Guinea Pig Tracheal Strips Research Articles

Discovery and Identification of Novel 5-Hydroxy-4H-benzo[1,4]oxazin-3-one Derivatives as Potent β2-Adrenoceptor Agonists through Structure-Based Design, Synthesis, and Biological Evaluation.

Although β2-agonists are crucial for treatment of chronic respiratory diseases, optimizing β2-agonistic activity and selectivity remains essential for achieving favorable therapeutic outcomes. A structure-based molecular design workflow was employed to discover a novel class of β2 agonists featuring a 5-hydroxy-4H-benzo[1,4]oxazin-3-one scaffold, which potently stimulated β2 adrenoceptors (β2-ARs). Screening for the β2-agonistic activity and selectivity led to the identification of compound A19 (EC50 = 3.7 pM), which functioned as a partial β2-agonist in HEK-293 cells containing endogenous β2-ARs. Compound A19 exhibited significant relaxant effects, rapid onset time (Ot50 = 2.14 min), and long duration of action (>12 h) on isolated guinea pig tracheal strips, as well as advantageous pharmacokinetic characteristics in vivo, rendering A19 suitable for inhalation administration. Moreover, A19 suppressed the upregulation of inflammatory cytokines and leukocytes and improved lung function in a rat model of COPD, thereby indicating that A19 is a potential β2 agonist candidate for further study.

8-Hydroxyquinolin-2(1H)-one analogues as potential β2-agonists: Design, synthesis and activity study.

β2-Agonists that bind to plasmalemmal β2-adrenoceptors causing cAMP accumulation are widely used as bronchodilators in chronic respiratory diseases. Here, we designed and synthesized a group of 8-hydroxyquinolin-2(1H)-one analogues and studied their β2-agonistic activities with a cellular cAMP assay. Compounds B05 and C08 were identified as potent (EC50<20 pM) and selective β2-agonists among the compounds tested. They behaved as partial β2-agonists in non-overexpressed HEK293cells, and possessed rapid smooth muscle relaxant actions and long duration of action in isolated guinea pig tracheal strip preparations. In summary, B05 and C08 are β2-agonists with potential applicability in chronic respiratory diseases.

Study on the mechanisms of the bronchodilator effects of Folium Eriobotryae and the selected active ingredient on isolated guinea pig tracheal strips

Context: Folium Eriobotryae (FE), the dry leaf of Eriobotrya japonica (Thunb.) Lindl. (Rosaceae), has been widely used to treat respiratory disorders.Objective: To examine the bronchodilatory activity of FE and the potential mechanisms involved.Materials and methods: The effects of ethyl acetate fraction of FE (EFE) (0.05–0.3 mg/mL) on the isolated tracheal strips, and ursolic acid (UA) (5–30 μg/mL) that was the main constituent of EFE, were tested in vitro. Meanwhile, acetylcholine (Ach) and histamine (His)-induced bronchospasm were conducted in vivo in guinea pig. Furthermore, mechanisms of relaxant effects of EFE and UA were evaluated in the absence and presence of specific inhibitors.Results: With in vitro studies, the contractile response evoked by Ach or His (EC50 = 0.21 and 0.16 mg/mL) was decreased by EFE, and UA caused a concentration-dependent relaxation precontracted by His (EC50 = 23.2 μg/mL). With in vivo studies, EFE strongly prolonged preconvulsive time similar to isoprenalin. The bronchodilator effects of EFE could be blocked by propranolol (1 μM), NG-nitro-l-arginine methyl ester (l-NAME) (100 μM) and 1H-[1,2,4]oxadiazolo[4,3-a]quinoxaline-1-one (ODQ) (1 μM). EFE also inhibited the contraction in Ca2+-free medium and produced rightward parallel displacement of CaCl2 curves. In addition, the relaxant effects of UA could only be blocked by l-NAME and ODQ.Discussion and conclusion: These results suggest that bronchodilator activities of EFE were related to activation of β-adrenoceptor and NO/cGMP pathway. Blockage of Ca2+ channels and inhibition of IP3R-mediated internal Ca2+ release were also involved. Additionally, UA produced relaxant effects by the NO/cGMP pathway.

