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(Hypertension. 1995;25:1260-1265.)
© 1995 American Heart Association, Inc.

Articles

Opposing Actions of Angiotensin-(1-7) and Angiotensin II in the Brain of Transgenic Hypertensive Rats

Presented in part at the Ninth Scientific Meeting of the American Society of Hypertension, New York, NY, May 11-14, 1994.

Atsushi Moriguchi; E. Ann Tallant; Kiyoshi Matsumura; Thomas M. Reilly; Harry Walton; Detlev Ganten; Carlos M. Ferrario

From The Hypertension Center, The Bowman Gray School of Medicine of Wake Forest University, Winston-Salem, NC (A.M., E.A.T., K.M., C.M.F.); DuPont Merck Pharmaceutical Co, Cardiovascular Disease Research, Wilmington, Del (T.M.R., H.W.); and the Max-Delbrück-Centrum für Molekulare Medizin (MDC), Berlin-Buch, Germany (D.G.).

	Abstract

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Abstract Lack of specific antagonists to the amino-terminal heptapeptide angiotensin-(1-7) [Ang-(1-7)] prompted us to evaluate the central effects of delivering a specific affinity-purified Ang-(1-7) antibody on the blood pressure and heart rate of 12-week-old conscious homozygous female rats (n=12) expressing the mouse submandibular Ren-2^d gene [(mRen-2^d)27] in their genome. Another group of transgenic hypertensive and strain-matched Sprague-Dawley controls were injected with a specific Ang II monoclonal antibody (KAA8). Cerebroventricular administration of the affinity-purified Ang-(1-7) antibody in conscious transgenic hypertensive rats caused significant dose-related elevations in blood pressure associated with tachycardia. The hypertensive response was augmented in transgenic rats studied 7 to 10 days after cessation of lisinopril therapy. Neutralization of Ang II with the Ang II antibody caused a hemodynamic response opposite to that obtained with the Ang-(1-7) antibody. All doses of the Ang II antibody produced hypotension and bradycardia. The magnitude of the depressor response was significantly augmented in transgenic rats weaned off lisinopril therapy. In contrast, central administration of either the Ang-(1-7) or Ang II antibodies had no effect on normotensive rats. Central injections of an affinity-purified IgG fraction were ineffective in both control and transgene-positive rats. These data suggest that in the brain of transgenic hypertensive rats, Ang-(1-7) opposes the action of Ang II on the central mechanism or mechanisms that contribute to the maintenance of this model of hypertension. In addition, these studies showed an important contribution of the brain renin-angiotensin system to the maintenance of this form of monogenetic hypertension.

Key Words: hypertension, genetic • renin-angiotensin system • renin • angiotensins • animals, transgenic • blood pressure

	Introduction

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It is generally accepted that hormones regulate endocrine function through multilevel feedback mechanisms. The renin-angiotensin system is no exception because the physiological actions of angiotensin II (Ang II) are restrained by inhibitory actions of the peptide on renin synthesis¹ ; angiotensin-converting enzyme expression² ; and stimulation of the synthesis and release of vasodilator prostaglandins, atrial natriuretic peptide, and endothelium-derived relaxing factors.³

Our laboratory has proposed the existence of an additional important mechanism in the feedback regulation of Ang II activity.⁴ This action is produced by the companion generation of the heptapeptide [Asp¹,Arg²,Val³,Tyr⁴,Ile⁵,His⁶,Pro⁷]angiotensin-(1-7) [Ang-(1-7)], which acts by stimulating the release of prostaglandins,⁴ nitric oxide,⁵ or both. Ang-(1-7) is generated by cleavage of the [Pro⁷,Phe⁸] bond of either Ang I or Ang II by tissue endopeptidases.^{6 7} While Ang-(1-7) is as potent as Ang II in stimulating vasopressin secretion,⁸ it behaves as a vasodilator⁹ and natriuretic agent,¹⁰ possibly because it is more potent than Ang II in augmenting prostacyclin secretion.^{11 12}

These data led us to suggest that the regulation of blood pressure (BP) by the renin-angiotensin system may be determined by the coordinated and reciprocal actions of Ang II and Ang-(1-7) on specific target-organ receptors.⁴ Recent studies of the characteristics of the renin-angiotensin system in the brain of a new genetic model of arterial hypertension provided us with the opportunity to evaluate this hypothesis. Mullins et al¹³ showed that insertion of the mouse submandibular gland Ren-2 gene into the rat genome causes a severe form of hypertension associated with vascular lesions reminiscent of those found in the malignant form of human hypertension and responsive to therapy with angiotensin-converting enzyme inhibitors or specific Ang II receptor antagonists.¹³ Recent studies from this laboratory implicated the participation of the brain renin-angiotensin system in the evolution of this form of transgene hypertension. We found the content of Ang II and Ang-(1-7) in the hypothalamus of adult transgenic hypertensive rats to be 10-fold and 6-fold higher, respectively, than in age-matched control rats.¹⁴ The large increases in the content of brain angiotensins were associated with central desensitization of angiotensin receptors because intracerebroventricular (ICV) administration of Ang II did not raise BP or stimulate release of arginine vasopressin in transgenic hypertensive rats.¹⁵ Similarly, application of similar doses of Ang-(1-7) ICV did not stimulate release of vasopressin in transgenic rats, whereas the response could be readily detected in control transgene-negative rats.¹⁵

