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Acetylcholine (Ach) causes vasodilatation by nitric oxide (NO) release from the vascular endothelium. Vasoconstrictors such as α-adrenoceptor agonists (phenylephrine) or thromboxane TxA2 mimetics (U46619) also release endothelial NO. Inhibition of nitric oxide synthase (NOS) with Nω-nitro-L-arginine (L-NAME) potentiates vasoconstriction by phenylephrine and the trace amine, β-phenylethylamine (PEA), indicating underlying opposing vasodilatation. However, the roles of the endothelium and NO in vasodilator and constrictor responses of guinea-pig aorta have not been examined and are the subject of this study. Guinea-pig thoracic aorta rings were set up in aerated Krebs solution (37 °C) and isometric tension recorded. Contractions to phenylephrine were fast onset, rapidly waned and antagonised by prazosin. PEA contractions were slow onset, sustained and not antagonised by prazosin and therefore not α1-adrenoceptor-mediated. PEA and phenylephrine contractions were enhanced by L-NAME whether endothelium was present or not. Ach produced only weak relaxation in a small proportion of endothelium intact U46619-constricted aortae, which were abolished by endothelium removal. In uncontracted aortae Ach caused small contractions, which like PEA contractions were potentiated by endothelium removal. α-Adrenoceptor agonists and trace amines release NO from non-endothelial sites causing underlying opposing vasodilatation. The endothelium plays only a minor role in vasodilator and vasoconstrictor responses of guinea-pigs aorta.

Background: Inhibitors of the extracellular calcium-sensing receptor (CaSR), termed calcilytics, were developed to evoke pulsatile changes in plasma parathyroid hormone (PTH) levels as a novel anti-osteoporotic treatment. Recently we have shown that they can suppress airway hyperresponsiveness (AHR) and inflammation in asthma models, providing a potential for a novel asthma treatment strategy1. Objective: To develop a new calcilytic specifically for the treatment of airway disease and test its efficacy in allergic asthma. Methods: The efficacy of a new chemical class of calcilytics to suppress CaSR signalling was tested in vitro in human embryonic kidney 293 cells expressing the CaSR. The most active stereoisomer of one compound (B2.1-E1) was tested for its biological activity in vivo by administering the calcilytic solution to mice via gavage, after which plasma PTH levels were measured. Finally, the efficacy of the inhaled calcilytics to suppress AHR and inflammation was tested in the poly-L-arginine-induced model of AHR and the ovalbumin-sensitised acute asthma model in mice1. Results: B2.1-E1 supressed CaSR signalling in vitro and increased PTH levels in vivo. Inhaled B2.1-E1 led to dose-dependent suppression of AHR evoked by poly-L-arginine, and diminished AHR and inflammation in the acute asthma model in vivo. Conclusions: The novel calcilytic B2.1-E1 suppressed CaSR in vitro, and elicited PTH release in vivo. In addition, B2.1-E1 suppressed AHR and inflammation in mice, providing a rationale for further development of this calcilytic and its chemical class for the treatment of inflammatory lung disease (IP GB1719023.2). References:1Yarova PL et al. (2015). Science TM 7: 284ra60.

Sympathomimetic and trace amines, including β-phenylethylamine (PEA) and amphetamine,\ud increase blood pressure and constrict isolated blood vessels. By convention this is regarded as\ud a sympathomimetic response, however, recent studies suggest trace amine-associated\ud receptor (TAAR) involvement. There is also uncertainty whether these amines also release\ud nitric oxide (NO) causing opposing vasodilatation. These questions were addressed in guineapig\ud isolated aorta, a species not previously examined. Guinea-pig aortic rings were set up to\ud measure contractile tension. Cumulative concentration-response curves were constructed for\ud the reference α-adrenoceptor agonist, phenylephrine, PEA or d-amphetamine before and in\ud the presence of vehicles, the α1-adrenoceptor antagonist, prazosin (1 μM), the nitric oxide\ud synthase inhibitor, Nω-nitro-L-arginine (L-NAME), or NO scavengers, curcumin and\ud astaxanthin. Prazosin inhibited phenylephrine contractions with low affinity consistent with\ud α1L-adrenoceptors. However, PEA and amphetamine were not antagonised, indicating nonadrenergic\ud responses probably via TAARs. L-NAME potentiated contractions to PEA both in\ud the absence and presence of prazosin, indicating that PEA releases NO to cause underlying\ud opposing vasodilatation, independent of α1-adrenoceptors. L-NAME also potentiated\ud amphetamine and phenylephrine. PEA was potentiated by the NO scavenger astaxanthin but\ud less effectively. Curcumin, an active component of turmeric, however, inhibited PEA. Trace\ud amines therefore constrict blood vessels non-adrenergically with an underlying NO-mediated\ud non-adrenergic vasodilatation. This has implications in the pressor actions of these amines\ud when NO is compromised.

Viral exacerbations of allergen-induced pulmonary inflammation in pre-clinical models reportedly reduce the efficacy of glucocorticoids to limit pulmonary inflammation and airways hyper-responsiveness to inhaled spasmogens. However, exacerbations of airway obstruction induced by allergen challenge have not yet been studied. hPIV-3 (human parainfluenza type 3 virus) inoculation of guinea-pigs increased inflammatory cell counts in BAL (bronchoalveolar lavage) fluid and caused hyper-responsiveness to inhaled histamine. Both responses were abolished by treatment with either dexamethasone (20 mg/kg of body weight, subcutaneous, once a day) or fluticasone propionate (a 0.5 mg/ml solution aerosolized and inhaled over 15 min, twice a day). In ovalbumin-sensitized guinea-pigs, allergen (ovalbumin) challenge caused two phases of airway obstruction [measured as changes in sGaw (specific airways conductance) using whole body plethysmography]: an immediate phase lasting between 4 and 6 h and a late phase at about 7 h. The late phase, airway hyper-responsiveness to histamine and inflammatory cell counts in BAL were all significantly reduced by either glucocorticoid. Inoculation of guinea-pigs sensitized to ovalbumin with hPIV-3 transformed the allergen-induced airway obstruction from two transient phases into a single sustained response lasting up to 12 h. This exacerbated airway obstruction and airway hyper-responsiveness to histamine were unaffected by treatment with either glucocorticoid whereas inflammatory cell counts in BAL were only partially inhibited. Virus- or allergen-induced pulmonary inflammation, individually, are glucocorticoid-sensitive, but in combination generate a phenotype where glucocorticoid efficacy is impaired. This suggests that during respiratory virus infection, glucocorticoids might be less effective in limiting pulmonary inflammation associated with asthma.

