bluuebunny/crossref_metadata_2025_split huggingface dataset:
Effects of the cysteinyl leukotriene receptor antagonists pranlukast and zafirlukast on tracheal mucus secretion in ovalbumin‐sensitized guinea‐pigs in vitro
Authors: Liu Yu‐Chih , Khawaja Aamir M , Rogers Duncan F 
Published: January 2005

We investigated the inhibitory effects of the cysteinyl leukotriene (CysLT1) receptor antagonists, pranlukast and zafirlukast, on 35SO4 labelled mucus output, in vitro, in guinea‐pig trachea, induced by leukotriene D4 (LTD4) or by antigen challenge of sensitized animals. Agonists and antagonists were administered mucosally, except in selected comparative experiments where drugs were administered both mucosally and serosally to assess the influence of the epithelium on evoked‐secretion.
LTD4 increased 35SO4 output in a concentration‐related manner with a maximal increase of 23 fold above controls at 100 μM and an approximate EC50 of 2 μM. Combined mucosal and serosal addition of LTD4 did not significantly affect the secretory response compared with mucosal addition alone. Neither LTC4 nor LTE4 (10 μM each) affected 35SO4 output. Pranlukast or zafirlukast significantly inhibited 10 μM LTD4‐evoked 35SO4 output in a concentration‐dependent fashion, with maximal inhibitions of 83% at 10 μM pranlukast and 78% at 10 μM zafirlukast, and IC50 values of 0.3 μM for pranlukast and 0.6 μM for zafirlukast. Combined mucosal and serosal administration of the antagonists (5 μM each) gave degrees of inhibition of mucosal‐serosal 10 μM LTD4‐evoked 35SO4 output similar to those of the drugs given mucosally. Pranlukast (0.5 μM) caused a parallel rightward shift of the LTD4 concentration‐response curve with a pKB of 7. Pranlukast did not inhibit ATP‐induced 35SO4 output.
Ovalbumin (10–500 μg ml−1) challenge of tracheae from guinea‐pigs actively sensitized with ovalbumin caused a concentration‐related increase in 35SO4 output with a maximal increase of 20 fold above vehicle controls at 200 μg ml−1. The combination of the antihistamines pyrilamine and cimetidine (0.1 mM each) did not inhibit ovalbumin‐induced 35SO4 output in sensitized guinea‐pigs. Neither mucosal (10 μM or 100 μM) nor mucosal‐serosal (100 μM) histamine had any significant effect on 35SO4 output.
Pranlukast or zafirlukast (5 μM each) significantly suppressed ovalbumin‐induced secretion in tracheae from sensitized guinea‐pigs by 70% and 65%, respectively.
We conclude that LTD4 or ovalbumin challenge of sensitized animals provokes mucus secretion from guinea‐pig trachea in vitro and this effect is inhibited by the CysLT1 receptor antagonists pranlukast and zafirlukast. These antagonists may be beneficial in the treatment of allergic airway diseases in which mucus hypersecretion is a clinical symptom, for example asthma and allergic rhinitis.
Adenosine monophosphate as a mediator of ATP effects at P1 purinoceptors
Authors: Ross Fiona M , Brodie Martin J , Stone Trevor W 
Published: January 2005

