scholarly journals Specific vasopressin binding to rat adrenal glomerulosa cells. Relationship to inositol lipid breakdown

1986 ◽  
Vol 235 (1) ◽  
pp. 209-214 ◽  
Author(s):  
G Guillon ◽  
N Gallo-Payet
Keyword(s):  
Inositol Phosphate ◽  
Binding Capacity ◽  
Free Ligand ◽  
Zona Glomerulosa ◽  
High Affinity ◽  
Rat Adrenals ◽  
Vasopressin Receptors ◽  
Inositol Phosphate Accumulation ◽  
Specific Receptors ◽  
Glomerulosa Cells

Cells from the zona glomerulosa of rat adrenals were isolated and maintained for 3 days in primary culture. Specific vasopressin binding was determined by using [3H]vasopressin. [3H]Vasopressin binding was time-dependent (half-time of about 2 min for 6 nM free ligand) and reversible on addition of unlabelled vasopressin (80% dissociation within 30 min). Dose-dependent [3H]vasopressin binding at equilibrium indicated that vasopressin interacted with two populations of sites: high-affinity sites (dissociation constant, Kd = 1.8 nM; maximal binding capacity = 10 fmol/10(6) cells) and low-affinity sites. Vasopressin increased the cellular content of labelled inositol mono-, bis- and tris-phosphate in cells prelabelled with myo-[3H]inositol. The vasopressin concentration eliciting half-maximal inositol phosphate accumulation was very close to the Kd value for vasopressin binding to high-affinity sites. Competition experiments using agonists and antagonists with enhanced selectivity for previously characterized vasopressin receptors indicated that vasopressin receptors from rat glomerulosa cells are V1 receptors of the vascular or hepatic subtype. The detected specific vasopressin-binding sites might represent the specific receptors mediating the mitogenic and steroidogenic effects of vasopressin on glomerulosa cells from rat adrenals.

The properties of adrenal zona glomerulosa cells after purification by gravitational sedimentation

1974 ◽  
Vol 185 (1081) ◽  
pp. 375-407 ◽  
Keyword(s):  
Angiotensin Ii ◽  
Cyclic Amp ◽  
Microscopic Examination ◽  
Zona Glomerulosa ◽  
Maximal Response ◽  
Zona Fasciculata ◽  
Gravitational Sedimentation ◽  
Rat Adrenals ◽  
Glomerulosa Cells ◽  
Zona Glomerulosa Cells

The densities of latex spheres and biological cells can be reliably determined from their sedimentation rate in an albumin gradient under unit gravitational force. The densities of zona glomerulosa and fasciculata cells of rat adrenals were found to be 1.072 ± 0.004 and 1.040 ± 0.001 respectively. Purified zona glomerulosa cells of rat adrenals can be prepared by gravitational sedimentation of dispersed cells from capsule strippings of the gland, which originally contain 3 to10% zona fasciculata contamination. Electron and phase microscopic examination of the sedimented glomerulosa cells and their steroidogenic response to ACTH and cyclic AMP indicate that they are reasonably free of contamination from zona fasciculata cells. Electron microscopic examination of the purified glomerulosa cells indicates that most of them are reasonably normal in structure. Their basal production of corticosterone is decreased after sedimentation. However, their maximal response of corticosterone output to serotonin and potassium and their response to all potassium concentrations is not significantly altered, indicating normal function for the cells producing steroids. Their maximal responses to ACTH, valine angiotensin II and cyclic AMP are decreased, but, at the doses used, steroidogenesis by the zona fasciculata contamination in the unfractionated preparation would be stimulated by these substances. Purified zona glomerulosa cells have about the same maximal response of corticosterone output (about twofold) to potassium, valine and isoleucine angiotensin II, serotonin and ACTH. The maximal response of the purified zona glomerulosa cells to cyclic AMP is similar to that elicited by valine and isoleucine angiotensin II, potassium, serotonin or ACTH. This indicates that if these stimuli act by increasing cyclic AMP output, then the maximal response of corticosterone output (about twofold) is defined by the limited response of the biosynthetic pathways to cyclic AMP.