Tryptamine is an endogenous and dietary indoleamine-based trace amine implicated in cardiovascular pathologies, including hypertension, migraine and myocardial infarction. This study aimed at identifying the signalling pathways for the vasoconstrictor response to tryptamine in rat isolated perfused mesenteric arterial beds and co-released vasodilator modulators of tryptamine-mediated vasoconstriction. Tryptamine caused concentration-dependent vasoconstriction of the mesenteric bed, measured as increases in perfusion pressure. These were inhibited by the 5-HT2A receptor antagonist, ritanserin, indicating mediation via 5-HT2A receptors. The response was inhibited by the phospholipase C (PLC) and phospholipase A2 (iPLA2) inhibitors, U-73122 and PACOCF3, suggesting involvement of phospholipase pathways. Activation of these pathways by tryptamine releases cyclooxygenase (COX) products since indomethacin (non-selective inhibitor of COX-1/2) and nimesulide (selective COX-2 inhibitor) reduced the vasoconstriction. The most likely COX vasoconstrictor product was prostaglandin PGE2 since the responses to tryptamine were reduced by AH-6809, a non-selective EP1 receptor antagonist. Involvement of the Rho-kinase pathway in the tryptamine-evoked vasoconstriction was also indicated by its reduction by the Rho-kinase inhibitors, Y-27,632 and fasudil. The tryptamine vasoconstriction is modulated by the co-released endothelial vasodilator, nitric oxide. Thus, circulating tryptamine can regulate mesenteric blood flow through a cascade of signalling pathways secondary to stimulation of 5-HT2A receptors., Graphical abstract

Magnesium sulphate is a potential treatment for acute severe asthma. However, the mechanisms and dose-response relationships are poorly understood. The first objective of this study was to examine whether inhaled magnesium sulphate exerts bronchodilator activity measured as bronchoprotection against histamine-induced bronchoconstriction in conscious guinea-pigs alone and combined with salbutamol. Secondly, we examined whether inhaled magnesium sulphate inhibits airways inflammation and function in models of neutrophilic and eosinophilic lung inflammation induced, respectively, by inhaled lipopolysaccharide or the inhaled antigen, ovalbumin (OVA). Airway function was measured in conscious guinea-pigs as specific airway conductance (sGaw) by whole-body plethysmography. Anti-inflammatory activity was measured against lung inflammatory cell influx induced by OVA inhalation in OVA-sensitised animals or by lipopolysaccharide (LPS) exposure of non-sensitised animals. Airway function (sGaw) was measured over 24 h after OVA exposure. Airway hyperresponsiveness to inhaled histamine and inflammatory cells in bronchoalveolar lavage fluid were recorded 24 h after OVA or LPS challenge. Histamine-induced bronchoconstriction was inhibited by inhaled magnesium sulphate or salbutamol alone and in combination, they produced synergistic bronchoprotection. LPS-induced neutrophil influx was inhibited by 6 days pretreatment with magnesium sulphate. Early and late asthmatic responses in OVA sensitised and challenged animals were attenuated by magnesium sulphate. Lung inflammatory cells were increased by OVA, macrophages being significantly reduced by magnesium sulphate. Nebulised magnesium sulphate protects against histamine-induced bronchoconstriction in conscious guinea-pigs and exerts anti-inflammatory activity against pulmonary inflammation induced by allergen (OVA) or LPS. These properties of magnesium sulphate explain its beneficial actions in acute asthma.

A series of 1′-(6-aminopurin-9-yl)-1′-deoxy-N-methyl-β-D-ribofuranuronamides that were characterised by 2-dialkylamino-7-methyloxazolo[4,5-b]pyridin-5-ylmethyl substituents on N6 of interest for screening as selective adenosine A3 receptor agonists, have been synthesised. This work involved the synthesis of 2-dialkylamino-5-aminomethyl-7-methyloxazolo[4,5-b]pyridines and analogues that were coupled with the known 1′-(6-chloropurin-9-yl)-1′-deoxy-N-methyl-β-D-ribofuranuronamide. The oxazolo[4,5-b]pyridines were synthesized by regioselective functionalisation of 2,4-dimethylpyridine N-oxides. The regioselectivities of these reactions were found to depend upon the nature of the heterocycle with 2-dimethylamino-5,7-dimethyloxazolo[4,5-b]pyridine-N-oxide undergoing regioselective functionalisation at the 7-methyl group on reaction with trifluoroacetic anhydride in contrast to the reaction of 4,6-dimethyl-3-hydroxypyridine-N-oxide with acetic anhydride that resulted in functionalisation of the 6-methyl group. To optimise selectivity for the A3 receptor, 5-aminomethyl-7-bromo-2-dimethylamino-4-[(3-methylisoxazol-5-yl)methoxy]benzo[d]oxazole was synthesised and coupled with the 1′-(6-chloropurin-9-yl)-1′-deoxy-N-methyl-β-D-ribofuranuronamide. The products were active as selective adenosine A3 agonists.

Syntheses of (1RS,2SR,6SR)-2-alkoxymethyl-, 2-hetaryl-, and 2-(hetarylmethyl)-7-arylmethyl-4,7-diaza-9-oxabicyclo[4.3.0]nonan-8-ones, of interest as potential muscarinic M1 receptor agonists, are described. A key step in the synthesis of (1RS,2SR,6SR)-7-benzyl-6-cyclobutyl-2-methoxymethyl-4,7-diaza-9-oxabicyclo[4.3.0]nonan-8-one, was the addition of isopropenylmagnesium bromide to 2-benzyloxycarbonylamino-3-tert-butyldimethylsilyloxy-2-cyclobutylpropanal. This gave the 4-tert-butyldimethylsilyloxymethyl-4-cyclobutyl-5-isopropenyloxazolidinone with the 5-isopropenyl and 4-tert-butyldimethylsilyloxymethyl groups cis-disposed about the five-membered ring by chelation controlled addition and in situ cyclisation. This reaction was useful for a range of organometallic reagents. The hydroboration-oxidation of (4SR,5RS)-3-benzyl-4-(tert-butyldimethylsilyloxymethyl)-4-cyclobutyl-5-(1-methoxyprop-2-en-2-yl)-1,3-oxazolidin-2-one gave (4SR,5RS)-3-benzyl-4-(tert-butyldimethylsilyloxymethyl)-4-cyclobutyl-5-[(SR)-1-hydroxy-3-methoxyprop-2-yl]-1,3-oxazolidin-2-one stereoselectively. 4,7-Diaza-9-oxabicyclo[4.3.0]nonan-8-ones with substituents at C2 that could facilitate C2 deprotonation were unstable with respect to oxazolidinone ring-opening and this restricted both the synthetic approach and choice of 2-heteroaryl substituent. The bicyclic system with a 2-furyl substituent at C2 was therefore identified as an important target. The addition of 1-lithio-1-(2-furyl)ethene to 2-benzyloxycarbonylamino-3-tert-butyldimethylsilyloxy-2-cyclobutylpropanal gave (4SR,5RS)-4-tert-butyldimethylsilyloxymethyl-4-cyclobutyl-5-[1-(2-furyl)ethenyl]-1,3-oxazolidinone after chelation controlled addition and in situ cyclisation. Following oxazolidinone N-benzylation, hydroboration at 35 °C, since hydroboration at 0 °C was unexpectedly selective for the undesired isomer, followed by oxidation gave a mixture of side-chain epimeric alcohols that were separated after SEM-protection and selective desilylation. Conversion of the neopentylic alcohols into the corresponding primary amines by reductive amination, was followed by N-nosylation, removal of the SEM-groups and cyclisation using a Mitsunobu reaction. Denosylation then gave the 2-furyloxazolidinonyl-fused piperidines, the (1RS,2SR,6SR)-epimer showing an allosteric agonistic effect on M1 receptors. Further studies resulted in the synthesis of other 2-substituted 4,7-diaza-9-oxabicyclo[4.3.0]nonan-8-ones and an analogous tetrahydropyran.

