bluuebunny/crossref_metadata_2025 huggingface dataset:
Regional haemodynamic responses to intravenous and intraarterial endothelin‐1 and big endothelin‐1 in conscious rats
Authors: Gardiner S.M. , Kemp P.A. , Bennett T.
Published: July 2012

In the same chronically‐instrumented conscious Long Evans rats, we assessed regional haemodynamic responses to i.v. and i.a. injections of endothelin‐1 (ET‐1) and big endothelin‐1 at doses of 0.05 (n = 4) and 0.5 nmol kg−1 (n = 7).

For ET‐1, the cardiovascular effects (initial hypotension and tachycardia with transient hindquarters vasodilatation, followed by hypertension, bradycardia and renal, mesenteric and hindquarters vasoconstrictions) of i.v. and i.a. injections were not different.

Similarly, for big ET‐1, the cardiovascular effects (hypertension, bradycardia and renal, mesenteric, and hindquarters vasoconstrictions) of i.v. and i.a. injections were not different.

At a dose of 0.5 nmol kg−1, i.v. or i.a., the pressor effects of ET‐1 and big ET‐1 were not different (area under curve [AUC0–30 min), ET‐1 i.v. = 786 ± 115 mmHg min; ET‐1 i.a. = 880 ± 165 mmHg min; big ET‐1 i.v. = 878 ± 93 mmHg min; big ET‐1 i.a. = 833 ± 103 mmHg min); but ET‐1 caused greater renal, and lesser hindquarters, vasoconstrictions than big ET‐1 (renal ET‐1 i.v., AOC = 1550 ± 211% min; ET‐1 i.a., AOC = 1746 ± 139% min; big ET‐1 i.v., AOC = 1097 ± 128% min; big ET‐1 i.a., AOC = 1041 ± 119% min; hindquarters, ET‐1 i.v., AOC = 758 ± 176% min; ET‐1 i.a. AOC = 787 ± 184% min; big ET‐1 i.v., AOC = 1270 ± 88% min; big ET‐1 i.a., AOC = 1173 ± 77% min), while the mesenteric vasoconstrictor effects of both peptides were not significantly different (ET‐1 i.v. AOC =1419 ± 132% min; ET‐1 i.a., AOC = 1526 ± 172% min; big ET‐1 i.v., AOC = 1099 ± 166% min; big ET‐1 i.a., AOC = 1155 ± 120% min).

Under the conditions of our experiments, pulmonary clearance of ET‐1, or pulmonary activation of big ET‐1, was not apparent from the haemodynamic responses to i.v. and i.a. administration of the peptides. The results are consistent with previous findings indicating that local, rather than systemic, conversion of exogenous big ET‐1 to ET‐1 is responsible for its regional haemodynamic actions.
Relevance of the use of [3H]‐clonidine to identify imidazoline receptors in the rabbit brainstem
Authors: Bricca G. , Zhang J. , Greney H. , Dontenwill M. , Stutzmann J. , Belcourt A. , Bousquet P.
Published: July 2012

[3H]‐clonidine binding was investigated in membranes isolated from the ventral medulla oblongata of the rabbit where clonidine produced a hypotensive effect which was not mediated by adrenoceptors. [3H]‐clonidine specific binding, as defined by the difference between the binding of [3H]‐clonidine in the presence and in the absence of 10 μm cirazoline, occurred at two sites: a high affinity site with a KD = 2.9 ± 0.7 nm and a Bmax of 40 ± 8 fmol mg−1 protein and a low affinity site with a KD = 18.2 ± 0.4 nm and a Bmax of 66 ± 14 fmol mg−1 protein.

The high affinity sites being catecholamine‐sensitive were identified as α2‐adrenoceptors. The low affinity binding of [3H]‐clonidine was insensitive to catecholamines, as well as to other α2‐adrenoceptor specific probes, and could be inhibited with high affinity only by compounds which lowered blood pressure when directly injected in the nucleus reticularis lateralis of the ventral brainstem, or by antagonists.

It was concluded that in the ventral medulla of the rabbit, [3H]‐clonidine labelled α2‐adrenoceptors and imidazoline receptors (IRs). Only the latter were related to the hypotensive effects of clonidine and rilmenidine directly injected into the rostroventrolateral medulla oblongata (RVLM) of the rabbit. The methodological problems regarding the study of IRs with [3H]‐clonidine are discussed.
5‐HT4 receptor mediated facilitation of the emptying phase of the peristaltic reflex in the guinea‐pig isolated ileum
Authors: Costall B. , Naylor R.J. , Tuladhar B.R.
Published: July 2012

The influence of 5‐hydroxytryptamine (5‐HT) receptor agonists and antagonists on the emptying phase (circular muscle contraction) of the peristaltic reflex was investigated in the guinea‐pig isolated ileum.

The effect of drug application to the serosal surface was measured as the changes in threshold pressure required to trigger the peristaltic reflex and the interval between the peristaltic strokes. A facilitation or inhibition of peristalsis was defined as a reduction or increase in threshold pressure respectively.

Peristalsis was not modified by the inclusion of methysergide (1 μm) and/or ondansetron (2 μm) in the bathing medium. 5‐HT (0.1–1.0 μm) caused a facilitation of the perstaltic reflex; the response curve to 5‐HT was not altered by the presence of methysergide (1 μm) and ondansetron (2 μm).

In the presence of methysergide (1 μm) plus ondansetron (2 μm), 5‐HT (7.36 ± 0.06), 5‐methoxytryptamine (7.01 ± 0.17), 5‐carboxamidotryptamine (5.43 ± 0.06), renzapride (6.09 ± 0.17), (S)‐zacopride (5.99 ± 0.11), (R)‐zacopride (5.61 ± 0.13) and metoclopramide (4.8 ± 0.65) caused a concentration‐related facilitation of the peristaltic reflex, the pEC50 values (mean ± s.e.mean) being shown in parentheses. 2‐Methyl‐5‐HT was ineffective up to 10 μm.

The administration of SDZ 205–557 (1 μm) alone failed to modify the peristaltic reflex, but caused a parallel dextral shift in the concentration‐effect curve to 5‐HT (apparent pKB 7.38 ± 0.30). It failed to modify the effect of acetylcholine to enhance the peristaltic reflex.

