Endothelium-independent relaxation by 17-alpha-estradiol of pig coronary arteries.

We have studied the effects of 17-alpha-estradiol, a non-estrogenic steroid, on pig coronary arteries contracted by K+, Ca2+ or serotonin. Experiments were performed on helicoidal strips and rings of left circumflex coronary arteries from freshly slaughtered white pigs and on helicoidal strips of rat thoracic aorta. The strips and rings were suspended inside a water-jacketed muscle chamber in an oxygenated Krebs solution at 37 degrees C. From the initial K(+)-evoked contraction, 17-alpha-estradiol caused a relaxation with an IC50 (15 microM) which was in the range of the IC50s obtained for nitroglycerin (1.3 microM) and nicorandil (50 microM). Contractions evoked by Ca2+ were inhibited by 17-alpha-estradiol, but full blockade could not be achieved. Serotonin-evoked contractions were also blocked by 17-alpha-estradiol with an IC50 of 2.1 microM; 17-beta-estradiol also inhibited the serotonin-evoked contractions. In the presence of 100 microM of the nitric oxide synthase inhibitor N-nitro-L-arginine methyl ester, the relaxing properties of 17-alpha-estradiol in pig coronary arteries and rat thoracic aorta were unaffected, suggesting that endothelial NO release was unrelated to these effects. 17-alpha-Estradiol also relaxed denuded pig coronary artery strips, suggesting that other endothelial-derived relaxing factors were not involved in its vascular effects. The results are compatible with the idea that 17-alpha-estradiol causes relaxation of coronary vessels by acting directly on the cell membrane of smooth muscle cells; these effects seem to be unrelated to the genomic physiological effects of estrogens.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of plasma catecholamines in eliciting cardiovascular changes seen during naloxone-precipitated withdrawal in conscious, unrestrained morphine-dependent rats.

Rats given morphine (8 mg/kg i.v.) followed after 2 hr by infusions of morphine (4 mg/kg i.v.) every 2 hr for 24 hr (total infusion time of 2 min for each infusion) became dependent on morphine. Injection of the opiate antagonist naloxone (5 mg/kg) precipitated a withdrawal response including an increase in mean arterial blood pressure (BP), biphasic heart rate response and an increase in plasma norepinephrine (NE) and Epinephrine (Epi). Plasma Epi was also higher after abrupt withdrawal from morphine. After removal of adrenal glands from morphine-dependent rats, naloxone injection produced no change in the BP or plasma Epi. However, naloxone injection to morphine-dependent rats treated with phentolamine to block the alpha receptor-mediated effects of circulating catecholamines led to a significant decrease in BP even though plasma Epi increased 8-fold. In morphine-dependent rats in which NE levels in sympathetic nerves have been reduced by prior exposure to 6-hydroxydopamine, naloxone produced a biphasic BP response, an initial decrease followed by an increase along with a 3-fold increase in plasma Epi. These results suggest that Epi released from the adrenal medulla of morphine-dependent rats mediates, in large part, the autonomic withdrawal responses elicited by naloxone. Naloxone injection to control and morphine-dependent rats produced similar increases in plasma NE (2-fold) indicating that the increase in plasma NE is not responsible for the withdrawal response.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of etorphine on nicotine- and muscarine-induced catecholamine secretion from perfused rat adrenal glands.

The effect of etorphine on nicotine and muscarine-mediated catecholamine (CA) release from isolated perfused rat adrenal glands was investigated. Nicotine increased CA secretion at the low concentration of 0.5 micrograms while higher concentrations of muscarine (5 micrograms) were required. Moreover, muscarine released primarily epinephrine (EP) from rat adrenal glands while nicotine released norepinephrine (NE) and Ep. Etorphine inhibited NE and EP release evoked by nicotine to the same extent, whereas, muscarine-mediated release of NE and EP was not affected. Mecamylamine and verapamil inhibited nicotine but not muscarine-induced CA secretion. Our results suggest that etorphine preferentially interacts with nicotinic receptors on rat adrenal chromaffin cell membranes.