Pre-clinical evaluation of asthma therapies requires animal models of chronic airways inflammation, airway hyperresponsiveness (AHR) and lung remodelling that accurately predict drug effectiveness in human asthma. However, most animal models focus on acute allergen challenges where chronic inflammation and airway remodelling are absent. Chronic allergen challenge models have been developed in mice but few studies use guinea-pigs which may be more relevant to humans. We tested the hypothesis that a chronic rather than acute pulmonary inflammation model would best predict clinical outcome for asthma treatments. Guinea-pigs sensitized with ovalbumin (OVA) received single (acute) or nine OVA inhalation challenges at 48 h intervals (chronic). Airways function was recorded as specific airways conductance (sG(aw)) in conscious animals for 12 h after OVA challenge. AHR to inhaled histamine, inflammatory cell influx and lung histology were determined 24 h after the single or 9th OVA exposure. The inhaled corticosteroid, fluticasone propionate (FP), the phosphodiesterase 4 inhibitor, roflumilast, and the inducible nitric oxide synthase (iNOS) inhibitor, GW274150, orally, were administered 24 and 0.5 h before and 6 h after the single or final chronic OVA exposure. Both models displayed early (EAR) and late (LAR) asthmatic responses to OVA challenge, as falls in sG(aw), AHR, as increased histamine-induced bronchoconstriction, and inflammatory cell influx. Tissue remodelling, seen as increased collagen and goblet cell hyperplasia, occurred after multiple OVA challenge. Treatment with FP and roflumilast inhibited the LAR, cell influx and AHR in both models, and the remodelling in the chronic model. GW274150 also inhibited the LAR, AHR and eosinophil influx in the acute model, but not, together with the remodelling, in the chronic model. In the clinical setting, inhaled corticosteroids and phosphodiesterase 4 inhibitors are relatively effective against most features of asthma whereas the iNOS inhibitor GW274150 was ineffective. Thus, while there remain certain differences between our data and clinical effectiveness of these antiasthma drugs, a chronic pulmonary inflammation guinea-pig model does appear to be a better pre-clinical predictor of potential asthma therapeutics than an acute model.

Aim of the study The present study evaluates the Philippine medicinal plant Artemisia vulgaris for antagonistic activity at selected biogenic amine receptors on smooth muscle of the airways and gastrointestinal tract in order to explain its traditional use in asthma and hyperactive gut. Materials and methods The antagonistic activity of chloroform crude extract (AV-CHCl3) and methanol crude extract (AV-MeOH) of Artemisia vulgaris was studied against concentration–response curves for contractions of the guinea pig ileum and trachea to 5-hydroxytrptamine (5-HT2 receptors), methacholine (M3 muscarinic receptors), histamine (H1 receptors) and β-phenylethylamine (trace amine-associated receptors, TAAR1). Results and discussion The Artemisia vulgaris chloroform (AV-CHCl3) and methanol (AV-MeOH) extract showed histamine H1 antagonism in the ileum and trachea. Further analysis of AV-CHCl3 isolated two major components, yomogin and 1,2,3,4-diepoxy-11(13)-eudesmen-12,8-olide. Yomogin, a sesquiterpene lactone, exhibited a novel histamine H1 receptor antagonism in the ileum. Conclusion The presence of a specific, competitive histamine receptor antagonist and smooth muscle relaxant activity in Artemisia vulgaris extracts on the smooth muscle in ileum and trachea explains its traditional use in the treatment of asthma and hyperactive gut.

Background: Seasonal influenza A infection affects a significant cohort of the global population annually, resulting in considerable morbidity and mortality. Therapeutic strategies are of limited efficacy, and during a pandemic outbreak would only be available to a minority of the global population. Over-the-counter medicines are routinely taken by individuals suffering from influenza, but few studies have been conducted to determine their effectiveness in reducing pulmonary immunopathology or the influence they exert upon the generation of protective immunity. Methods: A mouse model of influenza infection was utilised to assess the efficacy of paracetamol (acetaminophen) in reducing influenza-induced pathology and to examine whether paracetamol affects generation of protective immunity. Results: Administration (intraperitoneal) of paracetamol significantly decreased the infiltration of inflammatory cells into the airway spaces, reduced pulmonary immunopathology associated with acute infection and improved the overall lung function of mice, without adversely affecting the induction of virus-specific adaptive responses. Mice treated with paracetamol exhibited an ability to resist a second infection with heterologous virus comparable with that of untreated mice. Conclusions: Our results demonstrate that paracetamol dramatically reduces the morbidity associated with influenza but does not compromise the development of adaptive immune responses. Overall, these data support the utility of paracetamol for reducing the clinical symptoms associated with influenza virus infection.

Several α-hydroxyamides with (2,6-dialkoxyphenoxy)methyl substituents have been prepared and their activities as antagonists of the M3 muscarinic receptor in guinea pig ileum have been evaluated. N-{1-[(Phenyl)methyl]piperidin-4-yl}-2-{2-[(2,6-dimethoxyphenoxy)-methyl]phenyl}-2-hydroxypropanamide and N-(1-[{6-amino-4-[(1-propylpiperidin-4-yl)methyl]pyridin-2-yl}methyl]piperidin-4-yl)-2-cyclopentyl-2-hydroxy-2-phenylacetamide were the most potent compounds prepared, the micromolar potency of the latter indicating that it may be worth further investigation.

