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Vasopressin (AVP) receptors present in In-R1-G9 cells, a hamster glucagon-secreting [alpha]-pancreatic cell line, were characterized using SSR-149415, a selective nonpeptide [V.sub.1b] receptor antagonist, and reference AVP compounds. Binding experiments, using [[sup.3]H]AVP as a ligand, identified a single population of high-affinity binding sites. SSR-149415 competitively inhibited this binding and exhibited nanomolar and stereospecific affinity for these sites. The affinity of various AVP/oxytocin ligands confirmed a [V.sub.1b] binding profile. In functional studies, AVP was a potent stimulant in inducing intracellular [Ca.sup.2+] increase, glucagon secretion, and cell proliferation. These effects were fully antagonized by SSR-149415 with a nanomolar potency, whereas its diasteroisomer as well as two selective [V.sub.1a] and [V.sub.2] receptor antagonists were much less potent. Additionally, the order of potency of AVP agonists and antagonists was in agreement with [V.sub.1b]-mediated effects. By RT-PCR, we confirmed the presence of [V.sub.1b] receptor mRNA in both In-R1-G9 cells and in human pancreas. The distribution pattern of [V.sub.1b] receptors investigated in human pancreas by immunohistochemistry showed strong labeling in islets of Langerhans, and colocalization studies indicated that this receptor was expressed in [alpha]-glucagon, [beta]-insulin, and somatostatin pancreatic cells. Thus, in In-R1-G9 cells, AVP mediates intracellular [Ca.sup.2+] increase, glucagon secretion, and cell proliferation by activating [V.sub.1b] receptors, and these effects are potently antagonized by SSR-149415. Moreover, the presence of [V.sub.1b] receptors also found in human Langerhans islets could suggest hormonal control of AVP in human pancreas. arginine vasopressin [V.sub.1b] receptors; glucagon; SSR-149415

The limbic localization of the arginine vasopressin [V.sub.1b] receptor has prompted speculation as to a potential role of this receptor in the control of emotional processes. To investigate this possibility, we have studied the behavioral effects of SSR149415, the first selective and orally active non-peptide antagonist of vasopressin [V.sub.1b] receptors, in a variety of classical (punished drinking, elevated plus-maze, and light/dark tests) and atypical (fear/anxiety defense test battery and social defeat-induced anxiety) rodent models of anxiety, and in two models of depression [forced swimming and chronic mild stress (CMS)]. When tested in classical tests of anxiety, SSR149415 produced anxiolytic-like activity at doses that ranged from 1 to 30 mg/kg (i.p. or p.o.), but the magnitude of these effects was overall less than that of the benzodiazepine anxiolytic diazepam, which was used as a positive control. In contrast, SSR149415 produced clear-cut anxiolyticlike activity in models involving traumatic stress exposure, such as the social defeat paradigm and the defense test battery (1-30 mg/kg, p.o.). In the forced swimming test, SSR149415 (10-30 mg/kg, p.o.) produced antidepressent-like effects in both normal and hypophysectomized rats. Moreover, in the CMS model in mice, repeated administration of SSR149415 (10 and 30 mg/kg, i.p.) for 39 days improved the degradation of the physical state, anxiety, despair, and the loss of coping behavior produced by stress. These findings point to a role for vasopressin in the modulation of emotional processes via the [V.sub.1b] receptor, and suggest that its blockade may represent a novel avenue for the treatment of affective disorders.