Effect of naloxone on blood pressure responses and plasma catecholamine levels following clonidine injection in conscious, unrestrained rats.

Intravenous injection of clonidine produces a biphasic blood pressure response, a transient increase followed by a prolonged decrease. Significant increases in plasma norepinephrine (NE), epinephrine (EPI), and dopamine (DA) levels occur 5 min following injection, corresponding to the hypertensive phase. Thirty minutes after clonidine injection, blood pressure, NE, EPI, and DA levels had returned to control levels. However, at 60 min, when the blood pressure had decreased to below control levels, a significant decrease in plasma NE was found. This finding confirms that clonidine decreases blood pressure, in part, by reducing sympathetic nerve activity. Naloxone, an opiate antagonist, when given prior to clonidine, abolished the hypotensive phase as well as preventing the decrease in plasma NE levels. The hypertensive phase was potentiated. Naloxone, when given alone, increased plasma NE levels at 15, 40, and 70 min following injection, indicating that naloxone increases sympathetic nerve activity peripherally, and may prevent clonidine's hypotensive phase by also increasing sympathetic activity in the central nervous system through blockade of opiate receptors. Therefore, the hypotensive effect of clonidine may involve interactions with endogenous opiate peptides.

Modulation of adrenergic neurotransmission in the rat tail artery by dietary lipids.

The effects of dietary lipids on prejunctional alpha 2-adrenoceptor function were investigated in perfused/superfused caudal arteries from adult rats. The investigation was designed to study the effects of diet supplemented with saturated fatty acids (coconut oil diet) or unsaturated fatty acids (sunflower oil diet) on alpha 2-adrenoceptor neuronal function in the proximal rat tail artery. Pregnant rats were fed Purina Rodent Chow (reference diet) or a semisynthetic diet containing 16% (wt/wt) of either sunflower oil or coconut oil. Neonatal pups were exposed to the diet via maternal milk and weaned rats were maintained on the same diet throughout adulthood. Artery segments (5-6 cm) were prelabeled with [3H]norepinephrine and perfused/superfused with Krebs-bicarbonate solution at 37 degrees C. The release of endogenous norepinephrine, total 3H, and [3H]norepinephrine was measured during field stimulation (supramaximal voltage, 5 Hz, 1 ms duration, for a total of 1,200 pulses). Both test diets caused a significant increase in norepinephrine content when compared with the reference diet (p less than 0.01). The results show that field stimulation-evoked release of norepinephrine from arteries obtained from rats fed coconut oil diet was significantly less than that exhibited by either the sunflower oil group or the reference group (p less than 0.05). Phentolamine (3 microM) caused a significant increase in percent release of endogenous norepinephrine and total 3H (p less than 0.05) in all groups. However, the increase above control values for the sunflower oil group was higher than the coconut oil group which suggested that dietary manipulations altered alpha2-adrenoceptor sensitivity.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of etorphine on the secretion of endogenous catecholamines and total tritium evoked by nerve- and acetylcholine-stimulation in perfused rat adrenal glands.

Isolated perfused rat adrenal glands were prelabeled with 3H-norepinephrine and catecholamine secretion was evoked by nerve stimulation (10 Hz, supramaximal voltage for 30 seconds) or acetylcholine (ACh)(5.4 micrograms) injection. Nerve stimulation evoked significant increases in tritium (16371 +/- 2109 cpm) and catecholamine (11.5 +/- 1.0 ng norepinephrine [NE], 123.1 +/- 13.0 ng epinephrine [EP]) release from the adrenal medulla. ACh injection evoked catecholamine release, but failed to increase tritium release. In the presence of etorphine, the nerve stimulation-mediated release of tritium, NE and EP was inhibited. In contrast, the ACh-mediated release of NE but not EP was inhibited by etorphine. In a previous publication (1), we have shown that 3H-NE is taken up by sympathetic nerve endings contained in extra adrenal tissue removed along with the adrenal gland during the surgery, but not by chromaffin cells. Therefore, the inhibitory effect of etorphine on NE, EP and tritium release evoked by nerve stimulation suggests a functional role for opiate receptors on transmitter release from sympathetic and splanchnic nerve endings. However, the differential effect of etorphine on NE and EP release evoked by ACh injection indicates that opiate receptors on chromaffin cells modulates NE but not EP release.