The interaction of arginine vasopressin and oxytocin with bovine adrenal medulla cells

1989 ◽  
Vol 121 (1) ◽  
pp. 133-139 ◽  
Author(s):  
A. H. Taylor ◽  
G. St J. Whitley ◽  
S. S. Nussey
Keyword(s):  
Arginine Vasopressin ◽  
Chromaffin Cells ◽  
Inositol Phosphates ◽  
Inositol Phosphate ◽  
Binding Capacity ◽  
Camp Production ◽  
Low Concentrations ◽  
Inositol Phosphate Accumulation ◽  
Adrenal Chromaffin ◽  
Induced Changes

ABSTRACT Binding of [3H]arginine vasopressin (AVP) and [3H]oxytocin to primary monolayer cultures of bovine adrenal chromaffin cells was time-dependent, and the binding sites for each peptide were specific and saturable. Studies with the V1 AVP antagonist d(CH2)5Tyr(Me)2-AVP, the V2 agonist 1-deamino-8-d-AVP and the V2 antagonist d(CH2)5d-Leu2,Val4-AVP indicated that the AVP receptor was V1 in specificity. Scatchard plots showed that each ligand interacted with a single high-affinity, low-capacity binding site: oxytocin dissociation constant (Kd) 0·29 ± 0·02 nmol/l, maximum binding capacity (Bmax) 7·6 ± 0·2 fmol/106 cells (or 4500 ± 102 sites/cell) (n = 3); AVP Kd 0·09±0·02 nmol/l, Bmax 5·1±0·63 fmol/106 cells (or 3050 ± 318 sites/cell) (n = 3). Although forskolin in concentrations from 1 nmol/l to 1 mmol/l stimulated cyclic AMP (cAMP) production in isolated chromaffin cells, this did not result in detectable catecholamine release. Neither AVP nor oxytocin in concentrations between 10 pmol/l and 10 μmol/l stimulated cAMP production in these cells. Vasopressin in concentrations as low as 10 pmol/l stimulated a sixfold increase in total inositol phosphates; the dose–response curve was triphasic. Oxytocin had little effect on total inositol phosphate accumulation at low concentrations, but concentrations above micromolar stimulated total inositol phosphate production approximately fourfold. There was no measurable release of catecholamines in response to either peptide. The physiological consequences of these AVP-induced changes in inositol phosphate concentrations remain to be elucidated. Journal of Endocrinology (1989) 121, 133–139

scholarly journals High-affinity binding of oestradiol-17β by cytosols from testis interstitial tissue, pituitary, adrenal, liver and accessory sex glands of the male rat

1974 ◽  
Vol 140 (3) ◽  
pp. 495-502 ◽  
Author(s):  
Wilma M. O. Van Beurden-Lamers ◽  
Albert O. Brinkmann ◽  
Eppo Mulder ◽  
Henk J. Van Der Molen
Keyword(s):  
Binding Capacity ◽  
Gradient Centrifugation ◽  
Sedimentation Coefficient ◽  
Wide Distribution ◽  
Male Rat ◽  
Interstitial Tissue ◽  
Apparent Contradiction ◽  
High Affinity ◽  
Specific Receptors ◽  
Agar Gel Electrophoresis

The specificity of the binding of oestradiol-17β by cytoplasmic fractions of several tissues of the male rat was investigated. 1. Agar-gel electrophoresis, Sephadex chromatography, adsorption by dextran-coated charcoal and sucrose-gradient centrifugation were used to estimate the binding capacity and specificity. The four different methods all gave similar results for the capacity of the specific oestradiol-17β-binding macromolecules in the testis. 2. The presence of a specific saturable binding protein with a sedimentation coefficient of 8S was demonstrated in liver, adrenal, pituitary, prostate, epididymis and testis interstitial tissue. The highest concentration of oestradiol-17β-binding macromolecules was found in testis interstitial tissue (0.12pmol/mg of protein) and in the pituitary (0.075pmol/mg of protein). 3. The oestradiol-17β receptor in the testis cytosol showed the characteristics of a protein with respect to Pronase treatment and temperature sensitivity. In competition experiments with different steroids the receptor showed a high affinity for oestradiol-17β, a moderate affinity for diethylstilboestrol and oestradiol-17α and a low affinity for oestrone, oestriol, testosterone and 5α-dihydrotestosterone (17β-hydroxy-5α-androstan-3-one). 4. The wide distribution of oestradiol-17β receptors in the male rat is in apparent contradiction to the current concept of the specificity of steroid-hormone action. Further research is required to investigate a possible physiological meaning of the presence of specific receptors in the different tissues.

