AJP - Heart Fuel your research with LabChart
HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS
QUICK SEARCH:   [advanced]


     

Am J Physiol Heart Circ Physiol 279: H511-H519, 2000;
0363-6135/00 $5.00
This Article
Right arrow Abstract Freely available
Right arrow Full Text (PDF)
Right arrow Alert me when this article is cited
Right arrow Alert me if a correction is posted
Right arrow Citation Map
Services
Right arrow Email this article to a friend
Right arrow Similar articles in this journal
Right arrow Similar articles in ISI Web of Science
Right arrow Similar articles in PubMed
Right arrow Alert me to new issues of the journal
Right arrow Download to citation manager
Citing Articles
Right arrow Citing Articles via HighWire
Right arrow Citing Articles via ISI Web of Science (54)
Right arrow Citing Articles via Google Scholar
Google Scholar
Right arrow Articles by Geary, G. G.
Right arrow Articles by Duckles, S. P.
Right arrow Search for Related Content
PubMed
Right arrow PubMed Citation
Right arrow Articles by Geary, G. G.
Right arrow Articles by Duckles, S. P.
Vol. 279, Issue 2, H511-H519, August 2000

Estrogen reduces mouse cerebral artery tone through endothelial NOS- and cyclooxygenase-dependent mechanisms

Greg G. Geary, Diana N. Krause, and Sue P. Duckles

Department of Pharmacology, College of Medicine, University of California, Irvine, California 92697-4625


    ABSTRACT
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Gender and estrogen status are known to influence the incidence and severity of cerebrovascular disease. The vasoprotective effects of estrogen are thought to include both nitric oxide-dependent and independent mechanisms. Therefore, using small, resistance-sized arteries pressurized in vitro, the present study determined the effect of gender and estrogen status on myogenic reactivity of mouse cerebral arteries. Luminal diameter was measured in middle cerebral artery segments from males and from females that were either untreated, ovariectomized (OVX), or OVX with estrogen replacement (OVX + EST). The maximal passive diameters of arteries from all four groups were similar. In response to increases in transmural pressure, diameters of arteries from males and OVX females were smaller compared with diameters of arteries from either untreated or OVX + EST females. In the presence of NG-nitro-L-arginine methyl ester, artery diameters decreased in all groups, but diameters remained significantly smaller in arteries from males and OVX females compared with untreated and OVX + EST females. After endothelium removal or when inhibition of nitric oxide synthase and cyclooxygenase were combined, differences in diameters of arteries from OVX and OVX + EST were abolished. These data suggest that chronic estrogen treatment modulates myogenic reactivity of mouse cerebral arteries through both endothelium-derived cyclooxygenase- and nitric oxide synthase-dependent mechanisms.

cerebral arteries; gender; nitric oxide synthase


    INTRODUCTION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

GENDER IS KNOWN TO INFLUENCE the incidence and severity of cerebrovascular disease (3, 30, 41, 52). For instance, premenopausal females experience fewer and less severe strokes compared with males of similar age (41). The incidence of stroke increases rapidly following menopause, suggesting that estrogen protects cerebral blood vessels from disease (41). The role of estrogen as vasoprotective is further evidenced by findings that the occurrence of cerebrovascular disease is lower in postmenopausal females receiving hormone replacement therapy (17, 38), although not all studies have found a positive benefit of hormone therapy (40).

Despite the epidemiological evidence that suggests that plasma estrogen influences the course of cerebrovascular disease, only a few recently published studies have begun to describe the effects of estrogen on the function of cerebral blood vessels (20, 47). In these papers, estrogen and gender were found to modulate the level of vascular smooth muscle tone in rat cerebral arteries. This effect was dependent on nitric oxide (NO), a known modulator of cerebrovascular tone (27). In the rat peripheral circulation, estrogen has been shown to act through both NO-dependent (4, 25, 31, 33, 51) and independent mechanisms (11, 16, 29). Therefore, estrogen may modulate multiple mechanisms within the artery wall that contribute to the continual regulation of blood vessel diameter.

Cerebral blood flow is maintained within defined limits during changes in systemic pressure; this is termed autoregulation (15). Myogenic reactivity (one contributing mechanism) is an intrinsic characteristic of vascular smooth muscle resulting in constriction to increasing or dilation to decreasing transmural pressure (2, 10). Whereas the exact membrane and intracellular signaling events required to initiate and maintain the myogenic response have not been precisely determined, Ca2+ (both intracellular and extracellular) along with certain protein kinases (e.g., myosin light chain kinase and protein kinase C) are believed critically important to the response (10).

Cerebral blood vessels from the rat have been extensively used to characterize the myogenic mechanism of autoregulation; however, only the early in vivo study by Rosenblum et al. (44) has described the response of mouse cerebral arterioles to elevated transmural pressure. In this paper, mouse arterioles (diameter <= 65 µm) responded to a single step elevation of transmural pressure by either maintaining or decreasing their control diameter. These myogenic responses to increasing pressure in mouse cerebral arterioles appear similar to those of resistance-sized cerebral arterioles from rats (22), humans (49), and cats (32). However, there have been no further studies on myogenic reactivity of mouse cerebral arteries.

With the recent advances in development of genetically altered mice, increased use of the mouse as a model to investigate control mechanisms of the cerebral and systemic circulation can be predicted. For example, NO synthase (NOS)-mutant mice have been used to investigate the modulatory effects of NO on cerebral blood vessels (21, 24, 34, 37). In the peripheral circulation, estrogen receptor-deficient mice have greater vascular smooth muscle cell proliferation following vascular injury (28). Because one potential target of plasma estrogen is endothelial NOS (36), the potential effect of either NOS or estrogen receptor mutants on myogenic reactivity of cerebral arteries will require comprehensive investigation. Presently though, there is a complete absence of data on the effects of gender or estrogen on cerebral blood vessels in this species.

Therefore, the purpose of the present study was to determine the extent of myogenic regulation of vascular tone in resistance-sized cerebral arteries of male and female C57 Black mice. Additionally, because the vascular endothelium and vascular smooth muscle cells possess intrinsic mechanisms that modulate myogenic reactivity of rat cerebral arteries, we determined whether the myogenic mechanism in mouse cerebral arteries is also modulated by either endothelial or vascular smooth muscle mechanisms and if these mechanisms are dependent on gender. Finally, we investigated the effects of estrogen on the myogenic mechanism by comparing cerebral arteries from female mice that had been ovariectomized with or without estrogen replacement.