It is concluded that the rank order of potency of the 5‐HT agonists from the indole and substituted benzamide series to facilitate the emptying phase of the peristaltic reflex in the guinea‐pig ileum closely correlates with their published actions as 5‐HT4 agonists in other systems. An agonist action on the 5‐HT4 receptor is also supported by the potency of the 5‐HT3/5‐HT4 antagonist SDZ 205–557 (but not the 5‐HT3 antagonist ondansetron) to inhibit the effects of 5‐HT.
Expression of nitric oxide synthase in rat glomerular mesangial cells mediated by cyclic AMP
Authors: Mühl Heiko , Kunz Dieter , Pfeilschifter Josef
Published: July 2012

Treatment of rat mesangial cells with interleukin 1β (IL‐1β) or tumour necrosis factor α (TNFα) has been shown to induce a macrophage‐type of nitric oxide (NO) synthase. Here we report that adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) is another mediator that triggers induction of NO synthase in mesangial cells.

Incubation of mesangial cells with the β‐adrenoceptor agonist, salbutamol, forskolin or cholera toxin, which all activate adenylate cyclase and increase intracellular cyclic AMP concentration, increased nitrite formation in a dose‐dependent manner. Likewise, the addition of the membrane‐permeable cyclic AMP analogue, N6, 0–2′‐dibutyryladenosine 3′,5′‐phosphate (Bt2cyclic AMP) or the phosphodiesterase inhibitor, 3‐isobutyl‐1‐methylxanthine enhanced NO synthase activity in a dose‐dependent manner.

There was a lag period of about 8 h before a significantly enhanced secretion of nitrite could be detected upon exposure of cells to forskolin and for maximal stimulation, forskolin had to be present during the whole incubation period.

Treatment of mesangial cells with actinomycin D, cycloheximide or dexamethasone completely suppressed forskolin‐stimulated NO‐synthase activity, thus demonstrating that transcription and protein synthesis are necessary for nitrite formation.

Bt2 cyclic AMP, the most potent inducer of nitrite production, increased NO synthase mRNA levels in mesangial cells in a time− and dose‐dependent fashion. Dexamethasone completely inhibited the increase of NO synthase mRNA in response to Bt2 cyclic AMP.

Combination of Bt2 cyclic AMP and IL‐1β or TNFα revealed a strong synergy in terms of nitrite formation. Time‐course studies indicated that cyclic AMP needed to be increased during the whole period of IL‐1β stimulation for maximal nitrite production.

These observations suggest that cyclic AMP controls NO synthase expression in mesangial cells. Furthermore, the signalling cascades triggered by IL‐1β and TNFα synergize with the cyclic AMP pathway to stimulate NO synthase activity.
The effects of the endothelin ETA receptor antagonist, FR 139317, on infarct size in a rabbit model of acute myocardial ischaemia and reperfusion
Authors: McMurdo Lorraine , Thiemermann Christoph , Vane John R.
Published: July 2012

The effects were investigated of the ETA receptor antagonist, FR 139317, on endothelin‐1 (ET‐1)‐induced coronary vasoconstriction in the isolated perfused heart of the rabbit. In addition, this study examined whether FR 139317 reduced infarct size in a rabbit model of coronary artery occlusion and reperfusion.

In the rabbit isolated perfused heart, ET‐1 (1–100 pmol) elicited a dose‐dependent increase in coronary perfusion pressure (CPP). For example, 30 pmol ET‐1 caused CPP to rise by 22 ± 8 mmHg and 100 pmol ET‐1 by 47 ± 10 mmHg (n = 8). Infusion of FR 139317 (1 μm) significantly attenuated the increase in CPP caused by ET‐1 (30 pmol: 3 ± 1 mmHg, 100 pmol: 8 ± 2 mmHg; n = 8).

In the anaesthetized rabbit, infarct size (expressed as a percentage of the area at risk) after 45 or 60 min of coronary artery occlusion followed by 2 h of reperfusion was 47 ± 6% (n = 6) and 55 ± 7% (n = 5), respectively. A continuous infusion of FR 139317 (0.2 mg kg−1 min−1 preceded by a loading dose of 1.0 mg kg−1, i.v.; n = 5–6) had no effect on the extent of the myocardial infarct size (45 min: 47 ± 6%; 60 min: 49 ± 7%). Even a three‐times higher dose (0.6 mg kg−1 min−1 preceded by a loading dose of 3 mg kg−1, i.v.; n = 4) of FR 139317 had no effect on myocardial infarct size (48 ± 5%) after 45 min occlusion of the antero‐lateral branch of the left coronary artery (LAL) and 2 h reperfusion.

In a separate group of experiments, the LAL was occluded for 60 min and subsequently reperfused for 6 h. FR 139317 (0.6 mg kg−1 min−1 preceded by a loading dose of 3 mg kg−1, i.v.; n = 4) had no significant effect on infarct size even in this long reperfusion model (control: 48 ± 3%, FR 139317: 61 ± 6%).

Thus, the vasoconstrictor effects elicited by ET‐1 in the coronary vasculature of the rabbit are primarily mediated via the ETA receptor, for they were inhibited by the ETA receptor antagonist, FR 139317. However, an enhanced formation of endogenous ET‐1 does not play a major role in ischaemia/reperfusion injury of the rabbit heart, for FR 139317 had no effect on infarct size.
Anticonvulsant effects of the glycine/NMDA receptor ligands d‐cycloserine and d‐serine but not R‐(+)‐HA‐966 in amygdala‐kindled rats
Authors: Löscher Wolfgang , Wlaź Piotr , Rundfeldt Chris , Baran Halina , Hönack Dagmar
Published: July 2012

The effects of the glycine/NMDA receptor partial agonists, d‐cycloserine and (+)‐HA‐966 and the full agonist, d‐serine, on focal seizure threshold and behaviour have been determined in amygdala‐kindled rats, i.e. a model of focal (partial) epilepsy. The uncompetitive NMDA receptor antagonist, MK‐801, was used for comparison.

The high efficacy glycine partial agonist, d‐cycloserine, did not alter the threshold for induction of amygdaloid afterdischarges (ADT) at doses of 20–80 mg kg−1 i.p., but significant ADT increases were determined after application of higher doses (160 and 320 mg kg−1). The ADT increases after these high doses were long‐lasting; significant elevations were still observed 2 days after drug injection. Determination of d‐cycloserine in plasma and brain tissue showed that it was rapidly eliminated from plasma. Compared to peak levels in plasma, only relatively low concentrations of d‐cycloserine were measured in brain tissue.

