J Physiol Volume 514, Number 3, 875-884, February 1, 1999
The Journal of Physiology (1999), 514.3, pp. 875-884
© Copyright 1999 The Physiological Society
The effect of 17
-oestradiol on regional blood flow in anaesthetized pigs
G. Vacca, A. Battaglia, E. Grossini, D. A. S. G. Mary, C. Molinari and N. Surico
Laboratorio di Fisiologia, Dipartimento di Scienze Mediche, Facoltà di Medicina e Chirurgia di Novara, Università del Piemonte Orientale A. Avogadro, Italy
MS 8468 Received 10 July 1998; accepted after revision 14 October 1998.
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ABSTRACT |
- The present study was designed to investigate the effects of 17-oestradiol on the mesenteric, renal, iliac and coronary circulations and to determine the mechanisms involved.
- In pigs anaesthetized with sodium pentobarbitone, changes in blood flow in the superior mesenteric, left renal, left external iliac and left circumflex coronary arteries caused by intravenous infusion of 17-oestradiol at constant heart rate and arterial pressure were assessed using electromagnetic flowmeters.
- In eight pigs, infusion of 2 µg h-1 of the hormone caused an increase in renal, iliac and coronary blood flow without affecting mesenteric blood flow, left ventricular dP/dtmax (rate of change of left ventricular systolic pressure) and filling pressures of the heart. In four pigs, these vasodilator effects were enhanced by graded increases in the dose of the hormone between 1, 2 and 3 µg h-1; the highest dose also caused an increase in mesenteric blood flow.
- In five pigs, blockade of muscarinic cholinoceptors and adrenoceptors with the intravenous administration of atropine, propranolol and phentolamine did not affect the vasodilator responses caused by infusion of 2 µg h-1 of 17-oestradiol.
- The increases in renal, iliac and coronary blood flow caused by infusion of 2 µg h-1 of 17-oestradiol were prevented, respectively, by the injection of N
-nitro-L-arginine methyl ester (L-NAME) into the renal artery (five pigs), the iliac artery (five pigs) or the coronary artery (five pigs). In five pigs, all responses were prevented by injection of L-NAME into all three arteries. In two pigs, injection of L-NAME into the mesenteric, renal, iliac and coronary arteries abolished the vasodilator responses to the infusion of 3 µg h-1 of 17-oestradiol.
- The present study shows that intravenous infusion of 2 µg h-1 of 17-oestradiol primarily dilated renal, iliac and coronary circulations and that a higher dose of the hormone also caused vasodilatation in the mesenteric vascular bed. The mechanism of these responses was shown to be nitric oxide dependent.
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INTRODUCTION |
It is widely accepted that oestrogens have a vasodilator effect on the coronary, genito-urinary and limb vasculature (Magness et al. 1993; Volterrani et al. 1995; Riedel et al. 1995; Rosenfeld et al. 1996) and are thought to play an important role in the lower incidence of cardiovascular disease in women (Samaan & Crawford, 1995). Controlled experiments in animals have shown that acute administration of 17-oestradiol causes relaxation of coronary smooth muscle (Collins et al. 1994; Wellman et al. 1996; Lamping & Nuno, 1996; Thompson & Weiner, 1997) and vasodilatation (Eckstein et al. 1994; Gorodeski et al. 1995; Sudhir et al. 1995). However, little is known about the effect of this hormone on abdominal, renal and iliac vascular beds despite their known involvement in cardiovascular regulation. Indeed it has been reported in ewes that these vascular beds are not responsive to this hormone (Nuwayhid et al. 1975). Also, renal blood flow in the rat is suggested to increase following administration of 17-oestradiol (Kapitola et al. 1994) and to decrease in women at high plasma levels of the hormone (Oelkers, 1996).
The mechanisms of the known regional vasodilator effect of oestrogens have been controversial. In isolated vessels such as the aorta (Cheng et al. 1994; Paredes-Carbajal et al. 1995) and the coronary artery (Collins et al. 1994; Wellman et al. 1996; Thompson & Weiner, 1997) and in the isolated perfused heart (Gorodeski et al. 1995), the vasodilatation has been attributed mainly to nitric oxide. In anaesthetized dogs this mechanism was not confirmed as the cause of oestrogen-induced coronary vasodilatation (Sudhir et al. 1995).
The present investigation was planned to find out the effect of acute administration of 17-oestradiol on mesenteric, renal, iliac and coronary blood flow in anaesthetized female pigs and to determine whether the mechanism of this effect involved vascular muscarinic cholinoceptors and adrenoceptors and/or nitric oxide. For this purpose, experiments were performed under constant heart rate and arterial pressure to avoid interference from systemic reflex responses and local effects.
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METHODS |
The experiments were carried out in 39 pre-pubertal female pigs, weighing 65-70 kg. The animals, which were fasted overnight, were anaesthetized with intramuscular ketamine (20 mg kg-1; Parke-Davis, Milan, Italy) followed after about 15 min by intravenous sodium pentobarbitone (15 mg kg-1; Siegfried Zofingen, Switzerland), and artificially ventilated throughout all the experiments with oxygen-enriched air with a respiratory pump (Harvard 613; Harvard Apparatus, South Natick, MA, USA). Anaesthesia was maintained throughout the experiments by a continuous intravenous infusion of sodium pentobarbitone (7 mg kg-1 h-1) and assessed as previously reported (Linden & Mary, 1983) from responses of the animals to somatic stimuli. The experiments were carried out in accordance with the 'Decreto legislativo 27 gennaio 1992, n. 116 (Attuazione della direttiva n. 86/609/CEE in materia di protezione degli animali utilizzati a fini sperimentali o ad altri fini scientifici)' and supervised by the Veterinary Service of the University of Turin.
