Molecular analysis of beta-adrenergic receptor subtypes in rat collecting duct: effects on cell cAMP and Ca2+ levels

1995 ◽  
Vol 268 (6) ◽  
pp. F1070-F1080 ◽  
Author(s):  
B. Mandon ◽  
E. Siga ◽  
A. Champigneulle ◽  
M. Imbert-Teboul ◽  
J. M. Elalouf
Keyword(s):  
Rank Order ◽  
Collecting Duct ◽  
Receptor Subtypes ◽  
Thick Ascending Limb ◽  
Camp Accumulation ◽  
Beta Adrenergic ◽  
Beta Agonists ◽  
Fluorescence Measurements ◽  
Beta 2 ◽  
Collecting Tubules

Expression and functional properties of beta-adrenergic receptors (beta-ARs) were studied in rat collecting tubules isolated by microdissection. Reverse transcription-polymerase chain reaction experiments demonstrated that the beta 1- and beta 2-AR mRNAs, but not the beta 3-subtype, are expressed in the cortical collecting duct (CCD). Quantitation of mRNAs, carried out using mutant RNAs as internal standards, further showed that beta 1- and beta 2-ARs transcripts are present at comparable amounts in CCD (3,000–4,000 copies/mm of tubular length), but reach 6–8 times lower levels in the outer medullary collecting duct (OMCD: beta 1, 480 +/- 180; beta 2, 590 +/- 110 copies/mm of tubular length). Functional studies, carried out in CCD, corroborated the expression of these two receptor subtypes. The rank order of potency of beta-agonists for stimulating adenosine 3',5'-cyclic monophosphate (cAMP) accumulation was isoproterenol > norepinephrine = epinephrine, and similar efficiencies were found for a beta 1- and a beta 2-antagonist to inhibit isoproterenol-dependent cAMP formation. Fura 2 fluorescence measurements revealed that isoproterenol (10 microM) induces a biphasic rise of intracellular free Ca2+ concentration ([Ca2+]i), consisting of an initial fast increase (delta [Ca2+]i = 122 nM) followed by a plateau phase (delta [Ca2+]i = 58 nM). In the absence of basolateral Ca2+, the initial peak was still observed, suggesting intracellular Ca2+ release. Norepinephrine and epinephrine, as well as selective beta 1- and beta 2-agonists, also increased [Ca2+]i in CCD. Only slight [Ca2+]i variations were produced by isoproterenol in the OMCD (delta [Ca2+]i = 21 nM) and the cortical thick ascending limb (delta [Ca2+]i = 25 nM). These results show that both beta 1- and beta 2-ARs are expressed in the collecting tubule and that they predominate in the CCD. The two receptor subtypes contribute to cAMP accumulation induced by beta-agonists. They also trigger [Ca2+]i variations, indicating their possible coupling to several transduction pathways in the rat CCD.

scholarly journals Autoradiographic localization of beta-adrenoreceptors in rat uterus.

1988 ◽  
Vol 36 (12) ◽  
pp. 1475-1479 ◽  
Author(s):  
M Tolszczuk ◽  
G Pelletier
Keyword(s):  
Quantitative Estimation ◽  
Cell Types ◽  
Receptor Subtypes ◽  
High Concentration ◽  
Rat Uterus ◽  
Beta Adrenergic ◽  
Adrenergic Antagonist ◽  
Beta 2 ◽  
Different Cell Types

The inhibitory effects of catecholamines on uterine smooth muscle are known to be mediated through beta-adrenergic receptors. To investigate further the distribution of these receptors in the rat uterus, we utilized in vitro autoradiography using [125I]-cyanopindolol [CYP], a specific beta-receptor ligand that has equal activity for both beta 1- and beta 2-receptor subtypes. The specificity of the labeling and the characterization of receptor subtypes in different cell types were achieved by displacement of radioligand with increasing concentrations of zinterol, a beta-adrenergic agonist with preferential affinity for the beta 2-adrenoreceptor subtype, and practolol, a beta-adrenergic antagonist that binds preferentially to the beta 1-subtype. Quantitative estimation of ligand binding was performed by densitometry. It was shown that the vast majority of beta-adrenoreceptors were of the beta 2-subtype and were found in high concentration not only in the myometrium but also in the endometrial and serosal epithelia. Specific labeling was also observed in glandular elements. These results suggest that beta-adrenoreceptors might be involved in different functions in the uterus.