Although vasopressin V1B receptor (V1BR) mRNA has been detected in the kidney, the precise renal localization as well as pharmacological and physiological properties of this receptor remain unknown. Using the selective V1B agonist d[Leu4, Lys8]VP, either fluorescent or radioactive, we showed that V1BR is mainly present in principal cells of the inner medullary collecting duct (IMCD) in the male rat kidney. Protein and mRNA expression of V1BR were very low compared with the V2 receptor (V2R). On the microdissected IMCD, d[Leu4, Lys8]VP had no effect on cAMP production but induced a dose-dependent and saturable intracellular Ca2 + concentration increase mobilization with an EC50 value in the nanomolar range. This effect involved both intracellular Ca2 + mobilization and extracellular Ca2 + influx. The selective V1B antagonist SSR149415 strongly reduced the ability of vasopressin to increase intracellular Ca2+ concentration but also cAMP, suggesting a cooperation between V1BR and V2R in IMCD cells expressing both receptors. This cooperation arises from a cross talk between second messenger cascade involving PKC rather than receptor heterodimerization, as supported by potentiation of arginine vasopressin-stimulated cAMP production in human embryonic kidney-293 cells coexpressing the two receptor isoforms and negative results obtained by bioluminescence resonance energy transfer experiments. In vivo, only acute administration of high doses of V1B agonist triggered significant diuretic effects, in contrast with injection of selective V2 agonist. This study brings new data on the localization and signaling pathways of V1BR in the kidney, highlights a cross talk between V1BR and V2R in the IMCD, and suggests that V1BR may counterbalance in some pathophysiological conditions the antidiuretic effect triggered by V2R activation. [ABSTRACT FROM AUTHOR]

[³H]SSR-1494 15 is the first tritiated nonpeptide vasopressin V1b receptor (V1bR) antagonist Iigand. It was used for studying rodent (mouse, rat, hamster) and human V1bR from native or recombinant origin. Moreover, a close comparison between the human and the mouse V1bR was performed using SSR-149415/[³H]SSR-149415 in binding and functional studies in vitro. [³H]SSR-149415 binding was time-dependent, reversible, and saturable. Scatchard plot analysis gave a single class of high-affinity binding sites with apparent equilibrium dissociation constant (Kd) ~1 nM and maximum binding density (Bmax) values from 7,000 to 300,000 sites/cell according to the cell line. In competition experiments, [³H]SSR-149415 binding was stereospecific and dose-dependently displaced by reference peptide and nonpeptide arginine vasopressin (AVP)/OT ligands following a V1b rank order of affinity: SSR-149415 = AVP > dCha > dPen > dPal > dDavp > SSR-126768A > SR-49059 > SSR-149424 > OT > SR-121463B. Species differences between human, rat, mouse, and hamster V1bR were observed. Autoradiography studies with [³H]SSR-149415 on rat and human pituitary showed intense specific labeling confined to corticotroph cells and absence of labeling in the other tissues examined. SSR-149415 potently and stereospecifically antagonized the AVP-induced inositol phosphate production and intracellular Ca2+ increase (EC50 from 1.83 to 3.05 nM) in recombinant cell lines expressing either the mouse or the human V1bR. AVP (10-7 M) exposure of AtT20 cells expressing mouse or human EGFP-tagged V1bR induced their rapid internalization. Preincubation with 10-6 M SSR-149415 counteracted the internalization process. Moreover, recycling of internalized receptors was observed upon 10-6 M SSR-149415 treatment. Thus SSR-149415/[³H]SSR-149415 are unique tools for studying animal and human V1bR. [ABSTRACT FROM AUTHOR]