When perfused with a medium containing no added magnesium and 4‐aminopyridine (4AP) (50 μM) hippocampal slices generated epileptiform bursts of an interictal nature. We have shown in a previous study that adenosine 5′‐triphosphate (ATP) depressed epileptiform activity and that this effect was blocked by the adenosine A1 receptor antagonist cyclopentyltheophylline but was not affected by adenosine deaminase. This implied that ATP might act indirectly at P1 receptors or at a xanthine‐sensitive P2 receptor. The aim of the present study was to investigate further the action of ATP on epileptiform activity.
ATP can be metabolized by ecto‐nucleotidases to adenosine 5′‐diphosphate (ADP), adenosine 5′‐monophosphate (AMP) and adenosine, respectively. Each of these metabolites can activate receptors in its own right: P2 receptors for ADP and P1 receptors for AMP and adenosine.
We now show that both AMP and ATP (50 μM) significantly decrease epileptiform discharge rate in a rapid and reversible manner. 5′Adenylic acid deaminase (AMP deaminase, AMPase) (0.2 u ml−1), when perfused alone did not significantly alter the discharge rate over the 10 min superfusion period used for drug application. When perfused concurrently with AMP (50 μM), AMP deaminase prevented the depressant effect of AMP on discharge rate.
AMP deaminase, at a concentration of 0.2 u ml−1 which annulled the effect of AMP (50 μM), prevented the inhibitory activity of ATP (50 μM). A higher concentration of ATP (200 μM) depressed the frequency of spontaneous bursts to approximately 30% control and this response was also prevented by AMP deaminase.
Superfusion of the slices with 5′‐nucleotidase also prevented the inhibitory activity of ATP on epileptiform discharges.
The results suggest that AMP mediates the inhibitory effects of ATP on epileptiform activity, a conclusion which can explain the earlier finding that cyclopentyltheophylline but not adenosine deaminase inhibited the effect of ATP. A corollary to this is that, when examining the pharmacology of ATP, care must be taken to inactivate AMP with AMP deaminase, as well as adenosine with adenosine deaminase, before a direct action of ATP on P1 receptors can be postulated. Failure to do so may have led to erroneous conclusions in some previous studies of nucleotide activity on nucleotside receptors.
Pharmacological classification of adenosine receptors in the sinoatrial and atrioventricular nodes of the guinea‐pig
Authors: Meester B J , Shankley N P , Welsh N J , Wood J , Meijler F L , Black J W 
Published: January 2005

The effects of seven agonist and three antagonist adenosine receptor ligands were compared on the guinea‐pig sinoatrial (SA) node (isolated right atrium) and atrioventricular (AV) node (perfused whole heart). Single agonist concentration‐effect curves were obtained to 5′‐N‐ethylcarboxamidoadenosine (NECA), R(−)‐N6‐(2‐phenylisopropyl)adenosine (R‐PIA), N6‐cyclohexyladenosine (CHA), 2‐chloroadenosine (CADO),),S(+)‐N6‐(2‐phenylisopropyl)adenosine (L‐PIA), 2‐phenylaminoadenosine (CV 1808) and N6‐aminoadenosine (MeAdo). Adenosine and/or NECA curves were obtained in the absence and presence of the antagonists 8‐cyclopentyl‐1,3‐dipropylxanthine (DPCPX), 9‐chloro‐2–(2‐furanyl)‐5,6‐dihydro‐1,2,4‐triazolo[1,5‐c]quinazolin‐5‐imine (CGS15943) and N6‐(endonorbornan‐2‐yl)‐9‐methyladenine (N‐0861).
A formal comparison of the agonist and antagonist potency data was made by fitting the data to a straight line using a least squares procedure based on principal components analysis to account for the variance on both axes. The antagonist affinity estimates made on the two assays did not deviate significantly from the line of identity.
The agonist p[A]50 data obtained on the two assays did not deviate from the line of identity, indicating that there were no significant differences in potencies between the two assays. The p[A]50 ratio of R‐PIA and S‐PIA was 1.24±0.09 in the SA node and 1.36±0.11 in the AV node, indicating no difference in the stereoselectivity of the PIA isomers between the two tissues.
The agonist potency and antagonist affinity data obtained are consistent with previous findings showing that the AV and SA node data are pharmacologically indistinguishable and belong to the adenosine A1‐receptor class. No evidence for the reported A3‐receptor was found.
Pharmacological analysis of the activity of the adenosine uptake inhibitor, dipyridamole, on the sinoatrial and atrioventricular nodes of the guinea‐pig
Authors: Meester B J , Shankley N P , Welsh N J , Meijler F L , Black J W 
Published: January 2005