Pressures in the ascending aorta and in the right atrium were recorded via catheters connected to pressure transducers (Statham P23 XL; Gould, Valley View, OH, USA) inserted into the right femoral artery and the right external jugular vein, respectively. The abdomen was opened with a mid-line incision and a flowmeter probe (model BL 613; Biotronex Laboratory Inc., Chester, MD, USA) was placed near the origin of the superior mesenteric, left renal and left external iliac arteries. The chest was opened in the left fourth intercostal space, the pericardium was cut and an electromagnetic flowmeter probe was positioned around the proximal part of the left circumflex coronary artery. Distal to the probe, a plastic snare was placed around each artery for the assessment of zero blood flow. Each probe was calibrated in vitro at the end of each experiment. In some pigs, a catheter was inserted into a small artery originating in the marginal or oblique branch of the left circumflex coronary artery and used for the intracoronary injection of N-nitro-L-arginine methyl ester (L-NAME; Sigma). In some animals, L-NAME was administered in mesenteric, renal and iliac arteries using a catheter connected to a butterfly needle inserted into the arteries.
Left ventricular pressure was measured by means of a catheter connected to a pressure transducer (Statham P23 XL) inserted through the left atrium. The frequency response of the catheter- manometer system was found to be flat (± 5 %) up to 40 Hz. To pace the heart, electrodes were sewn on the left atrial appendage and connected to a stimulator (model S8800; Grass Instruments, Quincy, MA, USA) which delivered pulses of 3-5 V with 2 ms duration at the required frequency. Arterial blood samples were used to measure pH, PO2, PCO2 (with a gas analyser; IL 1304; IL Instrumentation Laboratory, Lexington, MA, USA), and the haematocrit. Normal values of pH, PO2 and PCO2 of 7·42 ± 0·02, 84·3 ± 3·9 mmHg and 39·8 ± 0·7 mmHg, respectively, have been reported in pre-pubertal pigs (Houpt, 1986). In the present study, the animals were artificially ventilated with oxygen-enriched air and values of pH and PCO2 were maintained within normal reported limits during the experiments by the infusion of a solution of 2·8 % sodium bicarbonate and by adjusting the respiratory stroke volume, when necessary (Linden & Mary, 1983).
To prevent changes in arterial blood pressure during the experiments, a cannula was introduced into the left internal mammary artery and connected to a reservoir containing Ringer solution (SIFRA-Società Italiana Farmaceutici Ravizza, Verona, Italy) kept at 38°C. The reservoir was pressurized using compressed air, which was controlled with a Starling resistance, and pressure within the reservoir was measured by a mercury manometer. This method has been shown in anaesthetized pigs to allow the aortic blood pressure to be maintained at steady levels without significant changes in left ventricular pressure or the haematocrit (e.g. Vacca et al. 1996a). Coagulation of the blood was avoided by the intravenous injection of heparin (Parke-Davis; initial doses of 500 i.u. kg-1, and subsequent doses of 50 i.u. kg-1 every 30 min). The rectal temperature of the pigs was monitored and kept between 38 and 40°C using an electric pad.
Mean and phasic aortic pressure, mean right atrial pressure, left ventricular pressure, and mean and phasic mesenteric, renal, iliac and coronary blood flows were monitored and recorded together with heart rate and the maximum rate of change of left ventricular systolic pressure (dP/dtmax), by using an electrostatic strip-chart recorder (Gould ES 2000). The heart rate was obtained from the electrocardiogram with a ratemeter (ECG/Biotach amplifier, model 13-4615-65 A; Gould). The frequency response of the differentiator used to obtain left ventricular dP/dtmax was found to be flat (± 5 %) up to 150 Hz.
At the end of the experiment, each animal was killed by an intravenous injection of 90 mg kg-1 sodium pentobarbitone.
Experimental protocol
The experiments were begun after at least 30 min of steady-state conditions with respect to measured haemodynamic parameters. In the 39 pigs, to avoid the interference of any possible changes in heart rate and arterial blood pressure during the experiments, the heart was paced to a frequency higher, by 19·6 ± 4 beats min-1 (range, 10-28), than that observed during the steady state and the arterial system connected to the pressurized reservoir. After at least 10 min of steady-state conditions, the experiments were carried out by intravenously infusing 2 µg of 17-oestradiol (Sigma) dissolved in saline or saline only, in a random order. The infusions were completed in a period of 1 h by using an infusion pump (Model 22; Harvard Apparatus) working at constant rate of 1 ml min-1. After the infusion was stopped, observations were continued for 2 h. The dose of 17-oestradiol was chosen because it corresponded to the hourly dose absorbed by women during replacement therapy with daily patches containing 50 µg of the hormone.
Recordings taken for 10 min during steady state before the infusion of 17-oestradiol were used as control. Measurements of haemodynamic variables were obtained during the last 10 min of infusion in the steady state and compared with control values. The effects of the infusion of 2 µg 17-oestradiol on measured blood flows were studied in eight pigs. In a further four pigs, the effects of graded administration of the hormone were examined. In these four animals, three subsequent doses of 17-oestradiol of 1, 2 and 3 µg, respectively, were cumulatively infused. Each infusion was completed in 1 h and changes in blood flow caused by each dose were compared with control values obtained before starting the infusion. The role of muscarinic cholinoceptors and adrenoceptors in the observed responses of measured blood flow was investigated in five pigs with experiments carried out by infusing 2 µg h-1 of 17-oestradiol after blockade of the receptors with intravenous administration of atropine (0·5 mg kg-1; Sigma), propranolol (0·5 mg kg-1; Sigma) and phentolamine (1 mg kg-1; Ciba-Geigy, Varese, Italy). The blocking agents were injected together. The role of nitric oxide in the responses of mesenteric, renal, iliac and coronary blood flows to 17-oestradiol was examined in a total of 22 pigs by giving the hormone after the intra-arterial administration of L-NAME. In 20 of these pigs, 2 µg h-1 of the hormone was infused after injection of L-NAME into the renal artery (five pigs), the iliac artery (five pigs), the coronary artery (five pigs) and into all three arteries (five pigs). In the remaining two pigs, the blocking agent was injected into the mesenteric, renal, iliac and coronary arteries and a subsequent infusion of 3 µg of 17-oestradiol in 1 h was performed.