Evidence for biochemical and functional overactivity of the renin-angiotensin system in the brain of transgenic hypertensive rats provided the opportunity for the assessment of the relative contributions of Ang II and Ang-(1-7) to BP regulation by central mechanisms. Although a variety of approaches are available for inhibiting Ang II activity, there are no specific antagonists that could be used to disentangle the actions of Ang-(1-7) from those of Ang II. To resolve this problem we evaluated an alternate way of documenting the physiological consequences of neutralization of high Ang-(1-7) levels in the brain of transgenic rats by using well-characterized specific affinity-purified antibodies to the peptide. The present study describes the effects of neutralization of either Ang II or Ang-(1-7) in the brain of (mRen-2)27 hypertensive transgenic rats.

	Methods

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Experiments were performed in 22 homozygous female transgenic hypertensive rats (body weight, 277±9 g) and 8 Sprague-Dawley (SD) controls (278±5 g) at 12 weeks of age. Transgene rats positive for the mouse Ren-2 gene were bred at our institution from founder breeders [(mRen-2)27] obtained from the German Institute for High Blood Pressure Research (Heidelberg, Germany). SD rats were obtained from the Zentralinstitut fur Versuchstierzucht (Hannover, Germany). These rats are descendants of the parent stock of rats used by Mullins et al¹³ to breed transgenic rats. Rats were housed in plastic cages within a room maintained at 22°C with a 12-hour dark/light cycle. The animals had free access to tap water and were fed a powdered rat chow (Rodent Laboratory Chow 5001, Purina Mills Inc) providing a daily intake of 17 mEq Na⁺ and 28 mEq K⁺ per 100 g of solid weight.

Twenty-four hours before experiments rats were anesthetized with sodium pentobarbital (50 mg/kg IP). A stainless steel cannula was inserted into a lateral cerebral ventricle (28-gauge stainless steel needle) under stereotaxic guidance with the use of coordinates described elsewhere.¹⁶ A polyethylene catheter (PE-50, Clay Adams, Becton Dickinson) was also inserted into the abdominal aorta via an incision in a femoral artery. The free end of the catheter was tunneled under the skin to the nape of the neck for arterial pressure recording, as described elsewhere.¹⁷ The surgical procedure was performed with the use of sterile conditions, and rats were given 30 000 U IM penicillin G at the completion of the procedure. On the day of the experiment, the arterial catheter was connected to a solid strain-gauge microtransducer (MP-150, Micron Instruments Inc), and tracings of arterial pressure and heart rate were recorded on a multichannel polygraph (model 2000, Gould Instruments). Beat-by-beat changes in heart rate were determined by a tachometer (Biotach, Gould Instruments).

Phasic and mean arterial pressure (MAP) and heart rate were measured in conscious, freely moving rats for a 1-hour habituation period and again during and after injection of antibodies or vehicle. At the completion of experiments, rats were killed with a lethal dose of sodium pentobarbital (1.5 g IV). The brain was examined histologically to verify the correct placement of the cannula into a lateral cerebral ventricle. Two of the 22 rats were excluded from the study because of incorrect placement of the ICV cannula.

Experimental Protocols
Twelve of 20 homozygous transgenic and 5 of 8 control SD rats were used for investigation of the effects of central administration of the Ang-(1-7) antibody. The antibody was given into the cannula leading to a brain ventricle at an infusion rate of 0.1 µL/s (CMA-100, Carnegie Medicine). Three separate doses of the Ang-(1-7) antibody (1.3 µg in 2 µL, 3.2 µg in 5 µL, and 6.3 µg in 10 µL) were injected in random order. Injections of either the Ang-(1-7) antibody or vehicle were spaced at least 30 minutes apart. Control studies included central administration of either similar amounts of an affinity-purified rabbit IgG preparation (phosphatase 2b¹⁸ ) or equal volumes of phosphate-buffered saline (vehicle). Six of the 12 transgenic rats were on lifetime therapy with lisinopril (10 mg/kg in the drinking water). In the other 6 transgenic rats, the medication was interrupted between 7 and 10 days before the experiment. Previous studies documented the necessity of maintaining homozygous transgenic hypertensive rats on lisinopril from an early age to prevent early demise caused by the development of fulminant hypertension.^{13 19}

A second group of 8 homozygous transgenic-positive and 3 age-matched SD rats were injected with an Ang II monoclonal antibody (KAA8). The Ang II antibody was dissolved in phosphate-buffered saline and delivered into a brain ventricle at concentrations and volumes (1.2 µg in 2 µL, 3.0 µg in 5 µL, and 6.0 µg in 10 µL) similar to those used with the Ang-(1-7) antibody. Four of the transgenic hypertensive rats were studied while on lifetime therapy with lisinopril; the drug was discontinued in 4 other rats for up to 2 weeks.