    METHODS
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Animals. Animal procedures were approved by the Animal Care and Use Committee of University of California, Irvine. Male and female 12-wk-old C57 Black mice were purchased from Charles River Laboratory (Wilmington, MA) and housed under a 12:12-h light-dark cycle with food and water available ad libitum. Four groups of mice were used in the present study: males (n = 6), untreated ovary-intact (cycling) females (n = 6), and ovariectomized females with (n = 18) and without (n = 18) estrogen replacement (OVX and OVX + EST, respectively). Ovariectomy and ovariectomy with estrogen replacement were performed with the mice under anesthesia (ketamine, 90 mg/kg, and xylazine, 10 mg/kg) (9). Estrogen was replaced at the time of ovariectomy by subcutaneous insertion of 1-mm silicone elastomer capsules made from Dow Corning Silastic medical-grade tubing (1.57 mm ID × 3.18 mm OD, Dow Corning), sealed with silicone elastomer adhesive type A (Dow Corning), and packed with 17beta -estradiol. Both OVX and OVX + EST animals were euthanized 1 mo after surgery.

Tissue preparation. At 16 wk of age, mice were decapitated in the middle of the day. The uterus was removed from each female animal and weighed. Brains were rapidly removed from the cranial cavity and placed in a dissecting dish with cold oxygenated physiological salt solution (PSS) containing (in mM) 118 NaCl, 4.8 KCl, 1.6 CaCl2, 1.2 KH2PO4, 25 NaHCO3, 1.2 MgSO4, 0.3 ascorbic acid, and 11.5 glucose. A 1- to 2-mm segment of the middle cerebral artery, taken about 1 mm from the circle of Willis, was carefully dissected and mounted in an arteriograph (Living Systems, Burlington, VT). Micropipettes were inserted into each end of the artery and secured in place with nylon ties. The proximal cannula was connected through a pressure transducer and windkessel to a reservoir of PSS equilibrated with 95% O2-5% CO2. The distal cannula was connected to a Luer-Lok that was open during the initial equilibration to gently flush the luminal contents. After the equilibration period, the Luer-Lok remained closed so all experiments were conducted under no-flow conditions. A constant-flow peristaltic pump continuously superfused (30 ml/min) the artery with PSS. During a 60-min equilibration period, a pressure servo system maintained transmural pressure at 30 mmHg. The artery was viewed with an inverted microscope equipped with a video camera and monitor. A video-electronic dimension analyzer was used to measure luminal diameter. Changes in transmural pressure and lumen diameter were digitized by a MacLab analog-to-digital converter and recorded on a Macintosh computer. All drugs, individually or in combination, were added to the superfusate in their final concentration.

Radioimmunoassay for serum levels of estradiol. Immediately after the mice were euthanized, blood was collected by cardiac puncture, placed in plain tubes, and centrifuged at 3,000 rpm for 10 min. The supernatant was decanted and frozen at -70°C. Serum 17beta -estradiol levels were determined by radioimmunoassay with commercially available kits (Diagnostic Products, Los Angeles, CA). Assay sensitivity for estradiol was 8 pg/ml.

Experimental protocols. In all protocols, changes in artery diameter in response to increased transmural pressure (no flow) were measured. After the 60-min equilibration period, one of the following four protocols were performed. In protocol 1, three separate series of pressure steps (each from 30 to 80 mmHg in 10-mmHg steps) were performed. The first series of pressure steps was in PSS, the second in the presence of NG-nitro-L-arginine-methyl ester (L-NAME; 10 µM), and the third in 0 Ca2+-EDTA (3 mM). Protocol 2 consisted of four separate series of pressure steps (each from 30 to 80 mmHg in 10-mmHg steps). The first series of pressure steps was in PSS, the second in the presence of indomethacin (10 µM), the third in the presence of indomethacin plus L-NAME, and the fourth in 0 Ca2+-EDTA (3 mM). In protocol 3, increasing pressure steps from 80 to 180 mmHg in 20-mmHg steps were performed in either arteries preconstricted with K+ (40-60 mM) in the presence of indomethacin, K+ channel blockers [tetraethylammonium, 1 mM (large conductance Ca2+-activated K+); barium, 50 µM (inward rectifier K+ channel); and apamin, 10 nM (small conductance Ca2+-activated K+)] and L-NAME, or 0 Ca2+-EDTA alone (3 mM). In protocol 4, four separate series of pressure steps (each from 30 to 80 mmHg in 10-mmHg steps) were performed. The first series of pressure steps was with an endothelium-intact artery in PSS, the second series followed endothelium removal, the third was with endothelium-damaged arteries in the presence of K+ channel blockers, and the fourth was with endothelium-denuded arteries in 0 Ca2+-EDTA (3 mM).

Endothelium removal was accomplished by perfusing 1 ml of air through the artery lumen. Successful removal of the endothelium was determined by complete inhibition of dilation to ADP (10 µM) in arteries preconstricted with Ba2+ (10-30 µM). All drugs were perfused for 20 min before the first pressure step, and each pressure step was maintained for 5-10 min to allow the vessel to reach a stable condition before diameter was measured. Control arteries showed consistent responses to four sequential series of pressure steps.

Myogenic tone was determined by subtracting the steady-state diameter at any given pressure either in PSS or L-NAME from the passive diameter (0 Ca2+ + 3 mM EDTA) at that same pressure. All drugs were purchased from Sigma Chemical (St. Louis, MO). Data are expressed as means ± SE. Statistical significance was determined using ANOVA with Scheffé's test. Acceptable level of significance was defined as P < 0.05.


    RESULTS
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

Body weights, uterine weights, and serum estrogen concentrations. Body weights, uterine weights, and serum estrogen concentrations for the various groups of animals studied are shown in Table 1. Weights of age-matched males, untreated females, and OVX females were significantly greater than OVX + EST females. Uterine weights of untreated and OVX + EST females were significantly greater (P < 0.001) than those of OVX females. Estrogen replacement resulted in serum levels significantly higher (P < 0.05) than OVX females.

                              
View this table:
[in this window]
[in a new window]
  		Table 1.   Effect of ovariectomy and estrogen treatment on body weight, uterine weight, and serum concentrations of estradiol

Myogenic responses in arteries from males and females. Passive responses measured in the absence of Ca2+ (EDTA, 3 mM) were not significantly different between arteries from male and female mice, except at a pressure of 30 mmHg, where passive diameters of arteries from males were significantly smaller than arteries from females. As shown in Fig. 1, arteries in PSS from males had significantly smaller diameters at all levels of pressure compared with their passive responses. In contrast, diameters of arteries in PSS from female mice were not significantly different from diameters in EDTA.


View larger version (27K):
[in this window]
[in a new window]
  		Fig. 1.   Response to increased intraluminal pressure in mouse middle cerebral arteries from four groups of mice. A: male; B: female; C: ovariectomized (OVX); D: OVX with estrogen replacement (OVX + EST). Mean steady-state luminal diameter is plotted as a function of pressure in either physiological salt solution (PSS), in the presence of NG-nitro-L-arginine-methyl ester (L-NAME; 100 µM), or 0 Ca2+ + 3 mM EDTA (passive response). Solutions of either L-NAME or 0 Ca2+-EDTA were superfused for 20 min before the step increases in pressure were performed. Values are means ± SE; n = 6. *P < 0.05, different from one other group. **P < 0.05, different from two other groups, comparisons among PSS, L-NAME, and EDTA.