The intracoronary dose of L-NAME was 100 mg. In anaesthetized pigs, this dose has been shown to reduce significantly the vasodilator effect of the intracoronary administration of acetylcholine at a dose of 1 µg (Vacca et al. 1996c). In the present investigation, in five of the pigs which received intracoronary L-NAME, this dose of acetylcholine increased coronary blood flow at constant heart rate and arterial pressure from 49·5 ± 6·7 ml min-1 (range, 38·6-55·4) to 95 ± 9 ml min-1 (79·6-102·6). When the same dose was given after L-NAME, coronary blood flow changed from 49·6 ± 6·4 ml min-1 (39·2-55·3) to 69·5 ± 9 ml min-1 (54·6-78·3), with a reduction in the response of 52·7 ± 11·1 % (43·4-66·9; P < 0·0005). This reduction in the acetylcholine-induced increase in coronary blood flow was considered a reliable marker of the inhibition of the release of nitric oxide (Parent et al. 1992). The dose of L-NAME used in the other arteries was assessed by comparing the mean values of blood flow measured in these arteries with that of coronary blood flow. The doses for renal and iliac arteries were 545 ± 119 mg (380-790) and 170 ± 19 mg (140-190), respectively. In the two pigs in which L-NAME was injected into the mesenteric artery, 1·5 g of the blocking agent was given.
In each group of pigs, changes in measured variables caused by the infusion of 17-oestradiol were analysed statistically using Student's parametric t test for paired data. Statistical analysis of responses caused by the infusion of the hormone in different groups of pigs was performed by comparing absolute changes (calculated relative to the data during the control period) using the Mann-Whitney U test for unpaired data. Analysis of variance and the Student-Newman-Keuls test were used to examine the effect of cumulative doses of 17-oestradiol on measured blood flows. A value of P < 0·05 was considered statistically significant. Group data are presented as means ± S.D. (range).
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RESULTS |
In the 39 pigs, recordings commenced approximately 5 h after the induction of anaesthesia. The mean pH, PO2 and PCO2 of arterial blood were 7·39 (7·36-7·41), 118 mmHg (105-138) and 39 mmHg (38-43), respectively, and the haematocrit was 36 % (32-39).
Effects of infusion of 2 µg h-1 of 17-oestradiol
In the eight pigs, infusion of the vehicle (60 ml of saline) did not cause any changes in the baseline values of measured blood flow. Group values of data and individual changes in measured blood flow caused by infusion of 17-oestradiol are shown in Table 1 and Fig. 1, respectively. Infusion of 17-oestradiol did not cause a significant change in mesenteric blood flow, but it caused an increase in renal, iliac and coronary blood flow. Changes in left ventricular dP/dtmax, mean right atrial pressure and left ventricular end-diastolic pressure were not significant (Table 1). The increases in renal, iliac and coronary blood flow amounted to 19·7 ± 7·9 % (11·1-33·3), 21·2 ± 14·7 % (9·1-55·7) and 22·9 ± 6 % (15·8-32·2) of the control values, respectively. Examples of the above responses elicited by 17-oestradiol in one pig are shown in Fig. 2. The increases in renal, iliac and coronary blood flow began within about 15 min after the start of the infusion and reached a steady state in about 25 min. In the eight pigs, renal, iliac and coronary blood flow returned to control values within 1·5 h after the end of the infusion.
Table 1. Changes in haemodynamic variables caused by infusion of 2 µg h-1 of 17-oestradiol at constant heart rate and arterial blood pressure in eight pigs
| Data | Control | Test | Change |
| HR (beats min-1) | 132·6 ± 22·8 | 132·6 ± 22·8 | 0·01 ± 0·1 |
| (100-171) | (100-171) | (-0·2 to 0·3) |
| ABP (mmHg) | 95·5 ± 11 | 95·6 ± 11·3 | 0·1 ± 1 |
| (83-112) | (83-112) | (-1 to 2) |
| dP/dt (mmHg s-1) | 2240 ± 218 | 2231 ± 221 | -9 ± 18 |
| (1962-2612) | (1962-2590) | (-36 to 13) |
| RAP (mmHg) | 3·1 ± 0·8 | 3·1 ± 0·8 | 0·01 ± 0·1 |
| (2·2-4·3) | (2·3-4·1) | (-0·3 to 0·1) |
| LVEDP (mmHg) | 5·6 ± 1 | 5·6 ± 1 | 0·04 ± 0·1 |
| (4·5-7·2) | (4·5-7·3) | (-0·1 to 0·2) |
| MBF (ml min-1) | 891 ± 172 | 890 ± 173 | -1 ± 5 |
| (650-1115) | (645-1117) | (-5 to 11) |
| RBF (ml min-1) | 333 ± 64 | 398 ± 79 | 65 ± 28  |
| (250-438) | (307-518) | (34-112) |
| IBF (ml min-1) | 98 ± 10 | 118 ± 18 | 20 ± 14 * |
| (81-110) | (92-148) | (10-53) |
| CBF (ml min-1) | 51·8 ± 8·7 | 63·3 ± 8·1 | 11·5 ± 1·8 |
| (41·2-64·1) | (53·2-75·6) | (9·2-14·8) |
Data are means ± S.D. (range). HR, heart rate; ABP, mean aortic blood pressure; dP/dt, left ventricular dP/dtmax; RAP, mean right atrial pressure; LVEDP, left ventricular end-diastolic pressure; MBF, mean mesenteric blood flow; RBF, mean renal blood flow; IBF, mean iliac blood flow; CBF, mean coronary blood flow. *P < 0·0025, P < 0·0005 vs. control.