The leaves of khat (Catha edulis Forsk.) are chewed as a social habit for the central stimulant action of their cathinone content. This review summarizes the prevalence of the habit worldwide, the actions, uses, constituents and adverse health effects of khat chewing. There is growing concern about the health hazards of chronic khat chewing and this review concentrates on the adverse effects on health in the peripheral systems of the body, including the cardiovascular system and gastrointestinal tract. Comparisons are made with amphetamine and ecstasy in particular on the detrimental effects on the cardiovascular system. The underlying mechanisms of action of khat and its main constituent, cathinone, on the cardiovascular system are discussed. Links have been proposed between khat chewing and the incidence of myocardial infarction, dilated cardiomyopathy, vascular disease such as hypertension, cerebrovascular ischaemia and thromboembolism, diabetes, sexual dysfunction, duodenal ulcer and hepatitis. The evidence, however, is often based on limited numbers of case reports and only few prospective controlled studies have been undertaken. There is therefore an urgent need for more thorough case-control studies to be performed. This review outlines the current knowledge on the adverse health effects of khat chewing on the cardiovascular system and other internal medical problems, it assesses the evidence and the limitations of the studies and identifies the questions that future studies should address.

Inhaled bradykinin causes bronchoconstriction in asthmatic subjects but not nonasthmatics. To date, animal studies with inhaled bradykinin have been performed only in anesthetized guinea pigs and rats, where it causes bronchoconstriction through sensory nerve pathways. In the present study, airway function was recorded in conscious guinea pigs by whole-body plethysmography. Inhaled bradykinin (1 mM, 20 s) caused bronchoconstriction and influx of inflammatory cells to the lungs, but only when the enzymatic breakdown of bradykinin by angiotensin-converting enzyme and neutral endopeptidase was inhibited by captopril (1 mg/kg i.p.) and phosphoramidon (10 mM, 20-min inhalation), respectively. The bronchoconstriction and cell influx were antagonized by the B2 kinin receptor antagonist 4-(S)-amino-5-(4-{4-[2, 4-dichloro-3-(2,4-dimethyl-8-quinolyloxymethyl)phenylsulfonamido]- tetrahydro-2H-4-pyranylcarbonyl}piperazino)-5- oxopentyl](trimethyl)ammonium chloride hydrochloride (MEN16132) when given by inhalation (1 and 10 μM, 20 min) and are therefore mediated via B2 kinin receptors. However, neither intraperitioneal MEN16132 nor the peptide B2 antagonist icatibant, by inhalation, antagonized these bradykinin responses. Sensitization of guinea pigs with ovalbumin was not sufficient to induce airway hyperreactivity (AHR) to the bronchoconstriction by inhaled bradykinin. However, ovalbumin challenge of sensitized guinea pigs caused AHR to bradykinin and histamine. Infection of guinea pigs by nasal instillation of parainfluenza-3 virus produced AHR to inhaled histamine and lung influx of inflammatory cells. These responses were attenuated by the bradykinin B2 receptor antagonist MEN16132 and H-(4-chloro)DPhe-2'(1-naphthylalanine)-( 3-aminopropyl)guanidine (VA999024), an inhibitor of tissue kallikrein, the enzyme responsible for lung synthesis of bradykinin. These results suggest that bradykinin is involved in virus-induced inflammatory cell influx and AHR

Inhaled corticosteroids are regularly co-administered with beta(2)-adrenoceptor agonists. This study evaluates in conscious guinea-pigs the bronchodilator effect, alone or combined with salbutamol, of TPI 1020, a novel anti-inflammatory corticosteroid and nitric oxide (NO) donor derived from budesonide. Guinea-pigs received inhaled histamine (3 mM) and specific airway conductance (sG(aw)) measured. Responses to histamine were measured before and on the next day 15 min after a 15 min inhalation of vehicle, salbutamol, TPI 1020, budesonide, the NO-donor, S-nitroso-N-acetylpenicillamine (SNAP), or combinations of these drugs. Salbutamol and TPI 1020 caused concentration-dependent bronchodilatation measured as inhibition of histamine-induced bronchoconstriction. TPI 1020-induced bronchodilatation was blocked by the guanylyl cyclise inhibitor, ODQ, indicating cGMP-dependence through released NO. While salbutamol at 80 microM did not exert significant bronchodilatation, significant inhibitions were observed when co-administered with TPI 1020, 0.11 and 0.33 mM. The combined effects of TPI 1020 and salbutamol lasted significantly longer than either drug alone. Inhaled budesonide was a weak bronchodilator and when co-administered with salbutamol there was enhanced bronchodilatation. Addition of the NO-donor, SNAP (0.1 mM), to the budesonide/salbutamol combination, also improved the inhibition of histamine-induced bronchoconstriction. This study has shown that TPI 1020 potentiates the bronchodilator activity of salbutamol, and their combination lasted longer than either drug administered individually. Both the corticosteroid and NO-releasing activities of TPI 1020 appear to be required for the potentiation of salbutamol. Combination of TPI 1020 with a beta(2)-adrenoceptor agonist may therefore be useful against acute bronchoconstriction episodes in asthma, and may offer an opportunity for reducing doses of inhaled beta(2)-adrenoceptor agonists.

Endogenous adenosine is released in asthmatic patients' lungs by inhaled allergen, however, its exact role in asthmatic responses or the receptors mediating these responses has not been determined. Our hypothesis was that adenosine released during allergen challenge contributes to the airways responses to inhaled allergen. The effects of selective antagonists of the four adenosine receptor subtypes were investigated on the airways responses of sensitized guinea-pigs to inhaled ovalbumin to ascertain the role of adenosine in these allergen responses, and compared with a corticosteroid, dexamethasone. Early (EAR) and late asthmatic responses (LAR) to inhaled ovalbumin (10 microg/ml) of sensitized, conscious guinea-pigs were recorded by whole body plethysmography following administration of selective adenosine receptor antagonists. Airway reactivity to inhaled histamine (1 mM) and inflammatory cell influx in bronchoalveolar lavage fluid were also determined 24 h after ovalbumin challenge. ZM241385 (A(2A) receptor antagonist) did not affect these responses, whereas DPCPX (A(1) receptor antagonist) exerted a small inhibition only of the LAR. MRS1706 (A(2B) receptor antagonist) inhibited the airways hyperreactivity and cellular influx and enhanced the EAR. MRS1220 (A(3) receptor antagonist) inhibited the airways hyperreactivity and cellular influx without affecting EAR and LAR. Dexamethasone inhibited the ovalbumin-induced late asthmatic responses, airways hyperreactivity and cellular influx. The blockade of airway hyperreactivity and inflammatory cell influx by A(2B) and A(3) receptor antagonists suggests that endogenous adenosine is released by inhaled allergen and these responses are mediated via A(2B) and A(3) receptors in guinea-pigs. The adenosine released by allergen inhalation does not, however, appear to be involved in the EAR, but it may contribute to the LAR via A(1) receptors.

