Effects of 17beta-estradiol on blood-brain barrier disruption during focal cerebral ischemia in younger and older rats.

This study was performed to compare the effects of 17beta-estradiol on blood-brain barrier disruption in focal cerebral ischemia between younger and older rats. Younger (three-month-old) and older (24-month-old) ovariectomized female Fischer 344 rats were studied. In one half of each age group, a 500 microg 17beta-estradiol 21-day release pellet and in another half, a vehicle pellet was implanted 21 days before the experiments. One hour after middle cerebral artery occlusion, the transfer coefficient (Ki) of 14C-alpha-aminoisobutyric acid and the volume of 3H-dextran distribution were determined to examine the degree of blood-brain barrier disruption. In all four groups, the Ki in the ischemic cortex was higher than in the corresponding contralateral cortex. There was no significant difference in the Ki in both cortices among the groups. The volume of dextran distribution of the ischemic cortex was only greater than in the corresponding contralateral cortex in the older 17beta-estradiol-treated group, and the volume of that group was greater than the younger 17beta-estradiol-treated group (4.00 +/- 1.29 VS. 2.13 +/- 0.88 ml/100 g). After analyzing the difference in Ki between the ischemic cortex and the contralateral cortex in each animal, the difference was significantly greater in the older 17beta-estradiol-treated rats than the older vehicle-treated rats (3.40 +/- 2.10 VS. 1.26 +/- 1.44 microl/g/min). In the younger rats, however, 17beta-estradiol did not significantly affect the difference. Our data showed that 17beta-estradiol treatment failed to attenuate the BBB disruption in the cerebral ischemic cortex in the older or younger Fischer 344 rats. However, our data also suggest the possibility that 17beta-estradiol could aggravate the BBB disruption in older rats.     

Effects of 17beta-estradiol on blood-brain barrier disruption in focal ischemia during GABA(A) receptor inhibition.

We performed this study to determine whether gamma-aminobutyric acid (GABA(A)) receptor inhibition could reverse the effect of 17beta-estradiol on blood-brain barrier (BBB) disruption in focal cerebral ischemia. Young ovariectomized rats were implanted with a 500 microg 17beta-estradiol 21-day release pellet or with a vehicle pellet 21 days before the experiments. Forty-five minutes after middle cerebral artery (MCA) occlusion, half of each group was infused with bicuculline (a GABA(A) receptor antagonist) 1 mg/kg/min for 2 min followed by 0.1 mg/kg/min up to the end of experiments. The other half was infused with the same volume of normal saline. The transfer coefficient (Ki) of 14C-alpha-aminoisobutyric acid and the volume of 3H-dextran distribution (70,000 Daltons) were determined to measure the degree of BBB disruption one hour after MCA occlusion. In the control vehicle-treated animals, the Ki in the ischemic cortex (7.2 +/- 2.6 microl/g/min) was higher than in the contralateral cortex (2.5 +/- 1.4 microl/g/min). After bicuculline infusion, the Ki in the ischemic cortex increased (10.6 +/- 5.4 microl/g/min) although the increase was not statistically significant. In the 17beta-estradiol treated animals, the Ki in the ischemic cortex (3.8 +/- 1.6 microl/g/min) was lower than control vehicle-treated rats. With bicuculline infusion, the Ki in the ischemic cortex (14.5 +/- 6.8 microl/g/min) was markedly increased. In the non-ischemic cortex, there was no significant difference in Ki among the experimental groups. The volume of dextran distribution was not significantly different between the experimental groups in the ischemic or non-ischemic cortex. Our data suggests that part of the reason for the decreased BBB disruption in the focal ischemic area after 17beta-estradiol treatment could be due to the interaction between GABA(A) receptors and 17beta-estradiol.     

Effects of 17beta-estradiol on blood-brain barrier disruption during focal ischemia in rats.