scholarly journals Cloning of Octopus cephalotocin receptor, a member of the oxytocin/vasopressin superfamily

2003 ◽  
Vol 179 (2) ◽  
pp. 281-291 ◽  
Author(s):  
A Kanda ◽  
K Takuwa-Kuroda ◽  
E Iwakoshi-Ukena ◽  
Y Furukawa ◽  
O Matsushima ◽  
...  
Keyword(s):  
Inositol Phosphate ◽  
Biological Activities ◽  
Binding Pocket ◽  
Orphan Receptor ◽  
Brain Extract ◽  
Octopus Vulgaris ◽  
Vasopressin Receptors ◽  
Specific Receptors ◽  
Hplc Fractionation ◽  
G Protein Coupled

We reported that the common octopus, Octopus vulgaris, in common with vertebrates, possesses two members of the oxytocin/vasopressin superfamily: octopressin (OP) and cephalotocin (CT). This was the first observation of its kind in invertebrates. As OP and CT have different biological activities, the presence of specific receptors has been proposed. We cloned the cDNA of an orphan receptor from Octopus brain and found it to encode a polypeptide of 397 amino acids that displays sequences characteristic of G-protein coupled receptors. The orphan receptor showed high homology to receptors of the oxytocin/vasopressin superfamily and seemed to conserve the agonist-binding pocket common to the oxytocin and vasopressin receptors. Xenopus oocytes that express the orphan receptor responded to the application of CT by an induction of membrane Cl(-) currents coupled to the inositol phosphate/Ca(2+) pathway. OP and the other members of the oxytocin/vasopressin superfamily did not activate this receptor. HPLC fractionation of the Octopus brain extract combined with an oocyte assay yielded a single substance that was identical to CT. On the basis of these results, we conclude that the cloned receptor is the CT receptor (CTR). Expression of CTR mRNA in Octopus was detected in the central and the peripheral nervous systems, the pancreas, the oviduct and the ovary. This receptor may mediate physiological functions of CT in Octopus such as neurotransmission, reproduction and metabolism.

scholarly journals Local production and action of adrenomedullin in the rat adrenal zona glomerulosa

1998 ◽  
Vol 156 (3) ◽  
pp. 477-484 ◽  
Author(s):  
S Kapas ◽  
A Martinez ◽  
F Cuttitta ◽  
JP Hinson
Keyword(s):  
In Situ Hybridization ◽  
Inositol Phosphate ◽  
Zona Glomerulosa ◽  
Binding Studies ◽  
Aldosterone Secretion ◽  
Glomerulosa Cells ◽  
Camp Formation ◽  
Zona Glomerulosa Cells

This study was designed to investigate the synthesis and action of adrenomedullin in the rat adrenal gland. The results obtained from in situ hybridization and immunocytochemical studies suggest that adrenomedullin is synthesized not only in the medulla, but also within the zona glomerulosa of the rat adrenal cortex. Findings from in situ hybridization and binding studies also suggested that specific adrenomedullin receptors are expressed in the zona glomerulosa, and that low levels are present in the inner zones of the cortex. The Kd of the zona glomerulosa adrenomedullin receptor (5.5 nmol/l) suggests that it may respond to locally produced adrenomedullin rather than circulating concentrations of the peptide, which are in a lower range. It was found that adrenomedullin acted on zona glomerulosa cells in vitro to stimulate aldosterone release and cAMP formation, but in this tissue did not stimulate inositol phosphate turnover. The effect of adrenomedullin on aldosterone secretion was significantly attenuated by a protein kinase A inhibitor, suggesting that cAMP mediates the effects of adrenomedullin on aldosterone secretion. Adrenomedullin did not significantly affect the response of zona glomerulosa cells to stimulation by either ACTH or angiotensin II. Adrenomedullin did not affect the release of catecholamines, either adrenaline or noradrenaline, by intact adrenal capsular tissue. These data suggest that both adrenomedullin and its specific receptor are expressed in the rat adrenal zona glomerulosa, leading to the hypothesis that adrenomedullin may have an autocrine/paracrine role in the regulation of the rat adrenal zona glomerulosa.