Trace amines, including tyramine and β-phenylethylamine (β-PEA), are constituents of many foods including chocolate, cheeses and wines and are generated by so-called ‘friendly’ bacteria such as Lactobacillus, Lactococcus and Enterococcus species, which are found in probiotics. We therefore examined whether these dietary amines could exert pharmacological effects on the gut and its vasculature. In the present study we examined the effects of tyramine and β-PEA on the contractile activity of guinea-pig and rat ileum and upon the isolated mesenteric vasculature and other blood vessels. Traditionally, these amines are regarded as sympathomimetic amines, exerting effects through the release of noradrenaline from sympathetic nerve endings, which should relax the gut. A secondary aim was therefore to confirm this mechanism of action. However, contractile effects were observed in the gut and these were independent of noradrenaline, acetylcholine, histamine and serotonin receptors. They were therefore probably due to the recently described trace amine-associated receptors. These amines relaxed the mesenteric vasculature. In contrast, the aorta and coronary arteries were constricted, a response that was also independent of a sympathomimetic action. From these results, we propose that after ingestion, trace amines could stimulate the gut and improve intestinal blood flow. Restriction of blood flow elsewhere diverts blood to the gut to aid digestion. Thus, trace amines in the diet may promote the digestive process through stimulation of the gut and improved gastrointestinal circulation.

Background and purpose: The dietary trace amines tyramine and β-phenylethylamine (β-PEA) can increase blood pressure. However, the mechanisms involved in the vascular effect of trace amines have not been fully established. The purpose of this study was to evaluate whether trace amine-dependent vasoconstriction was brought about by tyramine and β-PEA acting as indirect sympathomimetic agents, as previously assumed, or whether trace amine-dependent vasoconstriction could be mediated by recently discovered trace amine-associated (TAA) receptors. Experimental approach: The responses to p-tyramine and β-PEA were investigated in vitro in rings of the left anterior descending coronary arteries of pigs. Key results: p-Tyramine induced a concentration-dependent (0.1–3 mM) vasoconstriction. The maximum response and pD2 value for p-tyramine was unaffected by endothelium removal or pre-treatment with antagonists for adrenoceptors, histamine, dopamine or 5-HT receptors. β-PEA also produced a concentration-dependent (0.3–10 mM) vasoconstriction which was unaffected by endothelium removal, β-adrenoceptor or 5-HT receptor antagonists. A substantial, but reduced, response to β-PEA was obtained in the presence of prazosin (1-adrenoceptor antagonist), haloperidol (D2/D3 dopamine receptor antagonist) or mepyramine (H1 histamine receptor antagonist). The pD2 value for β-PEA was unaffected by any of the antagonists tested. Conclusions and implications: Vasoconstriction induced by p-tyramine does not involve an indirect sympathomimetic effect, although vasoconstriction caused by β-PEA may occur, in part, by this mechanism. We therefore propose that trace amine-dependent vasoconstriction is mediated by phenylethylamine-specific receptors, which are closely related to or identical to TAA receptors. These receptors could provide a target for new antihypertensive therapies

Background Endogenous adenosine levels are raised in the lungs during asthma attacks. 5′-adenosine monophosphate (5′-AMP) inhalation in asthmatics causes bronchoconstriction and in sensitized guinea-pigs induces early (EAR) and late asthmatic responses (LAR), airway hyper-reactivity (AHR) and inflammatory cell recruitment to the lungs. Objective The aim of this study was to investigate the roles of A1, A2A, A2B and A3 adenosine receptors in these responses to inhaled 5′-AMP in sensitized guinea-pigs. Comparisons were made with the effect of dexamethasone treatment on 5′-AMP-induced responses. Methods Functional airways responses to inhaled 5′-AMP (3 and 300 mm) of actively sensitized, conscious guinea-pigs were determined by whole-body plethysmography following administration of selective adenosine receptor antagonists or their vehicles. AHR to inhaled histamine (1 mm) and inflammatory cell influx in bronchoalveolar lavage fluid were determined. Results 5′-AMP at 3 mm caused an immediate bronchoconstriction (EAR), whereas 300 mm caused bronchodilatation. Both responses were followed at 6 h by a LAR, together with inflammatory cell influx and AHR to histamine. The A2A receptor antagonist, ZM241385, further enhanced cell influx after 5′-AMP inhalation (3 and 300 mm), and blocked the immediate bronchodilator response to 300 mm 5′-AMP, exposing an EAR. The A2B receptor antagonist, MRS1706 (in the presence of ZM241385), inhibited the LAR, AHR and cell influx, following inhalation of 5′-AMP (300 mm). The A3 receptor antagonist, MRS1220, inhibited 5′-AMP-induced inflammatory cell influx. The A1 receptor antagonist, DPCPX (in the presence of ZM241385), inhibited the EAR following 5′-AMP inhalation (300 mm). Dexamethasone inhibited the LAR, AHR and cell influx following inhalation of 5′-AMP (300 mm). Conclusion All four adenosine receptor subtypes play various roles in the airways responses to inhaled 5′-AMP in sensitized guinea-pigs.

Allergic asthma is a chronic disease of the airways, with superimposed acute inflammatory episodes which correspond to exacerbations of asthma. Two novel models of allergic asthma have been developed in mice receiving the same allergen sensitisation, but with acute or chronic allergen exposures, the latter to mimic the human situation more closely. Ovalbumin-sensitised mice were challenged by ovalbumin inhalation twice on the same day for the acute model, and 18 times over a period of 6 weeks for the chronic model. Lung function was monitored in conscious, unrestrained mice immediately after the last challenge for up to 12 h. Airway responsiveness to inhaled methacholine and serum antibody levels were determined 24 h after challenge. Bronchoalveolar inflammatory cell recruitment was determined at 2 or 24 h. Acute and chronically treated mice had similar early and late asthmatic responses peaking at 2 h and 7-8 h, respectively. IgE and IgG antibody levels, compared with naïve mice, and eosinophil infiltration, compared with naïve and saline challenge, were elevated. Airway hyperresponsiveness to methacholine was observed 24 h after challenge in both models. The acute model had higher levels of eosinophilia, whereas the chronic model showed hyperresponsiveness to lower doses of methacholine and had higher levels of total IgE and ovalbumin-specific IgG antibodies. Both novel murine models of allergic asthma bear a close resemblance to human asthma, each offering particular advantages for studying the mechanisms underlying asthma and for evaluating existing and novel therapeutic agents.