To determine whether NO contributes to the steady-state diameter, we added L-NAME (100 µM) to inhibit the synthesis of NO. In the presence of L-NAME, arteries from both males and females had significantly smaller diameters at each pressure step compared with PSS alone (Fig. 1), except at 30 and 40 mmHg for arteries from untreated females. Furthermore, in the presence of L-NAME, diameters of arteries from males were significantly smaller than diameters of arteries from untreated females at each pressure step (P < 0.05). As shown in Fig. 2, even in the presence of L-NAME, myogenic tone (difference in diameter between L-NAME and 0 Ca2+) was significantly greater in arteries from males compared with females.


View larger version (21K):
[in this window]
[in a new window]
  		Fig. 2.   Comparison of myogenic tone following nitric oxide synthase (NOS) inhibition in mouse middle cerebral arteries from males, females, OVX females, or OVX + EST females. Myogenic tone was calculated as the difference between diameters in 0 Ca2+ + EDTA and L-NAME and plotted as a function of pressure. Values are means ± SE; n = 6. *P < 0.05, compared with males and OVX females.

Effect of ovariectomy and estrogen replacement. Arteries from OVX mice and OVX + EST mice were then studied to determine whether circulating estradiol could account for the difference in myogenic tone between male and female cerebral arteries. Passive artery diameters at each pressure were not significantly different between arteries from OVX and OVX + EST mice (P > 0.05). As shown in Fig. 1, diameters of arteries in PSS from OVX mice were smaller compared with their passive responses, which contrasts with responses of arteries from untreated female mice (Fig. 1B). In OVX + EST mice, artery diameters in PSS were not significantly different from their passive diameters in EDTA at any of the pressure steps.

In the presence of L-NAME, diameters significantly decreased in arteries from both OVX and OVX + EST at all pressures, except at 30 mmHg for OVX + EST (Fig. 1). As shown in Fig. 2, even in the presence of L-NAME the level of myogenic tone was significantly greater in arteries from OVX compared with OVX + EST at all pressures (P < 0.01). Overall, either in PSS or in the presence of L-NAME, myogenic tone in arteries from OVX + EST was similar to that in arteries from intact females and was smaller than myogenic tone in arteries from males or OVX females.

Effect of inhibition of cyclooxygenase and NOS. Because diameters of arteries from OVX mice were significantly smaller following NOS inhibition compared with diameters of arteries from OVX + EST, we investigated whether a metabolite from the cyclooxygenase pathway might modulate myogenic reactivity in a manner sensitive to estrogen treatment. Therefore, arteries were treated with a cyclooxygenase inhibitor (indomethacin, 10 µM), and cerebral artery diameter was measured in response to step increases in pressure (Fig. 3). As before, maximal passive artery diameters (80 mmHg) were not significantly different between OVX (167 ± 6 µm) and OVX + EST (168 ± 5 µm) mice. Similar to the PSS data shown in Fig. 1, diameters of cerebral arteries from OVX mice were significantly smaller than their passive diameters in EDTA (Fig. 3). In contrast, diameters of arteries in PSS from OVX + EST mice were not significantly different from their passive diameters at pressures up to 80 mmHg (Fig. 3). Cyclooxygenase inhibition with indomethacin did not significantly affect diameter of arteries from either OVX or OVX + EST mice (Fig. 3). However, in the presence of indomethacin, L-NAME caused significantly greater constriction in arteries from OVX + EST animals compared with arteries from OVX animals (Fig. 3). Unlike the effect of L-NAME by itself, in the presence of indomethacin plus L-NAME, diameters of arteries from OVX and OVX + EST animals were no longer significantly different (Fig. 3). For example, at 60 mmHg in the presence of indomethacin and L-NAME, mean diameters were 123 ± 5 and 120 ± 3 µm for OVX and OVX + EST animals, respectively.


View larger version (14K):
[in this window]
[in a new window]
  		Fig. 3.   Effect of estrogen on myogenic response during inhibition of cyclooxygenase alone or together with NOS. Arteries from OVX females (A) or OVX + EST females (B) were studied during treatment with either 0 Ca2+ plus EDTA (3 mM), indomethacin (Indo, 10 µM), or Indo + L-NAME (10 µM). Drug solutions were superfused for 20 min before the step increases in pressure (30-80 mmHg) were performed. Values are means ± SE; n = 6. *P < 0.05, significantly different from one other group; **P < 0.05, significantly different from two other groups; ***P < 0.05, significantly different from three other groups, comparison among EDTA, PSS, Indo, and Indo + L-NAME.

Effect of endothelium removal. Although our data with NOS and cyclooxygenase inhibition suggest that estrogen affects NOS and cyclooxygenase metabolites, the source of these vasomodulators was not determined. Furthermore, the effect of estrogen on vascular smooth muscle cell reactivity to increasing pressure has not previously been determined in mouse cerebral arteries. Therefore, endothelium-intact and -denuded arteries from OVX and OVX + EST mice were exposed to multiple pressure steps. Again, maximal passive artery diameters (80 mmHg) were not significantly different between OVX (163 ± 3 µm) and OVX + EST (169 ± 3 µm) mice. Similar to the PSS data shown in Figs. 1 and 3, diameters of cerebral arteries from OVX mice decreased significantly compared with their passive responses as pressure increased (Fig. 4A). In contrast, diameters of arteries in PSS from OVX + EST mice were not significantly different from their passive responses at pressures up to 80 mmHg (Fig. 4B).


View larger version (17K):
[in this window]
[in a new window]
  		Fig. 4.   Effect of estrogen treatment on myogenic response with and without endothelial damage. Diameters of endothelium-intact (E+) and endothelium-damaged (E-) arteries from OVX females (A) or OVX + EST females (B) were studied during treatment with either PSS, a cocktail of K+ channel blockers (KCB) [tetraethylammonium, 1 mM; barium, 50 µM; and apamin, 10 nM], or with 0 Ca2+ plus EDTA (3 mM). Drug solutions were superfused for 20 min before the step increases in pressure were performed. Values are means ± SE; n = 6. **P < 0.05, different from two other groups; ***P < 0.05, different from three other groups, comparisons among EDTA, PSS (E+), PSS (E-), and KCB (E-).

To confirm damage to the endothelium, arteries were preconstricted with Ba2+ (10-30 µM), and the response to the endothelium-dependent dilator ADP (10 µM) was determined. In endothelium-intact arteries, dilation to ADP was not significantly different between groups (OVX, 38 ± 10%; OVX + EST, 59 ± 4%). However, following endothelial damage, dilation to ADP was abolished in arteries from both groups of animals (OVX, 0.5 ± 0.5%; OVX + EST, -6 ± 6%; n = 6).

