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K J Broadley

Publications and source records attributed to K J Broadley.

At least 19 recordsLinked to original sources
PubMed2006

Investigation of the mechanism for the relaxation of rat duodenum mediated via M1 muscarinic receptors.

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 mum antagonized the relaxant responses to alpha,beta-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 microm), 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 microm), but not those by McN-A-343. 4 Release of gamma-aminobutyric acid (GABA) was examined with the GABAA receptor antagonist, bicuculline (10 microm), 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 microm) 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.

(4-(m-Chlorophenylcarbamoyloxy)-2-butynyl)trimethy↗
PubMed2006

Radioligand binding and functional responses of ligands for human recombinant adenosine A(3) receptors.

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.

Adenosine↗
PubMed2005

Khat chewing is a risk factor for acute myocardial infarction: a case-control study.

AIM: Khat chewing is a common habit in Yemen and east African countries. Millions of people chew khat leaves daily for its euphoric and energetic effects and to increase alertness. Cathinone, the main active substance in fresh khat leaves, has sympathomimetic effects which increase heart rate and blood pressure. The aim was to examine the hypothesis that khat chewing is a risk factor for acute myocardial infarction (AMI) using a hospital-based matched case-control study. METHOD: Between 1997 and 1999, we selected 100 patients admitted to the Al-Thawra teaching hospital Sana'a ICU, Yemen with acute myocardial infarction. 100 control subjects, matched to cases for sex and age, were recruited from the outpatients clinics of the same hospital. A questionnaire was completed for case and control groups covering personal history of khat chewing, smoking, hypertension, diabetes and any family history of myocardial infarction. A blood sample was collected for performing lipid profiles. Cases and controls were compared by analysis conducted using conditional logistic regression which corrected for baseline imbalances leading to less biased estimations of odds ratio (OR). The risk associated with each classical factor and khat chewing habits was then investigated. OR values greater than 2.5 indicated a significant risk factor. RESULTS: Khat chewing was significantly higher among the AMI case group than control group (OR = 5.0, 95% CI 1.9-13.1). A dose-response relationship was observed, the heavy khat chewers having a 39-fold increased risk of AMI. CONCLUSION: This study indicates that khat chewing is associated with AMI and is an independent dose-related risk factor for the development of myocardial infarction.

Adult↗
PubMed2004

Early and late bronchoconstrictions, airway hyper-reactivity, leucocyte influx and lung histamine and nitric oxide after inhaled antigen: effects of dexamethasone and rolipram.

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 microg plus Al2(OH)3 100 mg, intraperitoneal (i.p.)) and 14 days later received inhaled OA (100 microg/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.

Administration, Inhalation↗
PubMed2004

The roles of alpha- and beta-adrenoceptor stimulation in myocardial ischaemia.

beta-Adrenoceptor (AR) ligands have been the mainstay of cardiovascular therapy for decades, with beta-AR antagonist being used for hypertension, angina and myocardial infarction and adrenaline in use for cardiopulmonary resuscitation for nearly 100 years. Ischaemia of the heart through coronary artery occlusion causes cell injury and death through necrosis and apoptosis. Reperfusion of the ischaemic myocardium results in cardiac dysfunction and infarction. Stimulation of alpha- and beta-ARs in the ischaemic heart have variable and inconsistent effects depending on when the agonist is applied. This review describes the different effects of stimulation of the three established beta-AR subtypes (beta(1)-, beta(2)- and beta(3)-ARs) either before ischaemia (preconditioning) or during ischaemia and reperfusion of the heart (postconditioning). Brief periods of ischaemia preceding a major ischaemic episode can have a protective effect against post-ischaemia-reperfusion damage, known as ischaemic preconditioning. This review considers the role of endogenous catecholamines released during preconditioning and the nature of the adrenoceptor subtypes that mediate these effects. The clinical significance of this to the use of beta-AR antagonists is considered. The transduction pathways and effects on apoptosis of the cardioprotective and deleterious effects of AR activation are considered. This commentary reviews the literature and attempts to bring together a unified synopsis of the effects of adrenoceptor stimulation in myocardial ischaemia and the potential clinical relevance.