Adrenaline (epinephrine) has been used for cardiopulmonary resuscitation (CPR) since 1896. The rationale behind its use is thought to be its alpha-adrenoceptor-mediated peripheral vasoconstriction, causing residual blood flow to be diverted to coronary and cerebral circulations. This protects these tissues from ischaemic damage and increases the likelihood of restoration of spontaneous circulation. Clinical trials have not demonstrated any benefit of adrenaline over placebo as an agent for resuscitation. Adrenaline has deleterious effects in the setting of resuscitation, predictable from its promiscuous pharmacological profile. This article discusses the relevant pharmacology of adrenaline in the context of CPR. Experimental and clinical evidences for the use of adrenaline and alternative vasopressor agents in resuscitation are given, and the properties of an ideal vasopressor are discussed.

3,4-methylenedioxymethamphetamine ('Ecstasy', MDMA) and cathinone, the active constituent of khat leaves, were examined on pig isolated left anterior descending coronary arteries to determine whether they cause vasoconstriction and whether this was an indirect sympathomimetic action. Coronary artery rings were set up in Krebs solution (37 degrees C) gassed with 5% CO2 in O2. Endothelium remained intact as indicated by relaxation by bradykinin. Isometric tension was recorded and cumulative concentration-response curves (CRCs) for noradrenaline, ecstasy or cathinone plotted as a percent of the constriction to KCl (60 mM). Noradrenaline-induced contractions of the coronary artery were enhanced by propranolol (1 microM) indicating beta-adrenoceptor-mediated opposing vasodilatation. Cocaine (10 microM) further potentiated, while prazosin (1 microM) virtually abolished the contractions to noradrenaline. Cathinone and ecstasy constricted the coronary artery rings, the peak contractions being 56.5+/-4.2% (n=4) and 37.3+/-2.4% (n=4), respectively. Higher concentrations relaxed. The vasoconstriction was not affected by cocaine (10 microM), prazosin (1 microM, in the presence of cocaine) or removal of the endothelium. There was no tachyphylaxis or desensitisation on repeated administration of single doses. Ecstasy- and cathinone-induced coronary vasoconstriction is therefore via mechanisms other than indirect sympathomimetic activity or alpha1 -adrenoceptors. This activity could explain the cardiac adverse effects following their excessive use.

Antigen inhalation in patients with atopic asthma results in an early asthmatic response (EAR), accompanied by a late asthmatic response (LAR) in 60% of patients, airway hyperresponsiveness (AHR) and inflammatory cell infiltration to the lungs. An ideal animal model of asthma should, therefore, provide at least these 4 features consistently and reproducibly. The aim of this study was to optimise the ovalbumin (OA) sensitisation conditions, for achieving EAR, LAR, AHR and cell influx, in a guinea-pig model of asthma. Animals were sensitised with 10 micro g or 100 micro g OA, as either a single or booster (day 1 and day 5) injection. Airway responses to inhaled OA (10 micro g, 1 h) of actively sensitised, conscious guinea pigs were determined by whole body plethysmography as the change in specific airways conductance (sG(aw)) over a 12 h period and at 24 h. Bronchoconstriction by inhaled histamine (1 mM) was used to investigate AHR, and inflammatory cell influx was determined by bronchoalveolar lavage (BAL), both at 24 h post-challenge. A single sensitisation with 10 micro g OA did not reveal an EAR, LAR or AHR following exposure to OA. However, total and differential cell counts (eosinophils and macrophages) were significantly greater 24 h post-challenge, when compared to saline-challenged sensitised animals. The addition of a booster injection of 10 micro g revealed an EAR, but no LAR or AHR after ovalbumin inhalation. However, there was a significant cell influx. Sensitisation with 100 micro g OA (single and booster injections) revealed all four parameters of the asthmatic response (EAR, LAR, AHR and cell influx). The incorporation of the booster sensitisation injection resulted in a prolongation of the LAR, and the AHR was more pronounced and cell influx increased significantly, when compared to all other sensitisation protocols. Thus, sensitisation with 100 micro g OA (with a booster injection) can reveal an EAR, LAR, AHR and cell influx following inhalation exposure to OA (10 micro g). Cellular infiltration to the lung may be a poor marker of the asthmatic response, as a threshold level of cell influx (eosinophils) appears to be required in order to elicit the LAR and AHR. There was an association between the LAR and AHR.

Airway hyperresponsiveness and inflammation are fundamental hallmarks of allergic asthma that are accompanied by increases in certain polycations, such as eosinophil cationic protein. Levels of these cations in body fluids correlate with asthma severity. We show that polycations and elevated extracellular calcium activate the human recombinant and native calcium-sensing receptor (CaSR), leading to intracellular calcium mobilization, cyclic adenosine monophosphate breakdown, and p38 mitogen-activated protein kinase phosphorylation in airway smooth muscle (ASM) cells. These effects can be prevented by CaSR antagonists, termed calcilytics. Moreover, asthmatic patients and allergen-sensitized mice expressed more CaSR in ASMs than did their healthy counterparts. Indeed, polycations induced hyper-reactivity in mouse bronchi, and this effect was prevented by calcilytics and absent in mice with CaSR ablation from ASM. Calcilytics also reduced airway hyperresponsiveness and inflammation in allergen-sensitized mice in vivo. These data show that a functional CaSR is up-regulated in asthmatic ASM and targeted by locally produced polycations to induce hyperresponsiveness and inflammation. Thus, calcilytics may represent effective asthma therapeutics.

Purpose: Asthma is associated with reversible airway obstruction, leucocyte infiltration, airways hyperresponsiveness (AHR) and airways remodelling. Fluid accumulation causes pulmonary oedema contributing to airways obstruction. We examined the temporal relationship between the late asthmatic response (LAR) following allergen challenge of sensitised guinea-pigs and pulmonary oedema measured by magnetic resonance imaging (MRI). Materials and Methods: Ovalbumin (OVA) sensitised guinea-pigs received either a single OVA inhalation (acute) or nine OVA inhalations at 48 h intervals (chronic). Airways obstruction was measured as specific airways conductance (sGaw) by whole body plethysmography. AHR to inhaled histamine and bronchoalveolar lavage for leucocyte counts were measured 24 h after a single or the final chronic ovalbumin challenges. MRI was performed at intervals after OVA challenge and high intensity oedemic signals quantified. Results: Ovalbumin caused early bronchoconstriction, followed at 7 h by a LAR and at 24 h AHR and leucocyte influx. The bright intensity MRI oedema signal, peaking at 7 h, was significantly (P