Various approaches might be employed in an effort to increase efficacy of the chemotherapeutic treatment of cancer. Recently, various modulators of anticancer therapy effectiveness have been studied. Antiproliferative effects of peripheral benzodiazepine receptor (PBR) ligands might be exploited to enhance cytotoxic effect of a chemotherapeutic drug towards cancer cells. In this work, we sought to enhance cytotoxic effect of etoposide (VP-16) by a PBR ligand, diazepam (DZ) in U-87 MG human glioma cells. Cytotoxicity of VP-16, DZ and their combinations was assessed by using the microculture MTT assay. Cell survival, effective concentrations (EC) and the onset of cytotoxic effect were determined. After 72 h of cultivation, survival of U-87 MG cells was reduced to 57 ± 7% in the presence of VP-16 at 12.5 µg/mL alone, whereas DZ at 10 [sup -4] mol/L alone caused 28 ± 6% reduction in cell survival. Coincubation of VP-16 at 12.5 µg/mL with DZ at 10[sup -4] mol/L led to a further decrease in cell survival to 45 ± 6%. Furthermore, DZ at 10[sup -4] mol/L significantly decreased effective concentrations, EC[sub 10], EC[sub 30] and EC[sub 50],of VP-16 and the dose-response curves were shifted to the left. Addition of DZ at 10[sup -4] mol/L to VP-16 also facilitated the onset of its cytotoxic effect. The same decrease in survival was thus achieved approximately 30 h earlier in comparison with VP-16 alone. However, DZ at 10[sup -9] mol/L failed both to exert any effect on glioma cells survival and enhance cytotoxic effect of VP-16. DZ at 10[sup -4] mol/L was capable of both reducing U-87 MG glioma cells survival when applied alone and also enhancing the cytotoxic effect of VP-16. No such observation was made for the lower concentrations of DZ. Potential implementation of diazepam in the antiglioma/anticancer armamentarium awaits further experimentation but phase I and phase II clinical trials could be suggested. [ABSTRACT FROM AUTHOR]

Vasopressin V2-receptor antagonists are promising agents for the use in water-retaining diseases. Potential renal mechanisms of action include effects on water permeability in the collecting duct as well as on electrolyte transport in the thick ascending limb of Henle's loop (TALH). To elucidate sites of action upstream of the distal tubule, e.g., in TALH, micropuncture experiments were performed in anesthetized rats during application of the V2-receptor antagonist SR 121463B. As compared to vehicle-treated rats, SR 121463B (0.3 mg/kg i.v.) did not affect mean arterial blood pressure (means ± SEM, n=10 rats per group: 108±4 mmHg vs. 107±4 mmHg), whole kidney GFR (1.1±0.1 ml/min vs. 1.1±0.1 ml/min), or whole kidney fractional reabsorption (FR) of potassium (66±5% vs. 68±4%). The drug, however, reduced whole kidney FR of fluid (92±1% vs. 99±1%), increased urinary flow rate (84±7 µl/min vs. 8±1 µl/min) and electrolyte-free-water clearance (72±8 µl/min vs. 2±1 µl/min), and reduced urinary osmolality (148±11 mosmol/kg vs. 1200±185 mosmol/kg). This pronounced diuretic response was associated with a minor reduction in whole kidney FR of sodium (99.6±0.1% vs. 99.9±0.1%) and chloride (98.3±0.2% vs. 98.9±0.1%). As compared to vehicle application, SR 121463B did not significantly alter single nephron GFR (39±2 nl/min vs. 39±1 nl/min, n=22 and 23 nephrons, respectively) or the FR up to the early distal tubule of fluid (76±2% vs. 76±1%), sodium (92±1% vs. 93±1%), potassium (91±1% vs. 90±1%) or chloride (90±1% vs. 91±1%). Together these data indicate a predominant aquaretic effect of SR 121463B which is located downstream of the early distal tubule. This response is compatible with blockade of vasopressin V2-receptors in the collecting duct and, as directly demonstrated by immunohistochemistry, subsequent retrieval of aquaporin-2 from apical plasma membrane, which inhibits water permeability and transport. [ABSTRACT FROM AUTHOR]