Adrenergic alpha-Agonists↗
PubMed2002

Nitric oxide in respiratory diseases.

The formation and modulation of nitric oxide (NO) in the lungs is reviewed. Its beneficial and deleterious roles in airways diseases, including asthma, chronic obstructive pulmonary disease, and cystic fibrosis, and in animal models is discussed. The pharmacological effects of agents that modulate NO production or act as NO donors are described. The clinical pharmacology of these agents is described and the therapeutic potential for their use in airways disease is considered.

Animals↗
PubMed2001

A3 receptors mediate rapid inflammatory cell influx into the lungs of sensitized guinea-pigs.

BACKGROUND: Inhaled adenosine causes bronchoconstriction in asthmatics and may modulate inflammatory cell activity. Elevated adenosine levels occur in the lungs after antigen challenge of asthmatics. OBJECTIVES: The aim of this study was to investigate whether the bronchoconstrictor effects of the adenosine derivative, 5'-AMP, were associated with altered migration of inflammatory cells into the airways using a sensitized atopic guinea-pig model previously shown to display a bronchoconstrictor response. Comparisons were made with the effects of inhaled antigen. METHODS: Airway responses of conscious sensitized guinea-pigs to inhalation exposures of 5'-AMP were determined by whole body plethysmography as the change in specific airway conductance (sGaw). Influx of leucocytes into the airways was determined by bronchoalveolar lavage (BAL). RESULTS: 5'-AMP caused bronchoconstrictor airway responses in sensitized animals. Dose-dependent infiltration of inflammatory cells into the lungs occurred 1 h after 5'-AMP exposure. No bronchoconstriction or cell influx was seen in unsensitized guinea-pigs. Exposure to ovalbumin (OA) also caused influx of inflammatory cells. Twenty-four hours after an OA exposure, 5'-AMP produced no bronchoconstriction. The P1-receptor antagonists, 8-PT and 8-SPT, inhibited the 5'-AMP-induced bronchoconstriction, indicating that the bronchoconstriction seen in sensitized animals is mediated by A1 or A2 receptors. They had no effect on the cell influx, whereas the A3 antagonist, MRS-1220, significantly inhibited cellular infiltration, suggesting mediation through A3 receptors. At 24 h after an OA challenge and accompanying the cellular influx, there was airway hyper-responsiveness to the bronchoconstriction by histamine. In contrast, no hyper-responsiveness to histamine was seen 1 h after 3 mM or 24 h after 300 mM 5'-AMP. CONCLUSIONS: 5'-AMP caused a rapid migration of eosinophils and macrophages into the airways only in sensitized guinea-pigs, and this was blocked by the A3 antagonist MRS-1220. This was not associated with bronchial hyper-reactivity to histamine.

Adenosine Monophosphate↗
PubMed2001

Inhibition of field stimulation-induced contractions of rabbit vas deferens by muscarinic receptor agonists: selectivity of McN-A-343 for M1 receptors.