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Figure 1. Responses of mesenteric blood flow (MBF), renal blood flow (RBF), iliac blood flow (IBF) and coronary blood flow (CBF) to the intravenous infusion of 2 µg h-1 of 17-oestradiol in eight pigs
The values of blood flow obtained in each animal during the test period are plotted against the corresponding control values before infusion. The continuous lines are the lines of equality.
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Figure 2. Example of experimental recordings showing the effects of the intravenous infusion of 2 µg h-1 of 17-oestradiol on regional blood flows in an anaesthetized pig
A, control period before infusion; B, last 10 min of infusion. From top to bottom: heart rate (HR), phasic and mean aortic blood pressure (ABP), left ventricular pressure (LVP), mean right atrial pressure (RAP), left ventricular dP/dtmax (dP/dt), mean and phasic mesenteric blood flow (MBF), mean and phasic renal blood flow (RBF), mean and phasic iliac blood flow (IBF), mean and phasic coronary blood flow (CBF).
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Effects of graded administration
In the four pigs examined, control values of mesenteric, renal, iliac and coronary blood flow were 900 ± 113 ml min-1 (760-1025), 366 ± 47 ml min-1 (316-423), 97 ± 7 ml min-1 (88-104) and 49·6 ± 5·9 ml min-1 (42·8-56·5), respectively. Graded administration of 17-oestradiol (three subsequent infusions of 1, 2 and 3 µg of the hormone, each infusion completed in a period of 1 h) caused graded increases in renal, iliac and coronary blood flow in each of the four pigs examined (Fig. 3). Group increases caused by the three infusions of the hormone, respectively, were 15 ± 5 ml min-1 (13-20, P < 0·005), 49 ± 10 ml min-1 (39-62, P < 0·0025) and 94 ± 17 ml min-1 (76-110, P < 0·0025) for renal blood flow; 7 ± 2 ml min-1 (6-10, P < 0·005), 18 ± 5 ml min-1 (12-23, P < 0·005) and 25 ± 5 ml min-1 (19-32, P < 0·0025) for iliac blood flow; and 5·6 ± 1·9 ml min-1 (3·1-7·6, P < 0·005), 12·4 ± 1·7 ml min-1 (10·2- 14·1, P < 0·0005) and 17·8 ± 3·8 ml min-1 (13·8-21·4, P < 0·0025) for coronary blood flow. These increases, respectively, amounted to 4·6 ± 1·3 % (3·5-6·3), 13·4 ± 2·2 % (10·2-14·7) and 25·8 ± 4·7 % (21·4-32·4) of the control values for renal blood flow; 7·6 ± 2·8 % (5·3-11·4), 18·4 ± 6 % (11·5-26·1) and 26·5 ± 7·5 % (18·3-36·4) for iliac blood flow; and 10·5 ± 5·8 % (6·6-17·8), 25·3 ± 4 % (19·7-28·3) and 36 ± 7·2 % (26·6-43·9) for coronary blood flow. Analysis of variance for repeated measurements showed a significant difference in the responses of renal, iliac and coronary blood flow (F = 74·8, P < 0·0005; F = 73·1, P < 0·0005; F = 43·8, P < 0·0005; respectively). The Student-Newman-Keuls test indicated that the responses obtained with the last dose of 17-oestradiol were significantly greater than those caused by the second dose, which elicited responses that were significantly greater than the responses caused by the initial dose.
In these experiments, changes in mesenteric blood flow caused by the first two doses of 17-oestradiol were not significant, amounting to -2 ± 4 ml min-1 (-8 to 2, P > 0·15) and 1 ± 6 ml min-1 (-8 to 5, P > 0·35), respectively. However, with the highest dose of the hormone an increase in mean mesenteric blood flow of 100 ± 17 ml min-1 (80-120, P < 0·0025; Fig. 3) or 11·1 ± 1·7 % (10·1-13·7) of the control value was obtained. Analysis of variance showed a significant difference in the responses of mesenteric blood flow to cumulative doses of the hormone (F = 166·7, P < 0·0005). The Student- Newman-Keuls test indicated that the response obtained with the last dose of 17-oestradiol was significantly greater than that obtained with the second and the initial doses. Changes in left ventricular dP/dtmax, mean right atrial pressure and left ventricular end-diastolic pressure during these experiments were not significant (at least, P > 0·10).
Experiments after blockade of muscarinic cholinoceptors and adrenoceptors
In the five pigs examined, administration of atropine, propranolol and phentolamine together caused decreases in heart rate, mean aortic blood pressure and left ventricular dP/dtmax of 15·4 ± 10·1 beats min-1 (-23 to 2, P < 0·025), 20 ± 13 mmHg (5-38, P < 0·025) and 319 ± 145 mmHg s-1 (154-512, P < 0·005), respectively, from control values of 100·9 ± 8·6 beats min-1 (92-110), 100·6 ± 4·9 mmHg (95-108) and 2330 ± 265 mmHg s-1 (1995-2715). These changes were accompanied by decreases in mesenteric, renal, iliac and coronary blood flow which were 125 ± 97 ml min-1 (59-270, P < 0·025), 78 ± 57 ml min-1 (12-156, P < 0·025), 14 ± 9 ml min-1 (4-28, P < 0·025) and 8·5 ± 5·3 ml min-1 (4·5-17·6, P < 0·0125), respectively, from control values of 848 ± 161 ml min-1 (750-1050), 342 ± 42 ml min-1 (292-400), 102 ± 10 ml min-1 (89-114) and 44·1 ± 4·7 ml min-1 (38·9-50·2).
