Demonstration of adrenergic catecholamine receptors in rat myometrium and their regulation by sex steroid hormones
Abstract
The molecular basis of sex steroid hormone-modulation of catecholamine-regulated smooth muscle cell contraction in the uterus was investigated at the level of the catecholamine receptor in rat myometrium. Myometrial membrane binding sites for 3H)-dihydroergocryptine bound α-but not β-adrenergic antagonists and stereospecifically bound the α-agonists (−)-norepinephrine > (−)-epinephrine > phenylephrine. Binding sites for (−) (3H)-dihydroalprenolol were specific for β-adrenergic antagonists and stereospecifically bound (−)-isoproterenol > epinephrine ⋍ norepinephrine. These results were consistent with the expected properties of the myometrial α- and β-adrenergic catecholamine receptors. Myometrial content of β- but not α-adrenergic catecholamine receptors was significantly elevated during proestrus and estrus, suggesting a role for sex steroid hormones in the regulation of these receptors. This posibility was substantiated in ovariectomized rats where castration resulted in a reduction in myometrial β-receptor content which was restored in a dose-dependent manner by estrodiol administration. We conclude: 1) rat uterus contains a substantial concentration of α- and β-adrenergic catecholamine receptors, 2) sex steroid hormones may modulated uterine contractility by regulation of these cell surface receptors; 3) modulation of cell responses to surface active hormones and agents by regulation of their cell surface receptors may be a major way in which sex steroids regulate target organ function.
References (27)
- S. Raz et al.
Am. J. Obstet. Gynecol.
(1971) - L.T. Williams et al.
J. Biol. Chem.
(1976) - C. Mukherjee et al.
J. Bio. Chem.
(1975) - L.T. Williams et al.
J. Biol. Chem.
(1976) - M.E. Maguire et al.
J. Biol. Chem.
(1976) - D.A. Greenberg et al.
Life Sci.
(1976) - J.N. Davis et al.
Brain Res.
(1977) - L.G. Nequin et al.
J. Ster. Biochem.
(1975) - R.D. Ahlquist
Am. J. Physiol.
(1948) - J.W. Miller
Ann. N.Y. Acad. Sci.
(1967)
Biol. Reproduc.
J. Physiol. (London)
Am. J. Physiol.
Cited by (62)
We investigated the role of estrogen and progesterone on rat uterine NmU receptor expression in both intact and ovariectomized animals and examined receptor expression through the estrous cycle. Chronic administration of β-estradiol 3-benzoate (E2) or progesterone in intact animals was devoid of any effect. RU486 caused a 2-fold up-regulation in NmU receptor density. Ovariectomy caused a 60% decrease in receptor density, but chronic E2 administration to ovariectomized rats significantly increased NmU receptor density. The estrous cycle had no significant effect on NmU receptor density. These results suggest that NmU receptor expression is estrogen-dependent, whereas progesterone or a progestin-induced factor is involved in the modulation of this expression.
Expression of β-adrenergic receptors in the rat uterus: Effects of puberty and oestrogen treatment during prepubertal development
1998, International Journal of Developmental NeuroscienceThe expression of β-adrenoceptors in the rat uterus has been analysed duringthe peripubertal transition and following acute and chronic oestradiol treatment duringprepubertal development. The distribution and density of β-adrenoceptors was assessedautoradiographically on cryostat tissue sections using [fn2H]-dihydroalprenolol ([3H]-DHA). Binding sites were localised in all the ages andexperimental situations examined and showed the following intensity of labelling : endometrialepithelium > longitudinal muscle layer > circular myometrial layer > endometrial stroma.Competition experiments with the selective antagonists ICI 118,551 and atenolol, showed thatmost of the β-adrenoceptors in the uterus belong to the β2 receptorsubclass. In prepubertal animals, the density of [3H]-DHA binding sites was extremelylow. Following puberty the density of binding sites showed a generalised increase. Acuteadmininstration of oestradiol at the end of the prepubertal period provoked an increase in thedensity of [3H]-DHA binding sites in all uterine regions, but the levels of labelling werelower than in peripubertal animals at proestrus and oestrus. Following chronic oestrogentreatment during postnatal development, oestradiol increased further the density of [3H]-DHA binding sites. Results are discussed considering both the endocrine and neuralchanges accompanying puberty and oestradiol treatment.