Insight into the bronchodilator activity of Vitex negundo

Context: Vitex negundo Linn. (Verbenaceae) is traditionally used in hyperactive respiratory disorders.Objective: This study explored the mechanisms underlying the effectiveness of Vitex negundo in hyperactive respiratory disorders.Materials and methods: Crude extract of V. negundo leaves was obtained. For in vivo bronchodilatory activity in anesthetized rats, different doses (1, 3, 10, 30, and 50 mg/kg) of the crude extract of V. negundo (Vn.Cr) were tested. The underlying mechanisms were studied in isolated guinea pig tracheal strips, suspended in organ baths at 37 °C.Results: Intravenous doses of the crude extract of Vn.Cr showed dose-dependent bronchodilatory effect (9–50%) against carbachol (CCh; 100 µg/kg)-induced bronchoconstriction, similar to aminophylline. In isolated guinea-pig tracheal strips, Vn.Crrelaxed CCh (1 µM) and high K+ pre-contractions with respective EC50 values of 0.72 (0.48–1.10; n = 5) and 3.38 mg/mL (1.84–6.21; n = 4), similar to papaverine. Diltiazem also relaxed both contractions with more potency against high K+ pre-contraction (p < 0.05). Pre-incubation of the tracheal strips with Vn.Cr potentiated the isoprenaline inhibitory concentration response curves (CRCs), similar to papaverine.Discussion: The inhibitory effect against CCh and high K+ suggests involvement of phosphodiesterase (PDE) inhibitory pathway(s), in addition to an inhibitory effect on Ca++ entry. This finding was further strengthened when pre-treatment of the tracheal strips potentiated the isoprenaline CRCs.Conclusion: Results suggest Vn.Cr possesses a combination of papaverine-like PDE inhibitor and diltiazem-like Ca++ entry blocking constituents, which partly explain its bronchodilatory effect, thus validating its medicinal importance in asthma.

Antihistamine Effect of a Pure Bioactive Compound Isolated from Slug (Diplosolenodes occidentalis) Material.

Folklore claims of the therapeutic effect of garden slug (Diplosolenodes occidentalis) extract used to relieve bronchoconstriction in asthmatic individuals were never validated scientifically. The aim of this study was to isolate the pure bioactive compound from slug extract causing this effect. The crude ground material was prepared in ethanol and after filtration, separation by flash column chromatography method was done. The structure was elucidated by data from hydrogen and carbon nuclear magnetic resonance (NMR) profiles. The bioactive compound was assessed for dose-dependent response effects on guinea pig tracheal smooth muscle pre-contracted with histamine. Receptor specificity studies were done by using HTMT dimaleate (H1 agonist). The type of antagonism was also identified. The pure component isolated from garden slug material was identified by spectral studies as glyceryl trilinolenate (GT). It caused dose-dependent relaxation in guinea pig tracheal smooth muscle strips pre-contracted with histamine, it acted via H1 type receptors and showed non-competitive antagonism. Glyceryl trilinolenate produced dose-dependent relaxation in tracheal smooth muscle strips in the presence of the agonist histamine. Glyceryl trilinolenate displayed non-competitive antagonism at H1 receptors in the trachea. This agent was able to alleviate bronchoconstriction in individuals presenting with atopic asthma in rural agricultural areas in Jamaica (verbal communications). It is possible that GT can be useful therapeutically to produce tracheal smooth muscle relaxation in individuals presenting with atopic asthma.

The Calcium Channel Blocking and Phosphodiesterase Inhibitory Activities of the Extract of Andropogon muricatus Explains its Medicinal Use in Airways Disorders

The present work was carried out to provide a pharmacological base for the medicinal use of Andropogon muricatus in airways disorders, such as asthma. In isolated guinea-pig tracheal strips, the crude extract of Andropogon muricatus exhibited a non-specific relaxant effect against carbachol (1 µM) and high K⁺ precontractions, with EC₅₀ values of 0.10 (0.07-0.11) and 0.15 mg/mL (0.11-0.18), respectively, similar to papaverine, while verapamil was more potent against high K⁺. This suggests the involvement of a non-specific relaxant effect, mediated possibly through Ca⁺⁺ channel blockade and phosphodiesterase inhibition. The functional nature of the relaxant effect was further confirmed through indirect evidence when pretreatment of the tissues with the plant extract caused potentiation of the isoprenaline inhibitory response curves, similar to papaverine, while the effect of verapamil remained unchanged. These data indicate that the crude extract of Andropogon muricatus contains constituent(s) that mediate the tracheal relaxant effect, possibly through dual inhibition of Ca⁺⁺ channels and phosphodiesterase and provide pharmacological evidence for its medicinal use in airways disorders, particularly asthma.