Blockade of muscarinic cholinoceptors and adrenoceptors did not affect the responses of measured blood flows to the infusion of 2 µg h-1 of 17-oestradiol. The increases in renal, iliac and coronary blood flow caused by infusion of the hormone in the five pigs at constant heart rate and arterial blood pressure were 42 ± 23 ml min-1 (21-73, P < 0·01), 20 ± 7 ml min-1 (14-30, P < 0·0025) and 10·4 ± 1·9 ml min-1 (8·1-12·9, P < 0·0005), respectively. These increases in renal, iliac and coronary blood flow were not significantly different from those obtained in the control group of eight pigs (Mann-Whitney U test, P > 0·05) and amounted to 16·2 ± 10·2 % (7·9-32·9), 21·9 ± 6·9 % (15·4-32·1) and 25·4 ± 6·3 % (18·6-32·9) of control values, respectively. In the same experiments, changes in mesenteric blood flow were not significant, amounting to -6 ± 16 ml min-1 (-33 to 8, P > 0·20).
Role of nitric oxide
In the 22 pigs, control values of mean aortic blood pressure, heart rate and left ventricular dP/dtmax were 95·4 ± 6·4 mmHg (88-109), 105·7 ± 9·9 beats min-1 (89-131) and 2350 ± 313 mmHg s-1 (1815-3012), respectively. Intra-arterial injection of L-NAME caused increases in these variables of 12·6 ± 4·9 mmHg (6-22, P < 0·0005), 5·8 ± 2·3 beats min-1 (4-11, P < 0·0005) and 93 ± 48 mmHg s-1 (P < 0·0005), respectively. Injection of the blocking agent into the renal artery (12 pigs, see Methods) caused a decrease in renal blood flow of 88 ± 47 ml min-1 (2-156, P < 0·0005) from a control value of 371 ± 68 ml min-1 (263-465). Injection of L-NAME into the iliac artery (12 pigs) caused a decrease in iliac blood flow of 4 ± 2·5 ml min-1 (0-8, P < 0·0005) from a control value of 96 ± 9 ml min-1 (82-110) and injection of the blocking agent into the coronary artery (12 pigs) caused a group decrease in coronary blood flow of 2·2 ± 3·4 ml min-1 (-8·1 to 4·9, P < 0·025) from a control value of 48 ± 5·3 ml min-1 (33·6-53·9). In the two pigs in which L-NAME was injected into all four vessels, control values of arterial blood pressure and mesenteric blood flow, respectively, were 91 and 108 mmHg, and 760 and 985 ml min-1. Injection of l-NAME increased arterial blood pressure by 11 and 21 mmHg and mesenteric blood flow by 24 and 10 ml min-1. Individual changes in measured blood flows caused by the infusion of 17-oestradiol in the 22 pigs are shown in Fig. 4.
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Figure 4. Responses of mesenteric blood flow, renal blood flow, iliac blood flow and coronary blood flow to the intravenous infusion of 17-oestradiol after the injection of L-NAME into regional circulations
The values obtained during the test period are plotted against the control values before infusion. The continuous lines are the lines of equality. Note that the responses were abolished in the regions in which L-NAME was injected.  , responses to infusion of 2 µg h-1 of 17-oestradiol after injection of L-NAME into the renal artery;  , responses to infusion of 2 µg h-1 of 17-oestradiol after injection of L-NAME into the iliac artery;  , responses to infusion of 2 µg h-1 of 17-oestradiol after injection of L-NAME into the coronary artery;  , responses to infusion of 2 µg h-1 of 17-oestradiol after injection of L-NAME into the renal, iliac and coronary arteries;  , responses to infusion of 3 µg h-1 of 17-oestradiol after injection of L-NAME into the mesenteric, renal, iliac and coronary arteries. Abbreviations as in Fig. 1.
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L-NAME into the renal artery. In the five pigs examined, control values of mesenteric, renal, iliac and coronary blood flow after the injection of L-NAME into the renal artery and before starting the infusion of 17-oestradiol were 888 ± 135 ml min-1 (718-1092), 299 ± 57 ml min-1 (210-365), 96 ± 8 ml min-1 (87-107) and 55·8 ± 6·3 ml min-1 (46·3-63·3), respectively. Injection of the blocking agent into the renal artery abolished the response of renal blood flow to the subsequent infusion of 2 µg h-1 of 17-oestradiol without affecting the responses of iliac and coronary blood flow (Fig. 4). Changes in these flows caused by the hormone were -1 ± 3 ml min-1 (-2 to 5, P > 0·20), 17 ± 5 ml min-1 (10-23, P < 0·0025) and 11·4 ± 2 ml min-1 (8-13·3, P < 0·0005), respectively. The increases in iliac and coronary blood flow obtained in these animals were not significantly different (Mann-Whitney U test, P > 0·05) from those obtained in the control group of eight pigs. In the same experiments, changes in mesenteric blood flow were not significant, amounting to -2 ± 7 ml min-1 (-8 to 9, P > 0·25; Fig. 4).
L-NAME into the iliac artery. In the five pigs examined, control values of mesenteric, renal, iliac and coronary blood flow after the injection of L-NAME into the iliac artery and before starting the infusion of 17-oestradiol were 877 ± 101 ml min-1 (743-1005), 352 ± 69 ml min-1 (260- 432), 89 ± 11 ml min-1 (78-103) and 55 ± 5·2 ml min-1 (49·3-62), respectively. Injection of the blocking agent into the iliac artery abolished the response of iliac blood flow to the subsequent infusion of 2 µg h-1 of 17-oestradiol without affecting the responses of renal and coronary blood flow (Fig. 4). Changes in these flows caused by the hormone were -1 ± 3 ml min-1 (-4 to 3, P > 0·20), 65 ± 16 ml min-1 (42-87, P < 0·0005) and 10·3 ± 2·9 ml min-1 (8·9-14·3, P < 0·0025), respectively. The increases in renal and coronary blood flow obtained in these animals were not significantly different (Mann-Whitney U test, P > 0·05) from those obtained in the control group of eight pigs. In the same experiments, changes in mesenteric blood flow were not significant, amounting to -3 ± 12 ml min-1 (-22 to 8, P > 0·30; Fig. 4).