Excitation-secretion coupling: involvement of potassium channels in ACTH-stimulated rat adrenocortical cells

1989 ◽  
Vol 120 (3) ◽  
pp. 409-421 ◽  
Author(s):  
N. Gallo-Payet ◽  
M. D. Payet
Keyword(s):  
Cyclic Amp ◽  
Calcium Influx ◽  
Inositol Phosphate ◽  
Aldosterone Secretion ◽  
Tetraethylammonium Chloride ◽  
Steroid Secretion ◽  
Phosphate Accumulation ◽  
Low Concentrations ◽  
Inositol Phosphate Accumulation ◽  
Glomerulosa Cells

ABSTRACT The mechanism by which ACTH stimulates calcium influx and steroid secretion was studied using rat adrenal glomerulosa cells, which were either freshly isolated or maintained in primary culture for 3 days. The potassium channel blocker tetraethylammonium chloride (TEA) stimulated twofold both corticosterone and aldosterone secretion; this stimulation was lower than that induced by ACTH at low concentrations (10 pmol/l). However, TEA and ACTH induced similar increases in Ca2+ influx and inositol phosphate accumulation. The three responses (steroid secretion, calcium influx and inositol phosphate accumulation) induced by TEA or low concentrations of ACTH were blocked by CoCl2. The greater stimulatory effect on steroid secretion of 10 nmol ACTH/l was decreased but not blocked by CoCl2. These data further document the complex mechanism of action of ACTH. It is postulated that, at low concentrations, ACTH binds preferentially to the high-affinity site of its receptor, leading to calcium influx by depolarization of the membrane potential, and to steroid secretion predominantly through an inositol phosphate- and Ca2+-stimulated pathway and also a cyclic AMP pathway. At higher concentrations, the hormone also binds to the low-affinity site of its receptor, largely stimulating cyclic AMP production and further increasing steroid secretion. Journal of Endocrinology (1989) 120, 409–421

scholarly journals Stimulation, by vasopressin and other agonists, of inositol-lipid breakdown and inositol phosphate accumulation in WRK 1 cells

1986 ◽  
Vol 240 (1) ◽  
pp. 197-204 ◽  
Author(s):  
C J Kirk ◽  
G Guillon ◽  
M N Balestre ◽  
S Jard
Keyword(s):  
Inositol Phosphates ◽  
Inositol Phosphate ◽  
Hormone Stimulation ◽  
Receptor Reserve ◽  
Phosphate Accumulation ◽  
Lipid Pool ◽  
Vasopressin Receptors ◽  
Inositol Phosphate Accumulation ◽  
Marked Accumulation

WRK 1 cells were labelled to equilibrium with 2-myo-[3H]inositol and stimulated with vasopressin. Within 3 s of hormone stimulation there was a marked accumulation of 3H-labelled InsP2 and InsP3 (inositol bis- and tris-phosphate), but not of InsP (inositol monophosphate). There was an associated, and rapid, depletion of 3H-labelled PtdInsP and PtdInsP2 (phosphatidylinositol mono- and bis-phosphates), but not of PtdIns (phosphatidylinositol), in these cells. Some 4% of the radioactivity in the total inositol lipid pool of WRK 1 cells was recovered in InsP2 and InsP3 after 10 s stimulation with the hormone. The selectivity of the vasopressin receptors of WRK 1 cells for a variety of vasopressin agonists and antagonists revealed these to be of the V1a subtype. There was no receptor reserve for vasopressin-stimulated inositol phosphate accumulation in WRK 1 cells. The accumulation of inositol phosphates was enhanced in the presence of Li+ions. Half-maximal accumulation of InsP, InsP2 and InsP3 in vasopressin-stimulated cells was observed with 0.9, 3.0 and 3.6 mM-Li+ respectively. Bradykinin and 5-hydroxytryptamine also provoked inositol phosphate accumulation in WRK 1 cells. The effects of sub-optimal concentrations of bradykinin and vasopressin upon inositol phosphate accumulation were additive, but those of optimal concentrations of the hormones were not.

scholarly journals Influence of bacterial toxins and forskolin upon vasopressin-induced inositol phosphate accumulation in WRK 1 cells

