypertrophic effect of angiotensin-(1-7). Increased expression of antioxidant enzymes superoxide dismutase 1 (SOD1 and catalase) and lower levels of reactive oxygen species and nuclear factor-κB (NF-κB) were observed after angiotensin-(1-7) treatment in vitro. Consistent with these results, transgenic rats overexpressing angiotensin-(1-7) displayed increased nuclear FOXO3 and SOD1 levels and reduced NF-κB levels in the heart. These results provide a new molecular mechanism responsible for the antihypertrophic effect of angiotensin-(1-7), which may contribute to future therapeutic targets.

Differential control of vasomotion by angiotensins in the rostral ventrolateral medulla of hypertensive rats.

The central and peripheral renin-angiotensin systems are known for playing a key role in cardiovascular control. In the present study, we evaluated the hemodynamic effects produced by nanoinjections of angiotensin II (Ang II) or angiotensin-(1-7) [Ang-(1-7)] into the rostral ventrolateral medulla (RVLM) of adult male normotensive (Wistar-WT) and spontaneously hypertensive rats (SHR). Animals were anesthetized (urethane 1.2g/kg) and instrumented for recording blood pressure (BP), heart rate (HR) and blood flow (BF) in the femoral, renal or mesenteric arteries. Afterwards, rats were positioned in a stereotaxic and prepared for nanoinjections (100 nl) of saline (NaCl 0.9%), Ang-(1-7) (40 ng) or Ang II (40 ng) into the RVLM. The vascular resistance (VR) was calculated by ΔMAP/ΔBF ratio. In WT, Ang-(1-7) or Ang II caused equipotent pressor effects that were not accompanied by changes in vascular resistance. However, MAP changes were greater in SHR. This strain also showed a concomitant increase in relative vascular resistance (ΔVR/VRbaseline) of renal (0.31 ± 0.07 and 0.3 ± 0.07 vs. 0.02 ± 0.01; Ang-(1-7), Ang II and Saline, respectively) and mesenteric beds (0.3 ± 0.06 and 0.33 ± 0.04 vs. 0.05 ± 0.02; Ang-(1-7), Ang II and saline, respectively). We conclude that Ang II and Ang-(1-7) at the RVLM control the vascular resistance of renal and mesenteric beds during hypertension.

Development of attenuated baroreflexes in obese Zucker rats coincides with impaired activation of nucleus tractus solitarius.

Adult obese Zucker rats (OZR; >12 wk) develop elevated sympathetic nerve activity (SNA) and mean arterial pressure (MAP) with impaired baroreflexes compared with adult lean Zucker rats (LZR) and juvenile OZR (6-7 wk). In adult OZR, baroreceptor afferent nerves respond normally to changes in MAP, whereas electrical stimulation of baroreceptor afferent fibers produces smaller reductions in SNA and MAP compared with LZR. We hypothesized that impaired baroreflexes in OZR are linked to reduced activation of brain stem sites that mediate baroreflexes. In conscious adult rats, a hydralazine (HDZ)-induced reduction in MAP evoked tachycardia that was initially blunted in OZR, but equivalent to LZR within 5 min. In agreement, HDZ-induced expression of c-Fos in the rostral ventrolateral medulla (RVLM) was comparable between groups. In contrast, phenylephrine (PE)-induced rise in MAP evoked markedly attenuated bradycardia with dramatically reduced c-Fos expression in the nucleus tractus solitarius (NTS) of adult OZR compared with LZR. However, in juvenile rats, PE-induced hypertension evoked comparable bradycardia in OZR and LZR with similar or augmented c-Fos expression in NTS of the OZR. In urethane-anesthetized rats, microinjections of glutamate into NTS evoked equivalent decreases in SNA, heart rate (HR), and MAP in juvenile OZR and LZR, but attenuated decreases in SNA and MAP in adult OZR. In contrast, microinjections of glutamate into the caudal ventrolateral medulla, a target of barosensitive NTS neurons, evoked comparable decreases in SNA, HR, and MAP in adult OZR and LZR. These data suggest that OZR develop impaired glutamatergic activation of the NTS, which likely contributes to attenuated baroreflexes in adult OZR.

The pressor effect of angiotensin-(1-7) in the rat rostral ventrolateral medulla involves multiple peripheral mechanisms.

