[PROSPECTS OF USING ADENOSINE, OPIOID AND BRADYKININ RECEPTOR AGONISTS FOR PHARMACOLOGICAL IMITATION OF HEART POSTCONDITIONING PHENOMENON.]

It is known that agonists of adenosine, opioid, and bradykinin receptors mimic the phenomenon of ischemic postconditioning. There is no commonly accepted notion of what adenosine receptor subtypes must be activated to increase cardiac resistance to reperfusion injury. Intravenous infusion of adenosine or intracoronary administration of adenosine produce infarct-limiting effect and contribute to a more complete restoration of coronary blood flow after recanalization of the infarct-related coronary artery. It was confirmed that opioids mimic the phenomenon of postconditioning. According to obtained data, the most promising compounds for the prevention of reperfusion injury of the heart are κ(1)- and δ(2)-opioid receptor agonists, as they produce the infarct-limiting effect, while not reducing the arterial pressure.

Trancriptomic approach reveals the molecular diversity of Hottentotta conspersus (Buthidae) venom.

Scorpion venom consists of a complex mixture of molecules including biologically active compounds. Because of their high potency and selectivity, toxins have medical applicability. In the last decades, scorpion toxins have thus gained considerable interest among scientist in the fields of pharmacology, biophysics and neurobiology. Identification of scorpion venom peptides and toxins can be achieved based on transcriptome approaches. We constructed the first cDNA library and Expressed Sequence Tag (EST) study to explore the transcriptomic composition of the telson from the southern African scorpion Hottentotta conspersus, belonging to the family Buthidae. We obtained 21 new venom-related sequences (8 contigs and 16 singlets) from a total of 98 ESTs analyzed, including putative neurotoxins (chloride, potassium, sodium and calcium channel toxins), bradykinin-potentiating peptides and other venom peptides without established function. These novel toxin-related sequences might serve as basis for further research both of pharmaceutical and phylogenetic nature.

Novel kinin B1 receptor agonists with improved pharmacological profiles.

There is some evidence to suggest that inducible kinin B1 receptors (B1R) may play beneficial and protecting roles in cardiovascular-related pathologies such as hypertension, diabetes, and ischemic organ diseases. Peptide B1R agonists bearing optimized pharmacological features (high potency, selectivity and stability toward proteolysis) hold promise as valuable therapeutic agents in the treatment of these diseases. In the present study, we used solid-phase methodology to synthesize a series of novel peptide analogues based on the sequence of Sar[dPhe(8)]desArg(9)-bradykinin, a relatively stable peptide agonist with moderate affinity for the human B1R. We evaluated the pharmacological properties of these peptides using (1) in vitro competitive binding experiments on recombinant human B1R and B2R (for index of selectivity determination) in transiently transfected human embryonic kidney 293 cells (HEK-293T cells), (2) ex vivo vasomotor assays on isolated human umbilical veins expressing endogenous human B1R, and (3) in vivo blood pressure tests using anesthetized lipopolysaccharide-immunostimulated rabbits. Key chemical modifications at the N-terminus, the positions 3 and 5 on Sar[dPhe(8)]desArg(9)-bradykinin led to potent analogues. For example, peptides 18 (SarLys[Hyp(3),Cha(5), dPhe(8)]desArg(9)-bradykinin) and 20 (SarLys[Hyp(3),Igl(5), dPhe(8)]desArg(9)-bradykinin) outperformed the parental molecule in terms of affinity, functional potency and duration of action in vitro and in vivo. These selective agonists should be valuable in future animal and human studies to investigate the potential benefits of B1R activation.

TNF-alpha and IL-1beta mediate inflammatory hypernociception in mice triggered by B1 but not B2 kinin receptor.