As shown in Fig. 4, removal of the endothelium resulted in significantly smaller diameters in arteries from both OVX and OVX + EST mice. More importantly, diameter differences between arteries from OVX and OVX + EST mice were abolished following endothelium removal. For example, at 60 mmHg in endothelium-damaged arteries, diameters were 123 ± 5 and 118 ± 5 µm for OVX and OVX + EST animals, respectively. Further treatment of endothelium-denuded arteries from OVX and OVX + EST mice with a cocktail of K+ channel blockers (KCB) [tetraethylammonium, 1 mM (large conductance Ca2+-activated K+); barium, 50 µM (inward rectifier K+ channel); and apamin, 10 nM (small conductance Ca2+-activated K+)] caused a significant reduction of artery diameter compared with denuding alone (Fig. 4). However, diameters of arteries were still not significantly different between the groups. For example, at 60 mmHg in endothelium-damaged arteries treated with indomethacin, mean arterial diameters were 96 ± 11 and 104 ± 11 µm for OVX and OVX + EST animals, respectively. These results clearly suggest that chronic estrogen treatment does not modify vascular smooth muscle cell response to increasing pressure. Furthermore, our data suggest that chronic estrogen treatment modulates myogenic reactivity through endothelium, NOS, and cyclooxygenase-dependent mechanisms.

Effect of estrogen on artery structure. In the next series of experiments we further explored the possibility that estrogen treatment affects structural properties of the vascular smooth muscle. Arteries were subjected to excessive pressures (80-180 mmHg) following preconstriction with K+ (40 to 60 mM) in the presence of indomethacin, KCB, and L-NAME. Wall thickness at 180 mmHg in the absence of Ca2+ plus EDTA (3 mM) was not significantly different between arteries from OVX and OVX + EST mice (OVX, 5.0 ± 0.4 µm; OVX + EST, 6.2 ± 0.8 µm; n = 14). As shown in Fig. 5, maximal passive artery diameters (180 mmHg) were not significantly different between OVX (168 ± 3 µm) and OVX + EST (167 ± 5 µm) mice. In K+-preconstricted arteries in the presence of indomethacin, KCB, and L-NAME, artery diameters at 80 mmHg from OVX and OVX + EST mice were not significantly different (OVX, 91 ± 2; OVX + EST, 88 ± 2 µm) (Fig. 5). Artery diameters from OVX and OVX + EST mice responded similarly to increasing transmural pressure to 180 mmHg. At intraluminal pressures between 140 and 180 mmHg, diameters of arteries from OVX and OVX + EST mice were not significantly different from their respective passive diameters (Fig. 5). For example, at 180 mmHg in K+-preconstricted arteries treated with indomethacin, L-NAME, and KCB, the artery diameters were 147 ± 16 and 160 ± 5 µm for OVX and OVX + EST animals, respectively.


View larger version (12K):
[in this window]
[in a new window]
  		Fig. 5.   Effect of excessive pressure on diameter of arteries from OVX females (A) or OVX + EST females (B) during treatment with either 0 Ca2+ plus EDTA (3 mM) or K+ preconstriction plus Indo (10 µM) and L-NAME (10 µM) combined with a cocktail of KCB [tetraethylammonium, 1 mM; barium, 50 µM; apamin, 10 nM]. Drug solutions were superfused for 20 min before the step increases in pressure (80-180 mmHg) were performed. Values are means ± SE; n = 6. *P < 0.001, compared with EDTA.


    DISCUSSION
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

The present study supports the conclusion that gender and estrogen status modulate the extent of myogenic tone in mouse cerebral arteries. Inhibition of NOS by itself caused significantly greater myogenic tone in arteries from male mice compared with arteries from untreated female mice. This gender difference was abolished by ovariectomy and restored by estrogen treatment. However, after inhibition of cyclooxygenase, the effect of L-NAME was greatly increased in arteries from OVX + EST mice, suggesting that a cyclooxygenase-derived vasodilator is able to nearly completely compensate for normal NOS-dependent modulation of myogenic tone. Myogenic tone differences between arteries from OVX and OVX + EST mice were abolished following either removal of the endothelium or combined NOS and cyclooxygenase inhibition. Estrogen status did not affect vascular smooth muscle cell growth, contractility, or K+ channel function. Combined, our findings suggest that estrogen status affects the synthesis and/or effect of endothelium-derived NOS and cyclooxygenase vasodilators, which together modulate myogenic tone of mouse cerebral arteries.

Gender, estrogen, and myogenic reactivity. Except for the study by Rosenblum et al. (44), little is known about the myogenic reactivity of mouse cerebral blood vessels. Thus the present study is the first in vitro attempt to investigate the myogenic reactivity of cerebral arteries from this species. Our data show that cerebral arteries (passive diameters = 154 ± 4 µm) from male mice gain significant myogenic tone in response to increasing pressure. These findings are consistent with previous studies in other species examining pial artery reactivity to pressure (20, 25, 51). Because gender has been shown to significantly influence the myogenic response of both peripheral and cerebral arteries (20, 25, 51), the present study also determined whether gender affects the development of myogenic tone in mouse cerebral blood vessels. Indeed, myogenic tone was significantly greater in cerebral arteries from male mice compared with similar-sized cerebral arteries from females. This gender-dependent, myogenic reactivity difference appeared dependent on the presence of estrogen because removal of estrogen (OVX) abolished and estrogen replacement (OVX + EST) restored myogenic tone. The effects of gender and estrogen status shown in the present study are consistent with our previously published study, which showed that gender and estrogen status modulate myogenic tone in rat cerebral arteries (20).

NOS-dependent mechanisms are well-known modulators of mouse cerebral artery diameter under normal physiological conditions (34, 37, 46). Because endothelial NOS is one important target of estrogen in myogenically reactive rat cerebral arteries (20, 36), we hypothesized that endothelial NOS would also be a primary target of estrogen in mouse cerebral blood vessels. It was quite surprising to find that NOS inhibition did not abolish myogenic tone differences among gender and estrogen treatment groups. In fact, in the absence of the modulatory influence of NO, myogenic tone of male and OVX mice remained significantly greater compared with either female or OVX + EST mice. Therefore, an estrogen-sensitive mechanism, independent of NOS, appeared to modulate myogenic tone of mouse cerebral arteries. Because estrogen could affect myogenic reactivity through multiple vascular smooth muscle and/or endothelial mechanisms (7, 9, 31, 53), we designed a series of experiments that would determine whether endothelial- or vascular smooth muscle-dependent mechanisms were responsible for the myogenic tone differences between arteries from OVX and OVX + EST mice.