Effect of etorphine on adrenergic neurotransmission in the rat and guinea pig heart.

The effect of etorphine (ET) on nerve stimulation-mediated release of norepinephrine (NE) was investigated in isolated rat and guinea pig hearts. Hearts were perfused with Krebs bicarbonate solution via the aorta and the overflow of NE was measured after stimulation of the heart. ET (0.001-0.1 mumol/l) caused a dose-dependent inhibition of NE release in both preparations. Inhibition of NE release from guinea pig hearts ranged from 13% at 0.001 mumol/l to 24% at 0.1 mumol/l. The same concentrations of ET decreased NE release by 10 and 36% in the rat heart. The inhibitory effect of ET was blocked by naloxone. It is concluded that presynaptic opioid receptors located on the adrenergic neuronal terminals may be involved in the regulation of adrenergic neurotransmission in the rat and guinea pig heart.

Effect of dietary lipids on myocardial norepinephrine content and field stimulation-mediated release of norepinephrine from perfused neonatal and adult rat hearts.

The effects of dietary lipids on the content and release of norepinephrine and on the overflow of norepinephrine after alpha-adrenoceptor blockade with phentolamine were investigated in isolated perfused rat hearts. Pregnant rats were fed Purina Rodent Chow (reference diet) or a semisynthetic diet containing 16% (wt/wt) of either coconut oil (saturated fatty acids) or sunflower oil (unsaturated fatty acids). Neonatal pups were exposed to the diet via maternal milk and weaned rats were maintained on the same dietary lipid supplementation. Coconut oil caused a significant decrease in cardiac norepinephrine in all age groups when compared with the reference diet (p less than 0.01). Sunflower oil caused a significant increase in cardiac norepinephrine at 14 and 21 days of age when compared with coconut oil (p less than 0.05). Hearts prelabeled with [3H]norepinephrine were stimulated with supramaximal voltage (5 Hz, 2 ms duration, 300 pulses). At 14 and 21 days, coconut oil caused a significant decrease in norepinephrine release when compared with sunflower oil (p less than 0.05). The release of norepinephrine from hearts exposed to sunflower oil diet and the reference diet were comparable. These alterations in neuronal storage and exocytotic release of norepinephrine may be due to dietary-induced membrane perturbations. Phentolamine (10(-8)-10(-6) M) caused a dose-related increase in norepinephrine release following stimulation (supramaximal voltage 2.5 Hz, 150 pulses) of adult rat hearts from all dietary groups. However, the increase above control values was highest for coconut oil and lowest for sunflower oil (p less than 0.01), suggesting changes in receptor sensitivity. It appears that dietary lipid supplementation in the developing and adult rat could affect the myocardial alpha-adrenoceptor microenvironment which could cause changes in the prejunctional alpha-adrenoceptor neuronal function.

Simultaneous secretion of catecholamines from the adrenal medulla and of [3H]norepinephrine from sympathetic nerves from a single test preparation: different effects of agents on the secretion.