Kinin receptors are involved in the genesis of inflammatory pain. However, there is controversy concerning the mechanism by which B(1) and B(2) kinin receptors mediate inflammatory hypernociception. In the present study, the role of these receptors on inflammatory hypernociception in mice was addressed. Mechanical hypernociception was detected with an electronic pressure meter paw test in mice and cytokines were measured by ELISA. It was observed that in naïve mice a B(2) (d-Arg-Hyp(3), d-Phe(7)-bradykinin) but not a B(1) kinin receptor antagonist (des-Arg(9)-[Leu(8)]-bradykinin, DALBK) inhibited bradykinin- and carrageenin-induced hypernociception. Bradykinin-induced hypernociception was inhibited by indomethacin (5 mg/kg) and guanethidine (30 mg/kg), while not affected by IL-1ra (10 mg/kg) or antibody against keratinocyte-derived chemokine (KC/CXCL-1, 500 ng/paw) or in TNFR1 knockout mice. By contrast, in previously lipopolysaccharide (LPS)-primed mouse paw, B(1) but not B(2) kinin receptor antagonist inhibited bradykinin hypernociception. Furthermore, B(1) kinin receptor agonist induced mechanical hypernociception in LPS-primed mice, which was inhibited by indomethacin, guanethidine, antiserum against TNF-alpha or IL-1ra. This was corroborated by the induction of TNF-alpha and IL-1beta release by B(1) kinin receptor agonist in LPS-primed mouse paws. Moreover, B(1) but not B(2) kinin receptor antagonist inhibited carrageenin-induced hypernociception, and TNF-alpha and IL-1beta release as well, in LPS-primed mice. These results suggest that in naïve mice the B(2) kinin receptor mediates inflammatory hypernociception dependent on prostanoids and sympathetic amines, through a cytokine-independent mechanism. On the other hand, in LPS-primed mice, the B(1) kinin receptor mediates hypernociception by a mechanism dependent on TNF-alpha and IL-1beta, which could stimulate prostanoid and sympathetic amine production.

Multifunctional effects of bradykinin on glial cells in relation to potential anti-inflammatory effects.

Kinins have been reported to be produced and act at the site of injury and inflammation. Despite many reports that they are likely to initiate a particular cascade of inflammatory events, bradykinin (BK) has anti-inflammatory effects in the brain mediated by glial cells. In the present review, we have attempted to describe the complex responses and immediate reaction of glial cells to BK. Glial cells express BK receptors and induce Ca(2+)-dependent signal cascades. Among them, production of prostaglandin E(2) (PGE(2)), via B(1) receptors in primary cultured microglia, has a negative feedback effect on lipopolysaccharide (LPS)-induced release of tumor necrosis factor-alpha (TNF-alpha) via increasing intracellular cyclic adenosine monophosphate (cAMP). In addition, BK up-regulates the production of neurotrophic factors such as nerve growth factor (NGF) via B(2) receptors in astrocytes. These results suggest that BK may have anti-inflammatory and neuroprotective effects in the brain through multiple functions on glial cells. These observations may help to understand the paradox on the role of kinins in the central nervous system and may be useful for therapeutic strategy.

Systemic inhibition of the angiotensin-converting enzyme limits lipopolysaccharide-induced lung neutrophil recruitment through both bradykinin and angiotensin II-regulated pathways.

Recruitment of neutrophils to the lung is a sentinel event in acute lung inflammation. Identifying mechanisms that regulate neutrophil recruitment to the lung may result in strategies to limit lung damage and improve clinical outcomes. Recently, the renin angiotensin system (RAS) has been shown to regulate neutrophil influx in acute inflammatory models of cardiac, neurologic, and gastrointestinal disease. As a role for the RAS in LPS-induced acute lung inflammation has not been described, we undertook this study to examine the possibility that the RAS regulates neutrophil recruitment to the lung after LPS exposure. Pretreatment of mice with the angiotensin-converting enzyme (ACE) inhibitor enalapril, but not the anti-hypertensive hydralazine, decreased pulmonary neutrophil recruitment after exposure to LPS. We hypothesize that inhibition of LPS-induced neutrophil accumulation to the lung with enalapril occurred through both an increase in bradykinin, and a decrease in angiotensin II (ATII), mediated signaling. Bradykinin receptor blockade reversed the inhibitory effect of enalapril on neutrophil recruitment. Similarly, pretreatment with bradykinin receptor agonists inhibited IL-8-induced neutrophil chemotaxis and LPS-induced neutrophil recruitment to the lung. Inhibition of ATII-mediated signaling, with the ATII receptor 1a inhibitor losartan, decreased LPS-induced pulmonary neutrophil recruitment, and this was suggested to occur through decreased PAI-1 levels. LPS-induced PAI-1 levels were diminished in animals pretreated with losartan and in those deficient for the ATII receptor 1a. Taken together, these results suggest that ACE regulates LPS-induced pulmonary neutrophil recruitment via modulation of both bradykinin- and ATII-mediated pathways, each regulating neutrophil recruitment by separate, but distinct, mechanisms.