Inhibition of the field stimulation-induced twitch responses of the rabbit vas deferens by the muscarinic receptor agonist, McN-A-343, has been attributed to presynaptic muscarinic receptors of the M1 subtype located on noradrenergic nerve terminals. Stimulation of these receptors causes inhibition of transmitter release and inhibition of the contractile response. However, the selectivity of McN-A-343 for M1 receptors has been questioned and this throws doubt on whether the prejunctional receptors of the rabbit vas deferens are of the M1 subtype. In this study we have undertaken a comprehensive re-evaluation of the inhibition of prostatic and epididymal portions of the rabbit isolated field-stimulated vas deferens by several agonists, including McN-A-343, and quantified the antagonism by M1-selective antagonists, pirenzepine and telenzepine. Prostatic and epididymal portions of vasa deferentia from New Zealand White rabbits were immersed in a low Ca2+ Krebs solution at 32+/-0.5 degrees C gassed with 5% CO2 in oxygen. Yohimbine (1.0mM) was present throughout to block prejunctional alpha2-adrenoceptors. Field stimulation was applied by repeated application of single pulses (30 V, 0.05 Hz, 0.5 ms) and isometric contractions recorded. Carbachol and oxotremorine initially potentiated the epididymal contractions but at higher concentrations there was inhibition. In the prostatic portion, oxotremorine only inhibited. McN-A-343 produced inhibitory responses only in both epididymal and prostatic portions. Pirenzepine shifted the concentration-response curves forthe inhibitory responses to oxotremorine to the right. However, the potentiation of the twitches also became more apparent with the lower concentrations of oxotremorine. Schild plots for the antagonism by pirenzepine yielded pA2 values of 7.96+/-0.004 and 7.7+/-0.02 for the epididymal and prostatic portions, respectively. The concentration-response curves for the inhibition of twitches by McN-A-343 were displaced to the right in a parallel manner by pirenzepine in both prostatic and epididymal portions with no potentiation of the twitches. The Schild plot for this antagonism generated pA2 values of 7.68+/-0.01 and 8.07+/-0.01, respectively. Telenzepine caused parallel shifts of the McN-A-343 concentration-response curves to the right in prostatic portions, the pA2 value being 8.70+/-0.13. Telenzepine (10(-7) M) abolished the inhibitory effect of carbachol to reveal only concentration-dependent potentiation of the contractions. The Schild plot for antagonism of this contractile effect yielded a pA2 value (7.07+/-0.09) that was significantly less by almost two orders of magnitude (1.70) than the value for the antagonism by telenzepine of the McN-A-343-induced inhibitory response. The pA2 values of pirenzepine and telenzepine against the inhibitory responses of the rabbit vas deferens are consistent with the involvement of M1 receptors. This leads to the conclusion that McN-A-343 causes inhibition through this receptor type. The doubts concerning the selectivity of McN-A-343 for M1 receptors are therefore unfounded. The fact that McN-A-343 does not display a selective binding profile suggests that its selectivity does not arise from affinity differences but probably resides in its intrinsic efficacy.

(4-(m-Chlorophenylcarbamoyloxy)-2-butynyl)trimethy↗
PubMed2001

Chronic lipopolysaccharide exposure on airway function, cell infiltration, and nitric oxide generation in conscious guinea pigs: effect of rolipram and dexamethasone.

This study investigated whether a correlation between airway hyperreactivity (AHR), leukocyte influx, and nitric oxide (NO) existed in guinea pigs chronically exposed to lipopolysaccharide (LPS). The effect of the corticosteroid, dexamethasone, or phosphodiesterase-4 (PDE4) inhibitor, rolipram, on these features was studied. Airway function was measured in conscious guinea pigs (specific airways conductance) before and after single, double, or chronic (nine) LPS (30 microg x ml(-1), 1 h) exposures. Airway reactivity to inhaled histamine (1 mM, 20 s) was assessed before and at various times after LPS challenges. Leukocytes and NO metabolites were measured in bronchoalveolar lavage fluid (BALF). AHR occurred at 1 h after a single LPS challenge and was resolved by 4 h. This coincided with reduction and recovery, respectively, of BALF NO levels. The AHR and NO deficiency were extended to 4 h, after a double LPS exposure. Chronic LPS exposures, 48 h apart, initially caused persistent bronchodilations, whereas later exposures produced progressively persistent bronchoconstrictions. There was AHR 24 h after the eighth challenge. Twenty-four hours after the ninth LPS exposure, macrophages, neutrophils, eosinophils, and NO metabolites were elevated in BALF. Dexamethasone (20 mg x kg(-1) i.p.) or rolipram (1 mg x kg(-1) i.p.) prevented single and chronic LPS-induced AHR, the respective deficiency and elevation in NO metabolites, and the chronic LPS-induced leukocyte influx. Dexamethasone exacerbated, whereas rolipram reversed, the chronic LPS-induced bronchoconstrictions. This study demonstrates for the first time that chronic LPS causes persistent bronchoconstriction, neutrophilic inflammation, AHR, and NO overproduction in guinea pig airways. These rolipram-sensitive features suggest the potential of PDE4 inhibitors in airway disease.

Animals↗
PubMed2001

Responses to adenosine of isolated transverse or spiral strips of sensitized guinea pig trachea: role of epithelium.