Estrogen and endothelial function. Endothelial cells from cerebral arteries secrete numerous vasoactive substances that function to regulate vascular smooth muscle tone (13, 23, 39, 54). However, in the mouse cerebral vasculature, only endothelium-derived NOS and a cyclooxygenase metabolite have been shown to modulate vascular tone in vivo (45). Therefore, we hypothesized that estrogen upregulates a cyclooxygenase-sensitive mechanism in mouse cerebral arteries, which modulates myogenic tone in the absence of normal NOS-dependent activity. Alone, cyclooxygenase inhibition did not affect artery diameters from either OVX or OVX + EST mice. However, when cyclooxygenase and NOS inhibitors were combined, diameter differences between groups were abolished, strongly suggesting that cyclooxygenase-dependent mechanisms are estrogen sensitive in mouse cerebral blood vessels. This cyclooxygenase substance appeared to originate from the endothelium because diameter differences between groups were also abolished following endothelium removal. Therefore, our data has clearly shown that endothelium-derived NO is primarily involved in modulating myogenic tone in arteries from OVX and OVX + EST animals. However, following pharmacological inhibition of NOS, cyclooxygenase mechanisms are unmasked and partially compensate for loss of normal NOS-dependent modulation of myogenic tone in mouse cerebral arteries.

Estrogen and interaction of endothelial factors. As we mentioned previously, agonist-mediated release of endothelium-derived NOS and cyclooxygenase metabolites modulates the diameter of mouse pial arterioles (45). But besides this previous study and data from our current work, little else is known about the relative contributions of NOS- and cyclooxygenase-dependent mechanisms in mouse cerebral blood vessels. In mouse skeletal muscle arterioles, cyclooxygenase- and NOS-dependent mechanisms are potent vasomodulators in response to flow (48). However, flow-dependent dilation is completely cyclooxygenase dependent in NOS mutant mice, suggesting that cyclooxygenase-dependent mechanisms are greatly upregulated in the absence of NOS. Also, in the mouse, dilation to acetylcholine in mesenteric blood vessels is not affected by either NOS or cyclooxygenase inhibition alone, but dilation is completely abolished when these two inhibitors are combined (8). Together, these previous studies show that NOS and cyclooxygenase substances interact to modulate mouse blood vessel tone.

Although the interactive effects of endothelium-derived NOS and cyclooxygenase in mouse blood vessels are of significant interest, one of the most important findings from our present study is that the vasomodulatory role of the cyclooxygenase pathway in estrogen-treated mice first required inhibition of NOS. Little experimental evidence currently exists concerning the effect of estrogen on interaction of NOS and cyclooxygenase pathways. A recently published article (6) using pressurized rat mesenteric arteries showed that chronic estrogen treatment tipped the balance between endothelium-derived NOS and cyclooxygenase substances toward NOS in response to agonist-mediated dilation. We (19) also found that during NOS inhibition in the perfused rat tail artery, estrogen upregulates an endothelium-derived cyclooxygenase substance(s) that mediates vasodilator responses to either flow or agonists. Therefore, our current data add to the emerging evidence that estrogen treatment upregulates an endothelium-derived cyclooxygenase vasodilator metabolite that, in the absence of NOS, modulates vascular smooth muscle tone.

As we have stated, the effect of estrogen on interactions between NOS and cyclooxygenase has not been extensively investigated. However, NOS inhibition or estrogen treatment have both been shown to influence endothelium-derived cyclooxygenase substances. For example, in arteries taken from dogs chronically treated with a NOS inhibitor, dilation to bradykinin shifts from NOS to cyclooxygenase dependent due to an upregulation of the endothelial constitutive isoform of cyclooxygenase (42). In cultured bovine aortic endothelial cells, high levels of NO inhibit the release of prostacyclin (14). Estrogen also has been shown to affect cyclooxygenase pathways. For example, estrogen upregulates cyclooxygenase gene expression as well as prostacyclin production in ovine fetal pulmonary artery endothelial cells (29). Estrogen also suppresses production of a cyclooxygenase-sensitive vasoconstrictor (11), which, if present in our current study, would reduce myogenic tone. Therefore, estrogen and NOS are clearly able to modify endothelium-dependent cyclooxygenase pathways.

Whereas our results suggest that estrogen modulates cyclooxygenase activity, they also support the hypothesis that estrogen modulates the function of NOS. The data of Fig. 1 suggest that, in contrast to the prevalent hypothesis, estrogen did not increase NOS function. However, upon closer examination this impression is incorrect. Indeed, when the modulatory effects of cyclooxygenase were inhibited, as shown in Fig. 3, the effect of NOS inhibition was greater in arteries from estrogen-treated animals. Because we (20) and others (25, 51) have previously demonstrated that estrogen increases NO-dependent modulation of rat cerebral and peripheral arteries, our current data support the existing hypothesis that one target of estrogen treatment is endothelial NOS (36).

Estrogen and physiological hormone levels. Creation of physiological hormone levels in OVX females undergoing estrogen replacement was essential to our study. Three important pieces of data suggest that our mode of estrogen replacement created physiological responses in the mouse. First, estrogen replacement resulted in serum levels significantly greater than OVX females. Although estrogen concentrations in OVX + EST mice are physiological (43, 50, 55), the ranges determined in the present study are more comparable to those seen during pregnancy (55). Second, uterine weights of untreated female mice were not significantly different from OVX + EST females, suggesting that estrogen replacement created physiological hormone levels (25). Third, in the absence of NOS, active diameters of cerebral arteries from OVX + EST females were not significantly different from normal, intact females, although they were larger than diameters of arteries from either males or OVX females. Together these observations suggest that estrogen replacement created hormone levels with effects comparable to those of estrogen levels in untreated females.

Myogenic reactivity: diameter and species differences. As we have mentioned, except for the study by Rosenblum et al. (44), little is known about the myogenic reactivity of mouse cerebral blood vessels. Our results show that cerebral arteries from male mice gain significant myogenic tone in response to increasing pressure. However, as pressure was increased, arterial diameter significantly increased, whereas the overall level of myogenic tone did not change. Findings in our study contrast with the earlier published work by Rosenblum et al. (44) in which diameters of mouse arterioles in situ (diameter <= 65 µm) responded to increasing pressure by maintaining, increasing, or decreasing diameter. Although the different experimental design between the study by Rosenblum et al. (44) and the present study (cranial window vs. organ bath) may explain some of these differences, it is also probable that a variation in myogenic responses with artery diameter may be an important factor (12). For example, it has been shown in the rat cerebral circulation that first- and second-order arterioles show smaller responses to changes in mean arterial pressure compared with third-order arterioles and small venules (22). Thus it is likely that the location of the artery segment within the intact cerebral circulation accounts for differences in degree of myogenic reactivity between our study and the smaller mouse arterioles studied in situ by Rosenblum et al. (44).