Pharmacologic targets and prototype therapeutics in the kallikrein-kinin system: bradykinin receptor agonists or antagonists.

The kallikrein-kinin system (KKS) is a complex system produced in various organs. This system includes kininogen (precursor for kinin), kallikreins, and pharmacologically active bradykinin (BK), which is considered to be proinflammatory and/or cardioprotective. It is a proinflammatory polypeptide that is involved in many pathological conditions and can cause pain, inflammation, increased vascular permeability, vasodilation, contraction of various smooth muscles, as well as cell proliferation. On the other hand, it has been shown that BK has cardioprotective effects, as all components of KKS are located in the cardiac muscles. Numerous observations have indicated that decreased activity of this system may lead to cardiovascular diseases, such as hypertension, cardiac failure, and myocardial infarction. BK acts on two receptors, B1 and B2, which are linked physiologically through their natural stimuli and their common participation in a variety of inflammatory responses. Recently, numerous BK antagonists have been developed in order to treat several diseases that are due to excessive BK formation. Although BK has many beneficial effects, it has been recognized to have some undesirable effects that can be reversed with BK antagonists. In addition, products of this system have multiple interactions with other important metabolic pathways, such as the renin-angiotensin system.

Targeting kinin receptors for the treatment of neurological diseases.

Kinins (bradykinin, kallidin and their active metabolites) are peptide autacoids with established functions in cardiovascular homeostasis, contraction and relaxation of smooth muscles, inflammation and nociception. They are believed to play a role in disease states like asthma, allergies, rheumatoid arthritis, cancer, diabetes, endotoxic and pancreatic shock, and to contribute to the therapeutic effects of ACE inhibitors in cardiovascular diseases. Although kinins are also neuromediators in the central nervous system, their involvement in neurological diseases has not been intensively investigated thus far. This review analyzes the potential of central kinin receptors as therapeutic targets for neurological disorders. Initial data highlight potential roles for B(1) receptor antagonists as antiepileptic agents, and for B(2) receptor antagonists (and/or B(1) agonists) in the treatment of stroke. Functional B(1) receptors located on T-lymphocytes and on the blood brain-barrier are also putative targets for the management of multiple sclerosis. However, successful elucidation of the therapeutic value of these new pharmacological approaches will require refinement of our knowledge on the physiology and cellular localization of central kinin receptors.

Therapeutic prospects for bradykinin receptor agonists in the treatment of cardiovascular diseases.

Kinins are located in the vascular smooth muscle and the heart, and are the most potent biologically active polypeptides. Pharmacological studies of cardiovascular disorders, including hypertension, cardiac failure, ischemia, myocardial infarction and left ventricular hypertrophy, indicate that reduced activity of the local kallikrein-kinin system (KKS) may be instrumental in the induction of these disorders. The ability of kallikrein gene delivery and bradykinin (BK) B2 receptor agonists to produce a wide spectrum of beneficial effects make them excellent candidate therapies for the treatment of hypertension, and cardiovascular and renal diseases. In addition, strategies that activate kinin receptors may be applicable to the treatment of cardiovascular and renal disorders. However, one major challenge of this approach is the unanswered question of whether there is a sufficiently safe therapeutic index between the potential cardioprotective and pro-inflammatory effects following administration of BK B2 receptor agonists.

Identification and characterisation of GPR100 as a novel human G-protein-coupled bradykinin receptor.