The low efficacy glycine partial agonist, (+)‐HA‐966, 10–40 mg kg−1 i.p., did not alter the ADT or seizure recordings (seizure severity, seizure duration, afterdischarge duration) at ADT currents. However, the drug dose‐dependently increased the duration of postictal behavioural and electroencephalographic depression in kindled rats. At the higher dose tested, postictal immobilization was dramatically increased from 3 min to about 120 min. This might indicate that glutamatergic activity is decreased postictally, which is potentiated or prolonged by (+)‐HA‐966.

Like d‐cycloserine, the glycine receptor full agonist, d‐serine, injected bilaterally into the lateral ventricles at a dose of 5 μmol, significantly increased the ADT, while no effect was seen at a lower dose (2.5 μmol).

The anticonvulsant effects observed with d‐cycloserine were completely antagonized by combined treatment with (+)‐HA‐966, indicating that the effects of d‐cycloserine were mediated by the glycine/NMDA receptor complex.

MK‐801, 0.1 mg kg−1, did not alter the focal seizure threshold or seizure recordings at ADT current, but induced marked phencyclidine(PCP)‐like behavioural alterations, such as hyperlocomotion, stereotypies and motor impairment. No PCP‐like behaviours were observed after d‐cycloserine, d‐serine or (+)‐HA‐966. High doses of (+)‐HA‐966 induced moderate motor impairment in kindled rats.

The long lasting increases in seizure threshold observed after the high efficacy glycine partial agonist, d‐cycloserine but not the low efficacy partial agonist, (+)‐HA‐966, may suggest that the effects of d‐cycloserine are mediated by adaptive changes in the NMDA receptor complex in response to glycine receptor stimulation.

Pharmacological intervention at the strychnine‐insensitive glycine receptor by high‐efficacy partial agonists with systemic bioavailability may be an effective means of increasing seizure threshold without concomitantly inducing PCP‐like adverse effects.
Effects of muscarinic M2 and M3 receptor stimulation and antagonism on responses to isoprenaline of guinea‐pig trachea in vitro
Authors: Watson N. , Eglen R.M.
Published: July 2012

In guinea‐pig and canine airway smooth muscle, there is reduced β‐adrenoceptor agonist sensitivity in tissues pre‐contracted with muscarinic agonists when compared to tissues pre‐contracted with other spasmogens, such as histamine or leukotriene D4. This reduced sensitivity may be the result of an interaction between muscarinic receptors and β‐adrenoceptors. In this study the effects of M2 receptor antagonism and stimulation have been investigated on the relaxant potency of isoprenaline in guinea‐pig isolated tracheal smooth muscle.

(+)‐cis‐Dioxolane contracted isolated tracheal strips in a concentration‐dependent manner (EC50 = 11.5 ± 0.9 nm). The rank order of antagonist apparent affinities (with pA2 values in parentheses) was atropine (9.4 ± 0.1) > zamifenacin (8.2 ± 0.1) >para‐fluoro‐hexahydro‐siladiphenidol (p‐F‐HHSiD, 7.2 ± 0.1) > pirenzepine (6.5 ± 0.1) > methoctramine (5.5 ± 0.1). Schild slopes were not significantly different from unity. This was consistent with a role of muscarinic M3 receptors in mediating contraction.

In tissues pre‐contracted to 3 g isometric tension using (+)‐cis‐dioxolane (0.2 μm, approximately EC80), the relaxant potency of isoprenaline was significantly (P < 0.05) increased by 0.3 μm methoctramine (control EC50 = 32.2 ± 4.3 nm, plus methoctramine EC50 = 19.1 ± 4.5 nm). This concentration of methoctramine had no effect on contractile responses to (+)‐cis‐dioxolane (control, EC50 = 17.6 ± 3.2 nm, plus methoctramine, EC50 = 21.0 ± 4.4 nm).

When acetylcholine (non‐selective), (+)‐cis‐dioxolane (non‐selective), L‐660,863 ((±)‐3‐(3‐amino‐1,2,4‐oxadiazole‐5‐yl)‐quinuclidine, M2‐selective) or SDZ ENS 163 (thiopilocarpine, mixed M2 antagonist, partial M3 agonist) were used to achieve isometric tensions of 3 g, the relaxant potency of isoprenaline ranged from 3.7 ± 0.3 nm (SDZ ENS 163) to 49.4 ± 3.2 nm ((+)‐cis‐dioxolane). Reducing the concentration of these agonists (and therefore the level of developed tension to 2 g), significantly (P < 0.05) increased the relaxant potency of isoprenaline. In contrast, when histamine was used to pre‐contract tissues to either 2 or 3 g (EC50 = 4.2 ± 0.6 and 3.8 ± 1.1 nm, respectively), there was no significant effect on the relaxant potency of isoprenaline.

There was a slight but significant (P < 0.05) reduction in the relaxant potency of isoprenaline, in tissues pre‐contracted to 3 g using histamine in combination with (+)‐cis‐dioxolane (30 nm). This effect was reversed by M2 receptor antagonism, using methoctramine (1 μm).

These data suggest that in guinea‐pig isolated trachea, the relaxant potency of isoprenaline may depend not only on the level of developed tension but also, on the level of muscarinic M2 receptor stimulation/blockade of the spasmogen inducing the tension. However, the lack of selective M2 agonist and the low M2/M3 selectivity of antagonists in this tissue do not permit definitive conclusions to be made about the role of these receptors in modulating isoprenaline potency.
Reversal of established responses to endothelin‐1 in vivo and in vitro by the endothelin receptor antagonists, BQ‐123 and PD 145065
Authors: Warner Timothy D. , Allcock Graham H. , Vane John R.
Published: July 2012

Endothelin‐1 binds almost irreversibly to its receptors and causes prolonged vasoconstrictions. Here we have studied the reversal of established responses to ET‐1 in vivo and in vitro by BQ‐123, an ETA receptor‐selective antagonist, and/or PD 145065, an ETA/ETB receptor non‐selective antagonist.

In anaesthetized rats pretreated with hexamethonium, infusion of ET‐1 (10−11 mol kg−1 min−1) increased the mean arterial pressure (MAP) from 93 ± 1.5 mmHg to 137 ± 2.4 mmHg after 70 min (n = 29). While the ET‐1 infusion was continued an additional infusion of BQ‐123 caused a gradual dose‐dependent reduction in the pressor effect of ET‐1. For instance, after a 60 min infusion of BQ‐123 (10−8 mol kg−1 min−1) the MAP was decreased by 29.3 ± 4.3 mmHg (n = 4).