This study was performed to test whether 17beta-estradiol could attenuate the blood-brain barrier disruption caused by middle cerebral artery occlusion in the ovariectomized rats. Rats aged twelve to fourteen weeks were used in this study. Their ovaries were removed one week prior to the implantation of the pellets. For the 17beta-estradiol group, a 500 microg 17beta-estradiol 21 day-release pellet was implanted and for the control group, a vehicle pellet was implanted 21 days before the experiments. One hour after middle cerebral artery occlusion under isoflurane anesthesia, the transfer coefficient of 14C-alpha-aminoisobutyric acid (104 Daltons) and the volume of 3H-dextran (70,000 Daltons) distribution were determined to represent the degree of blood-brain barrier disruption. Blood pressures and blood gases were similar between controls and 17beta-estradiol-treated rats. In both groups, the transfer coefficient of the ischemic cortex was higher than that of the corresponding contralateral cortex (control: Ischemic Cortex 12.5 +/- 5.9 microl/g/min, Contralateral Cortex 3.0 +/- 1.6, p < 0.001; 17beta-estradiol: Ischemic Cortex 6.7 +/- 3.3 micro l/g/min, Contralateral Cortex 2.2 +/- 0.9, p < 0.01). There was no significant difference in the transfer coefficient of the contralateral cortex between these two groups. However, the transfer coefficient of the Ischemic Cortex of the 17beta-estradiol-treated animals was 46 % lower than that of the control, vehicle-treated rats (p < 0.05). The increase of the volume of 3H-dextran distribution with middle cerebral artery occlusion was significant in the vehicle-treated rats (Ischemic Cortex: 6.4 +/- 2.7 ml/100 g, Contralateral Cortex: 3.8 +/- 0.8, p < 0.01) but not in the 17beta-estradiol-treated animals. Our data suggest that chronic 17beta-estradiol treatment was effective in reducing blood-brain barrier disruption during focal ischemia in the ovariectomized rats.     

Elevation of Striatal Dopamine Receptors by Estrogen: Dose and Time Studies

Administration to male rats of a single dose of 17 beta-estradiol valerate (8-500 micrograms/rat) or implantation of a pellet containing 17 beta-estradiol (0.5-50 mg/rat) increased serum 17 beta-estradiol levels in a dose-dependent relationship when measured on the sixth day after administration. At the same time, after these doses, the serum rat prolactin (rPRL) levels were doubled and the striatal 3,4-dihydroxyphenylethylamine (DA, dopamine) receptor densities were increased 20%. A single dose of 17 beta-estradiol valerate of 4 micrograms/rat or less did not alter serum 17 beta-estradiol or rPRL levels or the striatal DA receptor density. After the single injection of 17 beta-estradiol valerate (125 micrograms/rat) the serum 17 beta-estradiol levels peaked at 1 day, the serum rPRL levels peaked at 2 days, and the striatal DA receptor density elevation peaked from 4 to 8 days. Implantation of a pellet containing 17 beta-estradiol (25 mg/rat) produced a constant elevation of serum 17 beta-estradiol levels from 1 to 10 days. Whereas the serum rPRL levels were continuously elevated about two-fold, the densities of the striatal DA receptors were increased significantly by 20-25% only from 4 to 8 days after pellet implantation. These results indicate that striatal DA receptor density rises and returns to control levels during the constant elevation of serum 17 beta-estradiol and rPRL levels.(ABSTRACT TRUNCATED AT 250 WORDS)     

Effects of estrogen on venous function in rats with chronic heart failure

The effect of 17beta-estradiol on venous function was investigated in ovariectomized rats with heart failure. Rats (50-60 days old) were ovariectomized and implanted with 60-day-release pellets that contain 17beta-estradiol (1.5 mg) or vehicle. The left coronary artery was ligated 7 days later. Another group of ovariectomized rats was given vehicle pellets and then a sham operation was performed. The rats were studied while under pentobarbital anesthesia at 7 wk after ligation. Ligated rats, relative to sham groups, had lower mean arterial pressure (MAP, -34 mmHg) and cardiac output (CO, -38%); higher arterial resistance (R(A), +12%) and venous resistance (R(V), +116%); mean circulatory filling pressure (MCFP, +40%) and left ventricular end-diastolic pressure (LVEDP, +11 mmHg); and similar cardiovascular responses to norepinephrine (NE). Treatment of ligated rats with 17beta-estradiol increased CO (+16%); reduced R(A) (-16%), R(V) (-35%), MCFP (-23%), and LVEDP (-3 mmHg); and augmented MAP, R(V,) and MCFP responses to NE. Therefore, 17beta-estradiol reduced MCFP, and this reduced preload (LVEDP). 17beta-Estradiol decreased R(V), which, along with decreased R(A) (afterload), led to an increase in CO. 17beta-Estradiol likely augmented vasoconstriction to NE through an improvement on the cardiovascular status.     