G-protein-coupled receptor 100 (GPR100) was discovered by searching the human genome database for novel G-protein-coupled peptide receptors. Full-length GPR100 was amplified from a cDNA library of the neuroendocrine cell line BON, which is derived from a human pancreas carcinoid. The open-reading frame, present on a single exon, coded for a protein of 374 amino acids with highest sequence identity (43%) to the human orphan somatostatin- and angiotensin-like peptide receptor. The analysis of chromosomal localisation mapped the GPR100 gene to chromosome 1q21.2-q21.3. The stable expression of GPR100 in Chinese hamster ovary cells together with aequorin as calcium sensor and the promiscuous G-protein subunit alpha16 as signal transducer revealed bradykinin and kallidin as effectors to elicit a calcium response. Dose-response curves yielded EC50 values for both ligands in the low nanomolar range, while the respective analogues without arginine at the carboxy-terminus were inactive. Calcium mobilisation was inhibited by the phospholipase C blocker U73122, but not by pertussis toxin, suggesting the involvement of the G-protein subunit alphaq and not alphai or alphao in signal transduction. In line with the main function of kinins as peripheral hormones, we found that GPR100 was expressed predominantly in tissues like pancreas, heart, skeletal muscle, salivary gland, bladder, kidney, liver, placenta, stomach, jejunum, thyroid gland, ovary, and bone marrow, but smaller amounts were also detected in the brain and in cell lines derived from tumours of various origins.

Bradykinin receptor subtype 1 expression and function in prostate cancer.

Kinins exert multiple pathophysiological functions, including vascular permeability and mitogenesis, by activating their cognate receptors, bradykinin subtype 1 receptor (B1R) and bradykinin subtype 2 receptor (B2R), which belong to the superfamily of G protein-coupled receptors. Tissue-specific expression pattern or contribution of the individual kinin receptors to pathological prostate cell growth is not known. We report here the differential expression of B1R and B2R in human benign and malignant prostate specimens. Whereas B2R is ubiquitously expressed, the B1R is detected only in prostatic intraepithelial neoplasia and malignant lesions and not in benign prostate tissues. Using androgen-insensitive prostate cancer PC3 cells, we show that specific stimulation of endogenous B1R promotes cell growth, migration, and invasion. These findings identify B1R as an early marker for pathological growth of the prostate and suggest its potential utility as a drug target effective for the treatment of prostate cancer.

Overexpression of bradykinin type 2 receptors on glioma cells enhances bradykinin-mediated blood-brain tumor barrier permeability increase.

Variations in the expression levels of bradykinin (BK) type 2 receptors (B2R) in different brain tumors may explain variable increases in BK-mediated blood-brain tumor barrier (BTB) permeability. This study investigated whether elevation of the B2R expression levels on glioma cells enhances BK-mediated BTB permeability increases. Stable transfectants of C6 rat glioma cells overexpressing B2R were established by transfection with recombinant vectors harboring rat B2R cDNA sequence. Elevated B2R expression levels in transfectants were confirmed by quantitative real-time PCR, Western blots, and [3H]-BK binding studies. BTB permeability was quantified with autoradiography and expressed as a unidirectional transport constant, Ki, for [14C]-alpha-aminoisobutyric acid (AIB: Mr 103), using a rat brain tumor model. Baseline Ki values in tumors overexpressing B2R were not significantly higher than in control tumors. Ki values after BK treatment in tumors overexpressing B2R, however, were significantly higher than in control tumors. Western blots confirmed that B2R expression levels in vivo in tumors overexpressing B2R remained higher than in control tumors. These results suggested that alteration of B2R expression levels on tumor cells could modulate BK-mediated BTB permeability. Therefore, B2R expression levels in human glioma could be used to analyze the treatment results of patients undergoing treatment involving BK-modulated BTB permeability.

Characterization of bradykinin receptors in canine cultured corneal epithelial cells: pharmacological and functional studies.