The uptake and release of catecholamines was investigated in the isolated perfused adrenal gland of the rat after preloading the preparation with [3H]norepinephrine, and the effects of various agents were examined on the stimulation-evoked secretion of catecholamines and total tritium. Large quantities of tritium were found in the adrenal medulla after either intravenous injection of [3H]norepinephrine to the rat, or perfusion of the isolated adrenal gland with Krebs-bicarbonate solution containing [3H]norepinephrine. The retention of the tritium was inhibited 90% by desipramine. Acute treatment with guanethidine and chronic treatment with 6-hydroxydopamine abolished the secretion of tritium without affecting the secretion of catecholamines evoked at 1 Hz. Nicotine, muscarine and acetylcholine enhanced the secretion of catecholamines but not tritium, whereas tyramine and ephedrine enhanced the secretion of tritium but not catecholamines. It is concluded that chromaffin cells do not possess the norepinephrine uptake mechanism and that the uptake of [3H]norepinephrine occurs mainly in sympathetic nerve terminals present in the adrenal gland and the surrounding blood vessels (adrenal and renal veins). The differential localization of [3H]norepinephrine and catecholamines allowed us to test the effects of a variety of pharmacological agents that alter neurotransmitter release by acting on receptors on the neuronal membrane, acting on sodium and potassium channels, or acting to alter the intracellular concentrations of adenosine 3',5'-cyclic monophosphate and protein kinase C. Transmural stimulation (1 Hz for a total of 300 pulses) markedly enhanced the release of catecholamines and tritium which was blocked by tetrodotoxin (sodium channel-blocker) and potentiated by tetraethylammonium and gallamine (potassium channel-blockers). Phentolamine, an alpha adrenergic blocking agent which acts on both alpha-1 and alpha-2 receptors, caused a 3- to 4-fold facilitation of the tritium secretion while inhibiting catecholamine secretion by 45%. [Met]enkephalin almost completely inhibited the evoked-secretion of tritium but had very little effect on the secretion of catecholamines. Forskolin inhibited the tritium secretion by 80% but produced more than a 2-fold facilitation of catecholamine secretion. Phorbol 12,13-dibutyrate caused facilitation of evoked secretion of both catecholamines and tritium. A combination of phorbol ester and forskolin had a synergistic effect on stimulation-evoked secretion of catecholamines, whereas phorbol ester partially reversed the inhibitory effects of forskolin on the tritium secretion.(ABSTRACT TRUNCATED AT 400 WORDS)

Effect of continuous intraventricular estrogen or catechol estrogen treatment on catecholamine turnover in various brain regions.

The effect of 7-day i.v.t. administration of catechol estrogens (CE) or estrogens (5 micrograms/day) on the catecholamine turnover rate of various brain areas was examined in ovariectomized rats. Norepinephrine turnover was increased significantly in the hypothalamus and cerebral cortex by estradiol treatment but not by any CEs tested when compared to control values. However, the turnover rate of dopamine in the cerebral cortex was increased compared to control values only by the 2-hydroxyestrogens (2-hydroxyestradiol and 2-hydroxyestrone) and estradiol was without effect. Only estrogens and CEs with physiologically significant estrogen receptor binding affinities (17 beta-estradiol, moxestrol, 2-hydroxyestradiol and 4-hydroxyestradiol) decreased the turnover rate of dopamine in the corpus striatum compared to control values. Estrogens (17 alpha-estradiol and 2-hydroxyestrone) which are weak ligands for the estrogen receptor did not affect striatal dopamine turnover. In addition, body weight gain measured during estrogen treatment was reduced by CEs and estrogens which have significant estrogen receptor affinities. These results suggest that the CEs may play a role in central modulation of catecholaminergic function by estrogens either through direct actions of the catechol moiety or activation of estrogen receptors.

Effects of ouabain on the nerve stimulation-evoked release of norepinephrine from the isolated perfused guinea-pig heart.

