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Am J Physiol Heart Circ Physiol 288: H1063-H1070, 2005. First published November 18, 2004; doi:10.1152/ajpheart.01163.2003
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Estrogen receptor-{alpha} mediates estrogen facilitation of baroreflex heart rate responses in conscious mice

Jaya Pamidimukkala,1 Baojian Xue,1 Leslie G. Newton,4 Dennis B. Lubahn,3,4,5 and Meredith Hay1,2,3

1Dalton Cardiovascular Research Center, 2Department of Veterinary Biomedical Sciences, 3Center for Gender Physiology, 4Department of Biochemistry and Child Health, 5University of Missouri Center for Phytonutrient and Phytochemical Studies, University of Missouri, Columbia, Missouri

Submitted 9 December 2003 ; accepted in final form 6 November 2004


   ABSTRACT
TOP
 ABSTRACT
MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
Estrogen facilitates baroreflex heart rate responses evoked by intravenous infusion of ANG II and phenylephrine (PE) in ovariectomized female mice. The present study aims to identify the estrogen receptor subtype involved in mediating these effects of estrogen. Baroreflex responses to PE, ANG II, and sodium nitroprusside (SNP) were tested in intact and ovariectomized estrogen receptor-{alpha} knockout (ER{alpha}KO) with (OvxE+) or without (OvxE–) estrogen replacement. Wild-type (WT) females homozygous for the ER{alpha}+/+ were used as controls. Basal mean arterial pressures (MAP) and heart rates were comparable in all the groups except the ER{alpha}KO-OvxE+ mice. This group had significantly smaller resting MAP, suggesting an effect of estrogen on resting vascular tone possibly mediated by the ER{beta} subtype. Unlike the WT females, estrogen did not facilitate baroreflex heart rate responses to either PE or ANG II in the ER{alpha}KO-OvxE+ mice. The slope of the line relating baroreflex heart rate decreases with increases in MAP evoked by PE was comparable in ER{alpha}KO-OvxE– (–6.97 ± 1.4 beats·min–1·mmHg–1) and ER{alpha}KO-OvxE+ (–6.18 ± 1.3) mice. Likewise, the slope of the baroreflex bradycardic responses to ANG II was similar in ER{alpha}KO-OvxE– (–3.87 ± 0.5) and ER{alpha}KO-OvxE+(–2.60 ± 0.5) females. Data suggest that estrogen facilitation of baroreflex responses to PE and ANG II is predominantly mediated by ER{alpha} subtype. A second important observation in the present study is that the slope of ANG II-induced baroreflex bradycardia is significantly blunted compared with PE in the intact as well as the ER{alpha}KO-OvxE+ females. We have previously reported that this ANG II-mediated blunting of cardiac baroreflexes is observed only in WT males and not in ovariectomized WT females independent of their estrogen replacement status. The present data suggest that in females lacking ER{alpha}, ANG II causes blunting of cardiac baroreflexes similar to males and may be indicative of a direct modulatory effect of the ER{alpha} on those central mechanisms involved in ANG II-induced resetting of cardiac baroreflexes. These observations suggest an important role for ER{alpha} subtype in the central modulation of baroreflex responses. Lastly, estrogen did not significantly affect reflex tachycardic responses to SNP in both WT and ER{alpha}KO mice.

autonomic regulation; cardiac baroreflexes; hormone replacement; angiotensin II

SEX DIFFERENCES IN THE DEVELOPMENT of hypertension and coronary heart disease are well documented with cardiovascular disease manifesting in women mainly during postmenopausal years when estrogen levels are low (8, 27). Mechanisms by which estrogen protects against hypertension are incompletely understood but are thought to include improvement of lipid profile, decreases in vascular resistance, and modulation of central nuclei involved in cardiovascular regulation (14, 19, 37, 38, 42). In a number of experimental models of hypertension, it has been shown that ovariectomy exacerbates and estrogen replacement attenuates the course of hypertension (5, 12, 13, 28). Recent studies from our lab have also shown that chronic ANG II infusion causes greater increases in blood pressures in ovariectomized female mice compared with intact females suggesting that female sex hormones protect against the development of hypertension (46, 47). In most experimental models, development of hypertension is accompanied by either a reduction in baroreflex sensitivity and/or a resetting of the baroreflex curve toward higher pressures. In the ANG II-dependent hypertension model, development of hypertension is attributed to a centrally mediated increase in sympathetic outflow. ANG II has also been shown to inhibit reflex decreases in heart rate and sympathetic outflow that would normally accompany such large increases in blood pressure, thereby facilitating the maintenance of the elevated blood pressure (21, 24, 34). A potential mechanism by which estrogen could contribute to cardioprotection in females is by antagonizing these centrally mediated effects of ANG II on baroreflex function. We have previously shown that in chronically instrumented, awake mice ANG II-mediated blunting of baroreflex bradycardic responses is observed only in males but not in females (30). In this study, in the male mice ANG II-evoked baroreflex bradycardic responses were significantly blunted compared with those evoked by similar increases in blood pressure with phenylephrine (PE). In contrast, in ovariectomized female mice either with or without estrogen replacement, reflex heart rate responses to ANG II were similar to that of PE (30). These data suggested that ANG II-mediated blunting of the baroreflex heart rate responses is sex dependent and may be independent of the circulating estrogen levels. Although ANG II and PE responses were not different within the groups in this study, estrogen replacement did facilitate reflex bradycardic responses to both ANG II and PE compared with ovariectomized females without estrogen replacement (30). Very little is known about the receptors involved in mediating these effects of estrogen on baroreflex function. The biological actions of estradiol are thought to be mediated by two types of estrogen receptor (ER)-{alpha} and -{beta} (2, 10). ERs have been studied intensely in female reproductive physiology, but functional ERs are also present and physiologically important in other tissues, including the brain and the cardiovascular system (22). There is high sequence homology in the ligand binding domains of ER{alpha} and ER{beta}, making it difficult to develop ER antagonists specific to each subtype. A second strategy often used to delineate receptor function involves the use of transgenic mouse models lacking one or the other ER subtype to differentiate between ER{alpha}- and ER{beta}-mediated effects. The purpose of the present study is to determine the effects of estrogen on baroreflex function in mice lacking the ER{alpha} receptor (ER{alpha}KO) and to identify the receptor subtype(s) involved in mediating estrogen facilitation of PE and ANG II evoked reflex bradycardia.