Summary 1 Relaxation responses of the rat isolated duodenum to the putative M1 muscarinic receptor agonist, McN-A-343, were examined to determine whether the response was due to the release of known non-adrenergic, non-cholinergic relaxant neurotransmitters and to establish the involvement of M1 muscarinic receptors. 2 The role of ATP was examined with the P2 receptor antagonist, suramin, which at 30 μm antagonized the relaxant responses to α,β-methylene ATP. The same dose, however, failed to inhibit the relaxation by McN-A-343. 3 The role of nitric oxide (NO) was examined with the NO synthase inhibitor, NG-nitro-l-arginine methyl ester (l-NAME; 100 μm), which failed to inhibit the responses to McN-A-343. As NO mediates relaxation of the duodenum via cGMP generation through guanylyl cyclase, whether the relaxation by McN-A-343 was also via cGMP was examined with the guanylyl cyclase inhibitor, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ). The relaxation responses to the NO donor, S-nitroso-N-acetyl penicillamine, were inhibited in the presence of ODQ (3 μm), but not those by McN-A-343. 4 Release of γ-aminobutyric acid (GABA) was examined with the GABAA receptor antagonist, bicuculline (10 μm), which shifted the concentration–response curves for the relaxation of the duodenum by GABA to the right. There was a similar degree of shift in the concentration–response curve for McN-A-343 by bicuculline indicating that release of GABA from enteric neurones of the duodenum could explain the relaxation response to McN-A-343. 5 To test whether the muscarinic receptors mediating the relaxation of the duodenum were of the M1 subtype, the susceptibility to the selective competitive antagonist, pirenzepine and the selective muscarinic toxin from green mamba, MT7, was examined. Pirenzepine (1 μm) shifted the concentration–response for McN-A-343 to the right in a parallel fashion with a dose ratio of 33.3 ± 20.2. This yielded a pA2 value of 7.5, which concords with those for other responses reputed to be mediated via M1 muscarinic receptors. The toxin MT7 was used as an irreversible antagonist and following incubation with the duodenum was washed from the bath. An incubation time of 30 min with 100 nm of MT7 caused a significant parallel shift in the concentration–response to McN-A-343 confirming the involvement of M1 muscarinic receptors. 6 This study has confirmed that McN-A-343 relaxes the rat duodenum via muscarinic receptors of the M1 subtype and that these receptors are probably located on enteric neurones from which their stimulation releases GABA.

The effects of hypoxia on the vasodilator response of endothelium-denuded rat aortic rings to the calcium channel blocker, nifedipine, were examined. Under normoxic conditions, nifedipine (10(-8)-3x10(-6) M) attenuated the contractility of noradrenaline precontracted rings in a concentration-dependent manner, although the sensitivity was less than what occurs with K+ precontracted tissues. Under hypoxic conditions there was no relaxation by nifedipine. When a concentration-response curve to noradrenaline was constructed before and in the presence of a high concentration of nifedipine (10(-5) M), the response to noradrenaline was unaffected in both normoxic and hypoxic conditions. When noradrenaline was replaced by phenylephrine (10(-8)-10(-5) M), the maximum tension was reduced in the presence of nifedipine to 59+/-6% of the pre-nifedipine value. Repetition of the experiment in the presence of cocaine (10(-5) M) revealed the inhibitory effect of nifedipine on noradrenaline-induced contraction, the maximum contraction in the presence of nifedipine falling significantly (P0.005) to 67+/-6% of the pre-nifedipine response. When propranolol (10(-7) M) was present in the bath, the maximum contraction to noradrenaline was significantly (P0.05) reduced by nifedipine to 55+/-4% of its previous value. The fact that nifedipine was able to inhibit phenylephrine-induced contractions and relax noradrenaline-precontracted aortic rings confirms its calcium channel blocking activity. The failure to inhibit noradrenaline when added prior to the noradrenaline-induced contractions suggests an opposing effect in addition to calcium channel blockade, which cancels out the attenuation of noradrenaline--but not phenylephrine-induced contractions. When neuronal uptake of noradrenaline was blocked with cocaine or beta-adrenoceptors were blocked with propranolol, the inhibitory effect of nifedipine against noradrenaline-induced contractions was revealed. This suggests that the additional property was due to blockade of neuronal reuptake or antagonism at beta-adrenoceptors. This study also showed that nifedipine is ineffective as a vasodilator in the rat aorta under hypoxic conditions.

The binding and functional properties of adenosine receptor ligands were compared in Chinese hamster ovary cells transfected with human adenosine A(3) receptors. Inhibition of [(125)I]-aminobenzyl-5'-N-methylcarboamidoadenosine ([(125)I]-AB-MECA) binding by adenosine receptor ligands was examined in membrane preparations. Inhibition of forskolin-induced cAMP accumulation by agonists was measured using a cAMP enzyme immunoassay. The rank order of agonist potency for both assays was N(6)-(3-iodobenzyl)-adenosine-5'-N-methyluronamide (IB-MECA) > 5'-N-ethylcarboxamidoadenosine (NECA) > (-)-N(6)-[(R)-phenylisopropyl] adenosine (R-PIA) > 4-aminobenzyl-5'-N-methylcarboxamidoadenosine (AB-MECA) > N(6)-cyclopentyl adenosine (CPA) > adenosine. The radioligand binding rank order of antagonist potency was N-[9-chloro-2-(2-furanyl)[1,2,4]-triazolo[1,5-c]quinazolin-5-benzeneacetamide (MRS1220) > 1,3-dipropyl-8-cyclopentylxanthine (DPCPX) > 8-phenyltheophylline (8-PT) > 8-(p-sulfophenyl)-theophylline (8-SPT). MRS1220 competitively inhibited the effect of IB-MECA on cAMP production, with a K(B) value of 0.35 nm. These data are characteristic of adenosine A(3) receptors. The absence of Mg(2+) and presence of guanosine 5'-(gamma-thio)triphosphate (GTPgammaS) significantly reduced agonist binding inhibition potency, indicating binding to high- and low-affinity states. The IB-MECA, NECA and R-PIA IC(50) values were greater for the cAMP assay than for radioligand binding, suggesting an efficient stimulus-response transduction pathway.