Antibodies
Production and characterization of antibodies to Ang-(1-7) are described elsewhere.^{6 20 21 22} Briefly, rabbits were immunized by injection of synthetic Ang-(1-7) chemically coupled with glutaraldehyde to keyhole limpet hemocyanin. This antiserum (1:3500 dilution) was shown by radioimmunoassay to specifically recognize Ang-(1-7) and cross-react with Ang-(2-7) and Ang-(3-7) by an average of 10% and 18%, respectively. Cross-reactivity with Ang I, Ang-(1-6), Ang II, or Ang-(2-8) was less than 0.001%. In addition, the Ang-(1-7) antibody did not recognize bradykinin, arginine vasopressin, or substance P. The Ang-(1-7) antibody was affinity-purified on a Sepharose column prepared by coupling 5 mg Ang-(1-7) to 1 mL Affi-Gel 10, according to the manufacturer's directions. Coupling efficiency was 62%. Before affinity chromatography, the IgG fraction was extracted from 2 mL of whole antiserum with diethylaminoethyl-cellulose. The Ang-(1-7) IgG fraction was mixed overnight with the Ang-(1-7)–Sepharose in the presence of 0.5 mmol/L bacitracin. After extensive washing with phosphate-buffered saline (50 mmol/L HPO₄, 0.15 mol/L NaCl, pH 7.4), the antibody was eluted with a mixture of 50 mmol/L glycine and 50 mmol/L HCl at pH 2.2 and immediately neutralized with 1 mol/L Tris. Fractions containing antibody were pooled and dialyzed extensively against phosphate-buffered saline. After affinity purification, the slope of the log/logit plot was -0.934 for the affinity-purified antibody and -1.005 for whole-serum antibody. The total amount of binding at a 1:3500 dilution averaged 44.2% for whole serum and 18.6% for the affinity-purified antibody.

The Ang II monoclonal antibody (KAA8) was obtained from DuPont Merck Pharmaceutical Co. KAA8 is a monoclonal IgG displaying an enhanced affinity for Ang II, as demonstrated by Reilly and colleagues (Reilly et al²³ and Wong et al^{24 25} ), both in vitro and in vivo. Furthermore, substitution or deletion of the carboxy terminus of Ang II dramatically reduced the affinity of the resulting peptide for KAA8 (T.M. Reilly, unpublished observations, 1994).

Analysis of Data
All data are expressed as mean±SEM. ANOVA was used for multiple comparisons to evaluate the effects of the Ang-(1-7) and Ang II antibodies. Duncan's multiple range test was applied to assess differences whenever a level of significance was found. Student's t test and ANCOVA were used for paired and unpaired observations, respectively. The criterion for statistical significance was a value of P<.05.

	Results

Top Abstract Introduction Methods Results Discussion References

 
Baseline MAP and heart rate averaged 157±9 mm Hg and 484±20 beats per minute, respectively, in homozygous transgenic hypertensive rats weaned off lifetime lisinopril therapy. Transgenic rats maintained on lisinopril therapy had lower baseline MAP (121±9 mm Hg, P<.01) but not heart rates (441±20 beats per minute, P>.05) compared with transgenic rats off lisinopril therapy. Although transgenic hypertensive rats weighed less after cessation of lisinopril therapy, the difference was not statistically significant (266±5 g compared with 277±9 g on lisinopril, P>.05). Baseline MAP averaged 104±2 mm Hg and heart rate was 450±8 beats per minute in normotensive SD rats. The relationship between baseline MAP and heart rate with changes in these variables after administration of the antibodies was analyzed. No significant correlations were observed. ANCOVA with adjustment for baseline values produced similar results; therefore, Student's t test values are reported below.

Ang-(1-7) Antibody Elevates BP in Transgenic Rats
ICV injection of the Ang-(1-7) antibody in transgenic rats produced a prompt rise in arterial pressure that persisted for at least 5 minutes and was associated with tachycardia (Fig 1). For the group as a whole, the time to peak change in MAP averaged 22±3 seconds in transgenic rats off lisinopril and 32±5 seconds in those maintained on therapy (P>.05).

View larger version (13K): [in this window] [in a new window]   Figure 1. Tracings show pulsatile pressure (top), mean arterial pressure (middle), and heart rate (bottom) from a female homozygous transgenic hypertensive rat [(mRen-2)27] before and after injection of 3.2 µg (5 µL) of the affinity-purified antibody to angiotensin-(1-7) [Ang-(1-7)Ab] into a lateral cerebral ventricle.