Estrogen and vascular smooth muscle. Estrogen is known to influence both the passive mechanical components (collagen and elastin) of the vessel wall and to decrease injury-induced hyperplasia of the vascular smooth muscle cell (5, 9, 18). However, because the passive artery diameters (EDTA), wall thickness, and forced dilation responses of arteries from OVX mice were not significantly different compared with arteries from OVX + EST treated mice, we believe that estrogen did not alter the properties of vascular connective tissue or cause changes in vascular smooth muscle cell growth. Furthermore, because K+ channel blockers caused similar constriction between endothelium-denuded arteries from OVX and OVX + EST animals, it would seem that hyperpolarization mechanisms intrinsic to the smooth muscle cells were not altered by chronic estrogen treatment (53).

Estrogen and cerebral blood flow autoregulation. If gender and estrogen influence artery diameter and myogenic tone, then what are the benefits of estrogen on cerebral autoregulation? The primary role of cerebral autoregulation is to maintain a consistent cerebral blood flow during changes in mean arterial blood pressure (15). Therefore, in the normotensive and nondiseased state, exposure to estrogen probably has little or no effect on cerebral blood flow (26). In disease states, however, such as ischemic stroke due to thromboembolism, exposure to estrogen is likely to beneficially affect stroke outcome (1, 34). For example, female animals maintained higher tissue perfusion during cerebral artery occlusion (1). This gender-dependent flow preservation significantly reduced infarct size in female compared with male rats. Therefore, estrogen appears to influence cerebrovascular reserve capacity allowing flow to be preserved during conditions of artery occlusion.

In conclusion, gender and estrogen status affect myogenic reactivity of mouse cerebral arteries. Our data clearly show that inhibition of NOS caused significantly greater myogenic tone in arteries from male mice compared with arteries from untreated female mice. This gender difference was abolished by ovariectomy and was restored by estrogen treatment. Circulating estrogen appeared to affect an endothelium-derived cyclooxygenase product, which nearly completely compensated for normal NOS-dependent modulation of artery diameter. Estrogen status does not affect vascular smooth muscle growth, contractility, or K+ channel function. The physiological implications of the effects of estrogen on the cerebral circulation remain to be determined. However, myogenic tone is an important fundamental property of cerebral arteries relevant to the autoregulation of cerebral blood flow. Thus the current findings may help explain differences in incidence and severity of cerebrovascular diseases such as stroke and migraine that are associated with gender and estrogen status.


    ACKNOWLEDGEMENTS

This work was supported by National Heart, Lung, and Blood Institute Grant RO1 HL-50775.


    FOOTNOTES

Address for reprint requests and other correspondence: G. G. Geary, Dept. of Pharmacology, College of Medicine, Univ. of California, Irvine, Irvine, CA 92697-4625 (E-mail: gggeary{at}uci.edu).

The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. §1734 solely to indicate this fact.

Received 17 November 1999; accepted in final form 31 January 2000.


    REFERENCES
TOP
ABSTRACT
INTRODUCTION
METHODS
RESULTS
DISCUSSION
REFERENCES

1.   Alkayed, NJ, Harukuni I, Kimes AS, London ED, Traystman RJ, and Hurn PD. Gender-linked brain injury in experimental stroke. Stroke 29: 159-166, 1998[Abstract/Free Full Text].

2.   Bayliss, WM. On the local reaction of arterial wall to changes in internal pressure. J Physiol (Lond) 23: 220-231, 1902.

3.   Barrett-Conner, E, and Bush TL. Estrogen and coronary heart disease in women. JAMA 265: 1861-1867, 1991[Abstract].

4.   Bell, DR, Rensberger HJ, Koritnik DR, and Koshy A. Estrogen pretreatment directly potentiates endothelium-dependent vasorelaxation of porcine coronary arteries. Am J Physiol Heart Circ Physiol 268: H377-H383, 1995[Abstract/Free Full Text].

5.   Bhalla, RC, Toth KF, Bhatty RA, Thompson LP, and Sharma RV. Estrogen reduces proliferation and agonist-induced calcium increase in coronary artery smooth muscle cells. Am J Physiol Heart Circ Physiol 272: H1996-H2003, 1997[Abstract/Free Full Text].

6.   Case, J, and Davison CA. Estrogen alters relative contribution of nitric oxide and cyclooxygenase products to endothelium-dependent vasodilation. J Pharmacol Exp Ther 291: 524-530, 1999[Abstract/Free Full Text].

7.   Caulin-Glaser, T, Garcína-Cardeña G, Sarrel P, Sessa WC, and Bender JR. 17b-estradiol regulation of human endothelial cell basal nitric oxide release, independent of cytosolic Ca2+ mobilization. Circ Res 81: 885-892, 1997[Abstract/Free Full Text].

8.   Chataigneau, T, Félétou M, Huang PL, Fishman MC, Duhault J, and Vanhoutte PM. Acetylcholine-induced relaxation in blood vessels from endothelial nitric oxide synthase knockout mice. Br J Pharmacol 126: 219-226, 1999[ISI][Medline].

9.   Chen, S-J, Li H, Durand J, Oparil S, and Chen Y-L. Estrogen reduces myointimal proliferation after balloon injury of rat carotid artery. Circulation 93: 577-584, 1996[Abstract/Free Full Text].

10.   D'Angelo, G, and Meininger GA. Transduction mechanisms involved in the regulation of myogenic activity. Hypertension 23: 1096-1105, 1994[Abstract/Free Full Text].

11.   Davidge, ST, and Zhang Y. Estrogen replacement suppresses a prostaglandin H synthase-dependent vasoconstrictor in rat mesenteric arteries. Circ Res 83: 388-395, 1998[Abstract/Free Full Text].

12.   Davis, MJ. Myogenic response gradient in the arteriolar network. Am J Physiol Heart Circ Physiol 264: H2168-H2179, 1993[Abstract/Free Full Text].

13.   Dong, H, Waldron GJ, Galipeau D, Cole WC, and Triggle CR. NO/PGI2-independent vasorelaxation and the cytochrome P450 pathway in rabbit carotid artery. Br J Pharmacol 120: 695-701, 1997[ISI][Medline].

14.   Doni, MG, Whittle BJR, Palmer RMJ, and Moncada S. Actions of nitric oxide on the release of prostacyclin from bovine endothelial cells in culture. Eur J Pharmacol 151: 19-25, 1988[ISI][Medline].

15.   Edvinsson, L, MacKenzie ET, and McCulloch J. Cerebral blood flow and metabolism. In: Cerebral Blood Flow and Metabolism. New York: Raven, 1993, p. 553-580.

16.   Farhat, MY, Lavigne MC, and Ramwell PW. Vascular protective effects of estrogen. FASEB J 10: 615-624, 1996[Abstract].

17.   Finucane, FF, Madans JH, Bush TL, Wolf PH, and Kleinman JC. Decreased mortality in users of estrogen replacement therapy. Arch Intern Med 153: 73-79, 1993[Abstract].

