Vasopressin V2 Receptor/Bioligand Interactions

We predict some essential interactions between the V2 vasopressin renal receptor (V2R) and its agonists [Arg⁸]vasopressin (AVP) and [D-Arg⁸]vasopressin (DAVP), and the non-peptide antagonist OPC-31260. V2R controls antidiuresis and belongs to the superfamily of heptahelical transmembrane (7TM) G-protein-coupled receptors (GPCRs). The receptor was built, the ligands were docked and the structures relaxed using advanced molecular modeling techniques. Docked agonists and antagonists appear to prefer similar V2R compartments. A number of receptor amino acid residues are indicated, mainly in the TM3–TM7 helices, as potentially important in ligand binding. Many of these residues are invariant for either the GPCR superfamily or the subfamily of related (vasopressin V2R, V1aR and V1bR and oxytocin OR) receptors. Moreover, some of the equivalent residues in V1aR have already been found critical for ligand affinity [Mouillac et al., J. Biol. Chem., 270 (1995) 25771].     

Vasopressin V2 Receptor/Bioligand Interactions

Summary We predict some essential interactions between the V2 vasopressin renal receptor (V2R) and its agonists [Arg⁸]vasopressin (AVP) and [D-Arg⁸]vasopressin (DAVP), and the non-peptide antagonist OPC-31260. V2R controls antidiuresis and belongs to the superfamily of heptahelical transmembrane (7TM) G-protein-coupled receptors (GPCRs). The receptor was built, the ligands were docked and the structures relaxed using advanced molecular modeling techniques. Docked agonists and antagonists appear to prefer similar V2R compartments. A number of receptor amino acid residues are indicated, mainly in the TM3-TM7 helices, as potentially important in ligand binding. Many of these residues are invariant for either the GPCR superfamily or the subfamily of related (vasopressin V2R, V1aR and V1bR and oxytocin OR) receptors. Moreover, some of the equivalent residues in V1aR have already been found critical for ligand affinity [Mouillac et al., J. Biol. Chem., 270 (1995) 25771].     

Non-peptide oxytocin agonists

A library of compounds targeted to the vasopressin/oxytocin family of receptors was screened for activity at a cloned human oxytocin receptor using a reporter gene assay. Potency and selectivity were optimised to afford compound 39, EC50 = 33 nM. This series of compounds represents the first disclosed, non-peptide, low molecular weight agonists of the hormone oxytocin (OT).     

Synthesis and SAR studies of novel 2-(4-oxo-2-aryl-quinazolin-3(4H)-yl)acetamide vasopressin V1b receptor antagonists

Synthesis and structure-activity relationships (SAR) of a novel series of vasopressin V_1b (V₃) antagonists are described. 2-(4-Oxo-2-aryl-quinazolin-3(4H)-yl)acetamides have been identified with low nanomolar affinity for the V_1b receptor and good selectivity with respect to related receptors V_1a, V₂ and oxytocin (OT). Optimised compound 12j demonstrates a good pharmacokinetic profile and activity in a mechanistic model of HPA dysfunction.     

Micelle-bound conformations of neurohypophyseal hormone analogues modified with a Cα-disubstituted residue: NMR and molecular modelling studies

In this study, by applying a combined approach of NMR measurements and molecular modelling, the conformations and the interactions with membrane-like environment of five arginine vasopressin (AVP) or oxytocin (OT) analogues modified with Cα-disubstituted cis-1-amino-4-phenylcyclohexane-1-carboxylic acid in position 2 have been determined. In addition, the AVP analogues were prepared in N-acylated forms with various bulky acyl groups. All of the peptides studied interacted with the mixed dodecylphosphocholine:sodium dodecyl sulphate micelle, providing a model of biological membrane. A different polarities of the AVP- and OT-like peptides resulted in their different position relative to the micelle surface. Thus, the arrangement of the former was nearly perpendicular, whereas the latter was rather parallel to the micelle's surface. Moreover, the results of our studies have shown that the binding sites for antagonists may be overlapped with that for agonists, as well as it may be quite different. Nevertheless, the aromatic–aromatic contacts represent the most important interactions for antagonists, whereas the hydrophilic interactions seem to be crucial for agonists.     

