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Query: UNIPROT:P01185 (vasopressin)
 23,126 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Adenylate cyclase and the [8-lysine]vasopressin receptor were solubilized from pig kidney medulla membranes using the nonionic detergent Triton X-100. Optimal conditions for solubilization were under continuous stirring in a medium containing 0.5% (/v) Triton X-100, 100 mM Tris-HCl, pH 8, and 10 mM MgCl2. Both adenylate cyclase activity and [3H][8-lysine]vasopressin binding activity were recovered in a -26,000 X g supernatant of detergent-treated membranes. The yield of solubilized adenylate cyclase was nearly 100%. The soluble enzyme was no longer sensitive to antidiuretic hormone but was slightly activated by sodium fluoride. The affinity of the soluble receptor for [8-lysine]vasopresin was les than that of the membrane-bound receptor (mean apparent Km values, respectively 10(-7) M and 2 X 10(-8) M), however binding cooperativity was preserved. Hill coefficients were 1.42 for the soluble receptor and 1.50 for the membrane receptor. The soluble receptor discriminated as efficiently as did the membrane receptor between [8-lysine-a1vasopressin and oxytocin. The yield of spolubilized receptor was only 30% despite the fact that all binding activity had disappeared from the residual pellet of detergent-treated membranes. When the membranous receptors were occupied before solubilization and the latter was performed under conditions in which dissociation of the hormone-receptor comples is slow, i.e. at low temperature, 65% to 100% of the hormone-receptor complex was recovered in the soluble fraction. The soluble hormone-receptor complex partially dissociated on rewarming whereas the free hormone concentration was kept unchanged in the medium. The residual binding capacity, which was 30% of the initial value, was identical with that determined when the receptor was solubilized in free form before incubation with labeled hormone. It was concluded that (a) solubilization of the receptor molecules was complete, (b) during solubilization two forms of the receptor appear, of which only one is accessible to the hormone, (c) occupancy of the receptor by the hormone prevented the formation of the nonaccessible form, and (d) some component or components of the soluble fraction might be responsible for the loss in apparent affinity.
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PMID:Solubilization of the [8-lysine]vasopressin receptor and adenylate cyclase from pig kidney plasma membranes. 17 Feb 74

Recent data on the effects of neurohypophysial peptides at the cellular level are discussed with respect to the two basic processes involved in peptide hormone action--i.e., specific recognition of the information contained in the hormonal molecule and the transformation of this information into a stimulus leading to the final biological response. Four main aspects of this general problem are considered. A. Hormone-Receptor Interaction: Recent contributions in this field concern partial analysis of the three-dimensional conformation of oxytocin and vasopressin moleculal cells of the mammalian kidney. Conformational analysis of oxytocin and vasopressin molecules leads to the conclusion that, in solution, these peptides probably have a compact and highly stabilized three-dimensional configuration. Models have been proposed that provide a valuable clue to the interpretation of structure-activity relationships among natural hormones and many structural analogues. Binding studies with tritiated oxytocin and vasopressin have permitted determination of the kinetic parameters of hormone-receptor interaction in amphibian epithelial cells and mammalian kidney. B. Stimulus Generation: The nature of the primary stimulus generated by hormone-receptor interaction is still unknown. In the epithelial target cells of the amphibian skin and bladder and of the mammalian kidney, one of the first consequences of hormone-receptor interaction is the activation of membrane-bound adenylate cyclase. Analysis of the correlations between hormonal binding and adenylate cyclase activation suggests that activation is a function of receptor occupation rather than of the number of hormonal molecules interacting with the receptor per unit of time. On medullary adenylate cyclase of pig kidney, the relation between receptor occupancy and enzyme activation was found to be complex and nonlinear. The effects of several agents (calcium, nucleotides) on receptor occupancy and adenylate cyclase activation have been described. In mammalian uterus and other smooth muscle target cells, there is no evidence for direct involvement of cyclic AMP in the contractile response to oxytocin and other neurohypophysial peptides.
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PMID:Stimulus-response coupling in neurohypophysial peptide target cells. 17 91