Comparison of blood brain barrier permeability in normal and ovariectomized female rats that demonstrate right or left paw preference

We explored the relations among paw preference, cerebral asymmetry and asymmetrical disruption of blood-brain barrier (BBB) permeability in normal and ovariectomized female rats with known paw preference. A high dose of pentylenetetrazol was used to disrupt the BBB and induce acute hypertension. To determine the areas of macroscopic infarct, samples were stained with 2,3,5-triphenyltetrazolium chloride. Histological staining techniques were used to show the areas of infarct microscopically on paraffin sections. Sixty-two percent of the rats demonstrated right paw preference, 24% demonstrated left paw preference and 14% were ambidextrous. Areas of infarct, which indicated destruction of the BBB, were determined microscopically and macroscopically in rats that demonstrated right and left paw preference. We found a relation between permeability of the BBB and paw preference. There may be a relation between paw preference, cerebral asymmetry and asymmetrical destruction of the BBB in rats. Asymmetrical destruction of the BBB in experimental rats was similar to the control group, which had asymmetrically disrupted BBB with respect to paw preference. Like the control rats, asymmetrical areas of infarct consistent with cerebral asymmetry were observed in ovariectomized rats.     

Effects of sex and estrogen on myosin COOH-terminal isoforms and contractility in rat aorta

We reported that estrogen treatment of ovariectomized rats increased uterine smooth muscle contractility and the ratio of the COOH-terminal myosin heavy chain isoform SM1 (204 kDa) and SM2 [200 kDa; Hewett TE, Martin AF, Paul RJ. J Physiol (Lond) 460: 351-364, 1993]. We extended this model to study sex and estrogen effects on vascular contractility. Experimental groups included 10- to 14-wk-old male (M), female (F), ovariectomized female (OF), and OF treated with estrogen (OF&E) for 7 days with a subcutaneous pellet delivery system, resulting in 17beta-estradiol of 85 (OF&E) vs. 5 (OF or M) pg/ml. The SM1-to-SM2 ratio increased from 1.8 to 2.6 in thoracic aorta, similar to uterine muscle. Isometric force was measured in 5-mm segments of intact and endothelium-denuded (-endo) aorta. With KCl, the maximum forces were in the order OF approximately M > OF&E, and ED50 OF&E > OF approximately M. Differences in ED50 with estrogen persisted after endothelial denudation. The decreased force in -endo OF aorta was not seen in OF&E, suggesting that estrogen altered an endothelium-dependent effect. No differences in maximum forces were noted with norepinephrine: ED50 OF > OF&E > M. Estrogen treatment, in contrast to KCl, increased sensitivity. Endothelial denudation increased sensitivity but reduced the differences between groups. With ACh relaxation, males were more sensitive than females, and estrogen had no effect. In the abdominal aorta, there were no changes in SM1/SM2 with 17beta-estradiol, and differences in contractility were blunted. In summary, estrogen treatment decreased responses to KCl but increased sensitivity to norepinephrine; male rats always demonstrated the highest contractility. An increase in the COOH-terminal myosin heavy chain isoform SM1-to-SM2 ratio with 17beta-estradiol treatment may underlie the changes observed in contractility.     

Estrogen replacement increases coronary artery distensibility in ovariectomized rats.

The effect of estrogen on the passive characteristics of arteries is not known. We hypothesized that estrogen would increase arterial distensibility as part of its protective effect on the vasculature. Female Sprague-Dawley rats were ovariectomized at 11 weeks of age. One group received a placebo (n = 6), while two other groups (n = 5 each) of rats received a 17beta-estradiol pellet (0.15 mg or 0.5 mg with 60-day release). After 4 weeks of estrogen replacement, coronary and mesenteric arteries (<200 microm diameter) were dissected and mounted on a dual-chamber arteriograph. Lumen diameter and wall thickness were measured in pressurized arteries. The relative changes in diameter (distensibility) as well as wall thickness per unit change in pressure were significantly increased (p < 0.05) in the coronary arteries of the 0.5 mg estradiol replaced rats compared with the ovariectomized control animals and the 0.15 mg estradiol replaced rats. Surprisingly, in the mesenteric arteries from the same animals, there was no difference in distensibility or pressure - wall thickness among the groups. This study provides experimental data of a novel hypothesis that estrogen may afford part of its protection through vascular remodeling of the coronary circulation.     