Regulation of atrial autonomic receptors in experimental cyanotic heart disease

1991 ◽  
Vol 261 (4) ◽  
pp. H1135-H1140 ◽  
Author(s):  
R. Doshi ◽  
E. Strandness ◽  
D. Bernstein
Keyword(s):  
Heart Rate ◽  
Adrenergic Receptors ◽  
Receptor Subtypes ◽  
Receptor Density ◽  
Beta Adrenergic Receptors ◽  
Beta Adrenergic ◽  
Beta 2 ◽  
The Right ◽  
Chronic Hypoxemia ◽  
Inotropic Responses

During chronic hypoxemia, left ventricular beta-adrenergic receptor density is decreased and a dissociation occurs between increased chronotropic and decreased inotropic responses to chronically elevated sympathetic tone. To determine whether this dissociation was related to alterations in autonomic receptor populations in the right atrium, we studied right atrial cholinergic and beta-adrenergic receptors in chronically hypoxemic newborn lambs and in normoxemic controls. Heart rate response was determined by infusing isoproterenol at 0.1 or 0.5 microgram.kg-1.min-1. Muscarinic receptors were quantified with [3H]quinuclidinyl benzilate and beta-adrenergic receptors with [125I]iodocyanopindolol. Competition with ICI 118,551 was used to determine beta 1- vs. beta 2-receptor subtypes. In the hypoxemic lambs, isoproterenol resulted in a lesser percentage increase in heart rate (hypoxemic, 46 +/- 6% vs. control, 89 +/- 17%, P less than 0.05); however, because baseline heart rate was higher in the hypoxemic lambs (213 +/- 7 vs. 177 +/- 12 beats/min, P less than 0.05), maximal heart rate responses were similar (310 +/- 7 vs. 326 +/- 6 beats/min, NS). There was no change in receptor density or affinity of either muscarinic or beta-adrenergic receptors and no change in the proportion of beta 1- vs. beta 2-receptor subtypes. Thus the dissociation between the chronotropic and inotropic responses to chronic hypoxemia may be in part secondary to a differential regulation of beta-adrenergic receptors between the left ventricle and the right atrium.

Characterization of beta-adrenergic receptors in cultured bovine tracheal gland cells

1989 ◽  
Vol 256 (2) ◽  
pp. C310-C314 ◽  
Author(s):  
J. M. Madison ◽  
C. B. Basbaum ◽  
J. K. Brown ◽  
W. E. Finkbeiner
Keyword(s):  
Binding Sites ◽  
Adrenergic Receptors ◽  
Adrenergic Receptor ◽  
Rank Order ◽  
Beta Adrenergic Receptors ◽  
Binding Studies ◽  
Beta Adrenergic ◽  
High Affinity ◽  
Gland Cells ◽  
Beta 2

We characterized the beta-adrenergic receptors that mediate secretory responses to isoproterenol in cultured bovine tracheal submucosal gland cells. Previous studies have shown that these cells have morphological and biochemical features characteristic of serous cells. Isoproterenol, epinephrine, and norepinephrine each stimulated the secretion of 35SO4-labeled macromolecules from these cultured serous cells with a rank order of potency (isoproterenol greater than epinephrine greater than norepinephrine) consistent with the presence of beta 2-adrenergic receptors. These functional studies were supported by radioligand-binding studies using [I125]-iodocyanopindolol (125I-CYP) to identify beta-adrenergic receptors. 125I-CYP binding to membrane particulates prepared from cultured serous cells was saturable and of high affinity (equilibrium dissociation constant 20 +/- 3 pM; mean +/- SE, n = 6) and was antagonized stereoselectively by propranolol. Adrenergic agonists competed for 125I-CYP-binding sites with a rank order of potency characteristic of the beta 2-adrenergic receptor subtype. A specific beta 2-adrenergic receptor antagonist, ICI 118.551, competed for a single class of 125I-CYP-binding sites with high affinity (inhibition constant 1.8 +/- 0.3 nM, n = 3). We concluded that the secretory response of cultured tracheal gland cells to isoproterenol is a response mediated by beta-adrenergic receptors of the beta 2 subtype.