Fig 2 illustrates group changes in MAP and heart rate produced by brain ventricular administration of the Ang-(1-7) antibody. Injections of the Ang-(1-7) antibody into conscious SD rats were ineffective at all doses tested. In contrast, central injections of the Ang-(1-7) antibody in transgenic rats on lisinopril produced BP increases that were statistically significant at both the 5- and 10-µL doses (Fig 2). The pressor response was accompanied by significant tachycardia even at the lowest dose (Fig 2).

View larger version (26K): [in this window] [in a new window]   Figure 2. Bar graphs show average group values of the peak change in mean arterial pressure (top) and heart rate (bottom) obtained after administration of three doses of an angiotensin-(1-7) antibody into a lateral cerebral ventricle of normotensive Hannover Sprague-Dawley (SD, n=5) and hypertensive transgenic (TG) rats on (n=6) and off (n=6) lisinopril therapy. Protein concentrations for the antibody as in "Methods." For these subgroups of TG rats, baseline mean arterial pressure averaged 135±4 and 104±8 mm Hg off and on lisinopril, respectively (P<.05). Corresponding heart rate values were 407±22 and 463±27 beats per minute for TG rats on and off lisinopril, respectively (P>.05). Baseline mean arterial pressure and heart rate values for control SD rats averaged 104±3 mm Hg and 436±7 beats per minute, respectively. *P<.05, **P<.01 vs baseline values; #P<.05, ##P<.01 compared with control SD rats.

ICV injections of the Ang-(1-7) antibody had a more prominent effect in rats off lisinopril therapy (Fig 2). In these transgenic hypertensive rats, all three doses produced significant increases in arterial pressure. Peak changes in MAP at the 5- and 10-µL doses were significantly greater (P<.05) compared with corresponding values in transgenic rats on lisinopril therapy. However, discontinuation of lisinopril therapy did not potentiate the magnitude of the cardiac rate response produced by the Ang-(1-7) antibody. The pressor actions produced by ICV injection of the Ang-(1-7) antibody were specific because administration of 2 to 10 µL of either phosphate-buffered saline (vehicle) or an affinity-purified IgG fraction had no effect on arterial pressure or heart rate of SD and transgenic rats either on or off lisinopril. Likewise, intravenous injections of the Ang-(1-7) antibody at doses as high as 100 µL caused no change in arterial pressure or heart rate.

Ang II Antibody Lowers BP in Transgenic Rats
To gain further insight into the nature and specificity of the cardiovascular response produced by central neutralization of Ang-(1-7), we injected a separate group of transgenic hypertensive and normotensive SD rats with either vehicle or an Ang II monoclonal antibody (Fig 3). In contrast to the effects obtained by neutralization of Ang-(1-7), central administration of the Ang II antibody produced significant decreases in the arterial pressure and heart rate of transgenic rats and no changes in SD controls. All three doses of the Ang II antibody produced significant peak decreases in arterial pressure in rats on or off lisinopril therapy. In addition, the maximal BP fall produced by neutralization of Ang II was significantly augmented in transgenic rats no longer given lisinopril (Fig 3). The fall in heart rate associated with the depressor response was augmented in transgenic rats weaned off lisinopril therapy at the two largest doses tested. On the other hand, central injections of the same amounts of the Ang II antibody into conscious SD rats were ineffective (Fig 3).

View larger version (29K): [in this window] [in a new window]   Figure 3. Bar graphs show averages of peak depressor (top) and bradycardic (bottom) responses produced by intracerebroventricular administration of a monoclonal angiotensin II antibody (KAA8) in conscious Hannover Sprague-Dawley (SD, n=3) and hypertensive transgenic (TG, n=8) rats. Baseline mean arterial pressure before injections of either antibody or vehicle averaged 190±4 and 147±7 mm Hg in TG rats off and on lisinopril, respectively (P<.02). Baseline heart rate averaged 514±24 beats per minute in TG rats not receiving lisinopril and 490±19 beats per minute in TG rats maintained on the drug regimen (P>.05).*P<.05, **P<.01 vs baseline values; #P<.05, ##P<.01 compared with control SD rats.

We also evaluated the possibility that differences in baseline BP among the various treatment groups influenced the magnitude and direction of the peak changes in BP produced by central injection of either the Ang-(1-7) or the Ang II antibodies. Regression analysis of the peak change in MAP as a function of the level of arterial pressure before injection of the Ang-(1-7) antibody was not statistically significant (r=.44, P>.05) in transgenic hypertensive rats on and off lisinopril therapy. The r value for the relation between peak change and baseline MAP values averaged -0.34 (P>.05) for the group of transgenic rats injected with the Ang II antibody. Furthermore, ANCOVA showed that differences in baseline BP between rats given either the Ang-(1-7) or Ang II antibodies did not influence the magnitude or direction of the change in arterial pressure (F=0.04, P=.83).