The cation specific ionophore A23187 (Io) is a useful tool for studying the role of intracellular Ca++ (Ca++)i in physiologic processes. The present studies explore the role of (Ca++)i on Na transport in the toad bladder. Scraped bladder cells exposed to 1 muM Io for 60 min took up 100% more 45Ca than control cells. Io, 1 muM, added to the serosal side of bladders incubated in standard Ringers containing 2.5 mM Ca++ inhibited short circuit current (SCC) values by a mean of 30% at 60 min and 50% at 90 min. Io did not inhibit SCC significantly in bladders incubated in Ringers containing 0.2 mM Ca++. These data indicate that the effects of Io on SCC depend on the levels of external Ca++ and suggest that entry of Ca++ into cells mediates the inhibition of base-line SCC. PReincubation of the bladders with either lanthanum chloride or pentobarbital prevented the increased 45Ca uptake produced by ionophore as well as theinhibition of SCC caused by the antibiotic. Vasopressin, antidiuretic hormone (ADH). 10 MU/ml, increased peak SCC by 247% in bladders preincubated for 1 h in Ringers with 2.5 mM Ca++ and 1 muM Io and by 318% in control bladders (P less than 0.01). Bladders exposed to 1 muM Io in Ringers with 0.2 mM Ca++ had an increase in SCC after ADH comparable to that observed in controls. Since the effects of ADH on SCC are mediated by cyclic AMP, we tested the effects of Io on cAMP production by scraped toad bladder cells. ADH increased cAMP from 8 to 30 pmol/mg protein in controls but it did not increase cAMP over base-line values in the presence of Io when the Ringers contained 2.5 mM Ca++. Io did not inhibit cAMP production in response to ADH when the Ca++ in the Ringers was 0.2 mM. The results indicate that Io inhibits baseline and ADH stimulated SCC by increasing (Ca++)i or Ca++ bound to the cell membrane. It is suggested that: ()( (Ca++)i or membrane-bound Ca++ plays a key role in base-line and ADH stimulated Na transport in the toad bladder; (2) inhibition of ADH stimulated SCC may be due inpart to decreased cAMP generation in response to ADH when (Ca++)i or membrane-bound Ca++ levels are increased.
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PMID:Effects of ionophore A23187 on base-line and vasopressin-stimulated sodium transport in the toad bladder. 19 Feb 65

Two types of plasma membrane were purified from canine distal renal medulla by the techniques of differential and zonal density-gradient centrifugation followed by free-flow electrophoresis. One group of plasma membranes was identified as basal-laterally derived based on a 30-fold enrichment of Na-K-ATPase, a 20-fold enrichment of vasopressin-stimulated adenylate cyclase, and a 33-fold enrichment of [3H]vasopressin binding sites. The second type of plasma membrane was free of these markers, but had a cholesterol and phospholipid composition similar to them. Alkaline phosphatase also had a similar distribution in the two fractions. This lighter membrane fraction contained a membrane-bound cyclic AMP-dependent protein kinase as well as substrate for this kinase. In addition there was a 26-fold enrichment of specific activity of an anion (SO32-)-activated ATPase which was insensitive to mitochondrial ATPase inhibitor protein, in contrast to the mitochondrial fraction of the tissue. Based on the relative preponderance of collecting duct tissue in the distal medulla and the yield of membrane protein, these membranes are tentatively identified as containing apical membranes of the collecting duct.
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PMID:Purification of distinct plasma membranes from canine renal medulla. 20 99