Functional relations of crab molt-inhibiting hormone and neurohypophysial peptides

Biological and immunological relationships between molt-inhibiting hormone (MIH) activity in eyestalk ganglia extracts of the crab, Cancer antennarius Stimpson, and peptides of the vasopressin-oxytocin family were assessed. Lysine vasopressin (LVP), arginine vasopressin (AVP), vasotocin (VT), and oxytocin (OT) mimicked MIH action by inhibiting ecdysteroid production of Y-organ segments in vitro with the relative potencies LVP greater than AVP greater than VT much much greater than OT. The inhibitory effect was reversible and specific (6 other peptides did not alter Y-organ activity). MIH and LVP increased Y-organ cyclic adenosine 3',5' monophosphate (cAMP) levels dose-dependently and with identical time course in which the rise in cAMP preceded inhibition of ecdysteroid production. The synthetic vasopressin antidiuretic agonist 1-deamino-8-D-AVP (dDAVP) inhibited Y-organ steroidogenesis dose-dependently; the vasopressin analog ([1(B-mercapto-beta, beta-cyclopentamethylenepropionic acid), 2-(O-methyl)tyrosine[AVP) (d(CH2)5Tyr(Me)AVP), a vasopressor antagonist, had no effect on basal or MIH-suppressed steroidogenesis. AVP antiserum abolished the inhibitory action of MIH, LVP, and AVP. Competitive binding curves for MIH, LVP, AVP, VT, and OT with the AVP antiserum suggested that MIH is most closely related to LVP. MIH may be structurally related to the vasopressins and act on Y-organ cells via type V2 (cAMP-linked) receptors.     

Iodinated neurohypophyseal hormones as potential ligands for receptor binding and intermediates in synthesis of tritiated hormones.

[3-Iodo-Tyr2]oxytocin (MIOT), [3,5-diiodo-Tyr2]oxytocin (DIOT), [3-iodo-Tyr2,Lys8]vasopressin (MILVP), [3,5-diiodo-Tyr2,Lys8]vasopressin (DILVP), [3-iodo-Tyr2,Arg8]vasopressin (MIAVP), and [3,5-diiodo-Tyr2,Arg8]vasopressin (DIAVP) were synthesized by iodination of the respective hormones, pruified, and characterized. All the monoiodo hormones had to be freshly prepared prior to bioassays, since on storage they gave rise to hormonal-like biological activity. The biological activities of these iodo analogues were measured in an adenylate cyclase assay employing neurohypophyseal hormone (NHH) sensitive bovine renal medullary membranes, and/or the rat oxytocic assay. In the cyclase assay, DIOT, DILVP, and DIAVP were inactive as agonists or antagonists. MIOT shows no agonistic activity in the renal cyclase system and uterus, but is a weak reversible inhibitor of oxytocin (OT) in both systems. When MIOT (10(-4) M) was preincubated with renal membranes for 10 min at 37 degrees C before addition of OT, it behaved as a noncompetitive inhibitor of NHH-stimulated adenylate cyclase. MILVP and MIAVP appear to be partial agonists with Km (half maximal response) 3 X 10(-6) and 3 X 10(-7) M, respectively, as determined in the cyclase assay. Upon preincubation with renal medullary membranes, MILVP (10(-6) M) behaves as a more potent noncompetitive inhibitor of OT than MIOT. Accordingly, iodo derivatives of NHH do not exhibit sufficient affinity to serve an specific ligands to measure OT, LVP, or AVP receptors in the uterus and kidney. Study of the specificity of inhibition produced by MIOT revealed that this analogue does not act selectively upon NHH receptors. Thus, MIOT modified adenylate cyclase systems which do not have NHH receptors, e.g., the PTH-sensitive adenylate cyclase in bovine renal cortex and the glucagon-sensitive adenylate cyclase in rat liver. DIOT, DILVP, and DIAVP were subjected to catalytic tritiation (employing carrier free tritium) and were converted to [3H]OT (25, 31, and 25 Ci/mmol), [3H]LVP (26 and 23 Ci/mmol), and [3H]AVP (17 Ci/mmol), respectively. These tritiated ligands have been successfully used to measure NHH receptor sites both in kidney and uterine membranes as described in other studies.     