	Discussion

Top Abstract Introduction Methods Results Discussion References

 
The present study demonstrates the opposing effects of neutralization of Ang II and Ang-(1-7) in the brain of transgenic hypertensive rats. Central inhibition of endogenous Ang-(1-7) caused a hypertensive response that was augmented in rats in which hypertension was reestablished after interruption of antihypertensive therapy. The specific nature of the response produced by central neutralization of Ang-(1-7) was further underscored by the concurrent demonstration that blockade of Ang II with a specific Ang II antibody elicited a depressor response associated with bradycardia. Although past studies showed that Ang-(1-7) is biologically active,⁴ the absence of a specific antagonist hampered investigation of its role in the regulation of arterial pressure. Antibodies have proved to be important tools for assessing the actions of the renin-angiotensin system^{26 27} and verifying the effects of selective receptor antagonists.²⁸ The diametrically opposite nature of the effects produced by central neutralization of either Ang-(1-7) or Ang II now provides a compelling argument for a more intense analysis of the functions conveyed by Ang-(1-7) in the control of arterial pressure. In addition, this study showed that the angiotensin system in the brain of transgenic rats participates in the evolution of this genetic model of high BP.

The transgenic rat model of hypertension used in the present experiments depends on expression of an additional renin gene in tissues.^{13 19} As reviewed by Bader et al,¹⁹ Ang II plays a crucial role in the hypertensinogenic process in transgenic (mRen-2)27 rats by mechanisms that were not shown to be immediately obvious. The Ren-2 transgene is expressed most abundantly in the adrenal gland, but it is also present in the kidneys, vascular wall, brain, and pituitary.¹⁹ Although hypertension in transgenic rats is amenable to treatments that interfere with the formation or actions of Ang II,¹³ only plasma prorenin was found to be consistently augmented above normal levels.^{29 30} In contrast, plasma renin activity and Ang II concentrations have been reported to be unchanged¹³ or elevated above normal values.^{14 31} The earlier suggestion that the hypertension may result from overexpression of the renin gene in the adrenal glands³² has been questioned because the normalization of BP after adrenalectomy may be transient and explained by removal of steroid production.²⁹

Expression of the Ren-2 gene in the brain of transgenic hypertensive rats is associated with high concentrations of Ang II and Ang-(1-7).¹⁴ Moreover, ICV infusions of Ang II caused no increases in BP or release of vasopressin from the paraventricular and supraoptic nuclei of transgenic hypertensive rats.¹⁵ A similar desensitization of centrally mediated responses to Ang-(1-7) paralleled these findings.¹⁵ These data suggested that maintenance of high BP in this transgene model of hypertension is related to the high level of expression of the Ren-2 gene in the brain. The hemodynamic responses produced by central injections of antibodies to Ang II and Ang-(1-7) are in keeping with this interpretation. In addition, potentiation of the hemodynamic response to the antibodies after cessation of lisinopril administration favors this conclusion. The larger depressor response obtained in transgenic rats off lisinopril may be accounted for by restoration of higher levels of brain Ang II. Although we did not measure brain Ang II levels in transgenic rats off lisinopril therapy, other data support this explanation. Angiotensin-converting enzyme activity is inhibited in the cerebrospinal fluid of spontaneously hypertensive rats (SHR) and Wistar-Kyoto rats during oral administration of similar doses of lisinopril.⁶ This effect was associated with a reduction in the density of Ang II receptors in the dorsal medulla oblongata of SHR.³³

The Ang-(1-7) antisera used in the present experiments belongs to a bank of antibodies that was extensively characterized in our laboratory for their specificity to bind Ang-(1-7) and absence of cross-reactivity with Ang II and other centrally acting neuropeptides.³⁴ Moreover, central injections of another affinity-purified IgG fraction had no effects on the BP and heart rate of transgenic hypertensive rats. Likewise, central injections of the Ang-(1-7) antibody were ineffective when given into normotensive SD rats. In contrast, neutralization of Ang II in transgenic rats produced hemodynamic effects opposite to those obtained with the Ang-(1-7) antibody. The monoclonal Ang II antibody used in these experiments was also extensively characterized both in vivo and in vitro.^{23 24 25}