AIM: To determine the role of the epithelium in the responses to adenosine of isolated trachea from ovalbumin-sensitized guinea pigs. METHODS: Spirally cut tracheal preparations were superfused or immersed in organ baths and transversely cut strips were immersed. Epithelium was removed mechanically from some strips and confirmed by histological examination of a random sample. Tissues were from unsensitized or ovalbumin-sensitized guinea pigs. Isometric tension was measured and responses to adenosine recorded. RESULTS: In sensitized tissues, contractile responses to adenosine were evident as contractions of superfused spirals or as rightwards shift of the concentration-response curve compared with non-sensitized immersed spirals. Epithelium removal potentiated relaxation responses in both non-sensitized and sensitized strips indicating release of contractile mediators in both tissues. Dipyridamole potentiated relaxation responses in sensitized tissues with and without epithelium. CONCLUSION: Sensitization reveals a contractile response to adenosine. The epithelium is not involved in this contractile response nor is it the major site of uptake of adenosine in both sensitized and non-sensitized tissues.

Adenosine↗
PubMed2000

The ligand-receptor-G-protein ternary complex as a GTP-synthase. steady-state proton pumping and dose-response relationships for beta -adrenoceptors.

Steady-state solutions are developed for the rate of G alpha.GTP production in a synthase model of the ligand-receptor-G-protein ternary complex activated by a ligand-receptor proton pumping mechanism. The effective rate, k(31), defining the proton transfer, phosphorylation and G alpha.GTP release is a controlling rate of the synthase in the presence of a ligand with an efficient mode of signal activation, the ligand-receptor interaction taking place under effectively equilibrium conditions. The composite rate, however, becomes an amplifying factor in any dose-response relationship. The amplification is a triple product of the rate, k(31), the equilibrium constant associated with the activation of the proton signal, K(act)and the fraction of agonist conformer transmitting the signal, f(*). Where the rate of activation of the proton signal becomes critically inefficient, the rate of activation, k(act 1)replaces k(31)K(act). A correlation between beta(1)-adrenergic receptor-stimulated GDP release and adenylate cyclase activation shows that this correlation is not unique to an exchange reaction. Within the initiating Tyr-Arg-Tyr receptor proton shuttle mechanism, the position of Arg(r156) paralleldictates the high-(R(p)) and low-(R(u)) ligand-binding affinities. These states are close to R(*)and R(0)of the equilibrium model (De Lean et al., 1980, J. Biol. Chem.255, 7108-7117). An increased rate of hydrogen ion diffusion into a receptor mutant can give rise to constitutive activity while increased rates of G-protein release and changes in receptor state balance can contribute to the resultant level of action. Constitutive action will arise from a faster rate of G-protein release alone if proton diffusion in the wild-type receptor contributes to a basal level of G-protein activation. Competitive ligand-receptor occupancy for constitutive mutants shows that, where the rate of G-protein activation from the proportion of ligand-occupied receptors is less than the equivalent rate that would be generated from this fraction by proton diffusion, inverse agonism will occur. Rate-dependent dose-responses developed for the proposed synthase mechanism give explicit definition to the operational model for partial agonism (Black & Leff, 1983, Proc. Roy. Soc. Lond. B220, 141-162). When comparable ligands have effectively identical conformational states at the transition state for signal activation, the antagonist component of the binding "in vitro" can be derived by multiplying the apparent binding constant by (1-e) where e is the maximum stimulatory response. This component should be consistent throughout the tissues.

Dose-Response Relationship, Drug↗
PubMed2000

Airway reactivity, inflammatory cell influx and nitric oxide in guinea-pig airways after lipopolysaccharide inhalation.