Acute effects of 17beta -estradiol on femoral veins from adult gonadally intact and ovariectomized female pigs

Our experiments were designed to determine the acute effects of 17beta-estradiol on femoral veins from intact and ovariectomized female pigs. Rings of femoral veins with or without endothelium were suspended in organ chambers for measurement of isometric force. Concentration-response curves to 17beta-estradiol (10(-9) to 10(-5) M) were obtained in veins contracted with prostaglandin F(2alpha) in the absence and presence of inhibitors of either estrogen receptors (ICI-182780; 10(-5) M), nitric oxide synthase [N(G)-monomethyl-l-arginine (l-NMMA); 10(-4) M], soluble guanylate cyclase (1-H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one; 10(-5) M), or potassium channels (tetraethylammonium; 10(-2) M). Estrogen receptors were identified with the use of Western blotting and immunostaining in veins of both groups. 17beta-Estradiol caused acute endothelium-dependent relaxations in both groups. Relaxations to 17beta-estradiol were inhibited by l-NMMA and 1-H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one but not ICI-182780. Tetraethylammonium inhibited relaxations only in veins with endothelium from intact females. Results indicate that 17beta-estradiol causes acute endothelium-dependent relaxations in femoral veins. The relative contribution of nitric oxide and K(+) channels as mechanisms involved in relaxations to 17beta-estradiol in femoral veins is modulated by ovarian status.     

Regulation of leptin by steroid hormones in rat adipose tissue.

We investigated if steroid hormones regulate the secretion and the expression of leptin in female and male rat adipose tissue fragments in vitro. Dexamethasone time and dose-dependently increased the secretion and mRNA expression of leptin with a half-maximal stimulation of approximately 1 nM. A time-course revealed a maximal stimulatory effect of 17 beta-estradiol after 24 hours. In male adipose tissue 17 beta-estradiol increased leptin secretion (32% by 50 nM 17 beta-estradiol, P = 0.07 and 34% by 500 nM 17 beta-estradiol, P < 1780.05) after 24 hours. An additional effect of estrogen was seen in the dexamethasone (50 nM) stimulated cells (38% with 50 nM 17 beta-estradiol, P < 0.05 and 48% by 500 nM 17 beta-estradiol, P < 0.05). Basal secretion of leptin was equal in female and male adipose tissue, whereas the effects of 17 beta-estradiol (50 nM) and dexamethasone were significantly increased in female as compared with male adipose tissue. Progesterone, testosterone, dihydrotestosterone and dehydroepiandrostendione-sulfate neither affected leptin secretion in male nor female adipose tissue in vitro. Furthermore, to investigate the effect of estrogen female rats were ovariectomized (OVX) and the adipose tissue was incubated in vitro and compared with adipose tissue leptin secretion from sham operated rats (SHAM), and with ovariectomized rats treated with 17 beta-estradiol (EST). A decreased basal and dexamethasone-stimulated leptin secretion from OVX rats compared with SHAM rats was found (P < 0.005) whereas 17 beta-estradiol treatment of ovariectomized rats maintained a normal leptin secretion. However, the dexamethasone stimulation was equally increased above basal levels in SHAM, OVX and EST rats (3.7 +/- 1.2, 2.9 +/- 0.8, 4.2 +/- 1.4, NS, ANOVA) respectively.     

Effects of Exogenous Excitatory Amino Acid Neurotransmitters on Blood–Brain Barrier Disruption in Focal Cerebral Ischemia

This study was performed to determine whether exogenous N-methyl-d-aspartate (NMDA) or alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) would aggravate blood–brain barrier (BBB) disruption in focal cerebral ischemia in rats. Forty-five minutes after middle cerebral artery (MCA) occlusion, one of the following patches was applied to the exposed ischemic cerebral cortex of each rat: normal saline (control), 10⁻⁵ M AMPA, 10⁻⁴ M AMPA, 10⁻⁵ M NMDA, or 10⁻⁴ M NMDA. At 1 h after MCA occlusion, BBB permeability was determined by measuring the transfer coefficient (Ki) of ¹⁴C-α-aminoisobutyric acid (¹⁴C-AIB). In all experimental groups, the Ki of the ischemic cortex (IC) was higher than that of the corresponding contralateral cortex (CC). The Ki of the IC of the animals treated with 10⁻⁴ M AMPA or 10⁻⁴ M NMDA was higher (+41%: P < 0.05 and +33%: P < 0.05, respectively) than that of the control animals. Our data demonstrated that exogenous NMDA or AMPA could further aggravate the BBB disruption in focal cerebral ischemia. Any insult increasing the release of excitatory neurotransmitters could further aggravate BBB disruption and brain edema during the ischemic period.     