L-NAME into the coronary artery. In the five pigs examined, control values of mesenteric, renal, iliac and coronary blood flow after the injection of L-NAME into the coronary artery and before starting the infusion of 17-oestradiol were 898 ± 144 ml min-1 (710-1110), 333 ± 64 ml min-1 (228-402), 97 ± 10 ml min-1 (85-111) and 51·2 ± 7·8 ml min-1 (37·8-58·3), respectively. Injection of the blocking agent into the coronary artery abolished the response of coronary blood flow to the subsequent infusion of 2 µg h-1 of 17-oestradiol without affecting the responses of renal and iliac blood flow (Fig. 4). Changes in these flows caused by the hormone were 0·02 ± 0·3 ml min-1 (-0·3 to 0·4, P > 0·40), 54 ± 17 ml min-1 (40-80, P < 0·0025) and 19 ± 12 ml min-1 (8-33, P < 0·0125), respectively. The increases in renal and iliac blood flow obtained in these animals were not significantly different (Mann-Whitney U test, P > 0·05) from those obtained in the control group of eight pigs. In the same experiments, changes in mesenteric blood flow were not significant, amounting to -1 ± 10 ml min-1 (-18 to 8, P > 0·40; Fig. 4).
L-NAME into the renal, iliac and coronary arteries. In the five pigs examined, control values of mesenteric, renal, iliac and coronary blood flow after the injection of L-NAME into the renal, iliac and coronary arteries and before starting the infusion of 17-oestradiol were 872 ± 170 ml min-1 (702-1112), 285 ± 44 ml min-1 (227-343), 90 ± 11 ml min-1 (76-102) and 52·7 ± 4 ml min-1 (46·4-56·3), respectively. Injection of the blocking agent into the renal, iliac and coronary arteries abolished the responses of renal, iliac and coronary blood flow to the subsequent infusion of 2 µg h-1 of 17-oestradiol (Fig. 4). Changes in these flows caused by the hormone were -1 ± 6 ml min-1 (-7 to 8, P > 0·35), -1 ± 3 ml min-1 (-5 to 2, P > 0·20) and 0·2 ± 1·3 ml min-1 (-1·2 to 1·8, P > 0·35), respectively. In the same experiments, changes in mesenteric blood flow were not significant, amounting to -7 ± 19 ml min-1 (-32 to 14, P > 0·20; Fig. 4).
Infusion of 3 µg h-1 of 17-oestradiol after injection of L-NAME into the four arteries. In the two pigs examined, control values of mesenteric, renal, iliac and coronary blood flow after the injection of L-NAME into the four arteries and before starting the infusion of 17-oestradiol were 784 and 995, 245 and 327, 76 and 101, and 48·7 and 56·1 ml min-1, respectively. Injection of the blocking agent into the four arteries abolished the responses of renal, iliac and coronary blood flow to the subsequent infusion of 3 µg h-1 of 17-oestradiol (Fig. 4). Changes in these flows caused by the hormone in the two pigs were -17 and -5, 2 and -3, and 0·8 and -1·2 ml min-1, respectively. In the same experiments, despite the infusion of 3 µg h-1 of the hormone, changes in mesenteric blood flow were not significant, amounting to 12 and -17 ml min-1 (Fig. 4).
|
DISCUSSION |
The present investigation showed that infusion of 17-oestradiol primarily caused a steady-state increase in blood flow in the renal, external iliac and coronary arteries without significantly changing left ventricular contractility. Increasing the amount of infused 17-oestradiol in steps was accompanied by stepwise increments in regional blood flows, with the higher doses of the hormone causing an increase in mesenteric blood flow. These responses indicated a regional vasodilator effect of 17-oestradiol, which was shown to be nitric oxide dependent.
The design of the investigation ensured that 17-oestradiol and not other factors caused the observed responses. Prevention of changes in heart rate and arterial blood pressure and absence of changes in filling pressures of the heart and left ventricular dP/dtmax excluded any secondary interference from reflex and local metabolic and physical effects. In addition, infusion of the vehicle at the same rate as that of 17-oestradiol did not reproduce any of the hormonal effects. Although a vasodilator effect on the coronary circulation has been reported previously (Collins et al. 1994; Eckstein et al. 1994; Gorodeski et al. 1995; Sudhir et al. 1995; Wellman et al. 1996; Lamping & Nuno, 1996; Thompson & Weiner, 1997), our finding that the same hormone dilated the renal and iliac regions is new. This finding, together with previously reported findings on the effects of 17-oestradiol on genito-urinary and limb vasculature (Magness et al. 1993; Volterrani et al. 1995; Riedel et al. 1995; Rosenfeld et al. 1996) indicated that this hormone causes widespread vasodilatation. In the present study, wide ranges of control values of measured blood flows were observed. This finding, which reflects differences between animals, was expected and is consistent with previously reported findings obtained in the same experimental preparation (Vacca et al. 1996b).
The present finding on the effect of 17-oestradiol on left ventricular dP/dtmax as an indicator of left ventricular contractility of the heart is new. This was obtained in the absence of changes in heart rate, arterial pressure and filling pressures of the heart, which are known to influence ventricular contractility (Furnival et al. 1970). It could be argued that 17-oestradiol caused a decrease in left ventricular contractility which was masked by the improvement in this variable brought about by the concomitant increase in coronary blood flow. However, this was unlikely since graded increases in the dose of the hormone caused increases in coronary blood flow which averaged 11, 25 and 36 % without any significant influence on left ventricular dP/dtmax. It is of interest that oestrogens have been previously suggested either to increase ventricular contractility in experimental animals or women (Rozenberg et al. 1994; Samaan & Crawford, 1995) or to decrease it (Raddino et al. 1986; Jiang et al. 1992; Eckstein et al. 1994; Sitzler et al. 1996). The reasons for the differences between these reports and our results are not known. In the present investigation we found no significant changes in left ventricular contractility that could be attributed to 17-oestradiol.