   MATERIALS AND METHODS
TOP
 ABSTRACT
 MATERIALS AND METHODS
RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 
ER{alpha}KO mice were obtained by mating mice of a mixed c57BL6J/129SV background that were heterozygous for the ER{alpha} gene disruption as described previously (20). Briefly, the ER{alpha}KO mouse is generated by disrupting the start codon and amino terminal domain of the ER{alpha} gene yielding a small expression of incomplete ER{alpha} transcripts but no functional {alpha}-subtype receptors (1). After genotyping, only homozygous ER{alpha}KO (ER{alpha}–/–) females and homozygous (ER{alpha}+/+) wild-type (WT) females were used in these experiments. All the mice were obtained from a breeding colony maintained in the animal care facility at the University of Missouri. The mice were 28–32 wk old and on an average weighed 20–24 g. The mice were fed a soy- based Purina 5001 lab chow (PMI Feeds, St. Louis, MO). The University of Missouri Animal Care and Use Committee approved all the protocols.

Experimental Groups

All experiments were performed in female mice. Baroreflex heart rate responses to ANG II and PE were first tested in ER{alpha}KO mice (n = 6, age = 29 ± 2 wk, weight = 24 ± 3 g,) and WT (n = 7, age = 28 ± 2 wk, weight = 22 ± 1 g) controls with intact gonads. However, observations in intact female ER{alpha}KO mice are complicated by the presence of high circulating levels of estrogen and testosterone, an effect of ER deletion on the reproductive system and endocrine feedback mechanisms (2). Hence, subsequent experiments were carried out in ovariectomized ER{alpha}KO mice implanted with Silastic capsules containing 17{beta}-estradiol dissolved in corn oil (ER{alpha}KO-OvxE+, n = 7, 29 ± 4 wk, 23 ± 3 g) or corn oil alone (ER{alpha}KO-OvxE–, n = 6, 31 ± 2 wk, 22 ± 3 g). Control groups consisted of ovariectomized WT mice implanted with Silastic capsules containing 17{beta}-estradiol dissolved in corn oil (WT-OvxE+, n = 9, 30 ± 2 wk, 22 ± 1 g) or corn oil alone (WT-OvxE–, n = 6, 28 ± 3 wk, 23 ± 2 g).

Surgical Procedures

All surgical procedures were carried out in mice anesthetized with a mixture of ketamine (100 mg/kg ip) and xylazine (10 mg/kg ip) and supplemented with isoflurane when necessary.

Ovariectomy and Capsule Implantation

Ten days before catheter implantation, the mice were anesthetized, and the ovaries were exposed via a dorsal midline incision. After removal of the ovaries, a single Silastic capsule containing either 17{beta}-estradiol (10 µg) or the vehicle corn oil was implanted subcutaneously, and the incision was closed. For the purpose of making Silastic capsules, a 14-mm length of Silastic tubing (0.062 in. ID, 0.125 in. OD; Dow Corning) was filled with 20 µl solution of estradiol dissolved in tocopherol-stripped corn oil (0.5 µg/µl). Both ends were sealed with Silastic adhesive (Konigsberg Instruments), and the final length of the tubing containing estradiol or the vehicle was 10 mm. Capsules were incubated in 0.9% NaCl overnight at room temperature before being implanted in mice to provide a stable plasma estradiol concentration and to prevent a transitory peak in the plasma estrogen that would otherwise occur. These implants result in 21 ± 2 pg/ml levels of plasma estrogen in ovariectomized WT mice compared with the 8.6 ± 0.6 pg/ml in vehicle-treated mice (30). Ovariectomy reduces plasma estrogen levels in ER{alpha}KO below 5 pg/ml (32). Estrogen implants are expected to produce plasma estrogen levels similar to that measured in the WT mice. The plasma estrogen levels in intact WT and ER{alpha}KO mice previously have been reported to be ~20–40 pg/ml and 200–300 pg/ml, respectively (1, 3).