Dipyridamole potentiates the effects of adenosine on the heart by inhibiting adenosine uptake. The effects of dipyridamole on both adenosine and N‐ethylcarboxamidoadenosine (NECA) concentration‐effect (E/[A]) curves were compared on the AV node, in guinea‐pig isolated perfused hearts, and on the SA node, in isolated right atria, by measuring dromotropic and chronotropic responses, respectively. In the absence of dipyridamole, adenosine was significantly more potent on the AV node than SA node (AV p[A]50=4.95±0.10, SA p[A]50=3.62±0.10). In contrast, NECA and adenosine in the presence of dipyridamole were approximately equiactive in the two assays (NECA: AV p[A]50=7.07±0.07; SA p[A]50=7.30±0.08; adenosine: AV p[A]50=6.49±0.08; SA p[A]50=6.27±0.05). Dipyridamole was significantly more potent in enhancing the effects of adenosine on the SA node than on the AV node (pKi values estimated by Kenakin's method (1981): AV node=8.18±0.14; SA node=8.75±0.08).
The difference in pKi values did not appear to be due to dipyridamole expressing other actions because concentrations of dipyridamole which saturated the adenosine transporter had no effect on the NECA E/[A] curves in either assay. However, the test of another assumption of Kenakin's method, that adenosine taken up into cells is pharmacologically inactive, failed on the AV node assay because a significant potentiating interaction was found between adenosine and NECA. The interaction was concentration‐dependent, reciprocal to the extent that pre‐incubation with either agonist potentiated the other and was concluded to be due to an intracellular action of adenosine as the potentiation disappeared in the presence of dipyridamole.
An explanatory model was developed to account for the data obtained using existing pharmacological concepts of ligand action in isolated tissue bioassays. In the model, adenosine, but not NECA, was assumed to be subject to saturable agonist uptake, an uptake which was competitively blocked by dipyridamole. Adenosine and NECA were assumed to act extracellularly at adenosine A1‐receptors. In the AV node, but not the SA node, the adenosine transported into the cells was assumed to potentiate the effects of adenosine A1‐receptor activation. For the AV node assay, the model predicted that potentiation of adenosine by uptake blockade is offset by a simultaneous decrease in potentiation due to the intracellular action of adenosine. All of the experimental data obtained in the study could be accounted for by the model including the apparent differences in potency of adenosine in the absence of dipyridamole and the pKi values for dipyridamole.
Structural properties of phenylethylamine derivatives which inhibit transport‐P in peptidergic neurones
Authors: Al‐Damluji S , Kopin I J 
Published: January 2005

Transport‐P is an antidepressant‐sensitive, proton‐dependent, V‐ATPase‐linked uptake process for amines in peptidergic neurones of the hypothalamus. It is unusual in its anatomical location in postsynaptic neurones and in that it is activated by its substrate (prazosin). This study examined the structural properties of phenylethylamine derivatives which are substrates for transport‐P, as judged by competitive inhibition of the uptake of prazosin 10−6 M in immortalized hypothalamic peptidergic neurones.
A basic amine was essential for activity; absence of the amine or neutralization with a carboxyl group abolished activity. Primary, secondary and tertiary amines were active but quaternary and guanyl amines were inactive.
A phenyl group was essential for activity at transport‐P. Potency at transport‐P was reduced by phenolic hydroxyl groups and enhanced by phenolic halogens. Thus, for maximal potency, the phenyl group should be hydrophobic. Phenolic methoxyl groups had no effect on potency at transport‐P.
A side chain was necessary for activity at transport‐P. Potency at transport‐P was reduced by β‐hydroxyl and enhanced by α‐methyl groups.
These findings further distinguish transport‐P from other amine uptake processes in the brain.
Modulation of airway hyperresponsiveness and eosinophilia by selective histamine and 5‐HT receptor antagonists in a mouse model of allergic asthma
Authors: De Bie J J. , Henricks P A J. , Cruikshank W W. , Hofman G , Jonker E H. , Nijkamp F P. , Van Oosterhout A J M. 
Published: January 2005