A "random-hit" matrix model is proposed to account for the dynamic and steady state relationship between occupation of bovine renal medullary membrane receptors by [Lys8]vasopressin (LVP) and neurohypophyseal hormones (NHH) and the associated activation of membrane-bound adenylate cyclase. The model was developed by systematic introduction of specific rules concerning receptor coupling into a general structural model which consists of two square matrices of identical size, one composed of homogeneous R ("receptor") units, the second of homogeneous C ("cyclase") units. R units are either occupied (RO) or unoccupied (RU); C units are either active (CA) or inactive (CI). Hormone molecules are envisioned to "collide" with R units randomly; collision with RU leads to "binding", and occupation is maintained for a characteristic mean occupancy time, TO. In this structure, each R unit has an "interaction field" which consists of the "twin" unit in the "C" matrix, and the 4 nearest neighbor C units surrounding the twin. Occupation of an R unit leads to activation of all CI units in the interaction field of that R; CA units in the interaction field are refractory. Thus binding at a given R may "recruit" a variable number of inactive neighboring C units (5, 4, 3, 2, 1, or 0). The model requires that there be individual coupling delays between the moment of binding at a given R and subsequent activation of CI units (mean coupling delay (Td) approximately 10% To). Activation of C units persists as long as the "parent" R is occupied and is maintained for an additional short time interval (Tp) after RO reverts to RU, corresponding to hormone dissociation from receptor. The model accounts for the following previously demonstrated relations between LVP occupation of receptors and adenylate cyclase activation in bovine renal medullary membranes: 1) the shape of the nonlinear steady state relation between normalized (percentage maximal) receptor occupation (O) and cyclase activation (A), uniformly observed in different membrane preparations: 2) variable hormone concentration-dependent trajectories of approach to the final steady state A:O value (A:Oss) which may be either monophasic or biphasic; 3) the loss of intrinsic adenylate cyclase activity observed in bovine membranes for a series of NHH analogs with progressively diminishing affinity for receptors. The model represents an explicit theory of coupling where a successive series of temporal events are quantitatively related to each other and privide major constraints to any interpretation of the molecular organization of receptors and adenylate cyclase units in membranes. The model excludes a number of mechanistic proposals and suggests a new hypothesis for membrane coupling with features which may be generally applicable to other hormone-sensitive adenylate cyclase systems.
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PMID:Neurohypophyseal hormone-responsive renal adenylate cyclase. IV. A random-hit matrix model for coupline in a hormone-sensitive adenylate cyclase system. 64 Oct 67

Clomipramine inhibited pressor responses to potassium ions and vasopressin in the rat mesenteric vascular bed with an ID50 of about 1.8 microgram/ml against both pressor agents and the actions of indomethacin and PG2 on the clomipramine effect suggested that the drug may have been antagonising the action of an endogenous PG. This was supported by the inhibitory action of clomipramine on PG2 actions on guinea-pig ileum. A lower concentration also inhibited pressor responses to noradrenaline and angiotensin (ID50 about 9 ng/ml): inhibition was increased by PG2 and reduced by indomethacin. In this preparation potassium and vasopressin act primarily by stimulating calcium entry from the extracellular fluid whereas noradrenaline and angiotensin act primarily by releasing calcium from intracellular or membrane-bound stores. Our results can be explained by two actions: 1. a PG-antagonist action of clomipramine at the cell membrane and 2. a selectve inhibitory effect on release of intracellular calcium. Clomipramine may prove useful in studying PG and calcium-dependent mechanisms.
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PMID:Actions of the tricyclic antidepressant clomipramine on responses to pressor agents. Interactions with prostaglandin E2. 89 8

An antidiuretic hormone-inactivating peptidase located in renal plasma membranes of porcine kidney medulla has been studied. Treatment of antidiuretic hormone (lysine vasopressin) with renal plasma membranes resulted in a progressive loss of biological activity as measured by the rat pressor assay. The reaction of 2,4,6-trinitrobenzenesulfonic acid with released amino groups was employed to follow the peptidase-catalyzed hydrolysis of the hormone. An 83-fold purification of the membrane-bound peptidase was achieved by Lubrol PX solubilization of the membranes followed by DEAE-cellulose, hydroxylopatite, and 8% agarose column chromatography. The molecular weight of the peptidase was 442 000 as determined by 8% agarose gel filtration. An analysis of the antidiuretic hormone hydrolysis products by thin-layer chromatography revealed the presence of trinitrophenyl-glycinamide. The release of glycinamide from the hormone as a function of time was demonstrated. Mg2+ had a slight inhibitory effect and Ca2+ had a strong inhibitory effect on the peptidase activity.
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PMID:Partial purification and characterization of the antidiuretic hormone-inactivating enzyme from renal plasma membranes. 112 84