β-Adrenergic receptor subtype gene expression in timed-pregnant rat myometrium
1997, American Journal of Obstetrics and GynecologyOBJECTIVE: Classic radioligand binding techniques have suggested that β1- and β2-adrenergic receptor subtype proteins are expressed in myometrial tissue; however, to date these observations have not been confirmed at the level of the messenger ribonucleic acid for these clinically important membrane receptors. The studies described in this report sought to use quantitative reverse transcriptase–polymerase chain reaction techniques to confirm expression of messenger ribonucleic acid for the β1- and β2-adrenergic receptors in myometrial tissue and to determine whether messenger ribonucleic acid expression for these two adrenergic receptors is modulated during pregnancy.
STUDY DESIGN: For these studies total cellular ribonucleic acid was isolated from myometrial tissue obtained from timed-pregnant Sprague-Dawley rats by the guanidium thiocyanate–phenol–chloroform extraction technique; formaldehyde-agarose gels then confirmed isolation of intact ribonucleic acid. Random hexamer primers and reverse transcriptase were used to synthesize complementary deoxyribonucleic acid. Subsequently, polymerase chain reaction was performed with subtype specific 20-mer sense and antisense oligonucleotide primers specific for the rat β1- and β2-adrenergic receptors. Inclusion of internal standard deoxyribonucleic acid sequences allowed quantification of the reverse transcriptase–polymerase chain reaction results.
RESULTS: By use of total cellular ribonucleic acid isolated from myometrial tissue, reverse transcriptase–polymerase chain reaction generated the expected 328 bp product for the β1-receptor and the expected 559 bp product for the β2-receptor along with internal standard deoxyribonucleic acid sequences for both. The identity of the β1- and β2-adrenergic receptor polymerase chain reaction products was confirmed on the basis of restriction endonuclease digestions producing the expected deoxyribonucleic acid fragments and by Southern blots using β1- and β2-adrenergic receptor–specific complementary deoxyribonucleic acid probes. The reverse transcriptase–polymerase chain reaction studies confirmed a gradual decline in βl-receptor messenger ribonucleic acid and stable expression of β2-receptor messenger ribonucleic acid during the second half of gestation in pregnant rat myometrial tissue.
CONCLUSIONS: In summary, these studies have confirmed, at the messenger ribonucleic acid level, expression of the β1- and β2-adrenergic receptor subtypes in timed-pregnant rat myometrial tissue.(Am J Obstet Gynecol 1997;176:349-57.)
Potent inhibition of spontaneous rhythmic contraction by a novel β<inf>2</inf>-adrenoceptor agonist, HSR-81, in pregnant rat uterus
1996, European Journal of PharmacologyWe examined the effect of HSR-81 ((−)-(R)-α-[(tert-butylamino)methyl]-2-chloro-4-hydroxybenzyl alcohol l-tartrate), a newly developed, potent and selective β2-adrenoceptor agonist, as well as ritodrine and isoproterenol, on the spontaneous rhythmic contraction in uteri isolated from late pregnant, middle pregnant and non-pregnant (dioestrous and oestrous) rats. The three agonists inhibited the spontaneous rhythmic contraction at all the stages in a concentration-dependent manner. The pD2 value for HSR-81 was greater in late pregnancy than in dioestrus and oestrus. In the uterine preparations of late pregnancy and dioestrus, ICI-118,551 (1-(7-methylindan-4-yloxy)-3-isopropyl-aminobutan-2-ol, a selective β2-adrenoceptor antagonist) and atenolol (a selective β1-adrenoceptor antagonist) produced a parallel rightward shift of the concentration-response curves for HSR-81. The pKB values for ICI-118,551 and atenolol suggest that the inhibitory effect of HSR-81 was mediated through β2-adrenoceptors in the two stages. In the membranes prepared from rat uteri in late pregnancy and dioestrus, the equilibrium dissociation constant for [125I]iodocyanopindolol binding was not significantly different between the two stages. The three gb-adrenoceptor agonists and the two antagonists competed for the specific [125I]iodocyanopindolol binding and the pKi values were not significantly different between the two stages. However, the maximum number of binding sites was significantly greater in late pregnancy than in dioestrus. The configuration of the competition curves and the pKi values for the two antagonists confirmed the fact that these membranes contain predominantly β2-adrenoceptor subtype. These results indicate that the potent inhibition of the spontaneous rhythmic contraction by HSR-81 in the pregnant uterus may be due to the increased number of β2-adrenoceptors.