The pharmacological properties of bradykinin (BK) receptors were characterized in canine cultured corneal epithelial cells (CECs) using [(3)H]-BK as a radioligand. Analysis of binding isotherms gave an apparent equilibrium dissociation constant of 0.34 +/- 0.07 nM and a maximum receptor density of 179 +/- 23 fmol/mg protein. Neither a B(1) receptor-selective agonist (des-Arg(9)-BK) nor antagonist ([Leu(8), des-Arg(9)]-BK) significantly inhibited [(3)H]-BK binding to CECs, thus excluding the presence of B(1) receptors in canine CECs. The specific binding of [(3)H]-BK to CECs was inhibited by B(2) receptor-selective agonists (BK and kallidin) and antagonists (Hoe 140 and [D-Arg(0), Hyp(3), Thi(5,8), D-Phe(7)]-BK), with a best fit using a one-binding-site model. The order of potency for the inhibition of [(3)H]-BK binding was BK = Hoe 140 > kallidin > [D-Arg(0), Hyp(3), Thi(5,8), D-Phe(7)]-BK. Stimulation of CECs by BK produced a concentration-dependent accumulation of inositol phosphates (IP) and an initial transient peak of intracellular Ca(2+). B(2) receptor-selective antagonist ([D-Arg(0), Hyp(3), Thi(5,8), D-Phe(7)]-BK) significantly antagonized the BK-induced responses with dissociation constants of 6.0-6.1. Pretreatment of CECs with pertussis toxin (PTX) or cholera toxin did not alter the BK-induced IP accumulation. Incubation of CECs in the absence of external Ca(2+) led to a significant attenuation of the IP accumulation induced by BK. These results demonstrate that BK directly stimulates phospholipase C-mediated signal transduction through BK B(2) receptors via a PTX-insensitive G protein in canine CECs. This effect may function as the transducing mechanism for BK-mediated cellular responses.

Diabetes-induced changes in responsiveness of rat bladder and vas deferens to peptides in vitro: susceptibility to reversal by insulin.

Changes in responsiveness of the vas deferens and urinary bladder to bradykinin (BK) receptor agonists (Tyr8-BK and des-Arg9-BK), substance P (SP), and endothelin-1 (ET-1) were assessed 8 weeks after streptozotocin (STZ)-induced diabetes. Preparations from control or STZ-treated (60 mg/kg i.p.) male rats were tested for contractile and neurogenic twitch potentiating (TP, in VD only) effects of all four agonists (1 nM to 0.3 or 3 microM). In diabetic VD, contractile effects of Tyr8-BK, des-Arg9-BK, and SP were enhanced, but ET-1 effects were unchanged. In contrast, TP by des-Arg9-BK was unaffected, that by Tyr8-BK was decreased, and those by SP and ET-1 were increased. In diabetic UB, only contractions to des-Arg9-BK and SP were enhanced. Following insulin replacement (human, 1-3 U/day s.c.), starting 1 week after STZ, TP induced by Tyr8-BK and des-Arg9-BK in VD were further inhibited, but all other changes in both preparations were reversed at least partially. Insulin treatment of nondiabetic rats, however, also affected VD (but not UB) responsiveness, such that contractions to Tyr8-BK and TP by ET-1 were increased, but TP by Tyr8-BK was decreased. Thus, STZ-induced type I diabetes causes important alterations in responsiveness of non-vascular smooth muscle tissues of the rat to BK, SP, and ET-1. Long term insulin replacement, at doses normalising glycaemia, effectively reversed most changes in VD or UB responsiveness, but it is unclear if this is truly due to blocking of STZ-induced changes, since the treatment also affected responsiveness of nondiabetic tissues.

The role of migrating leukocytes in IL-1 beta-induced up-regulation of kinin B(1) receptors in rats.