Effects of Metabotropic Glutamate Receptor Stimulation on Blood–Brain Barrier Permeability During Focal Cerebral Ischemia

This investigation was performed to evaluate whether ACPD [(1S, 3R)-1-aminocyclopentane-1, 3-dicarboxylic acid], a metabotropic glutamate receptor agonist, would enhance the degree of increase in blood-brain barrier (BBB) permeability caused by focal cerebral ischemia. In this study, male Wistar rats were placed in control (n = 7) and ACPD (n = 7) groups under isoflurane anesthesia. Twenty minutes after middle cerebral artery (MCA) occlusion, patches of 10(-5) M ACPD or normal saline were placed on the ischemic cortex (IC) for a period of 40 min. Patches were changed every 10 min. One hour after MCA occlusion, BBB permeability was determined by measuring the transfer coefficient (Ki) of [alpha-14C] aminoisobutyric acid. There were no statistical differences in systemic blood pressures and heart rates between these groups. Blood gases were within normal limits. In the control group, the Ki of ischemic cortex (IC) was 2.1 times that of the contralateral cortex (CC) (3.7+/-0.9 vs. 1.8+/-0.3 microl/g/min). In the ACPD group, the Ki of the IC was 3.3 times that of the CC (5.0+/-0.7 vs. 1.5+/-0.4 microl/g/min). The increase in Ki of the ACPD group in the ischemic cortex was significantly greater than that in the control group. There was no significant difference in the Ki of the CC between these groups. Our data suggest that activation metabotropic glutamate receptors in the cortex can further augment the increase in BBB permeability caused by focal ischemia.     

Modulation of cardiac mast cell-mediated extracellular matrix degradation by estrogen

There are fundamental differences between males and females with regard to susceptibility to heart disease. Although numerous animal models of heart failure have demonstrated that premenopausal females are afforded cardioprotection and, therefore, fare better in the face of cardiac disease than their male counterparts, many questions as to how this occurs still exist. Recently, we showed that 1) increased mast cell density is associated with adverse ventricular remodeling and 2) chemically induced mast cell degranulation using compound 48/80 resulted in remarkable changes in matrix metalloproteinase (MMP) activity, cardiac collagen structure, and cardiac diastolic function in normal male rats. With the known gender differences in cardiac disease in mind, we sought to examine the effects of chemically induced cardiac mast cell degranulation in isolated, blood-perfused hearts of intact female rats, ovariectomized female rats, and ovariectomized female rats treated with 17beta-estradiol. In response to mast cell degranulation, no significant differences in cardiac function, MMP-2 activity, or collagen volume fraction were observed between intact female rats and ovariectomized female rats treated with estrogen. In the ovariectomized female group, a significant rightward shift in the left ventricular pressure-volume relation, accompanied by a marked 133% increase in active MMP-2 values over that in the intact female group, was noted after treatment with compound 48/80 (P < or = 0.05), along with a significant reduction in collagen volume fraction below control (0.46 +/- 0.23 vs. 0.73 +/- 0.13%, P < or = 0.05). These findings indicate that estrogen's cardioprotective role can be partially mediated by its effects on cardiac mast cells, MMPs, and the extracellular matrix.     

Subcellular distribution and properties of progesterone (delta4-steroid) 5alpha-reductase in rat medial basal hypothalamus.

The subcellular distribution and properties of rat hypothalamic progesterone 5 alpha-reductase, which accelerates the conversion of progesterone to 5 alpha-pregnane-3,20-dione, have been investigated by utilizing 3H-labeled substrate and a reverse isotopic dilution assay system. The enxymic activity was associated primarily with a cell debris-membranes fraction deribed from the 100 x g pellet. This fraction contained mainly membrane-like particulates and was free of nuclei. Little or no activity was associated with the purified nuclei. The hypothalamic 5 alpha-reductase was stimulated by NADPH but not by NADH. The reaction proceeded optimally over a pH range of 6.0 to 7.2 and at a temperaturhe substrate specificity of the enzyme for other delta 4-3-ketosteroids and the ability of these steroids to inhibit the 5 alpha reduction of [1,2-3H]progesterone as well as the effect of 17 beta-estradiol were also studied. 20 alpha-hydroxypregn-4-en-3-one was more reactive that progesterone, while testosterone was the least reactive. The estimated Km for 20 alpha-hydroxypregn-4-en-3-one was 8.6 +/- 1.9 x 10(-7) M, and for testosterone, 1.6 +/- 1.4 x 10(-5) M. The inhibition studies indicate that 20 alpha-hydroxypregn-4-en-3-one and 17 beta-estradiol are competitive and noncompetitive inhibitors, respectively, of the 5 alpha reduction of progesterone with Ki of 6.0 +/- 3.0 x 10(-8) M for 20 alpha-hydroxypregn-4-en-3-one and Kii (intercept inhibition constant) of 2.6 +/- 0.7 x 10(-5) M and Kis (slope inhibition constant) of 3.6 +/- 0.6 x 10(-5) M for 17 beta-estradiol. Testosterone is a poor competitive inhibitor of the reaction.     