中缝大核在刺激视上核镇痛中的作用

运用核团灌流液的放免测定和高压液相色谱法以及核团内注射拮抗剂,观察了化学刺激下丘脑视上核（ＳＯＮ）对中缝大核（ＮＲＭ）灌流液内催产素（ＯＴ）、精氨酸加压素（ＡＶＰ）和５－羟色胺（５－ＨＴ）含量的影响以及ＮＲＭ内注射ＡＶＰ、５－ＨＴ或ＯＴ受体拮抗剂对痛阈（ＰＴ）的影响。结果表明：ＳＯＮ内注射Ｌ－谷氨酸（Ｌ－Ｇｌｕ）１０μｇ后动物痛阈明显升高,ＮＲＭ灌流液中ＯＴ和５－ＨＴ的含量明显高于对照组水平,ＡＶＰ的含量仅有一过性增加。ＮＲＭ内注射ｏＴ或５－ＨＴ拮抗剂可逆转化学刺激ＳＯＮ引起的镇痛作用;而ＡＶＰ的Ｖ＿(１／２)受体拮抗剂也轻度抑制这种镇痛作用,但Ｖ＿１拮抗剂对此作用无影响。以上结果提示：在刺激ＳＯＮ镇痛中,ＯＴ起着重要作用,Ｌ－Ｇｌｕ刺激ＳＯＮ的ＯＴ细胞释放ＯＴ,作用于ＮＲＭ细胞的ＯＴ受体和Ｖ＿２受体而产生镇痛作用,５－ＨＴ在此过程中也发挥重要作用。     

Physiology of spontaneous [Ca2+]i oscillations in the isolated vasopressin and oxytocin neurones of the rat supraoptic nucleus

The magnocellular vasopressin (AVP) and oxytocin (OT) neurones exhibit specific electrophysiological behaviour, synthesise AVP and OT peptides and secrete them into the neurohypophysial system in response to various physiological stimulations. The activity of these neurones is regulated by the very same peptides released either somato-dendritically or when applied to supraoptic nucleus (SON) preparations in vitro. The AVP and OT, secreted somato-dendritically (i.e. in the SON proper) act through specific autoreceptors, induce distinct Ca²⁺ signals and regulate cellular events. Here, we demonstrate that about 70% of freshly isolated individual SON neurones from the adult non-transgenic or transgenic rats bearing AVP (AVP-eGFP) or OT (OT-mRFP1) markers, produce distinct spontaneous [Ca²⁺]_i oscillations. In the neurones identified (through specific fluorescence), about 80% of AVP neurones and about 60% of OT neurones exhibited these oscillations. Exposure to AVP triggered [Ca²⁺]_i oscillations in silent AVP neurones, or modified the oscillatory pattern in spontaneously active cells. Hyper- and hypo-osmotic stimuli (325 or 275 mOsmol/l) respectively intensified or inhibited spontaneous [Ca²⁺]_i dynamics. In rats dehydrated for 3 or 5 days almost 90% of neurones displayed spontaneous [Ca²⁺]_i oscillations. More than 80% of OT-mRFP1 neurones from 3 to 6-day-lactating rats were oscillatory vs. about 44% (OT-mRFP1 neurones) in virgins. Together, these results unveil for the first time that both AVP and OT neurones maintain, via Ca²⁺ signals, their remarkable intrinsic in vivo physiological properties in an isolated condition.     

Position Three in Vasopressin Antagonist Tolerates Conformationally Restricted and Aromatic Amino Acid Substitutions: A Striking Contrast with Vasopressin Agonists