1. The aim of this study was to investigate the relationship between airway reactivity, leukocyte influx and nitric oxide (NO), in conscious guinea-pigs after aerosolized lipopolysaccharide (LPS) exposure. 2. Inhaled histamine (1 mM, 20 s), causing no bronchoconstriction before LPS exposure (30 microg ml(-1), 1 h), caused bronchoconstriction at 0.5 and 1 h (P:<0.02) after LPS exposure. This airway hyperreactivity (AHR) recovered by 2 h. In contrast, 48 h after LPS exposure, the response from a previously bronchoconstrictor dose of histamine (3 mM, 20 s) was attenuated (P:<0.01) i.e. airway hyporeactivity (AHOR). 3. Investigation of the cellular content of bronchoalveolar lavage fluid (BALF) from these animals revealed a rapid (0.5 h: 691 fold increase) and progressive neutrophil influx after LPS exposure (24 h: 36.3+/-2.3x10(6) cells per sample), that subsided 48 h later. Macrophages and eosinophils also time-dependently increased (0.5 h: 4.6+/-0.4 and 0.1+/-0.05; 48 h: 31.0+/-6.0 and 1.8+/-0.3x10(6) cells per sample, respectively) after LPS, compared to vehicle exposure (24 h: neutrophils, eosinophils and macrophages: 0.28+/-0.19, 0.31+/-0.04 and 4.96+/-0. 43x10(6) cells per sample, respectively). 4. The combined NO metabolites in BALF, after vehicle (1 h), or LPS (1 h: AHR and 48 h: AHOR) exposure, were respectively increased (41%, P:<0.01), decreased (47%, P:<0.01) and further increased (80%, P:<0.001), compared with naïve animals. 5. Inhaled N(o)-nitro-L-arginine methyl ester (L-NAME: 1.2 and 12 mM, 15 min), reduced BALF NO metabolites 2 h later, but did not cause AHR to histamine (P:>0.05). When L-NAME inhalation followed LPS, AHR was prolonged from 1 h to at least 4 h (P:<0.01). 6. In summary, aerosolized LPS inhalation caused neutrophil and macrophage airways infiltration, and an early development of AHR followed 48 h later by AHOR to histamine. AHR and AHOR coincided with a respective reduction and elevation in airways NO (metabolites). Thus, NO may aid recovery from AHR, as inhibition of its production prolongs AHR. However, NO deficiency alone is not responsible for LPS-induced AHR.

Administration, Inhalation↗
PubMed1999

Resistance to antagonism of atrial P(1) purinoceptor responses in the presence of K(+) channel blockade.

The rate of onset of the negative inotropic responses of guinea-pig isolated paced atria to the adenosine receptor agonist, N(6)-cyclopentyladenosine, was significantly slowed by the K(+) channel inhibitor, 4-aminopyridine (10 mM). The concentration-dependent inhibition of developed tension by N(6)-cyclopentyladenosine, however, was unaffected by 4-aminopyridine (10 mM). Thus, K(+) efflux only governs the speed of onset of the negative inotropic response and does not appear to be a major component in the negative inotropy produced by the adenosine A(1) receptor agonist. The P(1) purinoceptor antagonist, 8(p-sulfophenyl) theophylline (1 x 10(-5) M) significantly shifted the concentration-response curve for N(6)-cyclopentyladenosine to the right (concentration-ratio, 7.1+/-1.5). In the presence of 4-aminopyridine (10 mM), 8(p-sulfophenyl) theophylline caused a non-parallel rightwards shift of the curve. At the IC(35) there was no significant shift, whereas at the IC(75) there was a small significant displacement of the curve. The adenosine A(1)/A(3) receptor agonist, N(6)-2-(4-aminophenyl) ethyladenosine (APNEA) yielded a biphasic concentration-response curve which was significantly shifted to the right by 8(p-sulfophenyl) theophylline (1 x 10(-5 ) M). In the presence 4-aminopyridine, however, there was no shift of the APNEA concentration-response curve by 8(p-sulfophenyl)theophylline. These results show that when K(+) channels are blocked by 4-aminopyridine, the residual response is resistant to antagonism by the P(1) purinoceptor antagonist, 8(p-sulfophenyl)theophylline. This residual component may involve L-type Ca(2+) channels, the adenosine A(1) receptor being possibly coupled to the two transduction pathways for negative inotropism via the different components of the G protein (receptor-transducer promiscuity).

4-Aminopyridine↗
PubMed1999

Temporal relationships between leukocytes, IL-5 and IL-8 in guinea pig lungs, plasma cortisol and airway function after antigen challenge.