Accumulating evidence suggests that Ang-(1-7) may oppose the actions of Ang II by mechanisms that are still a subject of investigation. Ang-(1-7) acts as a vasodilator in the mesenteric,³⁵ cerebral,³⁶ coronary,⁵ and systemic⁹ circulations. Long-term Ang-(1-7) infusions were reported recently to lower the BP of SHR for up to 2 days.³⁷ Previous studies showed that Ang-(1-7) stimulates the release of vasodilator prostaglandins in endothelial³⁸ and vascular smooth muscle³⁹ cells in culture, as well as the vas deferens in rabbits¹¹ and piglet pial arteries.³⁶ In felines and swine,⁵ however, alternate systems may be involved because the vasodilator actions of Ang-(1-7) were prevented after inhibition of nitric oxide synthase.³⁵ Although intermediate mechanisms for the actions of Ang-(1-7) in the brain have not been identified, the peptide is a selective and potent stimulus for the release of prostaglandin E₂ and the prostacyclin metabolite 6-ketoprostaglandin F₁ in human astrocytes in culture.¹² The demonstration that neutralization of Ang-(1-7) causes central activation of vasopressor activity in hypertensive animals with increased activity of the renin-angiotensin system provides a base for the further study of its action. On the basis of the evidence discussed above, it is possible that Ang-(1-7) may oppose the excitatory actions of Ang II in the brain by activation of a central prostaglandin or nitric oxide pathway engaged in a negative feedback regulation of the vasomotor actions of Ang II.

There is some foundation for the possibility that Ang-(1-7) may act in the brain to modulate BP. The peptide facilitates neuronal discharges both in vivo and in vitro.⁴⁰ In addition, Webb et al⁴¹ found that ICV administration of Z pro-prolinal, a specific inhibitor of prolyl endopeptidase,^{7 42 43} produced a substantial rise in the BP of SHR. Prolyl endopeptidase (EC 3.4.21.26) generated Ang-(1-7) from both Ang I and Ang II in dog brain.⁴³ Although it is recognized that mechanisms participating in the expression of SHR and Ren-2 gene hypertensions may differ, there is evidence for the participation of Ang II in the brain of SHR.⁴⁴

In conclusion, the (mRen-2)27 model of transgene hypertension provided a way for evaluation of the contribution of the brain renin-angiotensin system to the pathogenesis of arterial hypertension. In addition, the hemodynamic effects obtained with the Ang-(1-7) antibody confirmed that angiotensin peptides devoid of an amino acid in the eighth position of the Ang II molecule do not lose bioactivity and are able to interact with angiotensin receptors by still unknown mechanisms.²⁹ Discrimination of the mechanisms that account for the opposing actions of Ang II and Ang-(1-7) neutralization should define the contribution of brain angiotensins to the BP regulation in normotensive and hypertensive states.

	Acknowledgments

 
This research was supported in part by grants HL-50066 and 1PO1-HL-51952 from the National Heart, Lung, and Blood Institute of the National Institutes of Health, Bethesda, Md. We thank Dr Charles Sweet, PhD, and Merck Human Health Division for their generous assistance in supplying lisinopril for the breeding colony of transgenic hypertensive rats at the Bowman Gray School of Medicine. We thank Dr Timothy Morgan of the Section of Biostatistics, Department of Public Health Sciences at Bowman Gray School of Medicine, for his contribution to the analysis of the data.

	Footnotes

 
Reprint requests to Carlos M. Ferrario, MD, Hypertension Center, The Bowman Gray School of Medicine, Medical Center Boulevard, Winston-Salem, NC 27157-1032.

Received August 11, 1994; first decision January 17, 1995; accepted February 21, 1995.

	References

Top Abstract Introduction Methods Results Discussion References

 
1. Schunkert H, Ingelfinger JR, Jacob H, Jackson B, Bouyounes B, Dzau VJ. Reciprocal feedback regulation of kidney angiotensinogen and renin mRNA expressions by angiotensin II. Am J Physiol. 1992;263:E863-E869.

2. Schunkert H, Hirsch AT, Pinto M, Pelletier P, Jacob H, Remme WJ, Ingelfinger JR, Dzau VJ. Feedback regulation of angiotensin converting enzyme mRNA and activity by angiotensin II. Circulation. 1990;82(suppl III):III-230. Abstract.

3. Timmermans PBMWM, Benfield P, Chiu AT, Herblin WF, Wong PC, Smith RD. Angiotensin II receptors and functional correlates. Am J Hypertens. 1992;5:221S-235S. [Medline] [Order article via Infotrieve]

4. Ferrario CM. Biological roles of angiotensin-(1-7). Hypertens Res. 1992;15:61-66.

5. Porsti I, Bara AT, Busse R, Hecker M. Release of nitric oxide by angiotensin-(1-7) from porcine coronary endothelium: implications for a novel angiotensin receptor. Br J Pharmacol. 1994;111:652-654. [Medline] [Order article via Infotrieve]

6. Kohara K, Brosnihan KB, Ferrario CM. Angiotensin-(1-7) in the spontaneously hypertensive rat. Peptides. 1993;14:883-891. [Medline] [Order article via Infotrieve]

7. Santos RAS, Brosnihan KB, Jacobsen DW, DiCorleto PE, Ferrario CM. Production of angiotensin-(1-7) by human vascular endothelium. Hypertension. 1992;19(suppl II):II-56-II-61.