The isolated perfused guinea-pig heart prelabeled with (-)-[7-3H]-norepinephrine was used to examine the effects of increasing concentrations of ouabain on the sympathetic nerve stimulation-evoked release of endogenous norepinephrine and [3H]norepinephrine in the presence and absence of physostigmine or atropine. The overflow of norepinephrine and [3H]norepinephrine from guinea-pig hearts was measured during postganglionic stimulation of the cardiac accelerator fibers (5 Hz for 60 sec, 2 msec duration, for 300 pulses). Perfusion with 10(-7) M ouabain for 20 min had no effect on the release of norepinephrine or [3H]norepinephrine after nerve stimulation. However, perfusion with either 10(-6) or 3 X 10(-6) M ouabain for 15 min resulted in a significant decrease in the nerve stimulation-evoked release of norepinephrine (44.6 +/- 2.24 and 44.0 +/- 2.17%, respectively) and [3H]norepinephrine (43.8 +/- 1.62 and 44.9 +/- 2.16%, respectively) compared with previous control outputs. Perfusion of hearts with physostigmine (10(-6) M), an acetylcholinesterase inhibitor, or atropine (3 X 10(-6) M), a muscarinic blocking agent, did not alter the release of norepinephrine or [3H]norepinephrine after nerve stimulation. Perfusion with physostigmine during perfusion with 10(-6) M ouabain resulted in a decrease in the release of norepinephrine and [3H]norepinephrine only slightly greater than 10(-6) M ouabain alone, which was not significant, but the release of norepinephrine during stimulations performed after a 45-min washout of 10(-6) M ouabain was decreased significantly when 10(-6) M physostigmine was present. Perfusion of hearts with atropine during perfusion with either 10(-6) or 3 X 10(-6) M ouabain reversed the inhibitory effect of ouabain on the release of norepinephrine and [3H]norepinephrine.(ABSTRACT TRUNCATED AT 250 WORDS)

The effect of 17-alpha-estradiol, a possible endogenous opiate antagonist, on D-ala2-met5-enkephalinamide-induced blood pressure responses in conscious, unrestrained rats.

D-ala2-met5-enkephalinamide (DAME) produced a dose-related increase in the mean arterial blood pressure of conscious, unrestrained rats. Intravenous injection of DAME (0.5, 1, 2, and 4 mg/kg) resulted in mean systemic arterial blood pressures of 138 +/- 2, 146 +/- 5, 141 +/- 4, 156 +/- 5 mmHg, respectively. 17-alpha-estradiol and its derivatives are known to be inactive in target tissues responsive to estrogenic hormones such as 17-beta-estradiol. However, LaBella et al. (1978) found after testing a large number of steroid hormones and their metabolites that only 17-alpha-estradiol significantly inhibited binding of 3H-naloxone, an opiate antagonist, in rat-brain homogenates. The present study was designed to determine whether 17-alpha-estradiol could antagonize the cardiovascular responses elicited by intravenous injections of DAME. Intravenous infusion of 17-alpha-estradiol (1.5 mg/kg) every 2 hours for 24 hours (total infusion time was 2 minutes for each infusion) did not change the mean systemic arterial blood pressure (94 +/- 5 mmHg) compared to the blood pressure prior to infusion of 17-alpha-estradiol (99 +/- 7 mmHg). Intravenous infusion of 17-alpha-estradiol (1.5 mg/kg) 10 minutes prior to DAME (1 mg/kg, i.v.) resulted in a blood pressure of 106 +/- 9 mmHg, which is significantly less than the blood pressure of 146 +/- 5 mmHg seen with DAME (1 mg/kg, i.v.) alone. Intravenous injection of DAME (1 mg/kg) 8 hours after the last infusion of 17-alpha-estradiol produced an increase in mean systemic arterial blood pressure of 136 +/- 8 mmHg. These results indicate that 17-alpha-estradiol may function as an opiate antagonist.

Effect of intraventricular administration of catechol estrogens on catecholamine content in various brain regions.

Male rats treated for 7 days with either 2-hydroxyestradiol, 4-hydroxyestradiol or 2-hydroxyestrone had significantly lower striatal dihydroxy phenylacetic acid (DOPAC) levels when compared to the control group. After 14 days of treatment, groups which were treated with estradiol, 2-hydroxyestradiol or 4-hydroxyestradiol had significantly higher striatal dopamine levels and had gained significantly less weight when compared to the control group. These results indicate that estradiol and the catechol estrogens may act to reduce the activity of striatal dopaminergic neurons.