OBJECTIVE AND DESIGN: The aim was to determine the time courses for the changes in airway function, airway reactivity, influx of inflammatory cells and levels of the pro-inflammatory cytokines, interleukin (IL)-5 and IL-8 in bronchoalveolar lavage fluid (BALF), and the plasma levels of cortisol and ACTH after antigen challenge to determine whether a temporal link could be established between these events. METHODS: Airway function was measured as specific airway conductance (sGsw) in conscious ovalbumin (OvA)-sensitized guinea pigs using whole body plethysmography at intervals after an inhalation challenge with ovalbumin (0.5% for 10 min). Airway responses to the inhaled spasmogen, U46619 (30 ng/ml, 60 s), were measured at 3, 6 and 24 h after challenge. In separate animals, bronchoalveolar lavage fluid (BALF) was obtained after anaesthetic overdose either before challenge or at 1, 3, 6, 12, or 24 h after OvA challenge. Total and differential cell counts of eosinophils and neutrophils were performed on BALF and levels of IL-5 and IL-8 determined by scintillation proximity assays and ELISA, respectively. Plasma cortisol and ACTH levels were determined by RIA kits in blood removed by cardiac puncture at intervals after challenge. RESULTS: An early phase bronchoconstriction occurred which resolved by 3 h and was followed by a late phase between 17 and 24 h. Airway hyperresponsiveness to inhaled U46619, was evident at 3, 6 and 24 h after antigen challenge. Increased IL-5[BALF] was observed by 60 min post challenge implicating a performed storage site. In contrast, IL-8[BALF] was not raised until 3 h post challenge. There was a significant infiltration of neutrophils and eosinophils by 3 and 6 h, respectively. IL-5[BALF] further increased up to 24 h, during the appearance of the late phase of bronchoconstriction and whilst eosinophilia was maximal. Plasma cortisol levels were increased 1 and 3 hours after antigen challenge, thereafter returning to baseline levels. CONCLUSIONS: The hyperresponsiveness appears to be dissociated from the appearance of eosinophils in lavage fluid. The early appearance of IL-5, however, could be a trigger for the migration of eosinophils and development of hyper-responsiveness. The increased plasma cortisol levels occurring after antigen challenge were presumably due to the stress involved and these would be expected to exert an endogenous anti-inflammatory effect.

15-Hydroxy-11 alpha,9 alpha-(epoxymethano)prosta-5↗
PubMed1999

Effects of adenosine receptor agonists on induction of contractions to phenylephrine of guinea-pig aorta mediated via intra- or extracellular calcium.

The vasorelaxant actions of adenosine and its analogue, 5'-(N-ethylcarboxamido)-adenosine (NECA), were investigated in guinea-pig isolated aortic rings by addition to the tissue prior to induction of a contraction by the alpha1-adrenoceptor agonist phenylephrine (PE, 3x10(-6) M). The effect was calculated from the ratio (C2/C1) of the contraction to PE before (C1) and in the presence of adenosine or NECA (C2). This was compared with a control ratio obtained at the same time in which no vasorelaxant was present during C2. Experiments were performed in either "normal" or "Ca2+ -free" bathing medium. Both adenosine and NECA caused inhibition of contractions in "normal" and "Ca2+ -free" conditions, the latter indicating that the vasorelaxant action was due in part to inhibition of intracellular Ca2+ mobilization. To determine whether inhibition of influx of extracellular Ca2+ is a target for the vasorelaxation, contractions to PE were obtained in "normal" Ca2+ and in the presence of ryanodine (10(-5) M), which prevents the release of intracellular Ca2+. These contractions were inhibited by NECA indicating that stimulation of A2-receptors by NECA interferes with the influx of Ca2+ via the opening of receptor-operated Ca2+ channels (ROCs). This study has demonstrated that cell surface A2-receptor stimulation in the guinea-pig aorta inhibits phenylephrine-induced contractions by interfering with both the release of intracellular Ca2+ and the influx of extracellular Ca2+, presumably via ROCs.

Adenosine↗
PubMed1999

Analysis of the atypical characteristics of adenosine receptors mediating negative inotropic and chronotropic responses of guinea-pig isolated atria and papillary muscles.