18.   Fischer, GM, and Swain ML. Effects of sex hormones on blood pressure and vascular connective tissue in castrated and noncastrated male rats. Am J Physiol Heart Circ Physiol 232: H617-H621, 1977.

19.   Geary, GG, Duckles SP, and Krause DN. Estrogen enhances shear-induced release of NO in the perfused rat tail artery (Abstract). FASEB J 13: 422, 1999.

20.   Geary, GG, Krause DN, and Duckles SP. Estrogen reduces myogenic tone through a nitric oxide-dependent mechanism in rat cerebral arteries. Am J Physiol Heart Circ Physiol 275: H292-H300, 1998[Abstract/Free Full Text].

21.   Gödecke, A, Decking UKM, Ding Z, Hirchenhain J, Bidmon H-J, Gödecke S, and Schrader J. Coronary hemodynamics in endothelial NO synthase knockout mice. Circ Res 82: 186-194, 1998[Abstract/Free Full Text].

22.   Harper, SL, Bohlen HG, and Rubin MJ. Arterial and microvascular contributions to cerebral autoregulation in rats. Am J Physiol Heart Circ Physiol 246: H17-H24, 1984[Abstract/Free Full Text].

23.   Heinert, G, Nye PCG, and Paterson DJ. Nitric oxide and prostaglandin pathway in the regulation of hypercapnic cerebral vasodilation. Acta Physiol Scand 166: 183-193, 1999[ISI][Medline].

24.   Huang, PL, Huang Z, Mashimo H, Bloch KD, Moskowita MA, Bevan JA, and Fishman MC. Hypertension in mice lacking the gene for endothelial nitric oxide synthase. Nature 377: 239-242, 1995[Medline].

25.   Huang, A, Sun D, Koller A, and Kaley G. Gender differences in myogenic tone of rat arterioles is due to estrogen-induced, enhanced release of NO. Am J Physiol Heart Circ Physiol 272: H1804-H1809, 1997[Abstract/Free Full Text].

26.   Hurn, PD, Littleton-Kearney MT, Kirsch JR, Dharmarajan AM, and Traystman RJ. Postischemic cerebral blood flow recovery in the female: effect of 17beta -estradiol. J Cereb Blood Flow Metab 15: 666-672, 1995[ISI][Medline].

27.   Iadecola, C, Pelligrino DA, Moskowita MA, and Lassen NA. Nitric oxide synthase inhibition and cerebrovascular regulation. J Cereb Blood Flow Metab 14: 175-192, 1994[ISI][Medline].

28.   Iafrati, MD, Karas RH, Aronovitz M, Kim S, Sullivan TR, Jr, Lubahn DB, O'Donnell TF, Jr, Korach KS, and Mendelsohn ME. Estrogen inhibits the vascular injury response in estrogen receptor alpha -deficient mice. Nat Med 3: 545-548, 1997[ISI][Medline].

29.   Jun, SS, Chen Z, Pace MC, and Shaul PW. Estrogen upregulates cyclooxygenase-1 gene expression in ovine fetal pulmonary artery endothelium. J Clin Invest 102: 176-183, 1999[ISI][Medline].

30.   Kannel, WB, and Thom TJ. The incidence, prevalence and mortality of cardiovascular disease. In: The Heart, edited by Schlant RC, and Alexander RW.. New York: McGraw Hill, 1994, p. 185-197.

31.   Kauser, K, and Rubanyi GM. Gender differences in bioassayable endothelium-derived nitric oxide from isolated rat aortae. Am J Physiol Heart Circ Physiol 267: H2311-H2317, 1994[Abstract/Free Full Text].

32.   Kontos, HA, Wei EP, Navari RM, Levasseur JE, Rosenblum WI, and Patterson JL, Jr. Responses of cerebral arteries and arterioles to acute hypotension and hypertension. Am J Physiol Heart Circ Physiol 234: H371-H383, 1978[Abstract/Free Full Text].

33.   Latin-Hermoso, RL, Rosenfeld CR, Yuhanna IS, German Z, Chen Z, and Shaul PW. Estrogen acutely stimulates nitric oxide synthase activity in fetal pulmonary artery endothelium. Am J Physiol Lung Cell Mol Physiol 273: L119-L126, 1997[Abstract/Free Full Text].

34.   Ma, J, Ayata C, Huang P, Fishman MC, and Moskowitz MA. Regional cerebral blood flow response to vibrissal stimulation in mice lacking type I NOS gene expression. Am J Physiol Heart Circ Physiol 270: H1085-H1090, 1996[Abstract/Free Full Text].

35.   Matteis, M, Troisi E, Monalso BC, Caltagirone C, and Silvestrini M. Age and sex differences in cerebral hemodynamics. A transcranial doppler study. Stroke 29: 963-967, 1998[Abstract/Free Full Text].

36.   McNeill, AM, Kim N, Duckles SP, and Krause DN. Chronic estrogen treatment increases levels of endothelial nitric oxide synthase protein in rat cerebral microvessels. Stroke 30: 2186-2190, 1999[Abstract/Free Full Text].

37.   Meng, W, Ayata C, Waeber C, Huang PL, and Moskowitz MA. Neuronal NOS-cGMP-dependent ACh-induced relaxation in pial arterioles of endothelial NOS knockout mice. Am J Physiol Heart Circ Physiol 274: H411-H415, 1998[Abstract/Free Full Text].

38.   Paganini-Hill, A, Ross RK, and Henderson BE. Postmenopausal estrogen use and stroke: a prospective study. Br Med J 297: 519-522, 1998.

39.   Petersson, J, Zygmunt PR, and Högestött ED. Characterization of the potassium channels involved in EDHF-mediated relaxation in cerebral arteries. Br J Pharmacol 120: 1344-1350, 1997[ISI][Medline].

40.   Petitti, DB, Sidney S, Quesenberry CP, and Bernstein A. Ischemic stroke and use of estrogen and estrogen/progesterone as hormone replacement therapy. Stroke 29: 23-28, 1998[Abstract/Free Full Text].

41.   Prencipe, M, Ferretti C, Casini AR, Santini M, Giubilei F, and Culasso F. Stroke, disability, and dementia: results of a population survey. Stroke 28: 531-536, 1997[Abstract/Free Full Text].

42.   Puybasset, L, Béa M-L, Ghalen B, Giudicelli J-F, and Berdeauz A. Coronary and systemic hemodynamic effects of sustained inhibition of nitric oxide synthesis in conscious dogs. Evidence of cross talk between nitric oxide and cyclooxygenase in coronary vessels. Circ Res 79: 343-357, 1996[Abstract/Free Full Text].

43.   Raafat, AM, Hofseth LJ, Li S, Bennett JM, and Haslam SZ. A mouse model to study the effects of hormone replacement therapy on normal mammary gland during menopause: Enhanced proliferative response to estrogen in late postmenopausal mice. Endocrinology 140: 2570-2580, 1999[Abstract/Free Full Text].