Chronic Catheterization

At least 4 days before experiments, mice were again anesthetized with ketamine/xylazine mixture and surgically instrumented with intra-arterial catheters for direct measurement of pulsatile and mean arterial pressure (MAP) and heart rate. Intravenous catheters were inserted for administration of drugs. Catheters made of microrenethane tubing (catalog no. MRE25; Braintree Scientific, Braintree, MA) were inserted into the left femoral artery and right femoral vein. The catheters were then tunneled subcutaneously, exteriorized, and placed in a plastic protective tube sutured in place at the back of the neck. The exterior catheters were heat sealed until use. The mice were housed individually in autoclaved filter-top cages and allowed to recover from the surgery (3–4 days) before baroreflex function was tested. In the interim, catheters were flushed daily (100–200 µl) with dilute sterile heparinized saline (25 U/ml) to maintain patency and resealed. Surgery did not produce significant changes in body weight in the experimental animals. Average weights for all groups on days of the surgery and experiment were 22 ± 5 and 23 ± 2 g, before and after surgery, respectively. The mice were conscious and unrestrained in their individual cages, while the exteriorized catheters were being connected to polyethylene tubing filled with heparinized saline for flushing. For blood pressure recordings, the arterial catheter was connected via polyethylene tubing filled with heparinized saline to a pressure transducer (model MLT0698; AD Instruments) placed at the level of the heart. Drugs were infused intravenously by a calibrated Razel pump modified for infusion of small volumes. Infusion rates (0.002–0.003 ml/min, 1-ml syringe) were monitored such that blood pressure was increased or decreased 30–40 mmHg over a 30- to 45-s period. The mice remained in individual cages throughout the study and were handled only while replacing the bedding (twice/week). All protocols were carried out between 0900 and 1500 hrs.

Experimental Protocol

Resting MAPs and heart rates. Resting blood pressures and heart rates were measured daily in conscious, freely moving mice. Baroreflex testing was in general carried out 4–5 days after surgery when the blood pressure readings were stable.

Evaluation of cardiac baroreflexes. On the day of the experiment, before any experimental intervention, the mice were allowed to stabilize for at least 60 min after which a baseline recording of blood pressure was obtained. Arterial pressure was elevated with increasing doses of either intravenously administered PE (6 µg/min) or ANG II (0.6 µg/min) for 30- to 45-s periods, and baroreflex curves were constructed relating MAP and heart rate. Infusion of the pressor agent was terminated immediately after a 30-mmHg increase in blood pressure was achieved. After the blood pressure and heart rate returned to baseline, the venous catheter was flushed with a small volume (100–200 µl) of saline before another line was attached with a different pressor agent. The mice were allowed to recover for 45–60 min before subsequent drug administration, with care taken that the basal blood pressures and heart rates were within ±5% of the resting values. Infusions of the pressor agents were in random order. Reflex tachycardic responses to decreases in blood pressure with sodium nitroprusside (SNP, 6.0 µg/min) were measured at the end of the experiments. A PowerLab data acquisition system (AD instruments) with a sampling rate of 4,000/s was used to record blood pressures and heart rates during the entire baroreflex function curve.

Data Analysis

Baroreflex sensitivity was estimated by calculating the slope of regression line relating increases in arterial pressure with PE and ANG II to decreases in heart rate. Control values for arterial pressure and heart rate were taken as the 2-min average before the drug infusions. Seven points were used to construct each baroreflex curve. After averaging MAP and heart rate before infusion of the drug, the heart rates were obtained at 5-mmHg increments of MAP during drug infusion for a total of 30 mmHg increase (from the PowerLab chart traces). The heart rate changes were calculated and plotted in Excel and fitted with a linear regression line for each experiment, and the slope value was calculated. The heart rate and blood pressure changes were well correlated in all our experiments (r2 range = 0.85–0.93). After the slope value for each experiment was obtained, the average values for the groups were determined (as shown in Table 1). The data were represented as means ± SE. Slopes of baroreflex curves for PE and ANG II within groups and between groups were compared with one-way ANOVA. The probability level of P < 0.05 was considered to be statistically significant.


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  		Table 1. Slopes of baroreflex responses to PE, ANG II, and SNP in WT and ER{alpha}KO females

 

   RESULTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
DISCUSSION
 GRANTS
 REFERENCES
 
Resting MAP and heart rate in the intact ER{alpha}KO mice tended to be smaller compared with WT mice, but the differences were not statistically significant (Table 2). However, resting pressures in ER{alpha}KO-OvxE+ mice (95 ± 3 mmHg) were slightly but significantly (P < 0.05) smaller compared with WT-OvxE+ mice (107 ± 4 mmHg), which may be an indication of estrogen-mediated relaxation of vascular tone via ER{beta} subtype. MAP and heart rate in ER{alpha}KO-OvxE– were not different from WT.


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  		Table 2. Resting MAP and HR in WT and ER{alpha}KO females

 
Baroreflex Responses in Gonadally Intact WT and ER{alpha}KO Mice

Reflex decreases in heart rate evoked by progressive increases in pressure with intravenous infusions of PE and ANG II in intact WT and ER{alpha}KO mice are shown in Fig. 1, A and B, respectively. Increases in MAP with PE resulted in similar reflex decreases in heart rate and similar slope values of baroreflex curves (–5.70 ± 1.2 vs. –6.8 ± 1.4 beats·min–1·mmHg–1, P = 0.07, Table 1) in the intact WT and ER{alpha}KO mice, respectively. Baroreflex heart rate responses to ANG II were, however, significantly blunted in the ER{alpha}KO mice compared with WT mice. The regression lines fit through the baroreflex curves had significantly smaller slope values in the ER{alpha}KO (–3.5 ± 1.2) compared with the WT (–5.60 ± 0.3, P = 0.043) mice.