We report the solid-phase synthesis and some pharmacological properties of 12 position three modified analogues (peptides 1–12) of the potent non-selective antagonist of the antidiuretic (V₂-receptor), vasopressor (V_1a-receptor) responses to arginine vasopressin (AVP) and of the uterine contracting (OT-receptor) responses to oxytocin (OT), [1(-β mercapto-β,β-pentamethy lenepropionic acid)-2-O-ethyl-d -tyrosine 4-valine] arginine vasopressin [d(CH₂)₅D -Tyr(Et) ²VAVP] (A) and two analogues of ( B ) (peptides 13,14), the 1,2,3,4-tetrahydroisoquinoline-3-carboxylic acid³ (Tic³) analogue of ( A ). Peptides 1–12 have the following substituents at position three in ( A ): (1) Pro; (2) Oic; (3) Atc; (4) D -Atc; (5) Aic; (6) D -Phe; (7) Ile; (8) Leu; (9) Tyr; (10) Trp; (11) Hphe; (12) [HO]Tic; Peptide (13) is the Tyr-NH₂ ⁹ analogue of ( B ): Peptide (14) is the D -Cys ⁶ analogue of ( B ). All 14 new peptides were evaluated for agonistic and antagonistic activities in in vivo V₂ and V_1a assays and in in vitro (no Mg²⁺) _n oxytocic assays. With the exception of the D -Phe³ peptide (No. 6), which exhibits very weak V₂ agonism (…0.0017 u/mg), none of the remaining 13 peptides exhibit any agonistic activities in these assays. In striking contrast to their deleterious effects on agonistic activities in AVP, the Pro³, Oic³, Tyr³, Trp³ and Hphe³ substitutions in ( A ) are very well tolerated, leading to excellent retention of V₂, V_1a and OT antagonistic potencies. All are more potent as V₂ antagonists than the Ile³ and Leu³ analogues of ( A ). The Tyr-NH₂⁹ and D -Cys⁶ substitutions in ( B ) are also well tolerated. The anti-V₂ pA₂ values of peptides 1–5 and 7–14 are as follows (1) 7.77±0.03; (2) 7.41± 0.05; (3) 6.86±0.02; (4) 5.66±0.09; (5) …5.2; (7) 7.25± 0.08; (8) 6.82±0.06; (9) 7.58±0.05; (10) 7.61±0.08; (11) 7.59±0.07; (12) 7.20±0.05; (13) 7.57±0.1; (14) 7.52± 0.06. All analogues antagonize the vasopressor responses to AVP, with anti-V _1a pA₂ values ranging from 5.62 to 7.64, and the in vitro responses to OT, with anti-OT pA₂ values ranging from 5.79 to 7.94. With an anti-V₂ potency of 7.77±0.03, the Pro³ analogue of ( A ) is surprisingly equipotent with ( A ), (anti-V₂ pA₂=7.81±0.07). These findings clearly indicate that position three in AVP V₂/V_1a antagonists, in contrast to position three in AVP agonists, is much more amenable to structural modification than had heretofore been anticipated. Furthermore, the surprising retention of V₂ antagonism exhibited by the Pro³, Oic³, Tyr³, Trp³ and Hphe³ analogues of ( A ), together with the excellent retention of V₂ antagonism by the Tyr-NH₂⁹ and D -Cys⁶ analogues of ( B ) are promising new leads to the design of potent and possibly orally active V₂ antagonists for use as pharmacological tools and/or as radioiodinatable ligands and for development as potential therapeutic agents for the treatment of the hyponatremia caused by the syndrome of the inappropriate secretion of the antidiuretic hormone (SIADH). © 1997 European Peptide Society and John Wiley & Sons, Ltd.     

Molecular modeling of the neurohypophyseal receptor/atosiban complexes

The neurohypophyseal nonapeptide hormone oxytocin (OT) is the strongest uterotonic substance known and is responsible for the initiation of labor. Conversely, oxytocin antagonists blocking uterine OT receptor can suppress uterus contraction. In this paper we describe a computer simulated docking pertinent to affinity of an oxytocin antagonist atosiban towards OT receptor, versus vasopressin V1a and V2 receptors.     

Rhythmic activities of hypothalamic magnocellular neurons: Autocontrol mechanisms

Electrophysiological recordings in lactating rats show that oxytocin (OT) and vasopressin (AVP) neurons exhibit specific patterns of activities in relation to peripheral stimuli: periodic bursting firing for OT neurons during suckling, phasic firing for AVP neurons during hyperosmolarity (systemic injection of hypertonic saline). These activities are autocontrolled by OT and AVP released somato-dentritically within the hypothalamic magnocellular nuclei. In vivo, OT enhances the amplitude and frequency of bursts, an effect accompanied with an increase in basal firing rate. However, the characteristics of firing change as facilitation proceeds: the spike patterns become very irregular with clusters of spikes spaced by long silences; the firing rate is highly variable and clearly oscillates before facilitated bursts. This unstable behaviour dramatically decreases during intense tonic activation which temporarily interrupts bursting, and could therefore be a prerequisite for bursting. In vivo, the effects of AVP depend on the initial firing pattern of AVP neurons: AVP excites weakly active neurons (increasing duration of active periods and decreasing silences), inhibits highly active neurons, and does not affect neurons with intermediate phasic activity. AVP brings the entire population of AVP neurons to discharge with a medium phasic activity characterised by periods of firing and silence lasting 20–40 s, a pattern shown to optimise the release of AVP from the neurohypophysis. Each of the peptides (OT or AVP) induces an increase in intracellular Ca²⁺ concentration, specifically in the neurons containing either OT or AVP respectively. OT evokes the release of Ca²⁺ from IP3-sensitive intracellular stores. AVP induces an influx of Ca²⁺ through voltage-dependent Ca²⁺ channels of T-, L- and N-types. We postulate that the facilitatory autocontrol of OT and AVP neurons could be mediated by Ca²⁺ known to play a key role in the control of the patterns of phasic neurons.     