Inhalation by Design: Novel Tertiary Amine Muscarinic M3 Receptor Antagonists with Slow Off-Rate Binding Kinetics for Inhaled Once-Daily Treatment of Chronic Obstructive Pulmonary Disease

A novel tertiary amine series of potent muscarinic M(3) receptor antagonists are described that exhibit potential as inhaled long-acting bronchodilators for the treatment of chronic obstructive pulmonary disease. Geminal dimethyl functionality present in this series of compounds confers very long dissociative half-life (slow off-rate) from the M(3) receptor that mediates very long-lasting smooth muscle relaxation in guinea pig tracheal strips. Optimization of pharmacokinetic properties was achieved by combining rapid oxidative clearance with targeted introduction of a phenolic moiety to secure rapid glucuronidation. Together, these attributes minimize systemic exposure following inhalation, mitigate potential drug-drug interactions, and reduce systemically mediated adverse events. Compound 47 (PF-3635659) is identified as a Phase II clinical candidate from this series with in vivo duration of action studies confirming its potential for once-daily use in humans.

Inhibitory effect of CP-96,345, a non-peptide neurokinin-1-receptor antagonist, on neurogenic responses of guinea-pig isolated airway smooth muscle.

The actions of CP-96,345, a non-peptide neurokinin-1 receptor antagonist, on the responses evoked by electrical-field stimulation or by acetylcholine and substance P in guinea-pig tracheal and bronchial muscle strips were examined. Electrical-field stimulation evoked a biphasic response, consisting of a cholinergically-mediated fast contraction followed by non-adrenergically-mediated relaxation in tracheal muscle and by a non-cholinergically-mediated slow contraction in bronchial muscle. CP-96,345 (1-10 microM) caused a concentration-dependent and non-selective inhibition in the amplitude of these neurogenic responses, where non-cholinergic contractions were more profoundly inhibited. Submaximal contractions of tracheal and bronchial muscles evoked by exogenous substance P (0.1-3 microM) were partly inhibited by CP-96,345 (1-10 microM), but acetylcholine-induced contractions were hardly inhibited. The results indicate that in guinea-pig isolated airway smooth muscle CP-96,345 can non-selectively inhibit neurogenic responses probably via neurokinin-1 receptor-dependent and independent mechanisms.

Inhibitory effects of sympathomimetic drugs on cholinergically mediated contractions of guinea-pig isolated tracheal muscle.

The experiments examine the actions of sympathomimetic drugs on the responses evoked by electrical field stimulation or by acetylcholine in guinea-pig tracheal strip chains. Electrical field stimulation evoked contractions which were cholinergically mediated, in the presence of guanethidine (10 microM) and indomethacin (2 microM). All the sympathomimetic drugs tested caused a concentration-dependent reduction in the height of these contractions. Inhibitory effects of isoprenaline and terbutaline were largely prevented by propranolol (2 microM) alone, whereas those of clonidine, oxymetazoline, lidamidine and WHR1370 were prevented by yohimbine alone (2 microM). Treatments with both propranolol and yohimbine were required to prevent the inhibitory effects of noradrenaline, adrenaline and dopamine. Contractions evoked by exogenous acetylcholine (0.1-3 microM) were also inhibited by all catecholamines and terbutaline, but not by clonidine, oxymetazoline, lidamidine and WHR1370. The inhibitory effects were antagonized by propranolol (2 microM) alone. The results suggest that in guinea-pig isolated tracheal muscle, sympathomimetic drugs can inhibit cholinergic neurotransmission not only by postjunctional beta 2-adrenoceptors but also by prejunctional alpha 2-adrenoceptors.

Anticholinergic activity of antipsychotic drugs in relation to their extrapyramidal effects.

Antipsychotic drugs were evaluated with two indices of anticholinergic activity, mydriasis in mice in vivo and antagonism of carbamylcholine-induced contractions of guinea-pig tracheal strips in vitro. The drugs from most to least potent as oral mydriatic agents were mepazine, clozapine, thioridazine, promazine and chlorpromazine. Trifluoperazine, pimozide and haloperidol were inactive. These results were consistent with the hypothesis that anticholinergic activity of antipsychotic drugs is inversely related to their propensity to produce extrapyramidal effects in man. In vitro results appeared to predict the incidence of extraphyramidal effects less accurately than in vivo results.