OBJECTIVE: In the present study, the peripheral mechanism that mediates the pressor effect of angiotensin-(1-7) in the rostral ventrolateral medulla was investigated. METHOD: Angiotensin-(1-7) (25 pmol) was bilaterally microinjected in the rostral ventrolateral medulla near the ventral surface in urethane-anesthetized male Wistar rats that were untreated or treated (intravenously) with effective doses of selective autonomic receptor antagonists (atenolol, prazosin, methyl-atropine, and hexamethonium) or a vasopressin V1 receptor antagonist [d(CH2)5 -Tyr(Me)-AVP] given alone or in combination. RESULTS: Unexpectedly, the pressor response produced by angiotensin-(1-7) (16 ± 2 mmHg, n = 12), which was not associated with significant changes in heart rate, was not significantly altered by peripheral treatment with prazosin, the vasopressin V1 receptor antagonist, hexamethonium or methyl-atropine. Similar results were obtained in experiments that tested the association of prazosin and atenolol; methyl-atropine and the vasopressin V1 antagonist or methyl-atropine and prazosin. Peripheral treatment with the combination of prazosin, atenolol and the vasopressin V1 antagonist abolished the pressor effect of glutamate; however, this treatment produced only a small decrease in the pressor effect of angiotensin-(1-7) at the rostral ventrolateral medulla. The combination of hexamethonium with the vasopressin V1 receptor antagonist or the combination of prazosin, atenolol, the vasopressin V1 receptor antagonist and methyl-atropine was effective in blocking the effect of angiotensin-(1-7) at the rostral ventrolateral medulla. CONCLUSION: These results indicate that angiotensin-(1-7) triggers a complex pressor response at the rostral ventrolateral medulla that involves an increase in sympathetic tonus, release of vasopressin and possibly the inhibition of a vasodilatory mechanism.

The pressor effect of angiotensin-(1-7) in the rat rostral ventrolateral medulla involves multiple peripheral mechanisms

OBJECTIVE: In the present study, the peripheral mechanism that mediates the pressor effect of angiotensin-(1-7) in the rostral ventrolateral medulla was investigated. METHOD: Angiotensin-(1-7) (25 pmol) was bilaterally microinjected in the rostral ventrolateral medulla near the ventral surface in urethane-anesthetized male Wistar rats that were untreated or treated (intravenously) with effective doses of selective autonomic receptor antagonists (atenolol, prazosin, methyl-atropine, and hexamethonium) or a vasopressin V1 receptor antagonist [d(CH2)5 -Tyr(Me)-AVP] given alone or in combination. RESULTS: Unexpectedly, the pressor response produced by angiotensin-(1-7) (16 ± 2 mmHg, n = 12), which was not associated with significant changes in heart rate, was not significantly altered by peripheral treatment with prazosin, the vasopressin V1 receptor antagonist, hexamethonium or methyl-atropine. Similar results were obtained in experiments that tested the association of prazosin and atenolol; methyl-atropine and the vasopressin V1 antagonist or methyl-atropine and prazosin. Peripheral treatment with the combination of prazosin, atenolol and the vasopressin V1 antagonist abolished the pressor effect of glutamate; however, this treatment produced only a small decrease in the pressor effect of angiotensin-(1-7) at the rostral ventrolateral medulla. The combination of hexamethonium with the vasopressin V1 receptor antagonist or the combination of prazosin, atenolol, the vasopressin V1 receptor antagonist and methyl-atropine was effective in blocking the effect of angiotensin-(1-7) at the rostral ventrolateral medulla. CONCLUSION: These results indicate that angiotensin-(1-7) triggers a complex pressor response at the rostral ventrolateral medulla that involves an increase in sympathetic tonus, release of vasopressin and possibly the inhibition of a vasodilatory mechanism.

Lifetime overproduction of circulating Angiotensin-(1-7) attenuates deoxycorticosterone acetate-salt hypertension-induced cardiac dysfunction and remodeling.