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  		Fig. 1. Mean regression lines relating reflex decreases in heart rate (HR) to increases in mean atrial pressure (MAP) evoked by intravenous infusions of phenylephrine (PE) (A) and ANG II (B) in gonadally intact wild-type (WT) and estrogen receptor-{alpha} knockout (ER{alpha}KO) mice. Circles and triangles, mean HR changes for each group. {dagger}Significant differences in slope values between the two groups by ANOVA, P < 0.05.

 
Estrogen Modulation of Baroreflex Responses to PE in Ovariectomized ER{alpha}KO Mice

Estrogen replacement facilitated baroreflex heart rate responses to PE in the ovariectomized WT mice (Fig. 2A). The slope values in WT-OvxE+(–8.71 ± 1.6 beats·min–1·mmHg–1) are significantly greater compared with WT-OvxE– (–4.89 ± 0.3 beats·min–1·mmHg–1, P = 0.02, Table 1). On the other hand, baroreflex heart responses to PE in ER{alpha}KO-OvxE+ (slope: –6.18 ± 1.3 beats·min–1·mmHg–1) and ER{alpha}KO-OvxE– (slope: –6.97 ± 1.4) mice were very similar (Fig. 2B, Table 1), suggesting that ER{alpha} is involved in estrogen-mediated facilitation of reflex responses to PE.



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  		Fig. 2. Mean regression lines relating baroreflex decreases in HR evoked by PE in ovariectomized WT (A) and ER{alpha}KO (B) ovariectomized mice with estrogen replacement (OvxE+) or without (OvxE–) estrogen replacement. Open and filled circles, the mean HR changes in OvxE– and OvxE+ groups, respectively. {dagger}Significant differences in slope values between the two groups by ANOVA, P < 0.05.

 
Estrogen Modulation of Baroreflex Responses to ANG II in Ovariectomized ER{alpha}KO Mice

Estrogen replacement also facilitated reflex bradycardic responses to ANG II In ovariectomized WT mice (Fig. 3A). The slope of ANG II baroreflex curve in WT-OvxE+ (–7.26 ± 1.2 beats·min–1·mmHg–1) is significantly greater than that of WT-OvxE– (–4.2 ± 1.1, P = 0.02; Table 1). No such facilitation was observed in ovariectomized ER{alpha}KO mice (Fig. 3B), in which baroreflex responses to ANG II in ER{alpha}KO-OvxE+ (slope: –2.60 ± 0.5) and ER{alpha}KO-OvxE– (slope: –3.87 ± 0.5, Table 1) mice were similar. The data suggest that ER{alpha} expression is necessary for estrogen-mediated facilitation of ANG II evoked reflex bradycardic responses.



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  		Fig. 3. Mean regression lines relating baroreflex decreases in HR evoked by ANG II in ovariectomized WT (A) and ER{alpha}KO (B) OvxE+ and OvxE– mice. Open and filled triangles indicate the mean HR changes in OvxE– and OvxE+ groups, respectively. {dagger}Significant differences in slope values between the two groups by ANOVA, P < 0.05.

 
Baroreflex Responses to ANG II Compared With PE in ER{alpha}KO and WT

Figure 4, A and B, compares baroreflex decreases in heart rate evoked by ANG II to those evoked by PE in intact WT and ER{alpha}KO mice, respectively. In intact WT mice, increases in MAP with ANG II and PE resulted in similar reflex decreases in heart rate and similar slope values of baroreflex curves (–5.60 ± 0.3 vs. –5.70 ± 1.2 beats·min–1·mmHg–1, Table 1). These data show an absence of ANG II-mediated resetting of baroreflex control of heart rate responses in WT females. Similar observations were also made in ovariectomized WT females without estrogen replacement (Fig. 5A) or with estrogen replacement (Fig. 6A).



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  		Fig. 4. Comparison of baroreflex HR responses to increases in MAP evoked by PE and ANG II in gonadally intact WT (A) and ER{alpha}KO (B) mice. Circles and triangles, mean HR changes for PE and ANG II, respectively. Straight lines, average regression lines fit through the data points. {dagger}Significant differences in slope values between the two groups by ANOVA, P < 0.05.

 


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  		Fig. 5. Comparison of baroreflex HR responses to increases in MAP evoked by PE and ANG II in OvxE– WT (A) and ER{alpha}KO (B) mice. Circles and triangles, mean HR changes for PE and ANG II, respectively. Straight lines, average regression lines fit through the data points. {dagger}Significant differences in slope values between the two groups by ANOVA, P < 0.05.

 


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  		Fig. 6. Comparison of baroreflex HR responses to increases in MAP evoked by PE and ANG II in OvxE+ WT (A) and ER{alpha}KO (B) mice. Circles and triangles, mean HR changes for PE and ANG II, respectively. Straight lines, average regression lines fit through the data points. {dagger}Significant differences in slope values between the two groups by ANOVA, P < 0.05.

 
In contrast, in intact ER{alpha}KO mice, reflex decreases in heart rate evoked by ANG II were significantly smaller compared with PE and is reflected in the slope values (–3.5 ± 1.2 vs. –6.8 ± 1.4, P = 0.026, Table 1). This shows that in the absence of ER{alpha}, ANG II-mediated central resetting of cardiac baroreflexes becomes apparent in the female mice.