Blood pressure lowering, cardiovascular inhibitory and bronchodilatory actions of Achillea millefolium

Achillea millefolium Linn. (Asteraceae) is used in folk medicine for the treatment of overactive cardiovascular and respiratory ailments. This study describes its hypotensive, cardio-depressant, vasodilatory and bronchodilatory activities. The crude extract of Achillea millefolium (Am.Cr) caused a dose-dependent (1-100 mg/kg) fall in arterial blood pressure of rats under anaesthesia. In spontaneously beating guinea-pig atrial tissues, Am.Cr exhibited negative inotropic and chronotropic effects. In isolated rabbit aortic rings, Am.Cr at 0.3-10 mg/mL relaxed phenylephrine (PE, 1 µm) and high K(+) (80 mm)-induced contractions, as well as suppressed the PE (1 µm) control peaks obtained in Ca(++) -free medium, like that caused by verapamil. The vasodilator effect of Am.Cr was partially blocked by N(ω) -nitro-l-arginine methyl ester in endothelium intact preparations. In guinea-pig tracheal strips, Am.Cr inhibited carbachol (CCh, 1 µm) and K(+) -induced contractions. These results indicate that Achillea millefolium exhibits hypotensive, cardiovascular inhibitory and bronchodilatory effects, thus explaining its medicinal use in hyperactive cardiovascular and airway disorders, such as hypertension and asthma.

Inhalation by Design: Novel Ultra-Long-Acting β2-Adrenoreceptor Agonists for Inhaled Once-Daily Treatment of Asthma and Chronic Obstructive Pulmonary Disease That Utilize a Sulfonamide Agonist Headgroup

A novel series of potent and selective sulfonamide derived β(2)-adrenoreceptor agonists are described that exhibit potential as inhaled ultra-long-acting bronchodilators for the treatment of asthma and chronic obstructive pulmonary disease. Analogues from this series mediate very long-lasting smooth muscle relaxation in guinea pig tracheal strips. The sulfonamide agonist headgroup confers high levels of intrinsic crystallinity that could relate to the acidic sulfonamide motif supporting a zwitterionic form in the solid state. Optimization of pharmacokinetic properties was achieved through targeted introduction of a phenolic moiety to support rapid phase II clearance, thereby minimizing systemic exposure following inhalation and reducing systemically mediated adverse events. Compound 38 (PF-610355) is identified as a clinical candidate from this series, with in vivo duration of action studies confirming its potential for once-daily use in humans. Compound 38 is currently in advanced phase II clinical studies.

The 1.3 isoform of Na+–Ca2+ exchanger expressed in guinea pig tracheal smooth muscle is less sensitive to KB-R7943

The sodium-calcium exchanger (NCX) plays a major role in the regulation of cytosolic Ca(2+) in muscle cells. In this work, we performed force experiments to explore the role of NCX during contraction and relaxation of Cch-stimulated guinea pig tracheal smooth muscle strips. This tissue showed low sensitivity to NCX inhibitor KB-R7943 (IC50, 57 +/- 2 microM), although a complete relaxation was obtained by NCX inhibition at 100 microM. Interestingly, relaxation after washing the agonist was prolonged in the absence of external Na(+), whereas washing without Na(+) and in the presence of KB-R7943 resembled control conditions with physiological solution. Altogether, this suggests the reversal of NCX to a Ca(2+) influx mode by the manipulation on the Na(+) gradient, which can be inhibited by KB-R7943. In order to understand the low sensitivity to KB-R7943, we studied the molecular aspects of the NCX expressed in this tissue and found that the isoform of NCX expressed is 1.3, similar to that described in human tracheal smooth muscle. Sequencing revealed that amino acid 19 in exon B is phenylalanine, whereas in its human counterpart is leucine, and that the first amino acid after exon D is aspartate instead of glutamate in humans. Results herein presented are discussed in term of their possible functional implications in the exchanger activity and thus in airway physiology.

The Identification of Indacaterol as an Ultralong-Acting Inhaled β2-Adrenoceptor Agonist

Following a lipophilicity-based hypothesis, an 8-hydroxyquinolinone 2-aminoindan derived series of beta(2)-adrenoceptor agonists have been prepared and evaluated for their potential as inhaled ultralong-acting bronchodilators. Determination of their activities at the human beta(2)-adrenoceptor receptor showed symmetrical substitution of the 2-aminoindan moiety at the 5- and 6-positions delivered the targeted intermediate potency and intrinsic-efficacy profiles relative to a series of clinical reference beta(2)-adrenoceptor agonists. Further assessment with an in vitro superfused electrically stimulated guinea-pig tracheal-strip assay established the onset and duration of action time courses, which could be rationalized by considering the lipophilicity, potency, and intrinsic efficacy of the compounds. From these studies the 5,6-diethylindan analogue indacaterol 1c was shown to possess a unique profile of combining a rapid onset of action with a long duration of action. Further in vivo profiling of 1c supported the long duration of action and a wide therapeutic index following administration to the lung, which led to the compound being selected as a development candidate.