Mechanisms of 17beta-estradiol on the production of ET-1 in ovariectomized rats

In order to clarify the mechanism underlying the possible preventive effect of estrogen on atherogenesis, we investigated the role of 17beta-estradiol (E2) in the regulation of endothelin-1 (ET-1) production in ovariectomized rats, which may contribute to atherogenesis. Female Spragure-Dawly rats were randomly divided into three groups: sham-operated group (sham), ovariectomized group (OVX) and 17beta-estradiol replacement group (OVX + E2, 20 microg(-1).kg.d(-1),s.c.). 4 weeks after operation, the plasma concentration of ET-1, clearance of ET-1, functional ECE activity and preproET-1 mRNA expression in aorta were measured. Concentration of plasma ET-1 change from 107.8 +/- 18.3 pg/ml (sham) and 135.5 +/- 27.6 pg/ml (OVX + E2) to 190.7 +/- 25.5 pg/ml (OVX ) (n = 8, p < 0.05). There was no significant difference in the clearance of 125IET-1 among three groups (p > 0.05). Functional ECE activity was increased in OVX group in comparison to that in sham group (p < 0.05). The OVX increased the preproET-1 mRNA expression in sham, whereas treatment with estrogen reversed these changes (p < 0.05). The present study have shown that estrogen down-regulates plasma ET-1 levels by inhibiting the preproET-1 mRNA expression and functional ECE activity. Clearance of ET-1 was not affected. Inhibition of ET-1 production mediated by modulating ECE activity may be one of the novel mechanisms of the protective of estrogens on the cardiovascular system.     

Skin temperature rise induced by calcitonin gene-related peptide in gonadotropin-releasing hormone analogue-treated female rats and alleviation by Keishi-bukuryo-gan, a Japanese herbal medicine

The effects of Keishi-bukuryo-gan on calcitonin gene-related peptide (CGRP)-induced elevation of skin temperature were investigated in gonadotropin-releasing hormone (GnRH) analogue-treated female rats. Leupline (1.0 mg/kg) as the GnRH analogue was subcutaneously (s.c.) injected into female rats. After Keishi-bukuryo-gan (100-1,000 mg/kg, p.o.) or 17beta-estradiol (0.010 mg/kg, s.c.) was administered to GnRH analogue-treated rats for 14 days, CGRP-induced skin temperature elevation, concentration of plasma 17beta-estradiol and pituitary gonadotropin (luteinizing hormone; LH, and follicle stimulating hormone; FSH) were measured. In addition, effects of 17beta-estradiol and Keishi-bukuryo-gan on the proliferation of estrogen-dependent human breast cancer (MCF-7) cells were investigated under in vitro conditions. GnRH analogue significantly lowered the concentrations of plasma 17beta-estradiol and pituitary gonadotropins. Tissue weights of the ovaries and uterus were also decreased by the analogue. Under the condition of estrogen deficiency, intravenous (i.v.) injection of exogenous CGRP (10 microg/kg) elevated the skin temperature of the hind paws more significantly than it did in sham-treated control rats. Estrogen supplementation inhibited this elevation of skin temperature with restoration of both the lowered plasma estrogen level and the decreased uterine weight in GnRH analogue-treated rats. On the other hand, Keishi-bukuryo-gan inhibited the elevation of skin temperature in a dose-dependent manner without restoring the plasma estrogen level and uterine weight. In addition, in an in vitro study, MCF-7 cells proliferated in a dose-dependent manner by the addition of 17beta-estradiol (10(-13)-10(-8) M) to the medium. However, Keishi-bukuryo-gan (10(-6)-10(-4) mg/ml) did not activate the MCF-7 cell proliferation. These results suggest that Keishi-bukuryo-gan, which does not exhibit estrogen activity, may be useful for the treatment of hot flashes in women who are undergoing medical ovariectomy with a GnRH analogue.     