We evaluated the development of arterial hypertension, cardiac function, and collagen deposition, as well as the level of components of the renin-angiotensin system in the heart of transgenic rats that overexpress an angiotensin (Ang)-(1-7)-producing fusion protein, TGR(A1-7)3292 (TG), which induces a lifetime increase in circulating levels of this peptide. After 30 days of the induction of the deoxycorticosterone acetate (DOCA)-salt hypertension model, DOCA-TG rats were hypertensive but presented a lower systolic arterial pressure in comparison with DOCA-Sprague-Dawley (SD) rats. In contrast to DOCA-SD rats that presented left ventricle (LV) hypertrophy and diastolic dysfunction, DOCA-TG rats did not develop cardiac hypertrophy or changes in ventricular function. In addition, DOCA-TG rats showed attenuation in mRNA expression for collagen type I and III compared with the increased levels of DOCA-SD rats. Ang II plasma and LV levels were reduced in SD and TG hypertensive rats in comparison with normotensive animals. DOCA-TG rats presented a reduction in plasma Ang-(1-7) levels; however, there was a great increase in Ang-(1-7) ( approximately 3-fold) accompanied by a decrease in mRNA expression of both angiotensin-converting enzyme and angiotensin-converting enzyme 2 in the LV. The mRNA expression of Mas and Ang II type 1 receptors in the LV was not significantly changed in DOCA-SD or DOCA-TG rats. This study showed that TG rats with increased circulating levels of Ang-(1-7) are protected against cardiac dysfunction and fibrosis and also present an attenuated increase in blood pressure after DOCA-salt hypertension. In addition, DOCA-TG rats showed an important local increase in Ang-(1-7) levels in the LV, which might have contributed to the attenuation of cardiac dysfunction and prefibrotic lesions.

Angiotensin-(1-7) prevents cardiomyocyte pathological remodeling through a nitric oxide/guanosine 3',5'-cyclic monophosphate-dependent pathway.

The renin-angiotensin (Ang) system plays a pivotal role in the pathogenesis of cardiovascular disease, with Ang II being the major effector of this system. Multiple lines of evidence have shown that Ang-(1-7) exerts cardioprotective effects in the heart by counterregulating Ang II actions. The questions that remain are how and where Ang-(1-7) exerts its effects. By using a combination of molecular biology, confocal microscopy, and a transgenic rat model with increased levels of circulating Ang-(1-7) (TGR[A1-7]3292), we evaluated the signaling pathways involved in Ang-(1-7) cardioprotection against Ang II-induced pathological remodeling in ventricular cardiomyocytes. Rats were infused with Ang II for 2 weeks. We found that ventricular myocytes from TGR(A1-7)3292 rats are protected from Ang II pathological remodeling characterized by Ca(2+) signaling dysfunction, hypertrophic fetal gene expression, glycogen synthase kinase 3beta inactivation, and nuclear factor of activated T-cells nuclear accumulation. Moreover, cardiomyocytes from TGR(A1-7)3292 rats infused with Ang II presented increased expression levels of neuronal NO synthase. To provide a signaling pathway involved in the beneficial effects of Ang-(1-7), we treated neonatal cardiomyocytes with Ang-(1-7) and Ang II for 36 hours. Treatment of cardiomyocytes with Ang-(1-7) prevented Ang II-induced hypertrophy by modulating calcineurin/nuclear factor of activated T-cell signaling cascade. Importantly, antihypertrophic effects of Ang-(1-7) on Ang II-treated cardiomyocytes were prevented by N(G)-nitro-l-arginine methyl ester and 1H-1,2,4oxadiazolo4,2-aquinoxalin-1-one, suggesting that these effects are mediated by NO/cGMP. Taken together, these data reveal a key role for NO/cGMP as a mediator of Ang-(1-7) beneficial effects in cardiac cells.

Evidence for Mas-mediated bradykinin potentiation by the angiotensin-(1-7) nonpeptide mimic AVE 0991 in normotensive rats.