NMR studies of new arginine vasopressin analogs modified with α-2-indanylglycine enantiomers at position 2 bound to sodium dodecyl sulfate micelles

In this paper, we use NMR spectroscopy and molecular modeling to examine four new vasopressin analogs modified with α-2-indanylglycine (Igl) at position 2, [L-Igl²]AVP (I), [D-Igl²]AVP (II), [Mpa¹,L-Igl²]AVP (III) and [Mpa¹,D-Igl²]AVP (IV), embedded in a sodium dodecyl sulfate (SDS) micelle. All the analogs display antiuterotonic activity. In addition, the analogs with D-Igl reveal antipressor properties.Each analog exhibits the tendency to adopt β-turns at positions 2, 3 and/or 3, 4, which is characteristic of oxytocin-like peptides. Mutual arrangement of aromatic residues at positions 2 and 3 has been found to be crucial for binding antagonists with the OT and V_1a receptors. The orientation of the Gln⁴ side chain seems to be important for the V_1a receptor affinity. In each of the peptides studied, the Gln⁴ side chain is folded back over the ring moiety. However, it lies on the opposite face of the tocin moiety in analogs with L and D enantiomers of Igl.     

Quantitative study of [Tyr10]nociceptin/orphanin FQ (1-11) at NOP receptors in rat periaqueductal gray and expressed NOP receptors in HEK293 cells

AimThe nociceptin/orphanin FQ (N/OFQ) peptide (NOP) receptor was reported to be functionally heterogeneous. We investigated if [Tyr¹⁰]N/OFQ(1-11), a peptide ligand reported to selectively bind to the high affinity site of ¹²⁵I-[Tyr¹⁴]N/OFQ in rodent brains, can be a tool for revealing the NOP receptor heterogeneity. We have previously founded an NOP receptor subset insensitive to Ro 64-6198 and (+)-5a Compound, two non-peptide NOP agonists, in rat ventrolateral periaqueductal gray (vlPAG) neurons. Here, we examined if [Tyr¹⁰]N/OFQ(1-11) differentiated (+)-5a Compound-sensitive and -insensitive vlPAG neurons. Certain mu-opioid (MOP) receptor ligands highly competing with [Tyr¹⁰]N/OFQ(1-11) in binding studies also showed high affinity at expressed heteromeric NOP–MOP receptors. We also examined if [Tyr¹⁰]N/OFQ(1-11) distinguished heteromeric NOP–MOP receptors from homomeric NOP receptors.Main methodsThe NOP receptor activity was evaluated by G-protein coupled inwardly rectifying potassium (GIRK) currents in rat vlPAG slices, and by inhibition of cAMP accumulation in HEK293 cells expressing NOP receptors or co-expressing NOP and MOP receptors.Key findingsIn vlPAG neurons, [Tyr¹⁰]N/OFQ(1-11), like N/OFQ, induced GIRK currents through NOP receptors. It was less potent (EC₅₀: 8.98 μM) but equi-efficacious as N/OFQ. [Tyr¹⁰]N/OFQ(1-11) displayed different pharmacological profiles as (+)-5a Compound, and was effective in both (+)-5a Compound-sensitive and -insensitive neurons. In NOP-expressing HEK293 cells and NOP- and MOP-co-expressing cells, [Tyr¹⁰]N/OFQ(1-11) displayed similar concentration–response curves in decreasing cAMP accumulation.Significance[Tyr¹⁰]N/OFQ(1-11) is an NOP full agonist and less potent than N/OFQ. However, it can neither reveal the functional heterogeneity of NOP receptors in vlPAG neurons nor differentiate heteromeric NOP–MOP and homomeric NOP receptors.     

Theoretical conformational analysis of six arginine vasopressin analogs with the L-naphthylalanine in position 3.