1. Adenosine receptor(s) mediating negative inotropy of paced left atria, isoprenaline-stimulated paced left atria and papillary muscles, and negative chronotropy of spontaneously beating right atria were characterized. 2. Isometric tension of guinea-pig isolated paced left atria and left ventricular papillary muscles and rate of contraction of spontaneously beating right atria were recorded. Papillary muscles were pre-stimulated with isoprenaline (1x10-8 M). Concentration-response curves (CRCs) for tension or rate reduction by N6-cyclopentyladenosine (CPA), the stereoisomers of N6-(2-phenylisopropyl)adenosine ((+)-PIA and (-)-PIA), 5'-(N-carboxamido)adenosine (NECA), N6-2-(4-aminophenyl)ethyladenosine (APNEA) and N6-(3-iodobenzyl)adenosine-5'-N-methyuromide (IB-MECA) revealed a potency order of CPA=(-)-PIA>NECA in right atria and papillary muscles, which is consistent with involvement of A1-receptors. The potency order in left atria was CPA=NECA>(-)-PIA>(+)-PIA>APNEA, which is not typical of A1 adenosine receptors. Weak activity of APNEA and IB-MECA discounts involvement of A3 receptors. 3. pA2 values for the antagonism of CPA by 8(p-sulfophenyl)theophylline (8-SPT) were calculated from Schild plots (log concentration-ratio against log 8-SPT concentration), the unity slopes of which indicated competitive antagonism. The pA2 value in the papillary muscles was significantly higher than for atrial preparations, indicating a possible difference in receptor characteristics between atrial and papillary muscle responses. 4. In left and right atria there was a limit to the displacement of the CPA CRCs at higher concentrations of 8-SPT. The 8-SPT-resistant component of the response is suggested to arise from duality of coupling of a common A1 receptor through either different G proteins or G protein subunits to independent transduction pathways. The results with papillary muscles can be explained by a typical A1 receptor coupled to a single transduction pathway.

Animals↗
PubMed1999

In vivo demonstration of alpha-adrenoceptor-mediated positive inotropy in pithed rats: evidence that noradrenaline does not stimulate myocardial alpha-adrenoceptors.

1. This study examines whether positive inotropy via alpha-adrenoceptors could be observed in vivo in pithed rats. Cardiac contractility was measured as the maximum rate of rise of left ventricular pressure (dP/dt(max)). Heart rate and aortic blood pressure were also recorded. 2. The selective alpha1-adrenoceptor agonists, methoxamine, cirazoline, amidephrine and phenylephrine caused dose-related increases in dP/dt(max). This response was progressively reduced by increasing doses of the alpha1-adrenoceptor antagonist prazosin. However, since the concomitant increase in diastolic blood pressure (DBP) was also blocked, the changes in dP/dt(max) may have been a consequence of increased after load. 3. Adrenaline and noradrenaline also increased dP/dt(max), accompanied by pressor responses. Propranolol (1 mg kg(-1)) antagonized the increase in dP/dt(max) in response to noradrenaline, suggesting beta-adrenoceptor involvement, but not that to adrenaline. The additional presence of prazosin (1 mg kg(-1)) further shifted the dose-response curves for both noradrenaline and adrenaline to the right. 4. Analysis of the increases in dP/dt(max) at predetermined increases in DBP by each agonist revealed three groups of regression lines. Adrenaline in the presence of propranolol and the four selective alpha1-adrenoceptor agonists occupied a common central position. Above this group were adrenaline and noradrenaline in the absence of antagonists; their additional effects on contractility were beta-adrenoceptor-mediated since the regression lines were lowered by propranolol. Clearly below the main group of agonists was noradrenaline in the presence of propranolol. 5. Thus, for a given increase in DBP, adrenaline (in the presence of beta-blockade) and the alpha1-adrenoceptor agonists exert an additional inotropic effect to noradrenaline (also in the presence of beta-blockade). This is concluded to be an alpha-adrenoceptor-mediated increase in cardiac contractility which is not shared by noradrenaline.

Adrenergic alpha-Agonists↗