8. Schiavone MT, Santos RAS, Brosnihan KB, Khosla MC, Ferrario CM. Release of vasopressin from the rat hypothalamo-neurohypophysial system by angiotensin-(1-7) heptapeptide. Proc Natl Acad Sci U S A. 1988;85:4095-4098. [Abstract/Free Full Text]

9. Benter IF, Diz DI, Ferrario CM. Cardiovascular actions of angiotensin-(1-7). Peptides. 1993;14:679-684. [Medline] [Order article via Infotrieve]

10. DelliPizzi A, Hilchey SD, Bell-Quilley CP. Natriuretic actions of angiotensin-(1-7). Br J Pharmacol. 1994;111:1-3. [Medline] [Order article via Infotrieve]

11. Trachte GJ, Meixner K, Ferrario CM, Khosla MC. Prostaglandin production in response to angiotensin-(1-7) in rabbit isolated vasa deferentia. Prostaglandins. 1990;39:385-394. [Medline] [Order article via Infotrieve]

12. Tallant EA, Jaiswal N, Diz DI, Ferrario CM. Human astrocytes contain two distinct angiotensin receptor subtypes. Hypertension. 1991;18:32-39.[Abstract/Free Full Text]

13. Mullins JJ, Peters J, Ganten D. Fulminant hypertension in transgenic rats harbouring the mouse Ren-2 gene. Nature. 1990;344:541-544. [Medline] [Order article via Infotrieve]

14. Senanayake PD, Moriguchi A, Kumagai H, Ganten D, Ferrario CM, Brosnihan KB. Increased expression of angiotensin peptides in the brain of transgenic hypertensive rats. Peptides. 1994;15:919-926. [Medline] [Order article via Infotrieve]

15. Moriguchi A, Ferrario CM, Brosnihan KB, Ganten D, Morris M. Differential regulation of central vasopressin in transgenic rats harboring the mouse Ren-2 gene. Am J Physiol. 1994;267:R786-R791. [Abstract/Free Full Text]

16. Kawabe H, Husain A, Khosla MC, Smeby RR, Bumpus FM, Ferrario CM. Characterization of receptors for angiotensin-induced drinking and blood pressure responses in conscious rats using angiotensin analogs extended at the N-terminal. Neuroendocrinology. 1986;42:289-295. [Medline] [Order article via Infotrieve]

17. Kumagai H, Averill DB, Khosla MC, Ferrario CM. Role of nitric oxide and angiotensin II in the regulation of sympathetic nerve activity in spontaneously hypertensive rats. Hypertension. 1993;21:476-484. [Abstract/Free Full Text]

18. Wallace RW, Tallant EA, McManus MC. Human platelet calmodulin-binding proteins: identification and Ca²⁺-dependent proteolysis upon platelet activation. Biochemistry. 1987;26:2766-2773. [Medline] [Order article via Infotrieve]

19. Bader M, Lee MA, Zhao Y, Bohm M, Bachmann J, Sander M, Djavidani B, Bachmann S, Zimmermann F, Wilbertz J, Zeh K, Wagner J, Peters J, Ganten D. Renin gene expression and hypertension in transgenic animals. In: Raizada MK, Phillips MI, Sumners C, eds. Cellular and Molecular Biology of the Renin-Angiotensin System. Boca Raton, Fla: CRC Press; 1993:59-93.

20. Chappell MC, Milsted A, Diz DI, Brosnihan KB, Ferrario CM. Evidence for an intrinsic angiotensin system in the canine pancreas. J Hypertens. 1991;9:751-759. [Medline] [Order article via Infotrieve]

21. Chappell MC, Brosnihan KB, Diz DI, Ferrario CM. Identification of angiotensin-(1-7) in rat brain: evidence for differential processing of angiotensin peptides. J Biol Chem. 1989;264:16518-16523. [Abstract/Free Full Text]

22. Chappell MC, Brosnihan KB, Welches WR, Ferrario CM. Characterization by high performance liquid chromatography of angiotensin peptides in the plasma and cerebrospinal fluid of the dog. Peptides. 1987;8:939-942. [Medline] [Order article via Infotrieve]

23. Reilly TM, Chiu AT, Timmermans PBMWM. Monoclonal antibodies to angiotensin II. Biochem Biophys Res Commun. 1987;143:133-139. [Medline] [Order article via Infotrieve]

24. Wong PC, Price WA, Reilly TM, Duncia JV, Timmermans PBMWM. Antihypertensive mechanism of captopril in renal hypertensive rats: studies with a nonpeptide angiotensin II receptor antagonist and an angiotensin II monoclonal antibody. J Pharmacol Exp Ther. 1989;250:515-522. [Abstract/Free Full Text]

25. Wong PC, Reilly TM, Timmermans PBMWM. Effect of a monoclonal antibody to angiotensin II on hemodynamic responses to noradrenergic stimulation in pithed rats. Hypertension. 1989;14:488-497. [Abstract/Free Full Text]

26. Wong PC, Reilly TM, Timmermans PBMWM. Angiotensin II monoclonal antibody: blood pressure effects in normotensive and spontaneously hypertensive rats. Eur J Pharmacol. 1990;186:353-356. [Medline] [Order article via Infotrieve]