The synergistic anti-asthmatic effects of glycyrrhizin and salbutamol.

To investigate the efficacy of glycyrrhizin (GL) combined with salbutamol (SA) as an anti-asthma therapy. Rat lung beta2-adrenergic receptor (beta(2)-AR) mRNA level was measured by real-time RT PCR. Intracellular cAMP accumulation was evaluated with a reporter gene assay. An in vitro acetylcholine-induced guinea pig tracheal strip contraction model was used to test the relaxing effects of GL and SA. The anti-inflammatory effects of GL and SA were tested using tumor necrosis factor-alpha-induced NF-kappaB transcriptional activation reporter assay, I-kappaB Western blotting and interleukin-8 ELISA. An in vivo guinea pig asthma model was used to prove further the synergistic effect of GL and SA. GL (0.3 micromol/L) increased mRNA levels of beta(2)-AR in vivo and the accumulation of cAMP in vitro. The combination of GL and SA also resulted in significant complementary anti-inflammatory effects via inhibition of NF-kappaB activation, degradation of I-kappaB and production of interleukin-8. A significant synergistic effect of the combination was detected both in vitro and in vivo in a guinea pig mode. The results demonstrate that GL and SA have synergistic anti-asthmatic effects and offer the possibility of a therapeutic application of GL in combination with beta(2)-AR agonists in the treatment of asthma.

Effect of leukotriene C4 and D4 on isolated guinea pig tracheal strips (author's transl)

Effect of leukotriene C4 and D4 on isolated guinea pig tracheal strips (author's transl)

NT-702 (parogrelil hydrochloride, NM-702), a novel and potent phosphodiesterase 3 inhibitor, suppress the asthmatic response in guinea pigs, with both bronchodilating and anti-inflammatory effects

We evaluated the effects of NT-702 (parogrelil hydrochloride, NM-702, 4-bromo-6-[3-(4-chlorophenyl) propoxy]-5-[(pyridine-3-ylmethyl) amino] pyridazin-3(2H)-one hydrochloride), a selective phosphodiesterase 3 inhibitor, on the asthmatic response in guinea pigs. NT-702 at a concentration of 1 × 10 − 7 M elevated the cyclic adenosine monophosphate content in prostaglandin E 2-treated guinea pig tracheal smooth muscle cells. Leukotriene (LT) D 4- and histamine-induced contraction of isolated guinea pig tracheal strips was inhibited by NT-702, with EC 50 values of 3.2 × 10 − 7 and 2.5 × 10 − 7 M, respectively. In an in vivo study, NT-702 suppressed LTD 4-induced bronchoconstriction and the ovalbumin-induced immediate asthmatic response in guinea pigs through its bronchodilating effect. Furthermore, NT-702 also suppressed the ovalbumin-induced late asthmatic response, airway hyperresponsiveness, and the accumulation of inflammatory cells in the bronchoalveolar lavage fluid. These results suggest that NT-702 has an anti-inflammatory effect as well as a bronchodilating effect and might be useful as a novel potent therapeutic agent for the treatment of bronchial asthma, a new type of agent with both a bronchodilating and an anti-inflammatory effect.

Studies on cardio-suppressant, vasodilator and tracheal relaxant effects ofSarcococca saligna