IDENTIFICATION OF beta-LIKE ADRENOCEPTORS ASSOCIATED WITH BIOLUMINESCENCE IN THE SEA PANSY RENILLA KOELLIKERI

1993 ◽  
Vol 177 (1) ◽  
pp. 181-200 ◽  
Author(s):  
E. W. Awad ◽  
M. Anctil
Keyword(s):  
Radioligand Binding ◽  
Beta Blockers ◽  
Physiological Effect ◽  
The Body ◽  
Receptor Subtypes ◽  
Beta Adrenergic ◽  
Adrenergic Agents ◽  
Beta Agonists ◽  
Specificity Pattern ◽  
Renilla Koellikeri

Previous studies have reported pharmacological and biochemical evidence for the involvement of adrenergic substances in the regulation of neuroeffector activities in the bioluminescent cnidarian Renilla koellikeri (Cnidaria, Anthozoa). Therefore, direct radiobinding assays were developed to identify and characterize beta-adrenergic binding in membrane preparations from this species, using the two beta-antagonists [3H]dihydroalprenolol and [3H]CGP12177 as tracers. In addition, the effect of various beta-adrenergic agents on luminescence was examined. Binding of the radioligands at 25°C was rapid, reversible, saturable and specific. Saturation studies revealed the presence of two different and independent classes of binding site, site1 and site2, in the body of the colony (rachis). In contrast, homogeneous populations of binding sites corresponding to site1 were detected in autozooid polyps and to site2 in the peduncle. The pharmacological profile of beta-adrenergic binding in R. koellikeri membrane preparations displayed properties consistent with the presence of two sites and followed a pattern similar to beta2- and beta1-adrenergic receptor subtypes for site1 and site2, respectively. Bioluminescence in polyps was induced by beta-agonists as well as by one beta1-selective antagonist, atenolol, and was blocked by several beta-blockers including (+/−)CGP12177. The specificity pattern of the physiological effect of beta-adrenergic drugs on luminescence mirrors that of the radioligand interaction with site1. This suggests that radioligand binding to site1 represents binding to the receptor that mediates luminescence excitation in R. koellikeri. Blockade of the luminescent responses to site1 agonists by isotonic MgCl2 indicates that this beta-adrenergic mechanism must rely on interneuronal transmission. Collectively, these results suggest the evolutionary conservation of beta-adrenoceptors and of their dual character from coelenterates to higher vertebrates.

beta-Adrenergic receptors in the developing rabbit lung

1981 ◽  
Vol 240 (4) ◽  
pp. E351-E357 ◽  
Author(s):  
J. A. Whitsett ◽  
M. A. Manton ◽  
C. Darovec-Beckerman ◽  
K. G. Adams ◽  
J. J. Moore
Keyword(s):  
Adenylate Cyclase ◽  
Adrenergic Receptors ◽  
Adrenergic Receptor ◽  
Cyclase Activity ◽  
Adenylate Cyclase Activity ◽  
Receptor Subtypes ◽  
Beta Adrenergic Receptors ◽  
Rabbit Lung ◽  
Beta Adrenergic ◽  
Beta 2

beta-Adrenergic receptors and catecholamine-sensitive adenylate cyclase were identified and partially characterized in membrane fractions of rabbit lungs from day 25 of gestation to adulthood with the beta-adrenergic antagonists (--)-[3H]dihydroalprenolol [(--)-[3H]DHA] and (--)-[125I]iodohydroxybenzylpindolol [(--)-[125I]HYP]. beta-Adrenergic receptor number (Bmax) increased 11.5-fold during this time period, increasing progressively during the latter days of gestation and the early neonatal period, from 37 +/- 10 fmol/mg protein at 25 days gestation to 425 +/- 51 fmol/mg in the adult rabbit lung (mean +/- SD). Receptor affinity for (--)-[3H]DHA (KD = 1.8 nM) or (--)-[125I]HYP (KD - 0.104 nM) and the proportion of beta 1- and beta 2-adrenergic receptor subtypes (60% beta 1 and 40% beta 2) did not change with advancing age. Basal adenylate cyclase activity in lung homogenates decreased significantly with increasing age, whereas the activity in the presence of catecholamine or NaF remained nearly constant. Catecholamines stimulated adenylate cyclase activity at all ages studied supporting a role of the maturation of beta-adrenergic receptors in the regulation of pulmonary function.