Reflex bradycardic responses to ANG II were much smaller in ER{alpha}KO-OvxE– mice, in which the slope of ANG II (–3.87 ± 0.5) baroreflex curve was significantly blunted compared with PE (–6.97 ± 1.4, P = 0.026, Table 1). Similar responses were observed in ER{alpha}KO-OvxE+, the slope values for ANG II and PE baroreflex curves being –2.60 ± 0.5 and –6.18 ± 1.3, respectively (P = 0.035, Table 1, Fig. 6B). These data suggest that ANG II-mediated central resetting of cardiac baroreflexes observed in ER{alpha}KO mice is independent of the circulating levels of estrogen.

Baroreflex Responses to SNP

Slope values of baroreflex curves relating decreases in MAP to increases in heart rate during intravenous infusions of SNP in WT and ER{alpha}KO are shown in Table 1. No differences in reflex tachycardic responses to SNP were observed between the intact WT and ER{alpha}KO. Estrogen replacement had no effect on the reflex responses to SNP in either the ovariectomized WT or ER{alpha}KO.


   DISCUSSION
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
GRANTS
 REFERENCES
 
The main findings of this study are 1) estrogen-mediated facilitation of baroreflex heart rate responses to PE and ANG II is not observed in the ER{alpha}KO mice and 2) female ER{alpha}KO, unlike WT females, show ANG II-mediated central resetting of cardiac baroreflexes. These observations suggest an important role for ER{alpha} subtype in the central modulation of baroreflex responses in females.

We have previously reported that circulating estrogen levels influence cardiac baroreflex responses to PE and ANG II (30). Consistent with those observations, in the present study, reflex responses to PE and ANG II in WT-OvxE+ mice were also significantly facilitated compared with WT-OvxE–. Results from the present study in the ER{alpha}KO mouse suggest that estrogen facilitation of cardiac baroreflexes is mediated by the ER{alpha} subtype. Although the central mechanisms by which estrogen acts to produce these effects is not known, we and others (18, 29) have shown that estrogen is capable of altering central neuronal activity at important cardiovascular regulatory centers and can modulate autonomic regulation of heart rate. Microinjections of 17{beta}-estradiol decrease area postrema neuronal activity, and in primary cultures of area postrema neurons, estradiol facilitates calcium-activated K currents and decreases ANG II-mediated increases in intracellular calcium (18, 29). Microinjections of 17{beta}-estradiol into central nuclei involved in regulation of vagal tone, e.g., nucleus ambiguus or parabrachial nucleus (PBN), have been shown to increase parasympathetic activity and also facilitate PE-evoked increases in efferent vagal parasympathetic nerve activity (38, 39). Although these observations were made in rats, it is conceivable that estrogen has similar effects in mice. A recent report (25) documenting localization of the ER subtypes in the brain of ovariectomized mice shows robust expression of ER{alpha} subtype in the nucleus tractus solitarius, dorsal motor nucleus, PBN, area postrema, subfornical organ, and several other important cardiovascular regulatory centers. Estrogen could potentially facilitate baroreflex heart rate responses to PE and ANG II in ovariectomized WT mice by modulating neuronal activity in any of these central nuclei.

An important observation in the present study is the ANG II-mediated blunting of baroreflex heart rate responses in the ER{alpha}KO mice. Although both PE and ANG II are potent peripheral vasoconstrictors, ANG II also has centrally mediated effects on the cardiovascular system, including increases in arterial pressure, heart rate, and sympathetic nerve activity and decreases in parasympathetic activity (4, 7, 33, 35). One of the unique characteristics of ANG II is its ability to increase arterial blood pressure without causing a significant reflex bradycardia that is normally associated with such increases in pressure (21, 24, 34). We (30) have previously reported that this ANG II-mediated blunting of the baroreflex heart rate responses is observed only in male mice but not in ovariectomized and estrogen-replaced females, indicating sex differences in ANG II-mediated central resetting of cardiac baroreflex responses are not related to the circulating estrogen levels. Consistent with our previous observations, in the present study, PE and ANG II evoked comparable reflex bradycardia in intact WT females. However, reflex bradycardic responses to ANG II were significantly blunted in the ER{alpha}KO mice compared with PE. Similar observations were also made in ovariectomized ER{alpha}KO mice independent of their estrogen replacement status. The data suggest that in females lacking ER{alpha}, ANG II causes resetting of cardiac baroreflexes similar to males and may be indicative of a direct modulatory effect of the ER{alpha} on ANG II function.

Numerous laboratories have suggested that this central effect of ANG II is mediated predominantly by AT1 receptor subtype in circumventricular organs such as the area postrema (9, 44). The area postrema is located in the medulla oblongata with neuronal projections to central nuclei such as the nucleus ambiguus involved in the regulation of vagal tone (11, 41). Lesioning of the area postrema in several species, including mice, has been shown to prevent ANG II-mediated resetting of baroreflex heart rate responses (23, 45). The attenuation of the reflex bradycardic responses by ANG II has been partly attributed to an inhibition of the increase in vagal tone that normally occurs in response to an increase in arterial pressure (15, 17, 33). It is reasonable to hypothesize that alteration in AT1 receptor function or density in the area postrema could conceivably influence the effects of ANG II on cardiac baroreflexes. AT1 receptor density, along with other components of the renin-angiotensin system is modulated by the circulating estrogen levels. Chronic estrogen treatment is known to decrease AT1 receptor expression in the adrenal glands and the anterior pituitary by increasing posttranslational degradation of the receptor (16). In the absence of estrogen, a greater AT1 receptor expression can potentiate these central effects of ANG II. However, the observations in the present study suggest that it is the lack of ER{alpha} subtype and not circulating estrogen that leads to the functional expression of ANG II-mediated central resetting of cardiac baroreflexes in the females. There is some evidence to suggest that ER{alpha} has effects independent of its ligands. In the human breast cancer cell line, ER{alpha} has been shown to effect the transcription of retinoic acid receptor in the absence of estrogen or tamoxifen (36). However, further investigations are necessary to resolve the underlying mechanisms in the present model.