A potent non-peptide vasopressin (AVP) antagonist, SR-49059, displaying high stability and selective affinity for the V[sub 1a] AVP receptor subtype, has recently been described. The objective of this study was to assess the binding properties and the penetrability of this compound in the rat brain. Both in vitro and in vivo binding autoradiography experiments were performed. In all studies, the liver was used as a reference V[sub 1a] tissue. In vitro labelling of rat brain sections with [[sup 3] H]SR-49059 was similar to that previously detected with [[sup 3] H]AVP, which confirms that the majority of central AVP binding sites are V[sub 1a] sites similar to peripheral V[sub 1a] receptors. As expected, intense specific labelling occurred mainly in the lateral septum, the fundus striatum, the hypothalamic stigmoid nucleus and the area postrema-nucleus of the solitary tract complex. In vivo binding autoradiography showed that [[sup 3] H]SR-49059 injected intravenously did not enter the brain parenchyma. Specific labelling was however clearly detectable in brain regions with permeable hematoencephalic barrier, the choroid plexus and other circumventricular organs expressing V[sub 1a] receptors, namely the subfornical organ, the pineal gland and the area postrema. The specificity of [[sup 3] H]SR-49059 binding in the latter structures was confirmed by the fact that labelling was prevented by pretreatment of animals with high doses of nonradioactive SR-49059. In conclusion, our study shows that [[sup 3] H]SR-49059 is a suitable probe to investigate V[sub 1a] receptors in the rat brain. We also demonstrate that although this compound is not able to enter the brain tissue from the peripheral circulation, it does bind specifically to regions devoid of blood-brain barrier and known to be involved in autonomic regulations. [ABSTRACT FROM AUTHOR]

Although vasopressin V 1B receptor (V 1B R) mRNA has been detected in the kidney, the precise renal localization as well as pharmacological and physiological properties of this receptor remain unknown. Using the selective V 1B agonist d[Leu 4 , Lys 8 ]VP, either fluorescent or radioactive, we showed that V 1B R is mainly present in principal cells of the inner medullary collecting duct (IMCD) in the male rat kidney. Protein and mRNA expression of V 1B R were very low compared with the V 2 receptor (V 2 R). On the microdissected IMCD, d[Leu 4 , Lys 8 ]VP had no effect on cAMP production but induced a dose-dependent and saturable intracellular Ca 2+ concentration increase mobilization with an EC 50 value in the nanomolar range. This effect involved both intracellular Ca 2+ mobilization and extracellular Ca 2+ influx. The selective V 1B antagonist SSR149415 strongly reduced the ability of vasopressin to increase intracellular Ca 2+ concentration but also cAMP, suggesting a cooperation between V 1B R and V 2 R in IMCD cells expressing both receptors. This cooperation arises from a cross talk between second messenger cascade involving PKC rather than receptor heterodimerization, as supported by potentiation of arginine vasopressin-stimulated cAMP production in human embryonic kidney-293 cells coexpressing the two receptor isoforms and negative results obtained by bioluminescence resonance energy transfer experiments. In vivo, only acute administration of high doses of V 1B agonist triggered significant diuretic effects, in contrast with injection of selective V 2 agonist. This study brings new data on the localization and signaling pathways of V 1B R in the kidney, highlights a cross talk between V 1B R and V 2 R in the IMCD, and suggests that V 1B R may counterbalance in some pathophysiological conditions the antidiuretic effect triggered by V 2 R activation. NEW & NOTEWORTHY Although V 1B R mRNA has been detected in the kidney, the precise renal localization as well as pharmacological and physiological properties of this receptor remain unknown. Using original pharmaceutical tools, this study brings new data on the localization and signaling pathways of V 1B R, highlights a cross talk between V 1B R and V 2 receptor (V 2 R) in the inner medullary collecting duct, and suggests that V 1B R may counterbalance in some pathophysiological conditions the antidiuretic effect triggered by V 2 R activation.