1. The present study examines the role of migrating leukocytes in the ability of IL-1 beta to induce the functional up-regulation of B(1) receptors, as assessed by kinin B(1) agonist-induced oedema in the rat paw. 2. Pre-treatment with the PAF receptor antagonist WEB 2086 inhibited des-Arg(9)-BK-induced oedema in IL-1 beta-treated paws, while the LTB(4) receptor antagonist CP105696 had no effect. Des-Arg(9)-BK-induced paw oedema was also inhibited by pre-treatment with the selectin blocker fucoidin or by an anti-CD-18 monoclonal antibody. 3. I.d. injection of IL-1 beta produced a 5 - 10-fold increase of myeloperoxidase (MPO) activity in the rat paw. The increase in MPO activity was significantly inhibited by WEB 2086 (46 +/- 9%), fucoidin (68 +/- 5%) or the CD-18 antibody (84 +/- 3%). In contrast, i.d. injection of TNF alpha a dose known to upregulate the B(1) receptor functionally did not induce any significant increase in MPO activity. 4. Des-Arg(9)-BK alone had no effect in MPO activity but enhanced (by about 40%) the response induced by IL-1 beta, an effect prevented by the B(1) receptor antagonist des-Arg(9)-[Leu(8)]-BK. 5. The concentration of TNF-alpha was increased in the paws after i.d. injection of IL-1 beta. Pre-treatment with fucoidin, WEB 2086, anti-CD-18 or CP 105695, significantly reversed the local increases in TNF-alpha concentrations (80 +/- 2; 75 +/- 4, 73 +/- 3 and 40 +/- 2%), respectively. 6. Finally, IL-1 beta induced an increase of B(1) receptor mRNA levels in the rat paw, an effect which was prevented by fucoidin treatment. 7. Taken together, these results indicate that up-regulation of B(1) receptors in the rat paw following IL-1 beta seems to involve the local recruitment of neutrophils and subsequent local TNF-alpha production. The cross-talk between kinins, cytokines and leukocytes implicate B(1) receptors in chronic inflammatory diseases.

[30 years: Happy birthday, GPCR. The bradykinin B2 receptor: an alternative and antiproliferative pathway]. / 30 ans: happy birtday RCPG. Le récepteur B2 de la lbrady kinine: voie alternative et antiproliférative.

The bradykinin B2 receptor which belongs to the G-protein-coupled receptor (GPCR) family is known for its proliferative effects. The mitogenic signalling pathways used are "classical" pathways for GPCRs. Recent data from our laboratory show that bradykinin can also induce anti-mitogenic effects in proliferating cells using an "alternative" signal transduction pathway involving a protein tyrosine phosphatase. Besides this alternative signalling pathway for the B2 receptor, a novel concept of GPCR signalling is described.

The role of kinin B1 in the plasma extravasation of carrageenin-induced pleurisy.

The role of des-Arg9-bradykinin (des-Arg9-BK) and kinin B1 receptor in the plasma extravasation of rat carrageenin-induced pleurisy was investigated employing B1 receptor agonist and antagonists and kininogen-deficient rats. Expression of the B1 receptor mRNA in pleura was induced from 3 to 5 h after the injection of carrageenin into the pleural cavity of Sprague-Dawley rats. Exogenous injection of des-Arg9-BK into the pleural cavity provoked a significant increase in plasma extravasation in 5 h carrageenin-induced pleurisy, but not in 20 min kaolin-induced pleurisy. The level of immunoreactive des-Arg9-BK in the exudate of 5 h carrageenin-induced pleurisy was higher than that of bradykinin (BK). Administration of the B1 receptor antagonists, des-Arg9-[Leu8]-BK or des-Arg9-D-Arg-[Hyp3, Thi5, D-Tic7,Oic8]-BK significantly reduced the exudation rate. However, intrapleural administration of des-Arg9-BK to plasma kininogen-deficient. Brown Norway-Katholiek rats did not result in a further increase in the plasma extravasation. In conclusion, endogenously generated des-Arg9-BK could contribute to the plasma extravasation in carrageenin-induced pleurisy via mediation of the inducible B1 receptor.

Bradykinin increases Na(+)-K(+) pump activity in cultured guinea-pig tracheal smooth muscle cells.