Resting blood pressures in the ER{alpha}KO-OvxE+ mice were slightly but significantly lower compared with the WT mice. The cause for the lower resting blood pressures in this group is not clear and could be a direct effect of estrogen on vasculature. Estrogen is known to cause vasorelaxation by stimulating release of endothelium-derived nitric oxide (NO) or other vasodilatory substances or by acting directly on the vascular smooth muscle (43). Studies (6, 26, 31, 40) carried out in the ER{alpha}KO model suggest that estrogen-mediated increase in NO is independent of the ER{alpha} subtype. It is possible in the estrogen-replaced ER{alpha}KO mice, ER{beta}-mediated vasorelaxation predominates resulting in lower blood pressures. Recent reports (48) in mice lacking ER{beta} subtype show an increased vasoconstriction and development of hypertension with age, suggesting a role for ER{beta} subtype in maintenance of the basal vascular tone. A limitation of the ER{alpha}KO model is the presence of incomplete ER{alpha} transcripts with residual transcriptional activity (1). Initial studies suggested that these transcripts are not functional, since reproductive function is completely abolished in ER{alpha}KO. However, some effects of estradiol, such as uterine hypertrophy still persist albeit at a reduced level. Recent studies by Pendaries and colleagues (6, 32) comparing different models of ER{alpha}KO suggest that incomplete ER{alpha} transcripts in the ER{alpha}KO model used in the present study partially preserve some of the uterine and vascular responses to estradiol. Consequently, a role for the ER{alpha} subtype in mediating the vascular effects of estradiol cannot be ruled out in the present study.

In the present study, observations regarding the effects of absence of estrogen and ER{alpha} are limited to baroreflex sensitivity. Unlike generation of a typical baroreflex curve in which blood pressure is increased or decreased by 50–60 mmHg, the changes in blood pressure were restricted to 30 mmHg, mainly because the ANG II-mediated blunting of baroreflex heart rate responses is obvious at only smaller increases in blood pressure. In addition, responses to PE and SNP were also obtained separately, which does not allow us to generate a sigmoidal curve. In the absence of sigmoidal curve, it is not clear whether baroreflex function is reset in the ER{alpha}KO mice.

In conclusion, the present study suggests that estrogen facilitates baroreflex heart rate responses to ANG II and PE via the ER{alpha} subtype. In addition, independent of estrogen status, in the absence of this receptor function alone, protection against ANG II-mediated blunting of baroreflex responses is lost in female mice and could potentially lead to baroreflex resetting and maintenance of high blood pressures in ANG II dependent forms of hypertension.


   GRANTS
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
REFERENCES
 
This work was supported by National Heart, Lung, and Blood Institute Grant HL-62261 and American Heart Association Postdoctoral Grant 9920510Z.


   ACKNOWLEDGMENTS
 
The authors thank Hope Gole for excellent technical assistance and Leslie Newton for help with genotyping and maintenance of the mice colony.


   FOOTNOTES
 

Address for reprint requests and other correspondence: J. Pamidimukkala, Dalton Cardiovascular Research Center, Univ. of Missouri-Columbia, 134 Research Park, Columbia, MO 65211 (E-mail: PamidimukkalaJ{at}missouri.edu)

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   REFERENCES
TOP
 ABSTRACT
 MATERIALS AND METHODS
 RESULTS
 DISCUSSION
 GRANTS
 REFERENCES
 