We evaluated the effect of the nonpeptide mimic of angiotensin (Ang)-(1-7), AVE 0991, on the hypotensive effect of bradykinin (BK). Increasing doses of intra-arterial or intravenous BK were administered before and 30 minutes after the beginning of AVE 0991 infusion. The effect of AVE 0991 on plasma Ang-converting enzyme activity was tested using Hip-His-Leu as the substrate. The interaction of AVE 0991 with Ang-converting enzyme in vivo was tested by determining its effect on the pressor action of Ang I or Ang II. AVE 0991 produced a significant and similar potentiation of intra-arterial or intravenous bradykinin. AVE 0991 did not inhibit plasma Ang-converting enzyme activity in vitro or the pressor effect of Ang I in vivo. N(W)-nitro-l-arginine methyl ester or D-Ala(7)-Ang-(1-7) administration abolished the BK potentiating effect of AVE 0991. We further examined the BK-potentiating effect of AVE 0991, evaluating its effect on NO production in rabbit endothelial cells. The NO release was measured using the 4-amino-5-methylamino-2'-7'-difluorofluorescein diacetate. A synergistic effect of AVE 0991 and BK on NO release was observed. These results suggest that AVE 0991 potentiates bradykinin through an Ang-converting enzyme-independent, NO-dependent receptor Mas-mediated mechanism. This effect may contribute to the improvement of endothelial function by AVE 0991 in vivo.

Immunofluorescence localization of the receptor Mas in cardiovascular-related areas of the rat brain.

The G protein-coupled receptor Mas was recently described as an angiotensin-(1-7) [ANG-(1-7)] receptor. In the present study we evaluated the anatomical localization of Mas using immunofluorescence in the central nervous system of adult male Wistar rats. An abundant labeling was found in the hippocampus, amigdala, anterodorsal thalamic nucleus, cortex, and hypoglossal nucleus. More importantly, a dense ANG-(1-7) receptor Mas immunoreactivity was observed in cardiovascular-related areas of the medulla and forebrain, shown in several previous studies as sites for the action of ANG-(1-7) in the brain. A strong staining was found in the nucleus of the solitary tract, caudal and rostral ventrolateral medulla, inferior olive, parvo and magnocellular portions of the paraventricular hypothalamic nucleus, supraoptic nucleus, and lateral preoptic area. Furthermore, Mas staining was predominantly present in neurons. At the medullary sites, a specific and high-intensity binding for rhodamine-ANG-(1-7) was also shown. The specific ANG-(1-7) binding was completely displaced by the anti-Mas antibody or by the ANG-(1-7) antagonist, A-779. The data presented provide the first anatomical basis for the physiological role of ANG-(1-7)/Mas axis in the modulation of different cardiovascular functions and give new insights for clarifying the role of ANG-(1-7) in the central nervous system.

Expression of an angiotensin-(1-7)-producing fusion protein in rats induced marked changes in regional vascular resistance.

We have described a transgenic rat line that expresses an angiotensin-(1-7)-producing fusion protein, the TGR(A1-7)3292. In these rats, testis acts as an angiotensin-(1-7) biological pump, increasing its plasma concentration 2.5-fold. In this study, we performed hemodynamic measurements in TGR(A1-7)3292 and age-matched Hannover Sprague-Dawley (SD) control rats, using fluorescent microspheres. Urethane-anesthetized transgenic rats had similar levels of baseline blood pressure (99 +/- 3 mmHg) as did SD rats (101 +/- 3 mmHg). However, pronounced differences were observed in other hemodynamic measurements. TGR(A1-7)3292 rats presented a significant increase in stroke volume (0.29 +/- 0.01 vs. 0.25 +/- 0.01 ml in SD), increased cardiac index (24.6 +/- 0.91 vs. 21.9 +/- 0.65 ml.min(-1).kg) and decreased total peripheral resistance (3.9 +/- 0.13 vs. 4.5 +/- 0.13 mmHg.ml(-1).min.100 g). The increase in stroke volume in transgenic rats may be partially explained by the small decrease in heart rate (326 +/- 7.0 vs. 359 +/- 6.0 beats/min in SD). Strikingly, TGR(A1-7)3292 rats presented a substantial decrease in the vascular resistance in lung, spleen, kidney, adrenals, brain, testis and brown fat tissue with no significant differences in the left ventricle, mesentery, skin, gastrocnemius muscle and white fat tissue. These results corroborate and extend previous results observed after acute angiotensin-(1-7) infusion, showing that chronic increase in circulating angiotensin-(1-7) produces sustained and important changes in regional and systemic hemodynamics. Moreover, our data suggest a physiological role for angiotensin-(1-7) in the tonic control of regional blood flow.

Baroreflex modulation by angiotensins at the rat rostral and caudal ventrolateral medulla.