A further new finding was that this response of widespread vasodilatation was proportional to the infused dose of 17-oestradiol, and that 17-oestradiol increased mesenteric blood flow only at high infusion doses. This occurred at constant haemodynamic variables, indicating regional mesenteric vasodilatation. There was no evidence that the mesenteric vasodilatation was brought about by any factor other than the higher dose of 17-oestradiol. In these experiments, there were no changes in the filling pressures of the heart or aortic blood pressure, excluding any reflex effects from stimulation of receptor systems. We have previously shown in anaesthetized pigs that there is a tonic 2-adrenoceptor-induced mesenteric vasodilatation (Vacca et al. 1996b), which could be argued to attenuate the vasodilator effect of 17-oestradiol. The administration of propranolol or butoxamine caused a decrease in mesenteric blood flow (Vacca et al. 1996b). However, in the present study the interaction between 2-adrenoceptors and 17-oestradiol-mediated vasodilatation was not examined.
Blockade of muscarinic cholinoceptors and adrenoceptors did not affect the vasodilatation response to 17-oestradiol. The dose of atropine used in this study has been used previously in anaesthetized pigs to block coronary muscarinic cholinoceptors (e.g. Vacca et al. 1996a, c). The dose of propranolol used has also been used previously in anaesthetized pigs to block -adrenergic receptors in the coronary (Vacca et al. 1996a, c) and in the mesenteric, renal and iliac vascular beds (Vacca et al. 1996b). A dose of 1 mg kg-1 phentolamine was shown to abolish the reflex coronary vasoconstriction caused by distension of the gall bladder (Vacca et al. 1996a) and the reflex coronary, mesenteric, renal and iliac vasoconstriction caused by distension of the stomach (Vacca et al. 1996b). The present findings therefore excluded the involvement of muscarinic cholinoceptors and adrenoceptors in the responses of regional vasodilatation elicited by 17-oestradiol, or of any possible mechanism acting through efferent vagal and sympathetic effects.
The present investigations showed that the response of regional vasodilatation to 17-oestradiol was nitric oxide dependent. The responses were blocked by the local administration of L-NAME, which is known to inhibit the formation of nitric oxide (Henderson, 1991). The blocking effect of L-NAME was only local to the region examined, since this blockade was not accompanied by any changes in the responses of other regional blood flows to the infusion of 17-oestradiol. These findings are consistent with other reports implicating nitric oxide in the 17-oestradiol-induced vasodilatation in the uterine and the coronary circulation (Collins et al. 1994; Gorodeski et al. 1995; Rosenfeld et al. 1996; Wellman et al. 1996; Thompson & Weiner, 1997), but are new regarding the vasodilator response in the renal and the iliac circulation. Furthermore, the vasodilator effect of the high dose of 17-oestradiol on the mesenteric circulation was shown to be nitric oxide dependent. The dilator influence of 17-oestradiol on regional vascular beds obtained in the present study in anaesthetized pigs began within about 15 min after starting the infusion and reached its full effect in about 25 min, being slower than the action of other known dilators. This delay could be attributed to the low dose of 17-oestradiol used, since larger doses caused the dilator effect to start earlier.
These new findings support previous reports which proposed that vasodilatation is one of the beneficial effects on the cardiovascular system attributed to oestrogens. The present investigation examined the effects of acute administration of 17-oestradiol, and these may be different from those seen during chronic administration. However, the effect of acute administration can be more readily related solely to 17-oestradiol, thus avoiding interfering and compensatory events.
In conclusion, the present investigation has shown that acute administration of 17-oestradiol causes regional vasodilatation in the mesenteric, renal, iliac and coronary circulation, though the dose required to increase mesenteric blood flow was relatively higher. The mechanism of this effect was nitric oxide dependent.
|
REFERENCES |
| Cheng, D. Y., Feng, C. J., Kadowitz, P. J. & Gruetter, C. A. (1994). Effects of 17-estradiol on endothelium-dependent relaxation induced by acetylcholine in female rat aorta. Life Sciences 55, 187-191. |
|
| Collins, P., Shay, J., Jiang, C. & Moss, J. (1994). Nitric oxide accounts for dose-dependent estrogen-mediated coronary relaxation after acute estrogen withdrawal. Circulation 90, 1964-1968 |
[Abstract] |
| Eckstein, N., Nadler, E., Barnea, O., Shavit, G. & Ayalon, D. (1994). Acute effects of 17-estradiol on the rat heart. American Journal of Obstetrics and Gynecology 171, 844-848 |
[Medline] |
| Furnival, C. M., Linden, R. J. & Snow, H. M. (1970). Inotropic changes in the left ventricle: the effect of changes in heart rate, aortic pressure and end-diastolic pressure. The Journal of Physiology 211, 359-387 |
[Medline] |
| Gorodeski, G. I., Yang, T., Levy, M. N., Goldfarb, J. & Utian, W. H. (1995). Effects of estrogen in vivo on coronary vascular resistance in perfused rabbit hearts. American Journal of Physiology 269, R1333-1338 |
[Medline] |
| Henderson, A. H. (1991). Endothelium in control. British Heart Journal 65, 116-125. |
[Medline] |