  1. Couse JF, Curtis SW, Washburn TF, Lindzey J, Golding TS, Lubahn DB, Smithies O, and Korach, KS. Analysis of transcription and estrogen insensitivity in the female mouse after targeted disruption of the estrogen receptor gene. Mol Endocrinol 9: 1441–1454, 1995.[Abstract]
  2. Couse JF and Korach KS. Estrogen receptor null mice: what have we learned and where will they lead us? Endocr Rev 20: 358–417, 1999.[Abstract/Free Full Text]
  3. Couse JF, Yates MM, Walker VR, and Korach KS. Characterization of the hypothalamic-pituitary-gonadal axis in estrogen receptor (ER) null mice reveals hypergonadism and endocrine sex reversal in females lacking ER{alpha} but not ER{beta}. Mol Endocrinol 17: 1039–1053, 2003.[Abstract/Free Full Text]
  4. Cox BF and Bishop VS. Neural and humoral mechanisms of angiotensin-dependent hypertension. Am J Physiol Heart Circ Physiol 261: H1284–H1291, 1991.[Abstract/Free Full Text]
  5. Crofton JT and Share L. Gonadal hormones modulate deoxycorticosterone-salt hypertension in male and female rats. Hypertension 29: 494–499, 1997.[Abstract/Free Full Text]
  6. Darblade B, Pendaries C, Krust A, Dupont S, Fouque MJ, Rami J, Chambon P, Bayard F, and Arnal JF. Estradiol alters nitric oxide production in the mouse aorta through the {alpha}-, but not {beta}-, estrogen receptor. Circ Res 90: 413–419, 2002.[Abstract/Free Full Text]
  7. Ferrario CM. Neurogenic actions of angiotensin II. Hypertension 5: 73–79, 1983.
  8. Godsland IF, Wynn V, Crook D, and Miller NE. Sex, plasma lipoproteins and atherosclerosis: prevailing assumptions and outstanding questions. Am Heart J 114: 1467–1503, 1987.[CrossRef][ISI][Medline]
  9. Gorbea-Opplinger VJ and Fink GD. Cerebroventicular injection of angiotensin II antagonist: effects on blood pressure responses to central and systemic angiotensin II. J Pharmacol Exp Ther 273: 611–616, 1995.[Abstract/Free Full Text]
  10. Gustafsson JÅ. Estrogen receptor {beta}–a new dimension in estrogen mechanism of action. J Endocrinol 163: 379–383, 1999.[CrossRef][ISI][Medline]
  11. Hay M and Bishop VS. Interactions of the area postrema and the solitary tract in the nucleus tractus solitarius. Am J Physiol Heart Circ Physiol 260: H1466–H1473, 1991.[Abstract/Free Full Text]
  12. Haywood JR and Hinojosa-Laborde C. Sexual dimorphism of sodium-sensitive renal-wrap hypertension. Hypertension 30: 667–671, 1997.[Abstract/Free Full Text]
  13. Hinojosa-Laborde C, Lange DL, and Haywood JR. Role of female sex hormones in the development and reversal of Dahl hypertension. Hypertension 35: 484–489, 2000.[Abstract/Free Full Text]
  14. Hong MK, Romm PA, Reagan K, Green CE, and Rackley CE. Effects of estrogen replacement therapy on serum lipid values and angiographically defined coronary artery disease in postmenopausal women. Am J Cardiol 69: 176–178, 1992.[CrossRef][ISI][Medline]
  15. Ismay MJ, Lumbers ER, and Stevens AD. The action of angiotensin II on the baroreflex response of the conscious ewe and conscious fetus. J Physiol 288: 467–479, 1979.[Abstract/Free Full Text]
  16. Krishnamurthy K, Verbalis JG, Zheng W, Wu Z, Clerch LB, and Sandberg K. Estrogen regulates angiotensin At1 receptor expression via cytosolic proteins that bind the 5' leader sequence of the receptor mRNA. Endocrinology 140: 5435–5438, 1999.[Abstract/Free Full Text]
  17. Lee WB, Ismay MJ, and Lumbers ER. Mechanisms by which angiotensin II affects the heart rate of the conscious sheep. Circ Res 47: 289–292, 1980.
  18. Li ZC and Hay M. 17{beta}-estradiol modulation of area postrema potassium currents. J Neurophysiol 84: 1385–1391, 2000.[Abstract/Free Full Text]
  19. Lieberman EH, Gerhard MD, and Uehata A. Estrogen improves endothelium-dependent, flow-mediated vasodilation in postmenopausal women. Ann Intern Med 121: 936–941, 1994.[Abstract/Free Full Text]
  20. Lubahn DB, Moyer JS, Golding TS, Couse JF, Korach KS, and Smithies O. Alteration of reproductive function but not prenatal sexual development after insertional disruption of the mouse estrogen receptor gene. Proc Natl Acad Sci USA 90: 11162–11166, 1993.[Abstract/Free Full Text]
  21. Lumbers ER, McCloskey DI, and Potter EK. Inhibition by angiotensin II of baroreceptor-evoked activity in cardiac vagal efferent nerves in the dog. J Physiol 294: 69–80, 1979.[Abstract/Free Full Text]
  22. Manolagas SC and Kousteni S. Perspective: nonreproductive sites of action of reproductive hormones. Endocrinology 142: 2200–2204, 2001.[Free Full Text]
  23. Matsukawa S and Reid IA. Role of area postrema in the modulation of baroreflex control of heart rate by angiotensin II. Circ Res 67: 1462–1473, 1990.[Abstract/Free Full Text]
  24. Matsumura Y, Hasser EM, and Bishop V. Central effects of angiotensin II on baroreflex regulation in conscious rabbits. Am J Physiol Regul Integr Comp Physiol 256: R694–R700, 1989.[Abstract/Free Full Text]