PD 145065 was a much weaker antagonist of the established pressor effects of ET‐1. At 10−8 mol kg−1 min−1 it had no significant effect and even at 10−7 mol kg−1 min−1 the elevated blood pressure was only reduced by 11.8 ± 8.0 mmHg (n = 5) after 60 min. Co‐infusion of BQ‐123 and PD 145065 caused smaller reductions in the established response to ET‐1 than infusion of BQ‐123 alone.

Sustained contractions of rat aortic rings induced by ET‐1 (3 × 10−9 m) and mediated by ETA receptors were slowly reversed by addition of BQ‐123 (10−5 m) or PD 145065 (10−5 m). For instance, after 40 min the elevated tone was reduced 85.8 ± 5.6% (n = 6) by PD 145065, and 77.1 ± 6.7% (n = 6) by BQ‐123. Thus, on the rat aortic rings in vitro both antagonists were equally effective against established responses to ET‐1.

ET‐1 increased the perfusion pressure of the rat isolated perfused kidney by 138.1 ± 7.6 mmHg (n = 14). Subsequent co‐infusion of BQ‐123 or PD 145065 reversed this increase with PD 145065 being more active. For instance, PD 145065 (10−6 m) reversed the increase in perfusion pressure by 56.9 ± 8.8% (n = 5) and BQ‐123 (10−6 m) reversed it by 22.8 ± 8.0% (n = 5). This fits well with the vasoconstriction induced by ET‐1 in the rat kidney being mediated by ETA and ETB receptors.

Thus, sustained vasoconstrictions to ET‐1 in vitro, mediated by either ETA or ETB receptors, may be reversed slowly by the subsequent application of receptor antagonists. Similarly, endothelin antagonists reverse the pressor effects of ET‐1 in vivo although co‐antagonism of ETA and ETB receptors or the co‐administration of an ETA receptor antagonist, BQ‐123, and a mixed antagonist, PD 145065 produces less reversal than the application of an ETA receptor‐selective antagonist. This may be because PD 145065 also reduces vasodilatations induced by ET‐1 in vivo, or could suggest that because of its peptide structure PD 145065 affects the elimination of ET‐1.
The involvement of endothelial dysfunction, nitric oxide and prostanoids in the rat gastric microcirculatory responses to endothelin‐1
Authors: Lopez‐Belmonte J. , Whittle B.J.R
Published: July 2012

The role of endothelial dysfunction in the gastric microcirculatory responses during local endothelin‐1 (ET‐1) infusion has been investigated in the pentobarbitone‐anaesthetized rat. Furthermore, the involvement of prostanoids or nitric oxide (NO) in these actions has been investigated by the use of indomethacin to inhibit cyclo‐oxygenase and NG‐nitro‐l‐arginine methyl ester (l‐NAME) to inhibit NO synthase.

Close‐arterial infusion of ET‐1 (1–10 pmol kg−1 min−1 for 10 min) induced a dose‐dependent increase in the gastric leakage of radiolabeled albumin, used as an index of endothelial cell dysfunction.

Close‐arterial infusion of a submaximal dose of ET‐1 (5 pmol kg−1 min−1 for 10 min) significantly increased gastric albumin leakage after 2 min infusion, which reached maximal levels after 10 min, and only slowly declined during the 30 min observation period.

By contrast, gastric blood flow, as assessed by laser Doppler flowmetry, did not significantly increase until after 5 min of infusion of ET‐1 (5 pmol kg−1 min−1 for 10 min), reaching a maximum after 17 min, and was sustained for the 30 min observation period.

Pretreatment with l‐NAME (2 mg kg−1, i.v.) or indomethacin (5 mg kg−1, i.v.) significantly reduced both the hyperaemic response to ET‐1 and the increase in gastric albumin leakage, and in combination abolished these responses.

These results suggest that locally released NO and prostanoids mediate the gastric vasodilator response to close arterial infusion of ET‐1. This hyperaemia is preceded by changes in gastric albumin extravasation and hence may be initiated as a response to direct endothelial injury by ET‐1.
Effect of a 5‐lipoxygenase inhibitor and leukotriene antagonist (PF 5901) on antigen‐induced airway responses in neonatally immunized rabbits
Authors: Herd Caroline M. , Donigi‐Gale Donna , Shoupe T. Scott , Burroughs D.A. , Yeadon M. , Page Clive P.
Published: July 2012

The effect of a single intratracheal dose (10 mg) of PF 5901 (2‐[3(1‐hydroxyhexyl) phenoxymethyl] quinoline hydrochloride, a specific inhibitor of the 5‐lipoxygenase pathway of arachidonic acid metabolism and a leukotriene D4 antagonist) on airway changes induced in response to Alternaria tenuis aerosol challenge was assessed in adult rabbits neonatally immunized. Leukotriene generation was determined in vivo by measuring leukotriene B4 (LTB4) levels in bronchoalveolar lavage (BAL) fluid and ex vivo by measuring calcium ionophore‐stimulated production of LTB4 in whole blood.

While PF 5901 (10 mg) had no significant effect on the acute bronchoconstriction induced by antigen, this dose was sufficient to inhibit significantly the increase in airway responsiveness to inhaled histamine 24 h following antigen challenge (P < 0.05).

Total leucocyte infiltration into the airways induced by antigen, as assessed by bronchoalveolar lavage, was significantly inhibited by pretreatment with PF 5901 (10 mg). However, the pulmonary infiltration of neutrophils and eosinophils induced by antigen was unaltered by prior treatment with PF 5901 (10 mg).

PF 5901 (10 mg) had no effect on ex vivo LTB4 synthesis in whole blood. However, the antigen‐induced increase in LTB4 levels in BAL 24 h following challenge was significantly inhibited (P < 0.05).