Vasopressin and its receptors modulate several gut functions, but their role in intestinal inflammation is unknown. Our aims were to determine 1) the localization of V1b receptors in human and rodent colon, 2) the role of vasopressin and V1b receptors in experimental colitis using two approaches: V1b⁻(/)⁻ mice and a selective V1b receptor antagonist, SSR149415, and 3) the mechanisms involved. V1b receptors were localized in normal and inflamed colon from humans and rats. Experimental colitis was induced in rats and mice and some groups were treated before or after colitis induction with oral SSR149415 (3-30 mg/kg). Other groups of mice were submitted to dehydration to increase vasopressin plasma levels, prior to colitis induction. Body weight, damage scores, MPO, and TNF-α tissue levels were determined. Finally, colonic segments of wild-type (WT) and V1b⁻(/)⁻ mice were mounted in Ussing chambers and paracellular permeability in response to vasopressin was studied. V1b receptors were expressed in enterocytes and ganglia cells of the enteric nervous system of human and rat intestine. Expression levels were independent from inflammatory status. Colitis was less severe in rodents treated by either preventive or curative SSR149415 and in V1b⁻(/)⁻ mice. 2,4,6-Trinitrobenzene sulfonic acid induced a strong mortality in dehydrated animals that was reversed by preventive SSR149415 or mast cell stabilizer. Vasopressin significantly increased paracellular permeability in WT, but not in V1b⁻(/)⁻ mice. Preincubation of colon tissues with SSR149415 abolished the vasopressin effect. Similarly, vasopressin had no effect in colonic preparations from WT mice pretreated with mast cell stabilizers. Vasopressin, through V1b receptor interaction, has proinflammatory properties linked to mast cell activation and downstream alterations of the colonic epithelial barrier. These findings underline the potential interest of V1b receptor blockers in gut inflammatory diseases.

The hypothesis that vasopressin (VP) becomes the main mediator of pituitary corticotroph responsiveness during chronic hypothalamic pituitary adrenal (HPA) axis activation was tested by examining the effect of pharmacologic VP receptor blockade on the adrenocorticotropic hormone (ACTH) and corticosterone responses of 14-day repeatedly restrained rats. In spite of the increased vasopressinergic activity, repeatedly restrained rats showed lower ACTH and corticosterone responses to 10 min white noise compared with handled controls. These responses were unchanged by injection of the nonpeptide-selective V1b receptor antagonist SSR149415 i.v., 1 h before noise application. In contrast to noise stress, plasma ACTH responses to i.p. hypertonic saline injection were enhanced in the repeatedly restrained rats compared with handled controls, but responses were also unaffected by SSR149415 administered orally, daily 1 h before restraint. Since SSR149415 effectiveness was low, we used minipump infusion of the peptide V1 receptor antagonist, dGly[Phaa1,D-tyr(et), Lys, Arg]VP (V1-Ant) for 14 days, which effectively blocked ACTH responses to exogenous VP. Chronic V1-Ant infusion reduced plasma ACTH responses to i.p. hypertonic saline in handled controls but not in repeatedly restrained rats. These data suggest that the increased vasopressinergic activity characteristic of chronic stress plays roles other than mediating the hypersensitivity of the HPA axis to a novel stress.

[(3)H]SSR-149415 is the first tritiated nonpeptide vasopressin V(1b) receptor (V(1b)R) antagonist ligand. It was used for studying rodent (mouse, rat, hamster) and human V(1b)R from native or recombinant origin. Moreover, a close comparison between the human and the mouse V(1b)R was performed using SSR-149415/[(3)H]SSR-149415 in binding and functional studies in vitro. [(3)H]SSR-149415 binding was time-dependent, reversible, and saturable. Scatchard plot analysis gave a single class of high-affinity binding sites with apparent equilibrium dissociation constant (K(d)) approximately 1 nM and maximum binding density (B(max)) values from 7,000 to 300,000 sites/cell according to the cell line. In competition experiments, [(3)H]SSR-149415 binding was stereospecific and dose-dependently displaced by reference peptide and nonpeptide arginine vasopressin (AVP)/OT ligands following a V(1b) rank order of affinity: SSR-149415 = AVP > dCha > dPen > dPal > dDavp > SSR-126768A > SR-49059 > SSR-149424 > OT > SR-121463B. Species differences between human, rat, mouse, and hamster V(1b)R were observed. Autoradiography studies with [(3)H]SSR-149415 on rat and human pituitary showed intense specific labeling confined to corticotroph cells and absence of labeling in the other tissues examined. SSR-149415 potently and stereospecifically antagonized the AVP-induced inositol phosphate production and intracellular Ca(2+) increase (EC(50) from 1.83 to 3.05 nM) in recombinant cell lines expressing either the mouse or the human V(1b)R. AVP (10(-7) M) exposure of AtT20 cells expressing mouse or human EGFP-tagged V(1b)R induced their rapid internalization. Preincubation with 10(-6) M SSR-149415 counteracted the internalization process. Moreover, recycling of internalized receptors was observed upon 10(-6) M SSR-149415 treatment. Thus SSR-149415/[(3)H]SSR-149415 are unique tools for studying animal and human V(1b)R.