Since both histamine and 5‐hydroxytryptamine (5‐HT) can be released by murine mast cells, we investigated the possible role of these autacoids on airway hyperresponsiveness (AHR), eosinophil infiltration and serum‐IgE levels in a murine model of allergic asthma.
Ovalbumin‐sensitized mice were exposed to either ovalbumin (2 mg ml−1) or saline aerosols on 8 consecutive days. Starting one day before the challenge, animals were injected i.p. twice a day with a 5‐HT‐type 1 (5‐HT1) or type 2 (5‐HT2) receptor antagonist (methiotepine, 1.25 or 2.0 mg kg−1 and ketanserin, 12 mg kg−1, respectively) or a histamine‐type 1 (H1) or type 2 (H2) receptor antagonist (mepyramine, 12 or 20 mg kg−1 and cimetidine, 10 or 25 mg kg−1, respectively). Furthermore, animals were injected with a combination of cimetidine and ketanserin or with an α‐adrenoceptor antagonist (phentolamine, 5 mg kg−1).
In vehicle‐treated ovalbumin‐challenged animals airway responsiveness to intravenous injections of methacholine in vivo was significantly (9 fold increase, P<0.01) increased when compared to vehicle‐treated saline‐challenged animals. Furthermore, ovalbumin challenge of vehicle‐treated animals induced a significant increase in both eosinophil numbers in bronchoalveolar lavage (BAL) fluid (0±0, vehicle/saline and 15.0±5.9×104 cells vehicle/ovalbumin, P<0.05) and ovalbumin‐specific IgE levels in serum (157±69 and 617±171 units ml−1, respectively, P<0.05) compared to saline‐challenged mice. Virtually no eosinophils could be detected in saline‐challenged animals after all different treatments.
Treatment with ketanserin or cimetidine resulted in a partial but significant decrease of the ovalbumin‐induced AHR compared to ovalbumin‐challenged controls (P<0.05) and reduced eosinophil infiltration after ovalbumin challenge by 60% and 58%, respectively. The combination of cimetidine and ketanserin almost completely abolished AHR whereas eosinophilia was decreased by 49%. No effects of these antagonists were observed on IL‐16 levels in BAL fluid or on serum antigen‐specific IgE levels. Treatment with either the H1‐receptor, the 5‐HT1‐receptor or the α‐adrenoceptor antagonist, did not decrease the observed ovalbumin‐induced airway responsiveness or eosinophilia in vehicle‐treated animals. Higher doses of either methiotepine (2.0 mg kg−1) or mepyramine (20 mg kg−1) did decrease ovalbumin‐induced eosinophil infiltration (by 67%, P<0.05 and 73%, respectively), whereas no effects of these antagonists were observed on ovalbumin‐specific IgE levels in serum.
From these data it can be concluded that both histamine and 5‐HT play a role in antigen‐induced AHR and eosinophilia in the mouse.
Class specific inhibition of house dust mite proteinases which cleave cell adhesion, induce cell death and which increase the permeability of lung epithelium
Authors: Winton Helen L. , Wan Hong , Cannell Mark B. , Thompson Philip J. , Garrod David R. , Stewart Geoffrey A. , Robinson Clive 
Published: January 2005

House dust mite (HDM) allergens with cysteine and serine proteinase activity are risk factors for allergic sensitization and asthma. A simple method to fractionate proteinase activity from HDM faecal pellets into cysteine and serine class activity is described.
Both proteinase fractions increased the permeability of epithelial cell monolayers. The effects of the serine proteinase fraction were inhibited by 4‐(2‐aminoethyl)‐benzenesulphonyl fluoride hydrochloride (AEBSF) and soybean trypsin inhibitor (SBTI). The effects of the cysteine proteinase fraction could be inhibited by E‐64. No reciprocity of action was found.
Treatment of epithelial monolayers with either proteinase fraction caused breakdown of tight junctions (TJs). AEBSF inhibited TJ breakdown caused by the serine proteinase fraction, whereas E‐64 inhibited the cysteine proteinase fraction.
Agarose gel electrophoresis revealed that the proteinases induced DNA cleavage which was inhibited by the matrix metalloproteinase inhibitor BB‐250. Compound E‐64 inhibited DNA fragmentation caused by the cysteine proteinase fraction, but was without effect on the serine proteinase fraction. Staining of proteinase‐treated cells with annexin V (AV) and propidium iodide (PI) revealed a diversity of cellular responses. Some cells stained only with AV indicating early apoptosis, whilst others were dead and stained with both AV and PI.
HDM proteinases exert profound effects on epithelial cells which will promote allergic sensitization; namely disruption of intercellular adhesion, increased paracellular permeability and initiation of cell death. Attenuation of these actions by proteinase inhibitors leads to the conclusion that compounds designed to be selective for the HDM enzymes may represent a novel therapy for asthma.