44.   Rosenblum, WI, Donnenfeld H, and Aleu F. Effects of increased blood pressure on cerebral vessels in mice. Arch Neurol 14: 631-644, 1966[ISI][Medline].

45.   Rosenblum, WI, Nishimura H, Ellis EF, and Nelson G. The endothelium-dependent effects of thimersol on mouse pial arterioles in vivo: evidence for control of microvascular pressure by EDRF as well as prostaglandins. J Cereb Blood Flow Metab 12: 703-706, 1992[ISI][Medline].

46.   Rosenblum, WI, Nishimura H, and Nelson G. Endothelium-dependent L-Arg- and L-NMMA-sensitive mechanisms regulate tone of brain microvessels. Am J Physiol Heart Circ Physiol 259: H1396-H1401, 1990[Abstract/Free Full Text].

47.   Skarsgard, P, Van Breemen C, and Laher I. Estrogen regulates myogenic tone in pressurized cerebral arteries by enhanced basal release of nitric oxide. Am J Physiol Heart Circ Physiol 273: H2248-H2256, 1997[Abstract/Free Full Text].

48.   Sun, D, Huang A, Smith CJ, Stackpole CJ, Connetta JA, Shesely EG, Koller A, and Kaley G. Enhanced release of prostaglandins contribute to flow-induced arteriolar dilation in eNOS knockout mice. Circ Res 85: 288-293, 1999[Abstract/Free Full Text].

49.   Thorin-Trecases, N, Bartolotta T, Hyman N, Penar PL, Walters CL, Bevan RD, and Bevan JA. Diameter dependence of myogenic tone of human pial arteries. Possible relation to distensibility. Stroke 28: 2486-2492, 1997[Abstract/Free Full Text].

50.   Walmer, DK, Wrona MA, Hughes CL, and Nelson KG. Lactoferrin expression in the mouse reproductive tract during the natural estrous cycle: correlation with circulating estradiol and progesterone. Endocrinology 131: 1458-1466, 1992[Abstract].

51.   Wellman, GC, Bonev AD, Nelson MT, and Brayden JE. Gender differences in coronary artery diameter involve estrogen, nitric oxide, and the Ca2+-dependent K+ channel. Circ Res 79: 1024-1030, 1996[Abstract/Free Full Text].

52.   Wenger, NK, Speroff L, and Packard B. Cardiovascular health and disease women. N Engl J Med 329: 247-256, 1993[Free Full Text].

53.   White, RE, Darkow DJ, and Lang JLF Estrogen relaxes coronary arteries by opening BKCa channels through a cGMP-dependent mechanism. Circ Res 77: 936-942, 1995[Abstract/Free Full Text].

54.  Yamakawa N, Ohhashi M, Waga S, and Itoh T. Role of endothelium in regulation of smooth muscle membrane potential and tone in the rabbit middle cerebral artery. Br J Pharmacol 121: 1315-1322.

55.  Zhang L, Fishman MC, and Huang PL. Estrogen mediates the protective effects of pregnancy and chorionic gonadotropin in mouse model of vascular injury. Arterioscler Thromb Vasc Biol 19: 2059-2065.

Am J Physiol Heart Circ Physiol 279(2):H511-H519
0363-6135/00 $5.00 Copyright © 2000 the American Physiological Society


This article has been cited by other articles:


Home page
 Therapeutic Advances in Cardiovascular DiseaseHome page
P. D. Patel and R. R. Arora
Review: Endothelial dysfunction: A potential tool in gender related cardiovascular disease
Therapeutic Advances in Cardiovascular Disease, April 1, 2008; 2(2): 89 - 100.
[Abstract] [PDF]

Home page
 Am. J. Physiol. Heart Circ. Physiol.Home page
H. Girouard, A. Lessard, C. Capone, T. A. Milner, and C. Iadecola
The neurovascular dysfunction induced by angiotensin II in the mouse neocortex is sexually dimorphic
Am J Physiol Heart Circ Physiol, January 1, 2008; 294(1): H156 - H163.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Heart Circ. Physiol.Home page
O. Nevo, J. F. Soustiel, and I. Thaler
Cerebral blood flow is increased during controlled ovarian stimulation
Am J Physiol Heart Circ Physiol, December 1, 2007; 293(6): H3265 - H3269.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Gastrointest. Liver Physiol.Home page
T. Kawai, Y. Yokoyama, S. Kawai, S. Yokoyama, K. Oda, T. Nagasaka, M. Nagino, I. H. Chaudry, and Y. Nimura
Does estrogen contribute to the hepatic regeneration following portal branch ligation in rats?
Am J Physiol Gastrointest Liver Physiol, February 1, 2007; 292(2): G582 - G589.
[Abstract] [Full Text] [PDF]

Home page
 J. Appl. Physiol.Home page
D. N. Krause, S. P. Duckles, and D. A. Pelligrino
Influence of sex steroid hormones on cerebrovascular function
J Appl Physiol, October 1, 2006; 101(4): 1252 - 1261.
[Abstract] [Full Text] [PDF]

Home page
 Reproductive SciencesHome page
C. E. Wood and D. Giroux
Expression of Nitric Oxide Synthase Isoforms in the Ovine Fetal Brain: Alteration by Hormonal and Hemodynamic Stimuli
Reproductive Sciences, July 1, 2006; 13(5): 329 - 337.
[Abstract] [PDF]

Home page
 Am. J. Physiol. Heart Circ. Physiol.Home page
J. J. Andresen, N. I. Shafi, W. Durante, and R. M. Bryan Jr.
Effects of carbon monoxide and heme oxygenase inhibitors in cerebral vessels of rats and mice
Am J Physiol Heart Circ Physiol, July 1, 2006; 291(1): H223 - H230.
[Abstract] [Full Text] [PDF]

Home page
 Exp PhysiolHome page
K. Hayashi, M. Miyachi, N. Seno, K. Takahashi, K. Yamazaki, J. Sugawara, T. Yokoi, S. Onodera, and N. Mesaki
Variations in carotid arterial compliance during the menstrual cycle in young women
Exp Physiol, March 1, 2006; 91(2): 465 - 472.
[Abstract] [Full Text] [PDF]

Home page
 Am. J. Physiol. Heart Circ. Physiol.Home page
J. Ibrahim, A. McGee, D. Graham, J. C. McGrath, and A. F. Dominiczak
Sex-specific differences in cerebral arterial myogenic tone in hypertensive and normotensive rats
Am J Physiol Heart Circ Physiol, March 1, 2006; 290(3): H1081 - H1089.
[Abstract] [Full Text] [PDF]

Home page
 Mol. Interv.Home page
S. P. Duckles, D. N. Krause, C. Stirone, and V. Procaccio
Estrogen and mitochondria: a new paradigm for vascular protection?
Mol. Interv., February 1, 2006; 6(1): 26 - 35.