Lipopolysaccharide (LPS) inhalation (30 microg ml(-1), 1 h) caused airway hypereactivity (AHR) to histamine (1 mM, 20 s) 1 h later in conscious guinea-pigs. Bronchoalveolar lavage fluid (BALF) levels of neutrophils, myeloperoxidase (MPO) and protein were elevated whereas nitric oxide (NO) metabolites were reduced 1 h after LPS compared with saline challenge. 24 h after LPS, there was no AHR, but BALF neutrophils, eosinophils, macrophages, MPO, protein and NO metabolites were all raised. Budesonide (0.7 mM) and a molar equivalent concentration of the NO-donating budesonide derivative, NCX 1020, were inhaled (15 min) at 24 h and 45 min before LPS. The only change produced by budesonide was to reduce eosinophil influx at 24 h after LPS, compared with vehicle treated animals. NCX 1020, however, blocked AHR and reduced neutrophils (1 and 24 h) and MPO (1 and 24 h), while NO levels were raised at 1 and reduced at 24 h after LPS. The combined inhalation before LPS of the NO donor, SNAP (1.4 mM), with budesonide (0.7 mM) blocked the AHR to histamine and significantly reduced neutrophils (1 and 24 h) and MPO (1 and 24 h), while NO levels were raised at 1 h after LPS. Thus, NO and a corticosteroid co-administered as NCX 1020 or budesonide with a NO donor, have an additive effect against LPS-induced inflammatory responses and may have value in the treatment of neutrophil-driven airways disease such as COPD.

The effects of endogenous adenosine and adenosine receptor agonists were examined on hypoxia-induced myocardial stunning of guinea-pig isolated paced left atria and papillary muscles. Hypoxia (30 minutes) reduced developed tension and increased diastolic tension (contracture) of left atria (41.8 ′ 11.5%) and papillary muscles (17.7 6.2%). Developed tension recovered to 80.8 ′ 3.15 and 77.2 ′ 5.3% 15 minutes after reoxygenation (stunning). Recovery of left atria was unaffected by adenosine deaminase (1 IU mL - 1 ) but was depressed in papillary muscles (15 minutes, 48.6 ′ 4.3%) and contracture (46.1 ′ 7.5%) increased. Endogenous adenosine therefore protects from ventricular but not atrial stunning. Adenosine receptor agonists were introduced at 10 minutes into hypoxia. CPA (A 1 selective, 3 × 10 8 M) impaired left atrial recovery (5 minutes, 38.1 ′ 5.0%), through direct negative inotropy, but did not affect papillary muscles. CGS21680 (A 2 A selective, 3 × 10 7 M) did not affect recovery. APNEA (A 1 /A 3 receptor agonist, 10 8 M), increased recovery rate of left atria. Improved rate and extent of recovery of papillary muscles by APNEA (15 minutes, 94.8 ′ 3.1%) was prevented by the A 3 receptor antagonist, MRS-1220 (10 - 7 M). IB-MECA (A 3 selective, 3 × 10 - 7 M) increased atrial recovery rate but not the maximum developed tension reached in either tissue. However, when added at reoxygenation, IB-MECA caused complete recovery of both tissues, in the absence or presence of adenosine deaminase. Thus, A 3 receptor stimulation reverses myocardial stunning of isolated atria and papillary muscles.

Background Guinea-pig models can provide the essential features of asthma, including early- (EAR) and late- (LAR) phase asthmatic responses, airway hyper-reactivity (AHR) and inflammatory cell influx; however, these components are rarely demonstrated all in the same model. Objectives The aim of this study was to establish a conscious guinea-pig model with these essential features of asthma and to correlate these with bronchoalveolar lavage fluid (BALF) histamine and nitric oxide (NO) levels. The model would be validated from the susceptibility of these parameters to standard anti-asthmatic agents, the steroid, dexamethasone, and a phosphodiesterase-4 (PDE4) inhibitor, rolipram. Methods Guinea-pigs were sensitized with ovalbumen (OA) (10 μg plus Al2(OH)3 100 mg, intraperitoneal (i.p.)) and 14 days later received inhaled OA (100 μg/mL) or vehicle for 1 h. Airway function was measured by whole-body plethysmography as specific airway conductance (sGaw). Reactivity to inhaled histamine (nose-only, 1 mm, 20 s) was recorded 24 h before and at 6, 12 or 24 h after OA challenge. BALF was obtained to determine the total and differential cell counts, NO and histamine. Results Guinea-pigs challenged with OA showed an EAR as a fall in (sGaw) (−54.9±10.8%), which resolved by 6 h and was followed by an LAR between 7 and 11 h (−30.2±8.8%). No bronchoconstriction to inhaled histamine occurred before OA challenge but at 6, 12 or 24 h afterwards, sGaw fell significantly, indicating AHR. At 1 h after OA, macrophages, eosinophils and neutrophils significantly increased in BALF. Macrophages and eosinophils increased further up to 24 h (3- and 44-fold), but neutrophils declined to control levels. BALF histamine levels increased at 0.25 h after OA challenge and peaked at 6 h. BALF NO levels initially fell (44%) 1 h after OA exposure and then progressively rose above control levels. Dexamethasone (20 mg/kg, i.p.) and rolipram (1 mg/kg, i.p.) administered 24 and 0.5 h before and 6 h after OA challenge inhibited leucocyte influx, AHR and the early deficiency and later excess of NO. Dexamethasone but not rolipram attenuated the LAR. Conclusions This model displays many of the features of human asthma with predictable responses to dexamethasone and evidence of anti-asthmatic activity by the PDE4 inhibitor, rolipram.

The mediators of bronchoconstriction of isolated lungs and trachea from ovalbumen sensitized guinea-pigs to adenosine and ovalbumen were examined using relevant antagonists. Changes in perfusion pressure and tension of paired lung halves and tracheal spiral strips, respectively, were recorded in response to adenosine (1 mM lung, 300 microM trachea), histamine (10 microM), methacholine (10 microM) and ovalbumen (10 microg). One half was perfused with antagonist while the other received vehicle. Tracheal strips were superfused throughout with the P(1) receptor antagonist 8-phenyltheophylline, to examine 8-phenyltheophylline-resistant responses. The histamine H(1) receptor antagonist, mepyramine (1.5 mM), the cyclooxygenase inhibitors, indomethacin (5 mM) and diclofenac (5 mM), the leukotriene receptor antagonist, zafirlukast (1 mM), and the lipoxygenase inhibitor, zileuton (20 mM), alone failed to inhibit bronchoconstriction by adenosine and ovalbumen of the lung and trachea. When two antagonists were combined, only mepyramine and zafirlukast significantly reduced the lung responses to adenosine and ovalbumen. The tracheal adenosine response was substantially reduced, although not significantly, while ovalbumen was significantly reduced. When mepyramine, indomethacin and zafirlukast were combined, the lung constriction by adenosine and ovalbumen were virtually abolished. Similarly, the combination of mepyramine, diclofenac and zafirlukast significantly attenuated the lung responses to adenosine and ovalbumen. Thus, histamine, cyclooxygenase products and leukotrienes alone are not responsible for the bronchoconstriction of isolated sensitized lung tissues to adenosine or ovalbumen, which appears to be due to the release of all three mediators.