British Journal of Pharmacology (1998) 124, 1048–1059; doi:10.1038/sj.bjp.0701905
The 5‐HT1‐like receptors mediating inhibition of sympathetic vasopressor outflow in the pithed rat: operational correlation with the 5‐HT1A, 5‐HT1B and 5‐HT1D subtypes
Authors: Villalón Carlos M. , Centurión David , Rabelo Gonzalo , de Vries Peter , Saxena Pramod R. , Sánchez‐López Araceli 
Published: January 2005

It has been suggested that the inhibition of sympathetically‐induced vasopressor responses produced by 5‐hydroxytryptamine (5‐HT) in pithed rats is mediated by 5‐HT1‐like receptors. The present study has re‐analysed this suggestion with regard to the classification schemes recently proposed by the NC‐IUPHAR subcommittee on 5‐HT receptors.
Intravenous (i.v.) continuous infusions of 5‐HT and the 5‐HT1 receptor agonists, 8‐OH‐DPAT (5‐HT1A), indorenate (5‐HT1A), CP 93,129 (5‐HT1B) and sumatriptan (5‐HT1B/1D), resulted in a dose‐dependent inhibition of sympathetically‐induced vasopressor responses.
The sympatho‐inhibitory responses induced by 5‐HT, 8‐OH‐DPAT, indorenate, CP 93,129 or sumatriptan were analysed before and after i.v. treatment with blocking doses of the putative 5‐HT receptor antagonists, WAY 100635 (5‐HT1A), cyanopindolol (5‐HT1A/1B) or GR 127935 (5‐HT1B/1D). Thus, after WAY 100635, the responses to 5‐HT and indorenate, but not to 8‐OH‐DPAT, CP 93,129 and sumatriptan, were blocked. After cyanopindolol, the responses to 5‐HT, indorenate and CP 93,129 were abolished, whilst those to 8‐OH‐DPAT and sumatriptan (except at the lowest frequency of stimulation) remained unaltered. In contrast, after GR 127935, the responses to 5‐HT, CP 93,129 and sumatriptan, but not to 8‐OH‐DPAT and indorenate, were abolished.
In additional experiments, the inhibition induced by 5‐HT was not modified after 5‐HT7 receptor blocking doses of mesulergine.
The above results suggest that the 5‐HT1‐like receptors, which inhibit the sympathetic vasopressor outflow in pithed rats, display the pharmacological profile of the 5‐HT1A, 5‐HT1B and 5‐HT1D, but not that of 5‐HT7, receptors.
ATP and vasoactive intestinal polypeptide relaxant responses in hamster isolated proximal urethra
Authors: Pinna Christian , Puglisi Lina , Burnstock Geoffrey 
Published: January 2005