We suggest from the results of the present study that the antigen‐induced airway hyperresponsiveness to inhaled histamine in immunized rabbits is mediated, at least in part, by products of the 5‐lipoxygenase metabolic pathway, and is not dependent on the extent of eosinophil or neutrophil influx into the airway lumen.
The nonpeptide neurotensin antagonist, SR 48692, used as a tool to reveal putative neurotensin receptor subtypes
Authors: Dubuc I. , Costentin J. , Terranova J.P. , Barnouin M.C. , Soubrié P. , Fur G. Le , Rostène W. , Kitabgi P.
Published: July 2012

The nonpeptide neurotensin (NT) antagonist, SR 48692, was recently shown to inhibit NT binding to the cloned rat and human NT receptor and to antagonize NT effects in a variety of in vitro and in vivo assays. Here, we show that, in contrast to its antagonistic action on NT‐induced hypomotility in the rat, SR 48692 failed to antagonize NT‐induced hypothermia and analgesia in the mouse and rat. We suggest that these effects might be mediated through a subtype of SR 48692‐insensitive NT receptor.
Induction of nitric oxide synthase in cultured vascular smooth muscle cells: the role of cyclic AMP
Authors: Hirokawa K. , O'Shaughnessy K. , Moore K. , Ramrakha P. , Wilkins M.R.
Published: July 2012

Interleukin‐1β (IL‐1β) is a potent stimulant of inducible nitric oxide synthase (iNOS) mRNA and nitric oxide (NO) production in vascular smooth muscle (VSM) cells in culture. These studies investigate the role of adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) in this process.

Dibutyryl cyclic AMP (db cyclic AMP, 0.1–1 mm), forskolin (1–10 μm) and the phosphodiesterase inhibitor, Ro 20–1724 (1–10 μm), all of which increase intracellular cyclic AMP, had no effect on NO production when added alone but markedly enhanced NO production by IL‐1β‐stimulated VSM cells in a dose‐dependent manner. Consistent with a cyclic AMP‐mediated action, isoprenaline (1–10 μm) increased NO production from IL‐1β‐stimulated cells. Dibutyryl cyclic GMP (db cyclic GMP) had no effect at concentrations up to 1 mm.

Pursuing these observations, iNOS protein levels were examined by Western blot analysis and iNOS mRNA levels were measured by reverse transcription and amplification of the resultant cDNA using the polymerase chain reaction. In addition to enhancing NO production, db cyclic AMP increased iNOS protein and mRNA above that produced by IL‐1β alone.

These data demonstrate a major effect of cyclic AMP on cytokine‐induced NOS activity in VSM cells, mediated at least in part by regulating synthesis of iNOS, and has implications for the pathogenesis and management of septic shock.
Effect of topical administration of l‐arginine on formalin‐induced nociception in the mouse: a dual role of peripherally formed NO in pain modulation
Authors: Kawabata Atsufumi , Manabe Sachiko , Manabe Yuki , Takagi Hiroshi
Published: July 2012

We investigated the effects of intraplantar (i.pl.) administration of l‐arginine and NG‐nitro‐l‐arginine methyl ester (l‐NAME) on formalin‐induced behavioural nociception in the mouse.

l‐ but not d‐arginine, at 0.1 − 1 μg per paw, coadministered with i.pl. formalin, enhanced the second‐but not the first‐phase nociceptive responses, whereas it was without significant effects at 3 μg per paw, and conversely, produced antinociception at 10 μg per paw, resulting in a bell‐shaped dose‐response curve.

l‐NAME at 0.1 − 1 μg per paw, when administered i.pl., exhibited antinociceptive activity in the second phase in a dose‐dependent manner, although its d‐enantiomer produced no effect.

An antinociceptive dose (1 μg per paw) of l‐NAME (i.pl.) considerably reduced the increase in second‐phase nociception elicited by low doses (1 μg per paw) of i.pl. l‐arginine. The second‐phase nociception decrease induced by a large dose (10 μg per paw) of i.pl. l‐arginine was markedly reversed by i.pl. l‐NAME at 0.1 μg per paw, raising it to a level above that of the control (formalin only).

These results suggest that peripheral NO plays a dual role in nociceptive modulation, depending on the tissue level, inducing either nociceptive or antinociceptive responses.
Adrenoceptors mediating relaxation to catecholamines in rat isolated jejunum
Authors: MacDonald A. , Forbes I.J. , Gallacher D. , Heeps G. , McLaughlin D.P.
Published: July 2012

The characteristics of adrenoceptors mediating relaxation to catecholamines in rat isolated jejunum were investigated.

Catecholamines and BRL 37344 produced relaxation of the KCl‐contracted strips with an order of potency of isoprenaline (1.0) > BRL 37344 (0.63) > noradrenaline (0.1) > adrenaline (0.04).

In the presence of both prazosin (1 μm) and propranolol (1 μm) only small dextral shifts of the concentration‐response curves to agonists were observed and an order of potency of BRL 37344 (2.5) > isoprenaline (1.0) > noradrenaline (0.2) > adrenaline (0.1) was obtained.

In the presence of prazosin (1 μm) and propranolol (1 μm), cyanopindolol (0.1 − 10 μm) produced a concentration‐dependent rightward shift of the concentration‐response curve to adrenaline with a Schild slope not significantly different from unity and a mean pA2 value of 7.01.

The resistance of relaxant responses to propranolol, the relatively high potency of BRL 37344 compared to catecholamines and the competitive antagonism of relaxant responses to adrenaline by cyanopindolol suggest that β‐adrenoceptors in rat small intestine are mainly atypical in nature.
Agonistic and antagonistic properties of the bradykinin B2 receptor antagonist, Hoe 140, in isolated blood vessels from different species
Authors: Félétou Michel , Germain Martine , Thurieau Christophe , Fauchère Jean‐Luc , Canet Emmanuel
Published: July 2012

Hoe 140, a recently described bradykinin B2 antagonist, and NPC 567 from an earlier generation of bradykinin B2 antagonists, were tested in rabbit and sheep isolated blood vessels.

In rabbit jugular vein, a bradykinin B2 preparation, NPC 567 was an antagonist (apparent pA2: 8.67 ± 0.16) with marked residual agonistic activity (log[EC50]: −7.29 ± 0.13). Hoe 140 was a potent non‐competitive antagonist devoid of agonistic properties (slope of the Schild plot: 2.02; estimated pA2: 9.04).

In rabbit aorta, a bradykinin B1 preparation, NPC 567 was a competitive antagonist (pA2: 6.32 ± 0.13) but Hoe 140 was ineffective. The two antagonists did not show any agonistic properties in this tissue.

In sheep femoral artery without endothelium, bradykinin and Hoe 140 induced contractions with identical efficacy and similar potency (log[EC50]: −8.05 ± 0.12, −7.73 ± 0.10; maximal contraction in % of KCl [60 mm]: 59.5 ± 15.1, 62.0 ± 13.1; for bradykinin and Hoe 140, respectively). In contrast NPC 567 was an extremely weak agonist. The contractile responses to bradykinin and Hoe 140 were inhibited by NPC 567 (apparent pKB: 6.89 ± 0.22 and 6.58 ± 0.08 versus bradykinin and Hoe 140, respectively) but not by a B1 bradykinin antagonist, suggesting that the receptor involved was a bradykinin B2 receptor.