27. Reilly TM, Christ DD, Duncia JV, Pierce SK, Timmermans PBMWM. Monoclonal antibodies to the nonpeptide angiotensin II receptor antagonist, losartan. Eur J Pharmacol. 1992;226:179-182. [Medline] [Order article via Infotrieve]

28. Reilly TM, Wong PC, Price WA, Timmermans PBMWM. Characterization of the functional antagonism and antihypertensive activity displayed by a monoclonal antibody to angiotensin II. J Pharmacol Exp Ther. 1988;244:160-165. [Abstract/Free Full Text]

29. Bader M, Wagner J, Ganten D. The role of the renin-angiotensin system in cardiovascular disease. Hypertens Res. 1994;17:1-16.

30. Moriguchi A, Brosnihan KB, Kumagai H, Ganten D, Ferrario CM. Mechanisms of hypertension in transgenic rats expressing the mouse Ren-2 gene. Am J Physiol. 1994;266:R1273-R1278. [Abstract/Free Full Text]

31. Tokita Y, Franco-Saenz R, Mulrow PJ, Ganten D. Effects of nephrectomy and adrenalectomy on the renin-angiotensin system of transgenic rats TGR (mRen2) 27. Endocrinology. 1994;134:253-257. [Abstract/Free Full Text]

32. Sander M, Bader M, Djavidani B, Maser-Gluth C, Vecsel P, Mullins J, Ganten D, Peters J. The role of the adrenal gland in hypertensive transgenic rat TGR (mREN2)27. Endocrinology. 1992;131:807-814. [Abstract/Free Full Text]

33. Diz DI, Kohara K, Ferrario CM. Normalization of angiotensin (Ang) II receptors in the dorsal medulla oblongata of spontaneously hypertensive rats (SHR) follows converting enzyme inhibition and increases in plasma angiotensin-(1-7) concentrations. Am J Hypertens. 1992;5:16A. Abstract.

34. Kohara K, Tabuchi Y, Senanayake P, Brosnihan KB, Ferrario CM. Reassessment of plasma angiotensins measurement: effects of protease inhibitors and sample handling procedures. Peptides. 1991;12:1135-1141. [Medline] [Order article via Infotrieve]

35. Osei SY, Ahima RS, Minkes RK, Weaver JP, Khosla MC, Kadowitz PJ. Differential responses to angiotensin-(1-7) in the feline mesenteric and hindquarters vascular beds. Eur J Pharmacol. 1993;234:35-42. [Medline] [Order article via Infotrieve]

36. Meng W, Busija DW. Comparative effects of angiotensin-(1-7) and angiotensin II on piglet pial arterioles. Stroke. 1993;24:2041-2045. [Abstract/Free Full Text]

37. Benter IF, Ferrario CM, Morris M, Diz D. Chronic intravenous angiotensin-(1-7) infusions activate antihypertensive mechanisms in spontaneously hypertensive rats. Am J Hypertens. 1994;7:94A. Abstract.

38. Jaiswal N, Diz DI, Chappell MC, Khosla MC, Ferrario CM. Stimulation of endothelial cell prostaglandin production by angiotensin peptides: characterization of receptors. Hypertension. 1992;19(suppl II):II-49-II-55.

39. Jaiswal N, Jaiswal RK, Tallant EA, Diz DI, Ferrario CM. Alterations in prostaglandin production in spontaneously hypertensive rat smooth muscle cells. Hypertension. 1993;21:900-905. [Abstract/Free Full Text]

40. Barnes KL, Knowles WD, Ferrario CM. Angiotensin II and angiotensin-(1-7) excite neurons in the canine medulla in vitro. Brain Res Bull. 1990;24:275-280. [Medline] [Order article via Infotrieve]

41. Webb RL, Bazil MK, Graybill S, Lappe RW. Hemodynamic effect of inhibition of brain prolyl endopeptidase in spontaneously hypertensive rats. Pharmacologist. 1991;33:198. Abstract.

42. Welches WR, Brosnihan KB, Ferrario CM. A comparison of the properties, and enzymatic activity of three angiotensin processing enzymes: angiotensin converting enzyme, prolyl endopeptidase and neutral endopeptidase 24.11. Life Sci. 1993;52:1461-1480. [Medline] [Order article via Infotrieve]

43. Welches WR, Santos RAS, Chappell MC, Brosnihan KB, Greene LJ, Ferrario CM. Evidence that prolyl endopeptidase participates in the processing of brain angiotensin. J Hypertens. 1991;9:631-638. [Medline] [Order article via Infotrieve]

44. Phillips MI, Speakman EA, Kimura B. Levels of angiotensin and molecular biology of the tissue renin angiotensin systems. Regul Pept. 1993;43:1-20.[Medline] [Order article via Infotrieve]

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