Autoradiographic localization of beta-adrenergic receptors in rat kidney slices using [125I]iodocyanopindolol

1984 ◽  
Vol 246 (2) ◽  
pp. F240-F245
Author(s):  
P. A. Munzel ◽  
D. P. Healy ◽  
P. A. Insel
Keyword(s):  
Adrenergic Receptors ◽  
Rank Order ◽  
Collecting Duct ◽  
Rat Kidney ◽  
Distal Tubule ◽  
Cortical Collecting Duct ◽  
Beta Adrenergic Receptors ◽  
Beta Adrenergic ◽  
Kidney Slices ◽  
Adrenergic Antagonist

We have used autoradiography of [125I]iodocyanopindolol [( 125I]-ICYP) to define the distribution and localization of beta-adrenergic receptors in the rat kidney. [125I]ICYP, a radiolabeled beta-adrenergic antagonist, proved to be an excellent probe to identify these receptors in kidney slices in that specific [125I]ICYP binding was 1) saturable, 2) competed stereoselectively by the agonist isoproterenol and the antagonist propranolol, and 3) competed by agonists in a classical rank order of potency for beta 1-adrenergic receptors (isoproterenol greater than epinephrine congruent to norepinephrine). Autoradiographic studies demonstrated that specific [125I]ICYP sites were present exclusively in the cortex and outer medulla and were localized in glomeruli, ascending limb, and distal tubule-cortical collecting duct. Autoradiography of probes like [125I]ICYP appears to offer a simple and rapid method to assess distribution of adrenergic receptors in the kidney and presumably in other tissues as well.

Reserpine but not surgical denervation regulates rat renal beta-adrenergic receptors

1989 ◽  
Vol 256 (4) ◽  
pp. F532-F539
Author(s):  
T. L. Fortin ◽  
P. R. Sundaresan
Keyword(s):  
Adrenergic Receptors ◽  
Adrenergic Receptor ◽  
Kidney Cortex ◽  
Receptor Subtypes ◽  
Camp Accumulation ◽  
Beta Adrenergic Receptors ◽  
Beta Adrenergic ◽  
Beta Adrenoceptors ◽  
Beta Adrenoceptor ◽  
Reserpine Treatment

The effects of unilateral surgical denervation or reserpine administration on renal beta-adrenergic receptors were examined in rat kidney cortex. The specific binding of [125I]iodocyanopindolol was used to quantitate the beta-adrenoceptors. Denervation had no significant effect on beta-adrenoceptor concentration in denervated compared with contralateral control kidney, 7 days postsurgery. In contrast, reserpine treatment increased beta-adrenoceptor concentration 30% compared with control (P less than 0.05). Tissue norepinephrine levels were depleted to a significant extent with both manipulations. The reserpine effect was investigated further. Reserpine increased both beta 1- and beta 2-adrenergic receptor subtypes to the same extent. The effect of reserpine was primarily on tubular beta-adrenoceptors including those in the proximal tubules; glomerular beta-adrenoceptors were minimally affected by reserpine. Other adrenergic receptor subtypes (alpha 1- and alpha 2-) were also significantly increased by reserpine; however, angiotensin II receptors were not altered, indicating that the reserpine effect was not a general one affecting all membrane receptors. Reserpine treatment increased beta-adrenergic receptor-stimulated adenosine 3',5'-cyclic monophosphate (cAMP) accumulation by 49% over control in the renal cortex. Denervation had no significant effect on cAMP accumulation. Overall, our results suggest that, in addition to sympathetic nerve terminal norepinephrine, other factors may be involved in the regulation of renal beta-adrenergic receptors.

Diurnal and seasonal changes in sympathetic signal transduction in cardiac ventricles of European hamsters

1996 ◽  
Vol 270 (1) ◽  
pp. R304-R309 ◽  
Author(s):  
K. Pleschka ◽  
A. Heinrich ◽  
K. Witte ◽  
B. Lemmer
Keyword(s):  
Adenylyl Cyclase ◽  
Rank Order ◽  
Constant Darkness ◽  
Receptor Subtypes ◽  
Dark Phase ◽  
Light Cycle ◽  
Dark Light ◽  
Cardiac Ventricles ◽  
Beta 2 ◽  
The Relationship