Introduction of the naphthylalanine residue into either position 3 of arginine vasopressin (AVP), or its analogs results in peptides with interesting pharmacological properties. The single substituted analog of AVP with L-2-Nal in position 3 causes moderate antiduretic activity, whereas [Mpa1, (L-1-Nal)3, (D-Arg)8] VP and [Mpa1, (L-2-Nal)3, (D-Arg)8] VP are potent and selective V2 agonists. Moreover [(L-2-Nal)3, (D-Arg)8] VP is among the most potent and selective antagonists of V1a receptors. In this study we carried out conformational calculations on [(L-1-Nal)3] AVP, [(L-2-Nal)3] AVP, [(L-1-Nal)3, (D-Arg)8] VP, [(L-2-Nal)3, (D-Arg)8] VP, [Mpa1, (L-1-Nal)3, (D-Arg)8] VP, [Mpa1, (L-2-Nal)3, (D-Arg)8] VP, using the ECEPP/3 force field with and without including hydration to simulate aqueous and nonpolar environments. It was found that in all six compound studied, the low-energy conformations have common geometry and relative energies. Therefore, the modifications of the Phe in position 3 influence the binding to the receptor by changing the size of the third residue, rather than by changing the conformational space. The lowest-energy conformations in the presence and absence of water had beta-turns at residues Phe3-Gln4 and Gln4-Asn5 and Gln4-Asn5, respectively. The conformation at the Gln4-Asn5 turn was most similar to the crystal structure of the pressinoic acid (the cyclic moiety of vasopressin).     

Paradoxical effects of oxytocin and vasopressin on basal prolactin secretion and the estrogen-induced prolactin surge

The roles of oxytocin (OT) and vasopressin (AVP) on both basal and estrogen-induced prolactin (PRL) secretion were examined. Adult female Sprague-Dawley rats that were ovariectomized for 3 weeks and received estrogen treatment for 1 week were used. Intravenous administration of hormones and serial blood sampling were accomplished through indwelling intraatrial catheters which were implanted two days before. Plasma PRL levels were measured by radioimmunoassay. Oxytocin at a dose of 20 micrograms/rat stimulated a moderate PRL release in the morning and lower doses (5 and 10 micrograms) were without effect. Vasopressin was most effective at a dose of 5 micrograms/rat in stimulating PRL release, while consecutive injections of higher doses (10 and 20 micrograms) were less effective. In contrast, TRH, ranging from 1 to 8 micrograms/rat, induced a dose-dependent increases in PRL secretion. Using the effective dosages determined from the morning studies, repeated injections of either OT, AVP or their specific antagonists MPOMeOVT [( 1-(beta-mercapto-beta, beta-cyclopentamethylene propanoic acid), 2-(O-methyl)tyrosine, 8-ornithine]-vasotocin) and d (CH2)5Tyr(Me)AVP ([1-(beta-mercapto-beta, beta-cyclo-pentamethylene propionic acid), 2-(O-methyl)tyrosine, 8-arginine]-vasopressin), were given hourly between 1300 to 1800 h and blood samples were obtained hourly from 1100 to 1900 h. It was found that either OT or AVP significantly reduced the afternoon PRL surge, while their antagonists were not as effective. When OT or AVP were administered together with their specific antagonists, the inhibitory effects of either hormone on PRL surge were reversed. Thus it is concluded that both OT and AVP assume a non-specific stress-like effect on PRL release, in which basal secretion is stimulated and surge secretion is inhibited.     

Analysis of interactions responsible for vasopressin binding to human neurohypophyseal hormone receptors-molecular dynamics study of the activated receptor-vasopressin-G(alpha) systems.