1989 ◽  
Vol 260 (3) ◽  
pp. 665-672 ◽  
Author(s):  
G Guillon ◽  
M N Balestre ◽  
C Lombard ◽  
F Rassendren ◽  
C J Kirk
Keyword(s):  
Cyclic Amp ◽  
Cholera Toxin ◽  
Pertussis Toxin ◽  
Inositol Phosphates ◽  
Inositol Phosphate ◽  
Binding Capacity ◽  
Specific Binding ◽  
Phosphate Accumulation ◽  
Cyclic Amp Accumulation ◽  
Inositol Phosphate Accumulation

The accumulation of inositol phosphates in WRK 1 cells, stimulated with a range of vasopressin concentrations, was diminished by prior exposure to cholera toxin or forskolin, whilst that observed in the presence of maximal concentrations of the hormone was enhanced in pertussis-toxin-treated cells. In the presence of [32P]NAD+, both cholera toxin and pertussis toxin provoked the labelling of peptides with approximate Mrs of 45,000 and 41,000 respectively in the membranes of WRK 1 cells. Exposure to cholera toxin or forskolin for 15-18 h enhanced cyclic AMP accumulation in these cells. The concentrations of these agents which provoked half-maximal cyclic AMP accumulation were similar to those required to diminish receptor-mediated inositol phosphate accumulation by 50%. In contrast, half-maximal ADP-ribosylation of the 45,000Mr peptide needed 100-fold greater concentrations of the toxin than were effective in provoking half-maximal inhibition of inositol phosphate accumulation. Cholera toxin or forskolin also reduced the maximal specific binding, to intact WRK 1 cells, of both [3H][Arg8]vasopressin and the V1a antagonist [3H][beta-mercapto-beta,beta-cyclopentamethylenepropionic acid,O-methyl-Tyr2, Arg8]vasopressin. The kinetics for the loss of this binding capacity following cholera-toxin treatment were very similar to those describing the diminution of vasopressin-stimulated inositol phosphate accumulation in the same cells.

Intracellular signalling in the ovine pars tuberalis: an investigation using aluminium fluoride and melatonin

1991 ◽  
Vol 7 (2) ◽  
pp. 137-144 ◽  
Author(s):  
P. J. Morgan ◽  
M. H. Hastings ◽  
M. Thompson ◽  
P. Barrett ◽  
W. Lawson ◽  
...  
Keyword(s):  
Cyclic Amp ◽  
Inositol Phosphates ◽  
Inositol Phosphate ◽  
Pars Tuberalis ◽  
Phosphoinositide Metabolism ◽  
Inhibitory Pathway ◽  
High Affinity ◽  
Aluminium Fluoride ◽  
Inositol Phosphate Accumulation ◽  
Phosphate Response

ABSTRACT The effect of aluminium fluoride (AlF4−) has been studied on inositol phosphate accumulation, calcium mobilization, cyclic AMP production and [2-125I]iodomelatonin binding in ovine pars tuberalis cells. These cells have high-affinity receptors for, and respond to, melatonin through inhibition of forskolin-stimulated adenylate cyclase. In the presence of 10 mm LiCl, AlF4− stimulated the net accumulation of inositol monophosphate and inositol bisphosphate. Consistent with these findings, AlF4− increased intracellular calcium; although this response was attenuated in calcium-depleted medium, indicating that the calcium response comprises both intracellular and extracellular components. Melatonin was ineffective on either basal or AlF4−-stimulated turnover of inositol phosphates. In concordance with the inositol phosphate response, melatonin had no effect on either the AlF4−-stimulated or the basal calcium levels. AlF4− blocked the increase in cyclic AMP stimulation by l μm forskolin, being as effective as melatonin, achieving approximately 90% inhibition. AlF4− also attenuated the binding of [2-125I]iodomelatonin to ovine pars tuberalis membranes by 15%. At the concentration used, these results are consistent with the interpretation that AlF4− activates many G protein-mediated responses, and thus imply that the inhibitory pathway for cyclic AMP predominates over the stimulatory arm, whereas there can only be a stimulatory pathway linked to phosphoinositide metabolism in ovine pars tuberalis cells.

Sign in / Sign up

Export Citation Format

Share Document