The effect of hypertension on serum nitric oxide and vascular endothelial growth factor concentrations. A study in DOCA-Salt hypertensive ovariectomized rats

CardioVascular Disease (CVD) accounts for considerable mortality and morbidity in developed countries. Most of the common forms of CVD, such as hypertension, are caused by functional and structural changes in endothelial function. This study was designed to study the effect of hypertension on serum Nitric Oxide (NO) and Vascular Endothelial Growth Factor (VEGF) concentrations in DOCA-Salt hypertensive ovariectomized rats. Thirty female rats were ovariectomized. Blood samples were taken and the animals were divided into hypertensive and control groups. Hypertension was induced by DOCA-Salt method. DOCA was injected 30 mg/kg of body weight subcutaneously, twice a week with NaCl 1% instead of tap water for drinking throughout the experiment. The control group received normal saline injection with usual drinking water. Results showed that serum NO concentration in DOCA-Salt hypertensive rats was lower than the control group (18.35 +/- 5.31, 45.01 +/- 12.54 micromol/l, respectively) (p < 0.05). Also, the mean serum VEGF concentration was raised after induced hypertension (120.55 +/- 8.11 vs. 88.58 +/- 2.24 pg/ml) (p < 0.05). In conclusion, reduced serum NO and increased serum VEGF concentrations in hypertensive animals support the concept of endothelial dysfunction in hypertensive subjects.     

Aging and sex influence the permeability of the blood-brain barrier in the rat

The aim of the present study was to investigate the existence of aging- and sex-related alterations in the permeability of the blood-brain barrier (BBB) in the rat, by calculating a unidirectional blood-to-brain transfer constant (Ki) for the circulating tracer [14C]-alpha-aminoisobutyric acid. We observed that: a) the permeability of the BBB significantly increased within the frontal and temporo-parietal cortex, hypothalamus and cerebellum in 28-30 week old rats, in comparison with younger animals; b) in several brain areas of female intact rats higher Ki values (even though not significantly different) were calculated at oestrus than at proestrus; c) in 1-week ovariectomized rats there was a marked increase of Ki values at the level of the frontal, temporo-parietal and occipital cortex, cerebellum and brain-stem. One can speculate that aging- and sex-related alterations in the permeability of the BBB reflect respectively changes in brain neurochemical system activity and in plasma steroid hormone levels.     

Differential Modulation of D1 and D2 Dopamine-Sensitive Adenylate Cyclases by 17β-Estradiol in Cultured Striatal Neurons and Anterior Pituitary Cells

Primary cultures of anterior pituitary cells from female rats and of mouse embryonic striatal neurons were used to study the effects of 17 beta-estradiol on D1- and D2-dopamine (DA)-sensitive adenylate cyclase. 17 beta-Estradiol pretreatment (10(-9) M, 72 h) suppressed the D2-DA-induced inhibition of adenylate cyclase activity in anterior pituitary cells. The steroid (10(-9) M, 24 h) also blocked the D2-DA-evoked response in striatal neurons whereas it enhanced by twofold the D1-DA-induced stimulation of the enzyme activity in these neurons. All these effects of the steroid were dose dependent and specific, as neither 17 alpha-estradiol, dexamethasone, nor progesterone used at the same concentration (10(-9) M) was effective. Furthermore, the modulation of DA-sensitive adenylate cyclases by the steroid required long-term exposure of living cells to 17 beta-estradiol since neither 17 beta-estradiol pretreatment for 4 h nor its addition to broken cells directly into the adenylate cyclase assay induced any alteration in the DA-sensitive adenylate cyclase activity. These results are in agreement with a genomic effect of the steroid. Using both anterior pituitary cells and striatal neurons in culture, 17 beta-estradiol affected neither the total number of DA (D1 and D2) receptors nor the estimated number of adenylate cyclase catalytic units. Therefore, it is suggested that the steroid modifies the coupling process by a mechanism that still has to be elucidated. These results demonstrate an effect of 17 beta-estradiol on DA target cells in both systems.(ABSTRACT TRUNCATED AT 250 WORDS)