  25. Mitra SW, Hoskin E, Yudkovitz J, Pear L, Wilkinson HA, Hayashi S, Pfaff DW, Ogawa S, Rohrer SP, Schaeffer JM, McEwen BS, and Alves SE. Immunolocalization of estrogen receptor beta in the mouse brain: comparison with estrogen receptor alpha. Endocrinology 144: 2055–1067, 2003.[Abstract/Free Full Text]
  26. Muller-Delp JM, Lubahn DB, Nichol KE, Philips BJ, Price EM, Curran EM, Laughlin, and MH. Regulation of nitric oxide-dependent vasodilation in coronary arteries of estrogen receptor-{alpha}-deficient mice. Am J Physiol Heart Circ Physiol 285: H2150–H2157, 2003.[Abstract/Free Full Text]
  27. Nordby G, Haaland A, and Os I. Evidence of decreased fibrinolytic activity in hypertensive premenopausal women. Scand J Clin Lab Invest 52: 275–281, 1992.[ISI][Medline]
  28. Ouchi Y, Share L, Crofton JT, Iitake K, and Books DP. Sex differences in development of deoxycorticosterone-salt hypertension. Hypertension 9: 172–177, 1987.[Abstract/Free Full Text]
  29. Pamidimukkala J and Hay M. 17{beta}-Estradiol inhibits angiotensin II activation of area postrema neurons. Am J Physiol Heart Circ Physiol 285: H1515–H1520, 2003.[Abstract/Free Full Text]
  30. Pamidimukkala J, Taylor JA, Welshons WV, Lubahn DB, and Hay M. Estrogen modulation of baroreflex function in conscious mice. Am J Physiol Regul Integr Comp Physiol 284: R983–R989, 2003.[Abstract/Free Full Text]
  31. Pare G, Krust A, Karas RH, Dupont S, Aronovitz M, Chambon P, and Mendelsohn ME. Estrogen receptor-{alpha} mediates the protective effects of estrogen against vascular injury. Circ Res 90: 1087–1092, 2002.[Abstract/Free Full Text]
  32. Pendaries C, Darblade B, Rochaix P, Krust A, Chambon P, Korach KS, Bayard F, and Arnal JF. The AF-1 activation-function of ER{alpha} may be dispensable to mediate the effect of estradiol on endothelial NO production in mice. Proc Natl Acad Sci USA 99: 2205–2210, 2002.[Abstract/Free Full Text]
  33. Potter EK and Reid IA. Intravertebral angiotensin II inhibits cardiac vagal efferent activity in dogs. Neuroendocrinology 40: 493–496, 1985.[ISI][Medline]
  34. Reid I and Kumagai K. Angiotensin II exerts differential actions on renal nerve activity and heart rate. Hypertension 24: 451–456, 1994.[Abstract/Free Full Text]
  35. Reid IA. Actions of angiotensin II on the brain: mechanisms and physiologic role. Am J Physiol Renal Fluid Electrolyte Physiol 246: F533–F543, 1984.[Abstract/Free Full Text]
  36. Rousseau C, Pettersson F, Couture MC, Paquin A, Galipeau J, Mader S, and Miller WH. The N-terminal of the estrogen receptor (ER{alpha}) mediates transcriptional cross-talk with the retinoic acid receptor in human breast cancer cells. J Steroid Biochem Mol Biol 86: 1–14, 2003.[CrossRef][ISI][Medline]
  37. Rui He X, Wang W, Crofton JT, and Share L. Effects of 17{beta}-estradiol on sympathetic activity and pressor response to phenylephrine in ovariectomized rats. Am J Physiol Regul Integr Comp Physiol 275: R1202–R1208, 1998.[Abstract/Free Full Text]
  38. Saleh MC, Connell BJ, and Saleh TM. Autonomic and cardiovascular reflex responses to central estrogen injection in ovariectomized female rats. Brain Res 879: 105–114, 2000.[CrossRef][ISI][Medline]
  39. Saleh TM, Connell BJ, and Saleh MC. Acute injection of 17{beta}-estradiol enhances cardiovascular reflexes and autonomic tone in ovariectomized female rats. Auton Neuro Sci 84: 78–88, 2000.
  40. Sampei K, Goto S, Alkayed NJ, Crain BJ, Korach KS, Traystman RJ, Demas GE, Nelson RJ, and Hurn PD. Stroke in estrogen receptor-alpha-deficient mice. Stroke 31: 738–744, 2000.[Abstract/Free Full Text]
  41. Shapiro RE and Miselis RR. The central neural connections of the area postrema of the rat. J Comp Neurol 234: 344–364, 1985.[CrossRef][ISI][Medline]
  42. Walsh BW, Schiff I, Rosner B, Greinberg L, Ravnikar V, and Sacks F. Effects of postmenopausal estrogen replacement on the concentrations and metabolism of plasma lioproteins. N Engl J Med 324: 1196–1204, 1991.
  43. White RE. Estrogen and vascular function. Vascul Pharmacol 38: 73–80. 2002.[ISI][Medline]
  44. Wong J, Chou L, and Reid IA. Role of AT1 receptors in the resetting of the baroreflex control of heart rate by angiotensin II in rabbits. J Clin Invest 91: 1516–1520, 1993.[ISI][Medline]
  45. Xue B, Gole H, Pamidimukkala J, and Hay M. Role of the area postrema in angiotensin II modulation of baroreflex control of heart rate in conscious mice. Am J Physiol Heart Circ Physiol 284: H1003–H1007, 2003.[Abstract/Free Full Text]
  46. Xue B, Skala K, and Hay M. Gender difference in the development of angiotensin II-induced hypertension in conscious mice (Abstract). FASEB J 825: A1294, 2003.
  47. Xue B, Skala K, and Hay M. Sex differences in baroreflex function in the development of angiotensin II-induced hypertension in conscious mice (Abstract). Soc Neurosci Abstr 923: 15, 2003.
  48. Zhu Y, Bian Z, Lu P, Karas RH, Bao L, Cox D, Hodgin J, Shaul PW, Thoren P, Smithies O, Gustafsson JA, and Mendelsohn MA. Abnormal vascular function and hypertension in mice deficient in estrogen receptor beta. Science 295: 505–508, 2002.[Abstract/Free Full Text]



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