Mutations in PKD1 are associated with autosomal dominant polycystic kidney disease. Studies in mouse models suggest that the vasopressin (AVP) V2 receptor (V2R) pathway is involved in renal cyst progression, but potential changes before cystogenesis are unknown. This study used a noncystic mouse model to investigate the effect of Pkd1 haploinsufficiency on water handling and AVP signaling in the collecting duct (CD). In comparison with wild-type littermates, Pkd1(+/-) mice showed inappropriate antidiuresis with higher urine osmolality and lower plasma osmolality at baseline, despite similar renal function and water intake. The Pkd1(+/-) mice had a decreased aquaretic response to both a water load and a selective V2R antagonist, despite similar V2R distribution and affinity. They showed an inappropriate expression of AVP in brain, irrespective of the hypo-osmolality. The cAMP levels in kidney and urine were unchanged, as were the mRNA levels of aquaporin-2 (AQP2), V2R, and cAMP-dependent mediators in kidney. However, the (Ser256) phosphorylated AQP2 was upregulated in Pkd1(+/-) kidneys, with AQP2 recruitment to the apical plasma membrane of CD principal cells. The basal intracellular Ca(2+) concentration was significantly lower in isolated Pkd1(+/-) CD, with downregulated phosphorylated extracellular signal-regulated kinase 1/2 and decreased RhoA activity. Thus, in absence of cystic changes, reduced Pkd1 gene dosage is associated with a syndrome of inappropriate antidiuresis (positive water balance) reflecting decreased intracellular Ca(2+) concentration, decreased activity of RhoA, recruitment of AQP2 in the CD, and inappropriate expression of AVP in the brain. These data give new insights in the potential roles of polycystin-1 in the AVP and Ca(2+) signaling and the trafficking of AQP2 in the CD.

Vasopressin (AVP) and corticotropin-releasing factor (CRF) are key mediators in the organism's neuro-adaptive response to stress. Through pituitary and central vasopressin V(1b) receptors, AVP participates in the control of the hypothalamic-pituitary-adrenal axis (HPA) and is involved in various emotional processes. SSR149415 is the first selective, orally active vasopressin V(1b) receptor antagonist yet described. It is a competitive antagonist with nanomolar affinity for animal and human V(1b) receptors and displays a highly selective profile with regard to a large number of receptors or enzymes. In vitro, SSR149415 potently antagonizes functional cellular events associated with V(1b) receptor activation by AVP, such as intracellular Ca(2+) increase or proliferation in various cell systems. Pharmacological studies, performed by measuring ACTH secretion induced by various stimulants such as hormones (AVP or AVP + CRF) or physical stress (restraint or forced swimming stress and dehydration) in conscious rats or mice, confirm the antagonist profile of SSR149415 and its efficacy in normalizing ACTH secretion in vivo. SSR149415 is active by the oral route, at doses from 3 mg/kg, it potentiates CRF effect and displays a long-lasting oral effect in the different models. At 10 mg/kg p.o. its duration of action is longer than 4 h. This molecule also decreases anxiety and exerts marked antidepressant-like activity in several predictive animal models. The anxiolytic effects of SSR149415 have been demonstrated in various Generalized Anxiety Disorders (GAD) models (four-plate, punished drinking, elevated plus-maze, light dark, mouse defense test battery, fear-potentiated startle and social interaction tests). It is as effective as the benzodiazepine diazepam in the acute stress exposure test. SSR149415 has similar efficacy to the reference antidepressant drug, fluoxetine, in acute (forced-swimming) and chronic (chronic mild stress and subordination stress) situations in rodents. SSR149415 also reduces offensive aggression in the resident-intruder model in mice and hamsters. Depending on the model, the minimal effective doses are in the range of 1-10 mg/kg i.p. or 3-10 mg/kg p.o. SSR149415 is devoid of adverse effects on motor activity, sedation, memory or cognitive functions and produces no tachyphylaxis when administered repeatedly. It is well-tolerated in animals and humans and exhibits an adequate ADME profile. Thus, SSR149415 is a new dual anxiolytic/antidepressant compound, which appears to be free of the known side effects of classical anxiolytic/antidepressant drugs. Clinical trials are in progress, they will hopefully demonstrate its therapeutical potential for treating stress-related disorders.