In sheep femoral artery with endothelium, bradykinin induced a biphasic response: an endothelium‐dependent relaxation and a contraction which were both inhibited by NPC 567 (apparent pKB: 7.10 ± 0.15) and Hoe 140 (pA2: 8.38 ± 0.12). As bradykinin B2 receptor antagonists, Hoe 140 and NPC 567 were less potent in the sheep femoral artery than in the rabbit jugular vein. Neither Hoe 140 nor NPC 567 were agonists for the endothelial receptor.

This study demonstrates that Hoe 140, a new bradykinin B2 receptor antagonist, is more selective and more potent than NPC 567; however, it may possess, depending on the tissue studied, marked residual agonistic properties. Furthermore, bradykinin B2 receptors are subject to important species specificity. Finally, two different bradykinin B2 receptor subtypes may coexist in the sheep femoral artery with endothelium.
Comparative pharmacology of analogues of S‐nitroso‐N‐acetyl‐dl‐penicillamine on human platelets
Authors: Salas E. , Moro M.A. , Askew S. , Hodson H.F. , Butler A.R. , Radomski M.W. , Moncada S.
Published: July 2012

The effects of two new analogues of S‐nitroso‐N‐acetyl‐dl‐penicillamine (SNAP), S‐nitroso‐N‐formyl‐dl‐penicillamine (SNFP) and S‐nitroso‐dl‐penicillamine (SNPL), on platelet function were examined in vitro.

SNAP and its analogues were potent inhibitors of platelet aggregation and inducers of disaggregation.

All compounds inhibited fibrinogen binding to platelets.

They also decreased the release of P‐selectin from platelets.

Both inhibition of fibrinogen binding and release of P‐selectin correlated with an increase in intraplatelet cyclic GMP concentrations.

At concentrations sufficient to inhibit platelet function and induce cyclic GMP formation (0.01–3 μm), the release of NO could be detected from SNPL but not from SNAP and SNFP.

Release of NO from all compounds was detected at concentrations ≥ 10 μm.

Thus, the spontaneous release of NO from SNPL explains the actions of this compound on platelet function; however, platelet‐mediated mechanisms may be involved in the release of NO from SNAP and SNFP.
Effects of cyclopiazonic acid on rhythmic contractions in uterine smooth muscle bundles of the rat
Authors: Kasai Yasuyo , Iino Masamitsu , Tsutsumi Osamu , Taketani Yuji , Endo Makoto
Published: July 2012

We studied the effects of cyclopiazonic acid (CPA) on rhythmic contractions and on Ca2+ uptake by the intracellular stores in longitudinal muscle strips of the rat uterus at 30°C.

Oxytocin (1 μm) in Ca2+‐free solution induced a transient rise in the intracellular Ca2+ concentration ([Ca2+]i) and contraction after Ca2+ loading of the stores in high‐K+‐ and Ca2+‐containing solution. CPA inhibited oxytocin‐induced Ca2+ release and contraction, the half and full inhibitory concentrations of CPA being 0.3 and 10 μm, respectively. In contrast, addition of CPA after Ca2+ loading exerted no significant inhibitory effects.

Oxytocin (10 nM) applied in Ca2+‐containing solution induced rhythmic increases in both force and [Ca2+]i. CPA (10 μm) had no effect on oxytocin‐induced rhythmic contractions.

At a high concentration (300 μm), CPA inhibited the rhythmic contractions induced by 10 nM oxytocin; the frequency and the peak height were decreased, and in many bundles contractions were completely abolished. These inhibitory effects were reversed after CPA washout.

CPA (300 μm) inhibited the rate of rise of [Ca2+]i due to depolarization induced by high‐K+‐containing solution.

These results suggest that low concentrations of CPA inhibit the loading of Ca2+ into intracellular stores in intact tissue strips, and that the Ca2+ stores are not directly involved in the uterine rhythmic contractions. It is also suggested that a high concentration of CPA inhibits the mechanism that is responsible for the generation of rhythmic contractions as well as voltage‐dependent Ca2+ channels.
Potentiation by forskolin of both SNP‐ and ANP‐stimulated cyclic GMP accumulation in porcine isolated palmar lateral vein
Authors: Wright I.K. , Amirchetty‐Rao S. , Kendall D.A.
Published: July 2012

The aim of this study was to examine the effect of modulation of adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) levels (by using forskolin, a direct activator of adenylyl cyclase, or rolipram, a cyclic AMP selective phosphodiesterase inhibitor) on basal and stimulated guanosine 3′:5′‐cyclic monophosphate (cyclic GMP) levels in the porcine isolated palmer lateral vein by use of a [3H]‐guanine prelabelling technique.

Sodium nitroprusside (SNP; 10−5‐10−3M) and atrial natriuretic peptide (ANP; 10−8‐10−6M), produced concentration‐dependent increases in [3H]‐cyclic GMP levels via stimulation of soluble and particulate guanylyl cyclase respectively. The SNP‐stimulated [3H]‐cyclic GMP response peaked after 5 min in the presence and absence of the phosphodiesterase inhibitor, 3‐isobutyl‐1‐methylxanthine (IBMX).

In the absence of IBMX, forskolin (3 × 10−5 M) significantly increased [3H]‐cyclic GMP levels to 118.5 ± 8.7% of basal values (P < 0.05, n = 8), and significantly increased both the SNP‐ and ANP‐stimulated [3H]‐cyclic GMP accumulation at all concentrations of SNP and ANP used. For example, effects at the maximal SNP (10−3M) and ANP (10−6M) concentrations were: SNP: 154.7 ± 15.4% of basal; SNP + forskolin: 191.3 ± 14.8% of basal (P < 0.05, n = 4); ANP: 161.4 ± 17.4% of basal; ANP‐ + forskolin: 220.0 ± 20.0% of basal (P < 0.05, n = 4).