Nitric oxide (NO) is known from previous studies to be the principle transmitter in NANC inhibitory nerves supplying the hamster urethra. However, the identity of the cotransmitter(s) responsible for the responses remaining following block with l‐NG‐nitroarginine methyl ester (l‐NAME) is not known.
Electrical field stimulation (EFS) of circular strips of hamster proximal urethra precontracted with arginine vasopressin (AVP 10−8 m), and in the presence of phentolamine (10−6 m), propranolol (10−6 m) and atropine (10−6 m), caused frequency‐dependent relaxation, which was attenuated by suramin (10−4 m) and reactive blue 2 (RB2; 2×10−4 m), but not by pyridoxalphosphate‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS; 10−4 m), α‐chymotrypsin (10–50 u ml−1) or by the vasoactive intestinal polypeptide (VIP) antagonist, [Lys1, Pro2,5, Arg3,4, Tyr6]‐VIP, (5×10−7–10−6 m). In the presence of indomethacin (10−6 m) frequency‐dependent relaxations to EFS were enhanced, particularly at the lower frequencies of stimulation. EFS‐induced relaxation was blocked by tetrodotoxin (10−6 m), indicating its neurogenic origin.
Exogenous ATP (10−7–10−3 m) produced concentration‐related relaxations which were attenuated by the P2‐purinoceptor antagonists suramin (10−4 m) and RB2 (2×10−4 m) but not by PPADS (10−4 m). ATP‐induced relaxations were also reduced significantly by indomethacin (10−6 m). The inhibitory responses to ATP were urothelium‐ and NO‐independent, since they were not affected by either removal of urothelium or by l‐NAME (10−4 m).
Exogenous VIP (10−9–10−7 m) induced concentration‐related relaxations which were not affected by urothelium removal, l‐NAME (10−4 m), α‐chymotrypsin (10–50 u ml−1) or by [Lys1, Pro2,5, Arg3,4, Tyr6]‐VIP (3×10−7–10−6 m). Nevertheless, suramin (10−4 m) and RB2 (2×10−4 m) but not PPADS (10−4 m) antagonized the VIP‐induced relaxant responses. Calcitonin gene‐related peptide (CGRP: 10−9–10−7 m) was devoid of any effect or only elicited a small relaxant response in AVP‐precontracted strips.
Exogenous prostaglandin E2 (PGE2; 10−9–3×10−6 m) and the NO donor, sodium nitroprusside (SNP; 10−8–3×10−5 m) elicited concentration‐related relaxations on the hamster proximal urethra which were not attenuated by suramin (10−4 m), RB2 (2×10−4 m), or by PPADS (10−4 m), indicating a specific inhibitory effect of the antagonists used.
In summary, these results are consistent with the view that ATP is an inhibitory transmitter released from inhibitory nerves supplying the NANC relaxation of hamster proximal urethra. The relaxant effect of ATP is NO‐ and urothelium‐independent. The present study did not demonstrate whether VIP is released from parasympathetic nerves during EFS, since both α‐chymotrypsin and [Lys1, Pro2,5, Arg3,4, Tyr6]‐VIP were ineffective on neurogenic responses.

British Journal of Pharmacology (1998) 124, 1069–1074; doi:10.1038/sj.bjp.0701908
Autoradiographic visualization of group III metabotropic glutamate receptors using [3H]‐L‐2‐amino‐4‐phosphonobutyrate
Authors: Hudtloff Camilla , Thomsen Christian 
Published: January 2005

In vitroreceptor autoradiography using [3H]‐L‐2‐amino‐4‐phosphonobutyrate ([3H]‐L‐AP4) binding to sections of rat brain has been characterized and shown to most likely represent labelling of group III metabotropic glutamate receptors.Specific [3H]‐L‐AP4 binding to rat brain sections was observed at high densities in the molecular layer of the cerebellar cortex and the outer layer of the superior colliculus. Moderate levels were observed throughout the cerebral cortex, in the molecular layer of the hippocampal dentate gyrus, in thalamus, striatum, substantia nigra and in the medial geniculate nucleus. Low levels of [3H]‐L‐AP4 binding were found in other regions of the hippocampal formation, in the entorhinal cortex and the granule cell layer of cerebellum.Inhibitors of sodium‐ or calcium/chloride‐dependent glutamate uptake did not displace [3H]‐L‐AP4 binding to rat brain sections indicating that the observed binding does not represent [3H]‐L‐AP4 uptake via these carriers. Furthermore, in contrast to [3H]‐L‐AP4 uptake into cerebellar membranes, [3H]‐L‐AP4 binding to brain sections was sensitive to guanosine‐5′‐O‐(3‐thio)trisphosphate‐γ‐S.In the molecular layer of the cerebellar cortex, [3H]‐L‐AP4 binding showed a maximal binding density (Bmax) of 0.52±0.06 pmol mg−1tissue and an affinity (Kd) of 346 nM. The rank order of affinity for displacement of [3H]‐L‐AP4 binding to rat brain sections was:L‐AP4>L‐serine‐O‐phosphate>glutamate>(L)‐2‐aminomethyl‐4‐phosphonobutanoate>(1S,3R)‐1‐aminocyclopentane‐1,3‐dicarboxylate which is in agreement with a group III metabotropic glutamate receptor pharmacology.