| Houpt, T. R. (1986). The handling of swine in research. In Swine in Cardiovascular Research, vol. II, ed. Stanton, H. C. & Mersmann, H. J., pp. 49-71. CRC Press, Boca Raton, FL, USA. |
|
| Jiang, C., Poole-Wilson, P. A., Sarrel, P. M., Mochizuki, S., Collins, P. & MacLeod, K. T. (1992). Effect of 17-oestradiol on contraction, Ca2+ current and intracellular Ca2+ in guinea-pig isolated cardiac myocytes. British Journal of Pharmacology 119, 43-48. |
|
| Kapitola, J., Andrle, J. & Kubickova, J. (1994). Relationship between local circulatory changes in the tibia and kidney of male and female rats after castration and administration of estradiol or testosterone. Endocrine Regulations 28, 41-46. |
[Medline] |
| Lamping, K. G. & Nuno, D. W. (1996). Effects of 17-estradiol on coronary microvascular responses to endothelin-1. American Journal of Physiology 271, H1117-1124 |
[Medline] |
| Linden, R. J. & Mary, D. A. S. G. (1983). The preparation and maintenance of anaesthetized animals for the study of cardiovascular function. In Techniques in the Life Sciences , Cardiovascular Physiology, ed. Linden, R. J., pp. 1-22. Elsevier Science Publishers, Ireland. |
|
| Magness, R. R., Parker, C. R. & Rosenfeld, C. R. (1993). Systemic and uterine responses to chronic infusion of estradiol-17. American Journal of Physiology 265, E690-698 |
[Medline] |
| Nuwayhid, B., Brinkman, C. R., Woods, J. R. Jr, Martinek, H. & Assali, N. S. (1975). Effects of estrogens on systemic and regional circulations in normal and renal hypertensive sheep. American Journal of Obstetrics and Gynecology 123, 495-504 |
[Medline] |
| Oelkers, W. K. (1996). Effects of estrogens and progestogens on the renin-aldosterone system and blood pressure. Steroids 61, 166-171. |
[Medline] |
| Paredes-Carbajal, M. C., Juarez-Oropeza, M. A., Ortiz-Mendoza, C. M. & Mascher, D. (1995). Effects of acute and chronic estrogenic treatment on vasomotor responses of aortic rings from ovariectomized rats. Life Sciences 57, 473-486 |
[Medline] |
| Parent, R., Paré, R. & Lavallée, M. (1992). Contribution of nitric oxide to dilation of resistance coronary vessels in conscious dogs. American Journal of Physiology 262, H10-16 |
[Medline] |
| Raddino, R., Manca, C., Poli, E., Bolognesi, R. & Visioli, O. (1986). Effects of 17-estradiol on the isolated rabbit heart. Archives Internationales de Pharmacodynamie et de Therapie 281, 57-65 |
[Medline] |
| Riedel, M., Oeltermann, A., Mugge, A., Creutzig, A., Rafflenbeul, W. & Lichtlen, P. (1995). Vascular responses to 17-oestradiol in postmenopausal women. European Journal of Clinical Investigation 25, 44-47 |
[Medline] |
| Rosenfeld, C. R., Cox, B. E., Roy, T. & Magness, R. R. (1996). Nitric oxide contributes to estrogen-induced vasodilation of the ovine uterine circulation. Journal of Clinical Investigation 98, 2158-2166 |
[Abstract/Full Text] |
| Rozenberg, S., Liebens, I., Vandromme, J., Hotimsky, A. & Van Rijsselberge, M. (1994). Cardiovascular protection by estrogen: a hemodynamic mechanism? International Journal of Fertility and Menopausal Studies 39, 36-42. |
[Medline] |
| Samaan, S. A. & Crawford, M. H. (1995). Estrogen and cardiovascular function after menopause. Journal of the American College of Cardiology 26, 1403-1410 |
[Medline] |
| Sitzler, G., Lenz, O., Kilter, H., La Rosee, K. & Böhm, M. (1996). Investigation of the negative inotropic effects of 17-oestradiol in human isolated myocardial tissues. British Journal of Pharmacology 119, 43-48 |
[Medline] |
| Sudhir, K., Chou, T. M., Mullen, W. L., Hausmann, D., Collins, P., Yock, P. G. & Chatterjee, K. (1995). Mechanisms of estrogen-induced vasodilation: in vivo studies in canine coronary conductance and resistance arteries. Journal of the American College of Cardiology 26, 807-814 |
[Medline] |
| Thompson, L. P. & Weiner, C. P. (1997). Long-term estradiol replacement decreases contractility of guinea pig coronary arteries to the thromboxane mimetic U46619. Circulation 95, 709-714 |
[Abstract/Full Text] |
| Vacca, G., Battaglia, A., Grossini, E., Mary, D. A. S. G. & Molinari, C. (1996a). Reflex coronary vasoconstriction caused by gallbladder distension in anesthetized pigs. Circulation 94, 2201-2209 |
[Abstract/Full Text] |
| Vacca, G., Mary, D. A. S. G., Battaglia, A., Grossini, E. & Molinari, C. (1996b). The effect of distension of the stomach on peripheral blood flow in anaesthetized pigs. Experimental Physiology 81, 385-396 |
[Medline] |
| Vacca, G., Papillo, B., Battaglia, A., Grossini, E., Mary, D. A. S. G. & Pelosi, G. (1996c). The effects of hypertonic saline solution on coronary blood flow in anaesthetized pigs. The Journal of Physiology 491, 843-851 |
[Abstract] |
| Volterrani, M., Rosano, G., Coats, A., Beale, C. & Collins, P. (1995). Estrogen acutely increases peripheral blood flow in postmenopausal women. American Journal of Medicine 99, 112-122. |
|
| Wellman, G. C., Bonev, A. D., Nelson, M. T. & Brayden, J. E. (1996). Gender differences in coronary artery diameter involve estrogen, nitric oxide, and Ca2+-dependent K+ channels. Circulation Research 79, 1024-1030 |
[Abstract/Full Text] |
Acknowledgements
This research has received generous sponsorship from the University of Turin and Scuole Officine Serali di Torino.
Corresponding author
G. Vacca: Facoltà di Medicina e Chirurgia, Via Solaroli 17, I-28100 Novara, Italy.
Email: Vacca{at}scimed1.med.no.unipmn.it
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Top
Introduction
), 2 µg h-1 (
) and 3 µg h-1 (
) in four pigs (numbered 1 to 4). Each column represents the absolute change from the baseline. Abbreviations as in Fig. 1.