The cyclic AMP‐selective phosphodiesterase inhibitor, rolipram (10−5 M), had no effect on basal or SNP‐stimulated [3H]‐cyclic GMP levels; however, the combination of forskolin and rolipram produced an increase in the basal (158.7 ± 27.1% of basal) and SNP‐stimulated [3H]‐cyclic GMP accumulation (SNP (10−3M): 165.3 ± 8.7% of basal; SNP + forskolin + rolipram: 510.7 ± 64.8% of basal; P < 0.05, n = 5), greater than either forskolin or rolipram alone. The phosphodiesterase inhibitor, IBMX (10−3 M) significantly raised [3H]‐cyclic GMP levels, and forskolin (3 × 10−5 M) in the presence of IBMX had no significant effect on either basal or SNP‐stimulated [3H]‐cyclic GMP levels (e.g. in the presence of IBMX: SNP (10−3M): 660 ± 90% of basal; SNP + forskolin: 790 ± 86% of basal, n = 3).

The data indicate the presence of both soluble and particulate guanylyl cyclase in the porcine isolated palmar lateral vein. The ability of forskolin to potentiate SNP‐ and ANP‐stimulated [3H]‐cyclic GMP accumulation may suggest a cyclic AMP‐cyclic GMP interaction at the level of the phosphodiesterases. Further, the ability of cyclic AMP to influence cyclic GMP levels may indicate that the two nucleotides, as well as having independent mechanisms to induce smooth muscle relaxation, could produce vasodilatation via a common mechanism.
Mechanism of bile salt vasoactivity: dependence on calcium channels in vascular smooth muscle
Authors: Pak Jung‐Min , Adeagbo Ayotunde S.O. , Triggle Christopher R. , Shaffer Eldon A. , Lee Samuel S.
Published: July 2012

The vasoactive mechanisms of bile salts have been investigated in rat isolated portal venous and superior mesenteric arterial rings and perfused mesentery.

The isolated perfused mesentery was precontracted with a selective α1‐adrenoceptor agonist, cirazoline. Incremental doses of tauroursodeoxycholate (TUDC), taurochenodeoxycholate (TCDC) and taurodeoxycholate (TDC) caused a dose‐dependent vasorelaxation. The order of potency of the vasodilator effect was TDC > TCDC > TUDC.

The effect of TDC (1.9 × 10−8 −1.9 × 10−6 mol) was examined before and after propranolol (3 μm), tetraethylammonium (5 mM), ouabain (10−5 M), NG‐nitro‐l‐arginine methyl ester (10−4 M) and capsaicin (50 mg kg−1) to block, respectively, β‐adrenoceptors, K+‐channels, Na+, K+‐ATPase, nitric oxide synthase, and primary sensory nerves. The vasodilator effect of TDC was not affected by any of these blocking agents or by denuding vascular endothelium with distilled water.

Infusion of TDC (1.9 × 10−8 −1.9 × 10−6 mol) with K+‐free or high K+ (60 mM) physiological salt solution (PSS) did not affect the vasodilator effect of TDC.

Contractions induced by KCl (0.01–1.0 M), arginine vasopressin (AVP, 10−10 −10−7 M) or cirazoline (10−7 × 10−5 M) were all inhibited by TDC (300 μm).

TDC (10−6 to 10−3 M) also inhibited the basal tension and the development of spontaneous contractions in the isolated portal vein.

TDC (300 μm), however, did not affect noradrenaline‐induced phasic contractions elicited in Ca2+‐free PSS by Ca2+ release from intracellular stores.

We conclude that TDC inhibits Ca2+ entry through both voltage‐operated and receptor‐operated calcium channels, whereas intracellular Ca2+ release is not affected.
Antagonism by lipophilic quaternary ions of the K+ channel opener, levcromakalim, in vascular smooth muscle
Authors: McPherson Grant A. , Piekarska Anna E.
Published: July 2012

The aim of this study was to characterize the interaction between the K+ channel opener levcromakalim (LKM) and several quaternary ions, in vascular smooth muscle, in vitro. Segments of isolated, thoracic aorta of the rat were suspended in organ baths filled with Krebs solution at 37°C. Cumulative concentration‐response curves to LKM were obtained in the absence and in the presence of increasing concentrations of quaternary ions using a number of agents to pre‐constrict the vessel. The ions tested were tetraphenylphosphonium (chloride TPP‐Cl and bromide TPP‐Br salts), tetrapentylam‐monium (TPeA), tetraethylammonium (TEA), tetraphenylarsonium (TPAs) and tetraphenylboron (TPB).

For the compounds which antagonized the vasorelaxation responses of LKM, ‘apparent pKB’ values were estimated on the basis of a single concentration of antagonist. These were then used to obtain the following order of potency: TPP‐Br (7.22 ± 0.25) = TPAs (7.12 ± 0.04) = TPP‐Cl (7.11 ± 0.15) > TPeA (6.23 ± 0.20). TEA and TPB were both found to be inactive at the maximum concentrations used.

The interaction between the cationic TPP and anionic TPB was also investigated. The shift in the LKM concentration‐response curve constructed in the presence of both of these compounds was compared to that when each agent was present separately. We found that TPB, at concentrations greater than 1 μm, reversed the blockade of the LKM‐mediated relaxation induced by TPP (3 μm).

Similar experiments were undertaken combining TPB with either alinidine or glibenclamide (both functional antagonists of K+ channel openers). It was found that TPB (10 μm) partially reversed the antagonism induced by alinidine (30 and 100 μm) but had no effect on the action of glibenclamide (3 μm).

These studies show that lipophilic cations such as TPP and TPAs are potent antagonists of levcromakalim‐mediated vasorelaxation responses in the rat thoracic aorta. The mechanism by which these compounds cause their antagonism is not known. However, given the lipophilicity of these compounds, it is possible they may act at a number of sites including the KATP channel itself or possibly via some other intracellular mechanism.
Characterization of three non‐peptide endothelin receptor ligands using human cloned ETA and ETB receptors
Authors: Buchan Kevin W. , Alldus Carl , Christodoulou Chris , Clark Kenneth L. , Dykes Colin W. , Sumner Michael J. , Wallace Donald M. , White David G. , Watts Ian S.
Published: July 2012

A number of putative endothelin (ET) receptor ligands were synthesized with a view to assessing their relative affinity for human recombinant ET receptors.

Human (h) and endothelin ETA and ETB receptor open reading frames were cloned by reverse transcription‐polymerase chain reaction into the mammalian expression vector pcDNA1 and stable cell lines were created by transfection of Chinese hamster ovary cells.

Scatchard analyses of saturation isotherms for the specific binding of [125I]‐endothelin‐1 ([125I]‐ET‐1) to membranes, prepared from Chinese hamster ovary cells transfected with hETA or hETB receptors, yielded values for equilibrium dissociation constants (Kd) of 20.5 ± 1.8 pM and 25.5 ± 5.5 pM, respectively. Hill coefficients did not differ significantly from unity, suggesting binding to homogeneous, non‐interacting receptor populations.