Sarcococca saligna is a shrub that is traditionally used for its medicinal properties in Pakistan. In this study we report the cardio-suppressant, vasodilator and tracheal relaxant activities of the aqueous-methanolic extract (Ss.Cr) of the plant. Ss.Cr, that tested positive for the presence of saponins, flavonoids, tannins, phenols, and alkaloids, exhibited a dose-dependent (0.3-5 mg/mL) negative inotropic and chronotropic effect on the isolated guinea-pig atrium which was resistant to atropine (1 microM) and aminophylline (10 microM) pretreatment. In rabbit thoracic aorta, Ss.Cr dose-dependently (0.1-3 mg/mL) relaxed the high K+ (80 mM) and phenylephrine (PE, 1 microM)-induced contractions, indicating a possible Ca++ channel blocking (CCB) effect. When tested against PE (1 microM) control peaks in normal Ca++ and Ca++-free Kreb's solution, Ss.Cr exhibited dose-dependent (0.1-3 mg/mL) inhibition, being more potent in relaxing the PE responses in Ca++-free Kreb's solution, thus indicating specific blockade of Ca++ release from the intracellular stores. Ss.Cr also relaxed the agonist-induced contractions in: a) rat aorta irrespective of the presence of endothelium or nitric oxide synthase inhibitor L-NAME and b) rabbit and guinea-pig tracheal strips. The data shows that Ss.Cr possesses possible Ca++ channel blocking activity which might be responsible for its observed cardio-suppressant, vasodilator and tracheal relaxant effects though more tests are required to confirm this Ca++ channel blocking effect.

Fluorescent styryl dyes FM1‐43 and FM2‐10 are muscarinic receptor antagonists: intravital visualization of receptor occupancy

The fluorescent styryl dyes FM1-43 and FM2-10 have been used to visualize the endocytic and exocytic processes involved in neurotransmission in a variety of central and peripheral nerve preparations. Their utility is limited to some extent by a poorly understood vesicular-independent labelling of cells and tissues. We show here that one likely cause of this troublesome background labelling is that FM1-43 and FM2-10 are selective and competitive antagonists at both cloned and endogenously expressed muscarinic acetylcholine receptors. In radioligand binding studies, FM1-43 and FM2-10 bound with moderate affinity (23-220 nM) to membranes of Chinese hamster ovary (CHO) cells expressing cloned human muscarinic receptors (M1-M5). In functional studies in vitro, FM1-43 and FM2-10 inhibited electrical field stimulation (EFS) and acetylcholine-induced cholinergic contractions of guinea-pig tracheal strips (IC50: FM1-43, 0.4 +/- 0.1; FM2-10, 1.6 +/- 0.1 microM; concentration of antagonist producing a 2-fold leftward shift in the acetylcholine concentration-response curve (Kb): FM1-43, 0.3 +/- 0.1; FM2-10, 15.8 +/- 10.1 microM). Neither compound inhibited EFS-evoked, non-adrenergic non-cholinergic nerve-mediated relaxations or contractions of the airways, or contractions mediated by histamine H1 receptor or tachykinin NK2 receptor activation. Incubating freshly excised tracheal whole-mount preparations with 5 microM FM1-43 resulted in intense fluorescence labelling of the smooth muscle that was reduced by up to 90% in the presence of selective M2 and M3 receptor antagonists. The potency of the FM dyes as muscarinic receptor antagonists is within the concentration range used to study vesicular cycling at nerve terminals. Given that muscarinic receptors play a key role in the regulation of neurotransmitter release from a variety of neurones, the anticholinergic properties of FM dyes may have important implications when studying vesicular events in the nervous system. In addition, these dyes may provide a novel tool for visualizing muscarinic receptor occupancy in living tissue or cell preparations.

New 8-substituted xanthiene derivatives as potent bronchodilators

The synthesis and structure determination of 8-aryl /alkyl aryl 1, 3-dimethyl-3, 7-dihydropurin-2, 6-dione derivatives (1-13), was carried out in this study. Bronchodilator activity is investigated using isolated guinea-pig tracheal strips, pre-contracted by acetylcholine and histamine. Spasmolytic activity of the compounds was compared to theophylline. Synthesized compounds (1-13) did not inhibit the acetylcholine-induced pre-contractions except compound (8) at 10 −5 M concentration. In contrast, some of the compounds, especially (7), (11), (12) at 10 −5 M and (3), (4), (9) and (11) in 10 −4 M displayed inhibitory activity on the tracheal strips pre-contracted by histamine. The potency of the compounds at human adenosine receptors was evaluated using radioligand binding assay and a cyclic AMP functional assay in CHO cells expressing these receptors. Compound (11) displayed the greatest activity against radioligand binding of specific agonists to A 2 A and A 2B receptors. The compounds were relatively selective for both A 2 A and A 2B compared with A 1 and A 3 receptors. All compounds were also tested for the inhibition of NECA-induced cAMP accumulation mediated by the A 2B adenosine receptor and compound (11) was found to be the most effective. Our results showed that these compounds are acting as selective adenosine antagonists, especially for adenosine A 2B receptors, and are promising as potent anti-inflammatory agents rather than bronchodilator for the treatment of asthma.