By activating three distinct vasopressin receptor isoforms called V1a-R, V1b-R (V3-R) and V2-R, vasopressin (VP) mediates a wide number of biological effects in mammals and may be involved in several pathological states. Up to now only specific V1a and V2 receptor agonists and antagonists have been successfully designed. The role of the V1b-R still remains partially unknown, due to the lack of selective V1b-R ligands and orally-active molecules, which are crucial tools for investigating the central and peripheral functions or pathological disorders associated with this receptor. In this review, we report the biological and pharmacological properties of the first two specific V1b-R ligands: d[Cha4] AVP, a high affinity V1b-R agonist and SSR149415, a potent orally-active V1b-R antagonist with good selectivity with respect to other VP/OT receptor isoforms and able to control ACTH secretion in vitro and in vivo. Indeed, these molecules constitute invaluable tools for exploring the central and peripheral roles of VP mediated via V1b receptors. Interestingly, SSR149415 displays potent anxiolytic and antidepressant-like activities, indicating that this new class of drugs has a promising therapeutical potential in the treatment of stress-related disorders, anxiety and depression.

(2S,4R)-1-[5-Chloro-1-[(2,4-dimethoxyphenyl)sulfonyl]-3-(2-methoxy-phenyl)-2-oxo-2,3-dihydro-1H-indol-3-yl]-4-hydroxy-N,N-dimethyl-2-pyrrolidine carboxamide (SSR149415), the first selective, nonpeptide vasopressin V1b receptor antagonist yet described, has been characterized in vitro and in vivo. SSR149415 showed competitive nanomolar affinity for animal and human V1b receptors and exhibited much lower affinity for rat and human V1a, V2, and oxytocin receptors. Moreover, this compound did not interact with a large number of other receptors, enzymes, or ion channels. In vitro, SSR149415 behaved as a full antagonist and potently inhibited arginine vasopressin (AVP)-induced Ca2+ increase in Chinese hamster ovary cells expressing rat or human V1b receptors. The in vivo activity of SSR149415 has been studied in several models of elevated corticotropin secretion in conscious rats. SSR149415 inhibited exogenous AVP-induced increase in plasma corticotropin, from 3 mg/kg i.p. and 10 mg/kg p.o. upwards. Similarly, this compound antagonized AVP-potentiated corticotropin release provoked by exogenous corticoliberin at 3 mg/kg p.o. The effect lasted for more than 4 h at 10 mg/kg p.o. showing a long-lasting oral effect. SSR149415 (10 mg/kg p.o.) also blocked corticotropin secretion induced by endogenous AVP increase subsequent to body water loss. Moreover, 10 mg/kg i.p SSR149415 inhibited plasma corticotropin elevation after restraint-stress in rats by 50%. In the four-plate test, a mouse model of anxiety, SSR149415 (3 mg/kg p.o. upwards) displayed anxiolytic-like activity after acute and 7-day repeated administrations. Thus, SSR149415 is a potent, selective, and orally active V1b receptor antagonist. It represents a unique tool for exploring the functional role of V1b receptors and deserves to be clinically investigated in the field of stress and anxiety.