This investigation of the relationship between cardiac beta-adrenoceptors and adenosine 3',5'-cyclic monophosphate (cAMP) formation in cardiac ventricles of the nocturnally active European hamster both during euthermia under a 12:12-h dark-light cycle and during hibernation under constant-darkness conditions showed that neither the densities, affinities, nor distribution of the beta 1- and beta 2-receptor subtypes differed between the dark phase, light phase, and hibernation. Basal formation of cAMP by the cardiac adenylyl cyclase of euthermic hamsters was higher in ventricles obtained at night [core temperature (Tcore) = 37.8 degrees C] than in ventricles obtained during the day (Tcore = 36.4 degrees C). Basal formation of cAMP was also significantly lower in hibernating hamsters (Tcore = 7.0 degrees C) than in euthermic hamsters. When adenylyl cyclase activity was stimulated by isoprenaline, guanylylimidodiphosphate [Gpp(NH)p], or forskolin, the rank order of potency was the same in euthermic hamsters and hibernating hamsters: isoprenaline < Gpp(NH)p < forskolin. Functional competition curves indicated that in the euthermic hamsters beta 1-receptors were responsible for 67% of the response to isoprenaline at night and 62% of the response during the day. In hibernating hamsters, in contrast, most of the response to isoprenaline (58%) was mediated via beta 2-receptors. This shift in the relative importance of the receptor subtypes may facilitate arousal from hibernation by making the heart more sensitive to circulating epinephrine.

scholarly journals Genetic regulation of β2-adrenergic receptors in 3T3-L1 fibroblasts

1989 ◽  
Vol 260 (1) ◽  
pp. 53-59 ◽  
Author(s):  
M T Nakada ◽  
K M Haskell ◽  
D J Ecker ◽  
J M Stadel ◽  
S T Crooke
Keyword(s):  
Phorbol Ester ◽  
Adrenergic Receptors ◽  
Adrenergic Receptor ◽  
Genetic Regulation ◽  
Receptor Subtype ◽  
Receptor Subtypes ◽  
Regulatory Sequences ◽  
Beta Adrenergic Receptor ◽  
Beta Adrenergic ◽  
Beta 2

The beta 2-adrenergic receptor from mouse 3T3-L1 cells is up-regulated through genetic mechanisms by glucocorticoids and butyrate. To study the genetic regulation of these receptors, we sequenced a 5 kb region of genomic DNA from 3T3-L1 cells, containing the beta-adrenergic receptor gene and approx. 1.5 kb of both 5′ and 3′ flanking sequences. The sequence contained one copy of an 8 bp consensus sequence which can confer phorbol ester-responsiveness to genes. Phorbol esters attenuated the up-regulation of beta 2-adrenergic receptors by glucocorticoids but not by butyrate. This effect was probably due to a phorbol ester-induced decrease in glucocorticoid receptor number. Using methylation-sensitive restriction enzymes, we examined the methylation of a CG-rich region occurring 5′ to the gene and did not detect any changes in methylation of this region upon dexamethasone or butyrate treatment. A total of 16 putative glucocorticoid response elements were found which may mediate the glucocorticoid-induced increase in beta 2-adrenergic receptors. A comparison of the regulatory sequences of the two beta-adrenergic receptor subtypes from human and mouse confirms the observed physiological controls of receptor subtype expression and offers an explanation as to why the subtypes differ in genetic regulation.

Segmental expression of somatostatin receptor subtypes sst1 and sst2 in tubules and glomeruli of human kidney

2001 ◽  
Vol 280 (3) ◽  
pp. F457-F465 ◽  
Author(s):  
Douglas A. Balster ◽  
M. Sue O'Dorisio ◽  
Monica A. Summers ◽  
Martin A. Turman
Keyword(s):  
Cell Function ◽  
Collecting Duct ◽  
Somatostatin Receptor ◽  
Human Kidney ◽  
Distal Tubule ◽  
Receptor Subtypes ◽  
Thick Ascending Limb ◽  
Somatostatin Receptor Subtypes ◽  
Normal Human Kidney ◽  
Normal Human

Somatostatin is known to modulate mesangial and tubular cell function and growth, but the somatostatin receptor (sst) subtypes responsible for these effects have not been defined. There are at least five different sst receptor subtypes (sst1-sst5). We used RT-PCR to demonstrate that normal human kidney consistently expresses mRNA for sst1 and sst2 (9 of 9 donors). Some donors expressed sst4 or sst5mRNA, but none expressed sst3 mRNA. Expression of sst1 and sst2 was further assessed by staining serial sections of normal human kidney with sst1 and sst2 antisera, Arachis hypogaea (AH) lectin (to define distal tubule/collecting duct cells), Phaseolus vulgaris lectin (proximal tubules), and Tamm-Horsfall protein (THP) antiserum (thick ascending limb of the loop of Henle). Specificity of antisera was demonstrated by transfection and absorption studies. Sst2, but not sst1, was expressed in glomeruli. Intense sst1 and sst2 staining localized exclusively to AH+ and THP+ tubules. Thus sst1 and sst2 subtype-selective analogs may be useful to beneficially modulate renal cell function in pathological conditions.

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