Identification of uterine-related sympathetic neurons in the rat inferior mesenteric ganglion: Neurotransmitter content and afferent input
1996, Journal of the Autonomic Nervous SystemThe rat uterus is innervated by sensory and autonomic nerves. Sensory and sympathetic fibers travel in the hypogastric nerves and are associated with the thoracolumbar spinal cord levels T13-L3. The inferior mesenteric ganglion (IMG) contains the somata of sympathetic postganglionic neurons and some of these may project axons to the uterus. Sensory and parasympathetic fibers travel in the pelvic nerve and are associated with the lumbosacral cord levels L6-S1 and pelvic ganglion (PG). We previously reported data concerning the neurochemical anatomy of the PG with regard to the uterine innervation; the present study was undertaken to characterize the neurochemical anatomy of the IMG with regard to its involvement in uterine innervation. A retrograde axonal tracer was used to verify projections of axons of IMG neurons to the uterus. Immunostaining of cryostat sections of the IMG revealed neurons immunoreactive for neuropeptide Y (NPY) and for tyrosine hydroxylase (TH). Immunostaining for the synaptic terminal protein synapsin I (SYN) revealed numerous fine terminals immediately surrounding the principal neurons and in the interneuronal spaces. Varicosities immunoreactive for calcitonin gene-related peptide (CGRP), vasoactive intestinal polypeptide (VIP), enkephalin (ENK), substance P (SP) and galanin (GAL) appeared to be associated with principal neurons. Additional varicosities stained for nicotinamide adenine dinucleotide phosphate (reduced)-diaphorase (NADPH-d) and nitric oxide synthase (NOS), thus indicating sites of neuronal nitric oxide synthesis. This study revealed that the IMG contains uterine-related neurons and that some of the retrogradely labeled uterine-related neurons contain NPY, TH or both NPY/TH. In addition, uterine-related neurons received abundant afferent inputs indicated by SYN-immunoreactive (-ir) terminals and some of these varicosities labeled for GAL, CGRP, VIP, ENK, or NADPH-d/NOS.
Effects of androgen on α- and β-adrenergic receptors in membranes from the rat seminal vesicle
1992, BBA - Molecular Cell ResearchThe effects of castration and androgen-replacement on adrenergic receptors in membranes from the rat seminal vesicle were studied. Membranes from seminal vesicles showed saturable and high-affinity binding sites for the β-adrenergic receptor antagonist, [3H]dihydroalprenolol ([3H]DHA), and the receptor antagonist, [3H]prazosin. Castration markedly reduced β-adrenergic receptors with decreasing the effect of GTP modulating the receptor-ligand affinity, suggesting defects in both the receptor per se and the guanine-nucleotides-regulating mechanism after castration. In contrast, castration increased receptors and androgen-replacement reversed this change. The effects of GTP decreasing the binding affinity to the radioligand were observed to a similar extent in the castrated and control membranes. These results demonstrate an inverse regulation by androgen on β- and receptors in membranes of the rat seminal vesicle.