Pharmacological characterization of the transfected hETA and hETB receptors was undertaken by measuring the relative abilities of ETA and ETB receptor‐selective peptide ligands to inhibit binding of [125I]‐ET‐1. For interaction with hETA receptors, the relative order of potency was ET‐1 > ET‐3 = FR139317 = BQ123 > [Ala1,3,11,15]‐ET‐1 = sarafotoxin S6c (S6c). In contrast, the relative order of potency, at hETB receptors, was ET‐1 = ET‐3 = [Ala1,3,11,15]‐ET‐1 = S6c >> FR139317 = BQ123.

The novel non‐peptide ligands, Ro 46–2005, SB 209670 and BMS 182874, were found to inhibit [125I]‐ET‐1 binding to human recombinant ETA and ETB receptors. At hETA receptors, the calculated pIC50 values were 6.7 (Ro 46–2005), 8.7 (SB 209670) and 5.8 (BMS 182874), while at hETB receptors, the corresponding pIC50 values were 6.8, 7.5 and < 5, respectively.

In conclusion, we have characterized the pharmacology of human cloned ETA and ETB receptors and used these in membrane binding assays to determine the affinity and selectivity of three structurally‐diverse non‐peptide ET receptor ligands. SB 209670 is, to date, the highest affinity non‐peptide ligand to be described for ET receptors. As such, it may prove to be a valuable tool in further examination of the physiological and pathophysiological roles of endothelins.
Natriuretic action of angiotensin(1–7)
Authors: DelliPizzi AnnMarie , Hilchey Sean D. , Bell‐Quilley Caroline P.
Published: July 2012

Evidence that angiotensin (1–7) (Ang(1–7)) is biologically active and can be synthesized by the kidney prompted us to examine its actions in the rat, isolated kidney. Ang(1–7) had three major effects producing, (1) a substantial natriuresis and diuresis, (2) an increase in urinary sodium concentration associated with a fall in potassium concentration and (3) an increase in glomerular filtration rate without affecting renal vascular resistance. Thus, Ang(1–7) may participate in the renal effects of the renin‐angiotensin system.
Release of nitric oxide by angiotensin‐(1–7) from porcine coronary endothelium: implications for a novel angiotensin receptor
Authors: Pörsti Ilkka , Bara Agnieszka T. , Busse Rudi , Hecker Markus
Published: July 2012

The angiotensin I (AI) metabolite, A(1–7), elicited a concentration‐dependent dilator response (ED50 ≥ 2 μm) in porcine coronary artery rings which was markedly attenuated by the nitric oxide (NO) synthase inhibitor, NG‐nitro‐l‐arginine, and abolished after removal of the endothelium. This effect of the heptapeptide was not mimicked by AII, AIII or A(3–8) at comparable concentrations. The A(1–7)‐induced relaxation was not affected by AT1 or AT2 receptor blockade or cyclo‐oxygenase inhibition, but was attenuated by the B2 receptor antagonist, Hoe 140, and augmented by the angiotensin‐converting enzyme (ACE) inhibitor, quinaprilat. These findings suggest that the relaxation to A(1–7) was mediated by the release of NO from the coronary endothelium through activation of an, as yet unidentified, AT receptor, the occupation of which also seems to stimulate the release of vasoactive kinins. Since A(1–7) accumulates during ACE inhibition, this mechanism may contribute to the coronary dilator effect of ACE inhibitors in vivo.
Activation of BKCa channels in acutely dissociated neurones from the rat ventromedial hypothalamus by NS 1619
Authors: Sellers Andrew J. , Ashford Michael L.J.
Published: July 2012

NS 1619 activates the large‐conductance Ca2+‐dependent K+ channel (BKCa) in membrane patches isolated from rat ventromedial hypothalamic neurones. The activation is concentration dependent, with a maximal effect at less than 30 μm, reversible and can be inhibited by application of iberiotoxin to the extracellular membrane. NS 1619 does not activate ATP‐K+ channels present in the same neurones.
Cyclo‐oxygenase and nitric oxide synthase isoforms in rat carrageenin‐induced pleurisy
Authors: Tomlinson Annette , Appleton Ian , Moore Adrian R. , Gilroy Derek W. , Willis Dean , Mitchell Jane A. , Willoughby Derek A.
Published: July 2012

The profiles of cyclo‐oxygenase (COX) and nitric oxide synthase (NOS) isoforms were determined in the rat carrageenin‐induced pleurisy model of acute inflammation.

The enzymes were assessed in peripheral blood leucocyte (PBL) cell pellets taken from untreated animals and at 2, 6 and 24 h after injection of the irritant in pleural exudate cell pellets and lung homogenates.

COX activity was assessed by the generation of prostacyclin (PGI2, measured as the stable metabolite, 6‐keto prostaglandin F1α) and prostaglandin E2 (PGE2). Western blot analysis and immunohistochemistry were also carried out.

NOS activity was based on the conversion of [3H]‐L‐arginine to [3H]‐L‐citrulline in the presence (total NOS activity) or absence of Ca2+ (inducible NOS; iNOS).

Peripheral blood leucocyte samples contained low levels of COX activity. In pleural exudate cell pellets, COX activity peaked at 2 to 6 h after injection of the carrageenin. At 24 h, COX activity was significantly reduced.

Western blot analysis demonstrated that the inducible isoform of COX (COX‐2), was the predominant enzyme at all time points. Low levels of COX‐2 were seen in PBLs. In pleural exudate cell pellets maximal COX‐2 protein levels were seen at 2h.

Immunohistochemistry confirmed the findings of Western blot studies. Approximately 10% of polymorphonuclear neutrophils (PMNs) in PBLs from untreated animals were immunopositive for COX‐2. In cell pellet smears from carrageenin‐induced pleurisy taken 2 h after injection of the irritant, PMNs were also the major source of COX‐2 immunoreactivity. A small proportion of macrophages and mesothelial cells were also immunolabelled for COX‐2.

Low levels of NOS activity were seen in PBLs. In pleural exudates NOS activity was maximum at 6h and greatly reduced by 24 h. This activity was solely attributable to iNOS.

The present results illustrated a similar profile of COX and NOS activity in the carrageenin‐induced pleurisy model of acute inflammation. It was demonstrated that COX‐2 and iNOS were the predominant isoforms of their respective enzymes.