We determined the effect of microinjection of ANG-(1-7) and ANG II into two key regions of the medulla that control the circulation [rostral and caudal ventrolateral medulla (RVLM and CVLM, respectively)] on baroreflex control of heart rate (HR) in anesthetized rats. Reflex bradycardia and tachycardia were induced by increases and decreases in mean arterial pressure produced by intravenous phenylephrine and sodium nitroprusside, respectively. The pressor effects of ANG-(1-7) and ANG II (25 pmol) after RVLM microinjection (11 +/- 0.8 and 10 +/- 2 mmHg, respectively) were not accompanied by consistent changes in HR. In addition, RVLM microinjection of these angiotensin peptides did not alter the bradycardic or tachycardic component of the baroreflex. CVLM microinjections of ANG-(1-7) and ANG II produced hypotension (-11 +/- 1.5 and -11 +/- 1.9 mmHg, respectively) that was similarly not accompanied by significant changes in HR. However, CVLM microinjections of angiotensins induced differential changes in the baroreflex control of HR. ANG-(1-7) attenuated the baroreflex bradycardia (0.26 +/- 0.06 ms/mmHg vs. 0.42 +/- 0.08 ms/mmHg before treatment) and facilitated the baroreflex tachycardia (0.86 +/- 0.19 ms/mmHg vs. 0.42 +/- 0.10 ms/mmHg before treatment); ANG II produced the opposite effect, attenuating baroreflex tachycardia (0.09 +/- 0.06 ms/mmHg vs. 0.31 +/- 0.07 ms/mmHg before treatment) and facilitating the baroreflex bradycardia (0.67 +/- 0.16 ms/mmHg vs. 0.41 +/- 0.05 ms/mmHg before treatment). The modulatory effect of ANG II and ANG-(1-7) on baroreflex sensitivity was completely abolished by peripheral administration of methylatropine. These results suggest that ANG II and ANG-(1-7) at the CVLM produce a differential modulation of the baroreflex control of HR, probably through distinct effects on the parasympathetic drive to the heart.

Differential regulation of central vasopressin receptors in transgenic rats with low brain angiotensinogen.

The consequences of permanent alteration to the brain renin-angiotensin system (RAS) on central vasopressinergic system was studied in transgenic rats with low brain angiotensinogen [TGR(ASrAOGEN)]. Levels of vasopressin (AVP) and V1a receptor mRNAs were measured by ribonuclease protection assay (RPA) and AVP by radioimmunoassay (RIA). AVP (100 pmol/50 nl) was microinjected into the nucleus tractus solitarii (NTS) of urethane-anesthetized TGR(ASrAOGEN) and Sprague-Dawley (SD) rats and the mean arterial pressure (MAP) and heart rate (HR) baroreflex induced by phenylephrine were evaluated. AVP but not its mRNA levels were significantly lower in the hypothalamus and hypophysis of TGR(ASrAOGEN) rats. Brainstem V1a mRNA levels were significantly higher in TGR(ASrAOGEN) in comparison to SD rats (5.2+/-0.4% vs. 3.3+/-0.2% of beta-actin mRNA, P<0.05). In contrast, the hypothalamic V1a mRNA levels in TGR(ASrAOGEN) were not different from those found in SD rats. AVP microinjections induced a greater decrease in MAP in TGR(ASrAOGEN) in comparison with SD rats (-19.9+/-5.2 vs. -7.5+/-0.7 mm Hg, P<0.01). The significantly higher baroreflex sensitivity observed in TGR compared to that of SD rats was normalized after AVP microinjection. The increased brainstem V1a mRNA levels and sensitivity to AVP in TGR(ASrAOGEN) rats indicates a functional upregulation of AVP receptors in the NTS. The fact that the hypothalamic V1a mRNA levels are not altered indicates that these receptors are differentially regulated in different brain regions. This study demonstrates that a permanent deficit in brain angiotensinogen synthesis can alter the functionality of central vasopressinergic system.

Differential effects of angiotensin II and angiotensin-(1-7) at the nucleus tractus solitarii of transgenic rats with low brain angiotensinogen.