Vasopressin (CYFQNCPRG-NH(2), AVP) is a semicyclic endogenous peptide, which exerts a variety of biological effects in mammals. The main physiological roles of AVP are the regulation of water balance and the control of blood pressure and adrenocorticotropin hormone (ACTH) secretion, mediated via three different subtypes of vasopressin receptors: V1a, V1b and V2 receptors (V1aR, V1bR and V2R, respectively). They are the members of the class A, G-protein-coupled receptors (GPCRs). AVP also modulates several behavioral and social functions. In this study, the interactions responsible for AVP binding to vasopressin V1a and V2 receptors versus the closely related oxytocin ([I3,L8]AVP, OT) receptor (OTR) have been investigated. Three-dimensional models of the activated receptors were constructed using multiple sequence alignment, followed by homology modeling using the complex of activated rhodopsin with Gt(alpha) C-terminal peptide of transducin MII-Gt(338-350) prototype as a template. AVP was docked into the receptor-G(alpha) systems. The three lowest-energy pairs of receptor-AVP-G(alpha) (two complexes per each receptor) were selected. The 1-ns unconstrained molecular dynamics (MD) of complexes embedded into the fully hydrated 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphatidylcholine (POPC) lipid bilayer was conducted in the AMBER 7.0 force field. Six relaxed receptor-AVP-G(alpha) models were obtained. The residues responsible for AVP binding to vasopressin receptors have been identified and a different mechanism of AVP binding to V2R than to V1aR has been proposed.     

Theoretical conformational analysis of a μ-selective cyclic opioid peptide analog

The allowed conformations of the μ-receptor-selective cyclic opioid peptide analog were determined using a grid search through the entire conformational space. Energy minimization of the 13-membered ring structure lacking the exocyclic Tyr¹ residue and the Phe³ side chain using the molecular mechanics program Maximin resulted in only four low-energy conformations. These four ring structures served as templates for a further energy minimization study with the Tyr¹ residue and Phe³ side chain added to the molecule. The results indicated that the Tyr¹ and Phe³ side chains enjoy considerable orientational freedom, but nevertheless, only a limited number of low-energy side-chain configurations were found. The obtained low-energy conformers are discussed in relation to various proposed models of the bioactive conformation of enkephalins and morphiceptin.     

Effects of oxytocin,vasopressin and vasotocin and their agonists on steroid secretion by bovine granulosa cells

The effects of oxytocin and vasopressin and their agonists on the secretion of progesterone and oestradiol-17β by bovine luteinised granulosa cells cultured in a serum-supplemented medium were analysed. The effects of oxytocin (OT), its long-acting agonist 2-0-methyl-tyrosin (deamino-karba)-oxytocin (DK-OT), arginine-8-vasopressin (AVP), 1-desamino-arginine-8-vasopressin (D-AVP, a vasopressin analogue with high antidiuretic and without vasopressor properties) and arginine-8-vasotocin (AVT) were investigated. It was found that OT and DK-OT had a stimulatory effect on progesterone release, while AVP, D-AVP and AVT had an inhibitory effect. All peptide hormones investigated significantly increased oestradiol-17β secretion. The results suggest the involvement of nonapeptide hormones of both oxytocin and vasopressin groups in the regulation of steroidogenesis by granulosa cells from bovine ovarian follicles.     

Conformational studies of vasopressin analogues modified with N-methylphenylalanine enantiomers in dimethyl sulfoxide solution

The conformations of [Arg8]vasopressin (AVP) analogues substituted at positions 2 and 3 with N-methylphenylalanine (MePhe) enantiomers were earlier investigated by using nuclear magnetic resonance (NMR) spectroscopy in aqueous solution. A comparison of the results obtained in H2O/D2O (9:1) and DMSO-d6 has shown the structures in the first solution to be more flexible than those in DMSO-d6. This is manifested by a higher percentage of minor conformations in H2O/D2O. The largest differences between the NMR spectra in both solvents were noticed for [MePhe2, D-MePhe3]AVP (II) and [D-Cys1,MePhe2,D-MePhe3]AVP (III). Namely, in the ROESY spectra in aqueous solution, the cis/trans isomerization between MePhe2-DMePhe3 and D-Cys1-MePhe2 for II and III, respectively, is observed, while in DMSO-d6, the appropriate cross peaks indicate isomerization across the Cys6-Pro7 peptide bond. In the case of the remaining peptides, the position of cis/trans isomerization is the same in aqueous solution and in dimethyl sulfoxide. [D-MePhe2,MePhe3]AVP (V) displays low antiuterotonic and antipressor activities, while [D-MePhe2,)]AVP (IV) is a weak but selective blocker of oxytocin (OT) receptors in the uterus. The former shows similar conformational preferences as another antagonist of V1a and OT receptors-namely, [Acc2,D-Arg8]VP (Acc: 1-aminocyclohexane-1-carboxylic acid)-investigated by us. In the case of IV, the cis peptide bond between residues at positions 2 and 3 might be the reason for selectivity.