1. The effect of bradykinin on the Na+-K+ pump of airway smooth muscle was investigated by measuring ouabain-sensitive (86)Rb(+) uptake in cultured guinea-pig tracheal smooth muscle cells. 2. Bradykinin induced a concentration-dependent increase in ouabain-sensitive (86)Rb(+) uptake, with an EC(50) of 3 nM (pD(2) = 8.50+/-0.10). Stimulation was not affected by indomethacin (1 microM) suggesting that it is not mediated by cycloxygenase products of arachidonic acid. 3. The B(1) receptor agonists Lys-des-Arg(9)-bradykinin and des-Arg(9)-bradykinin had no effect on ouabain-sensitive (86)Rb(+) uptake. In contrast, the B(1) and B(2) receptor agonist Lys-bradykinin induced a concentration-dependent increase in ouabain-sensitive (86)Rb(+) uptake with an EC(50) of 6 nM (pD(2) = 8.21 +/- 0.20). 4. The B(1) receptor antagonist des-Arg(10)-HOE 140 (1 microM) had no effect on bradykinin-stimulated ouabain-sensitive (86)Rb(+) uptake. The B(2) receptor antagonists HOE 140 and WIN 64338 antagonized bradykinin-stimulated ouabain-sensitive (86)Rb(+) uptake with pK(B) values (-log M) of 8.20 +/- 0.08 and 8.11 +/- 0.20 respectively. 5. Reducing extracellular Na+ from 146 mM to 11 mM caused a 53.5% decrease in basal ouabain-sensitive (86)Rb+ uptake and abolished bradykinin-induced uptake. Two inhibitors of the Na(+)-H(+) exchanger, methylisobutyl-amiloride (MIA; 1 - 100 microM) and ethylisopropyl-amiloride (EIPA; 0.1 - 10 microM), inhibited bradykinin-stimulated ouabain-sensitive (86)Rb(+) uptake without affecting basal uptake. 6. These results suggest that bradykinin increases Na+-K+ pump activity of guinea-pig tracheal smooth muscle via stimulation of B(2) receptors and activation of the Na+-H+ exchanger.

Syntheses of 1,4-benzothiazepines and 1,4-benzoxazepines via cyclizations of 1-[2-arylthio(oxy)ethyl]-5-benzotriazolyl-2-pyrrolidinones and 3-benzotriazolyl-2-[2-arylthio(oxy)ethyl]-1-isoindolinones.

1-[2-Arylthio(oxy)ethyl]-5-benzotriazolyl-2-pyrrolidinones 6a-e, 12 and 3-benzotriazolyl-2-[2-arylthio(oxy)ethyl]-1-isoindolinones 9a-f, 14 are readily available from reactions of benzotriazole (4), 2-(arylsulfanyl)ethylamines 3, or 2-phenoxyethylamine (11) with 2,5-dimethoxy-2,5-dihydrofuran (5) or 2-formylbenzoic acid (8). Lewis acid mediated cyclizations of 6 and 9 produced novel 1,4-benzothiazepines 7a-e and 10a-f, respectively. Cyclizations of 12 and 14 gave 1,4-benzoxazepines 13 and 15, respectively.

Structural characterization of lipopeptide agonists for the bradykinin B2 receptor.

The conformational features of Pam-Lys(0)-Arg(1)-Pro(2)-Pro(3)-Gly(4)-Phe(5)-Ser(6)-Pro(7)-Phe(8)-Arg(9)-OH (PKD) and Pam-Gly(-1)-Lys(0)-Arg(1)-Pro(2)-Pro(3)-Gly(4)-Phe(5)-Ser(6)-Pro(7)-Phe(8)-Arg(9)-OH (PGKD), the Pam-Lys and Pam-Gly-Lys analogues of bradykinin, have been determined by high-resolution NMR in a zwitterionic lipoid environment. Radical-induced relaxation of the (1)H NMR signals was used to probe the topological orientation of the peptides with respect to the zwitterionic lipid interface. The radical-induced relaxation and molecular dynamics (MD) data indicated that the palmitic acid and N-terminal amino acid residues embed into the micelles, while the rest of the polypeptide chain is closely associated with the water-micelle interface. Throughout the entire nuclear Overhauser effect restrained MD simulation, a nonideal type I beta-turn was observed in the C-terminus of PKD between residues 6 and 9, and a gamma-turn was observed in the C-terminus of PGKD between residues 6 and 7. Therefore, the additional glycine has a dramatic effect on the structural preferences of the biologically important C-terminus, an effect brought about by the interaction with the lipid environment. These structural features are correlated to the biological activity at the bradykinin B2 receptor.