OBJECTIVES: In this study, we investigated the effects of angiotensin II and angiotensin-(1-7) at the nucleus tractus solitarii (nTS) in transgenic rats with a severe deficit in brain angiotensinogen production, TGR(ASrAOGEN) (TGR). METHODS: Angiotensin II (10 pmol), angiotensin-(1-7) (10 pmol) or NaCl (0.9%/50 nl) were microinjected into the nTS of urethane-anaesthetized TGR (n = 36) and Sprague Dawley (SD) (n = 34) rats. Mean arterial pressure (MAP) and heart rate were measured via a femoral artery catheter and the baroreflex control of heart rate was evaluated after increases in MAP induced by phenylephrine (baroreflex bradycardia). RESULTS: Angiotensin II microinjections into the nTS of the TGR induced a higher decrease in MAP and heart rate (-37 +/- 5 mmHg and -69 +/- 12 b.p.m., respectively) in comparison to SD rats (-18 +/- 1 mmHg and -43 +/- 5 b.p.m., respectively). In contrast, changes after angiotensin-(1-7) microinjections into the nTS of TGR (-6 +/- 1 mmHg and -13 +/- 4 b.p.m.) were significantly smaller than that induced in SD (-11 +/- 2 mmHg and -24 +/- 6 b.p.m.). The baseline baroreflex sensitivity to phenylephrine of TGR was accentuated in comparison to SD rats (0.70 +/- 0.06 versus 0.44 +/- 0.03 ms/mmHg). Angiotensin II microinjection into the nTS produced similar attenuation in the baroreflex bradycardia in both SD (0.28 +/- 0.07 versus 0.5 +/- 0.07 ms/mmHg, before injection) and TGR (0.44 +/- 0.1 versus 0.82 +/- 0.1 ms/mmHg, before injection). In contrast, angiotensin-(1-7) microinjection elicited a facilitation of the baroreflex bradycardia in SD (0.68 +/- 0.12 versus 0.41 +/- 0.03 ms/mmHg, before injection), while in TGR, angiotensin-(1-7) induced an attenuation of baroreflex bradycardia (0.34 +/- 0.07 ms/mmHg versus 0.55 +/- 0.05 ms/mmHg, before injection). CONCLUSIONS: These results indicate that a permanent inhibition of angiotensinogen synthesis in the brain can lead to an increase in the sensitivity of the baroreflex control of heart rate (baroreflex bradycardia) and an increase in angiotensin II responsiveness at the nTS. However, the nTS effect of angiotensin-(1-7) was significantly attenuated in these transgenic rats. These data further indicate that the decrease in brain angiotensins in the transgenic rats may be functionally relevant and support the concept of differential regulatory mechanisms for the effects of the two angiotensin peptides.

Angiotensin-(1-7) is involved in the endothelium-dependent modulation of phenylephrine-induced contraction in the aorta of mRen-2 transgenic rats.

1. The contribution of the local vascular production of angiotensin-(1-7) [Ang-(1-7)] to the control of alpha-adrenergic-induced contractions in the aorta of Sprague-Dawley (SD) and TGR(mRen-2)27 [mRen-2] rats was studied. 2. In mRen-2 rats, contractile responses to phenylephrine were diminished as compared to control SD rats in endothelium containing but not in endothelium-denuded vessels. L-NAME increased contractile responses to phenylephrine in mRen-2 rats and, after nitric oxide synthase blockade, responses to phenylephrine became comparable in both strains. 3. Inhibition of angiotensin-converting enzyme (ACE) by captopril potentiated contractile responses in mRen-2 rats and diminished contractile responses in SD rats, both effects being dependent on the presence of a functional endothelium. The effect of captopril in mRen-2 rats was abolished in vessels pre-incubated with Ang-(1-7). 4. Blockade of Ang-(1-7) and bradykinin (BK) receptors by A-779 and HOE 140 respectively, increased phenylephrine-induced contraction in mRen-2, but not in SD rats. This effect was seen only in endothelium-containing vessels. 5. Angiotensin II AT(1) and AT(2) receptor blockade by CV 11974 and PD 123319 did not affect the contractile responses to phenylephrine in aortas of transgenic animals but diminished the response in SD rats. This effect was only seen in the presence of a functional endothelium. 6. It is concluded that the decreased contractile responses to phenylephrine in aortas of mRen-2 rats was dependent on an intact endothelium, the local release and action of Ang-(1-7) and bradykinin.