Rat embryo fibroblasts (REF52 cells) and the simian virus 40 transformed derivative (WT6 Ag6) were employed to characterize phospholipase D (PLD) activity in normal and transformed cells. In cells prelabeled with [3H]myristic acid or [3H]glycerol and treated with 12-O-tetradecanoylphorbol-13-acetate (TPA, 50 ng/ml medium) or vasopressin (VP, 100 ng/ml medium) in the presence of ethanol, the formation of labeled phosphatidylethanol (PEt) was 3- to 5-fold higher in REF52 cells than in the transformed cells. The transphosphatidylation of phosphatidylcholine (PC) to PEt was further examined in cell-free assay systems. Results demonstrated that the formation of PEt in the cell-free assays was dependent on the mode of substrate presentation and the source of the PC. With endogenous membrane-bound substrate, the formation of [3H]myristoyl-PEt was 5-fold higher in homogenates derived from normal cells as compared to transformed cell homogenates. In experiments using exogenous labeled PC isolated from either REF52 or transformed cells as substrate, cell-free PLD activity differed greatly with regard to the source of the PC. The formation of PEt from REF52-derived PC was approx. 4-fold higher as compared to PEt formed with PC derived from the transformed cells, irrespective of enzyme source. The results demonstrate that PLD in intact nontransformed fibroblasts is activatable by TPA and VP to a greater extent than in the transformed counterpart. The results from cell-free assays suggest that PLD activity is more dependent on the type of PC substrate than on the source of the enzyme.
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PMID:Phospholipase D activity in nontransformed and transformed fibroblasts. 132 32

1. A vasopressin (AVP) binding protein was purified from rat liver membranes by an improved method using [125I][d(CH2)5'Sarcosine7]AVP, a selective V1 AVP radioligand and a combination of CHAPS solubilization, gel filtration, lectin affinity and FPLC ion exchange chromatography. 2. The purified protein exhibited a maximum binding activity of 2480 pmol/mg protein with a KD of 4.5 nmol/L, which corresponds to a purification of approximately 26,700-fold. The molecular weight of this protein was 70,000 Da. 3. The binding of [125I][d(CH2)5'Sarcosine7]AVP to the solubilized membranes was dependent on the protein concentration, and was inhibited by the unlabelled peptides [d(CH2)5'Sarcosine7]AVP, AVP, and to a lesser degree by peptides with high V2 receptor affinity, such as 1-desamino-D-AVP and [d(CH2)5'D-Ileu2-Ileu4]AVP. 4. In addition, an AVP anti-idiotypic monoclonal antibody bound to both the partially purified and purified lectin affinity AVP binding protein in a concentration-dependent manner. These results indicate that the purified protein displays similar characteristics to the liver membrane-bound AVP V1 receptor.
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PMID:Isolation and characterization of the rat liver AVP receptor using [125I][d(CH2)5'sarcosine7]AVP. 151 73

The relationship between cell proliferation and inositol lipid turnover has been studied by comparing the steady state of inositol derivative metabolism in quiescent and regenerating rat hepatocytes isolated at 4 h (G1 phase of first cell cycle) and 24 h (onset of M phase) after partial hepatectomy. The effect of two hormones able to regulate hepatic regeneration, insulin and vasopressin, has been considered, and the results can be summarized as follows: (i) at 4 h after partial hepatectomy, the precursor incorporation into inositol polyphosphates and the particulate phospholipase C activity increase with respect to quiescent hepatocytes, whereas the content of 11, 4, 5P3 does not change, suggesting an increased turnover of this molecule in this step of cell cycle priming; (ii) 24 h after partial hepatectomy, the radioactivity linked to IP3 and IP4, as well as soluble and particulate phospholipase C activity, and IP3 content increase, suggesting the presence, at the onset of M phase, of second messenger accumulation; (iii) only 24 h after partial hepatectomy, the inositol derivative metabolism is affected by vasopressin; and (iv) insulin exerts a modulatory role on inositol polyphosphate production without involving membrane-bound PLC activity or phosphoinositide hydrolysis. These data suggest that inositol-derived signal molecules are associated with hepatic regeneration; moreover, the metabolic pathway of such compounds seems to be regulated so that only specific inositol phosphates are present in each step of the cell cycle.
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PMID:Signal transduction during liver regeneration: role of insulin and vasopressin. 163 71


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