bluuebunny/crossref_metadata_2025 huggingface dataset:
Binding kinetics and antiplatelet activities of picotamide, a thromboxane A2 receptor antagonist
Authors: Modesti Pietro Amedeo , Cecioni Ilaria , Colella Andrea , Costoli Alessandro , Paniccia Rita , Serneri Gian Gastone Neri
Published: July 2012

Picotamide was shown to inhibit platelet binding of thromboxane A2 (TxA2)‐mimetics and to cause a reduction of TxA2 platelet receptors after in vivo administration. The present study aimed to investigate directly [3H]‐picotamide binding to human platelets and in particular the relationship between binding kinetics and antiaggregating properties.

[3H]‐picotamide time‐dependently bound to a single class of platelet TxA2 receptors with a KD of 325 nmol l−1 at equilibrium. The binding was displaceable by TxA2 analogues U46619 and ONO11120 (Ki 19 and 28 nmol l−1 respectively) but not by prostacyclin (PGI2), prostaglandin E2 (PGE2) and TxB2. Antiaggregating activity and TxA2 formation inhibition paralleled with binding kinetics.

By prolonging the incubation time from 30 to 120 min, picotamide showed a progressively increasing non‐displaceable binding, whereas specific displaceable binding decreased in comparison to the values reached at 30 min. Non displaceable binding was specific, temperature‐dependent, saturable and followed a Michaelis‐Menten kinetic (Vmaxapp = 130 fmol per 108 platelets h−1, KMapp = 330 nmol l−1). Picotamide progressively underwent a specific stable interaction with its platelet receptor.

In conclusion, after an initial reversible binding, a progressive stabilization of picotamide binding takes place resulting in a progressively more stable interaction with platelets.
The effect of nitric oxide synthase inhibition on the plasma fibrinolytic system in septic shock in rats
Authors: Korbut Richard , Warner Tim D. , Gryglewski Richard J. , Vane John R.
Published: July 2012

We have investigated the effect of pretreatment of rats with nitric oxide (NO) synthase inhibitor, NG‐nitro‐l‐arginine methyl ester (l‐NAME) on the E. coli lipopolysaccharide (LPS)‐induced changes in the plasma fibrinolytic system, platelet count, fibrinogen level, as well as in gross and microscopic pathophysiological changes indicative of disseminated intravascular coagulation (DIC) in rats.

E. coli LPS (6 mg kg−1, i.p.) produced a decrease in the levels of plasma fibrinogen and a drop in the blood platelet count 6 h after administration. The decrease in fibrinogen but not the drop in platelet count was reversed by pretreatment with l‐NAME (30 mg kg−1, i.p., 24 h and 15 min before administration of LPS).

Pretreatment with l‐NAME antagonized the LPS‐induced activation of fibrinolysis as measured by changes in the euglobulin clot lysis time (ECLT) and enhanced the LPS‐induced rise in the plasma level of plasminogen activator inhibitor (PAI). In animals pretreated with l‐NAME there was also a marked reduction in the histological changes indicative of DIC.

We propose that l‐NAME can act as a protective agent in LPS‐induced DIC, and this protection is due to an increased generation of PAI following inhibition of NO synthase.
Biological actions of purines on rat megakaryocytes: potentiation by adenine of the purinoceptor‐operated cytoplasmic Ca2+ oscillation
Authors: Uneyama Chikako , Uneyama Hisayuki , Takahashi Michihito , Akaike Norio
Published: July 2012

We have found that adenine enhanced the purinoceptor‐operated cytoplasmic Ca2+ oscillation in rat megakaryocytes at submillimolar concentrations. Guanine and other nucleic acid bases had no effect on this system. Adenine enhanced the reaction intensity but had no effect on the threshold concentration of ATP to evoke the oscillation.
Inhibition by spermine of the induction of nitric oxide synthase in J774.2 macrophages: requirement of a serum factor
Authors: Szabó Csaba , Southan Garry J. , Wood Elizabeth , Thiemermann Christoph , Vane John R.
Published: July 2012

Polyamines are endogenous regulators of various cell functions. Nitric oxide (NO) is a cytostatic and cytotoxic free radical which is produced by the inducible NO synthase (iNOS) in immuno‐stimulated macrophages. We tested whether spermine modulates the induction of iNOS in J774.2 macrophages. Stimulation of macrophages by bacterial lipopolysaccharide (LPS) or γ‐interferon increased the accumulation of nitrite in the culture medium. Spermine (10−6 − 10−4 m) inhibited nitrite production without causing cytotoxicity. This inhibition of NO formation by spermine was significantly reduced when it was given 6 h after LPS. Spermine did not inhibit nitrite accumulation when foetal calf serum was omitted from the tissue culture medium. Thus, spermine is an inhibitor of the induction of iNOS, and its inhibitory activity requires the presence of a serum factor.
Comparison of the ability of nicardipine, theophylline and zaprinast to restore cardiovascular haemodynamics following inhibition of nitric oxide synthesis
Authors: Herity N.A. , Allen J.D. , Silke B. , Adgey A.A.J.
Published: July 2012

The use of pharmacological inhibitors of nitric oxide (NO) synthesis to treat patients with septic shock is limited by the observation that they cause a fall in cardiac output in some subjects. The aim of this work was to investigate this fall and to test whether it was reversible by subsequent administration of nicardipine, theophylline or the cyclic GMP‐selective phosphodiesterase inhibitor, zaprinast (M&B 22948).

In pentobarbitone‐anaesthetized pigs, haemodynamic indices were measured before and after intravenous administration of NG‐nitro‐l‐arginine methyl ester (l‐NAME) in a dose‐response protocol (0.2–20 mg kg−1; n = 6) and as a single bolus of 10 mg kg−1 either alone or followed by increasing doses of nicardipine, theophylline or zaprinast (n = 8 in each group).

l‐NAME caused a dose‐dependent rise in systemic vascular resistance and mean systemic arterial pressure and a dose‐dependent fall in cardiac output. A single bolus of l‐NAME (10 mg kg−1) produced these effects within 15 min.

Subsequent administration of nicardipine (0.05–0.2 mg kg−1) caused complete reversal of systemic vasoconstriction and hypertension and in doing so completely restored cardiac output. Theophylline (7.5–10 mg kg−1) partially reversed the rise in systemic vascular resistance and partially restored cardiac output but the effect was small compared to that of nicardipine. Zaprinast (1–5 mg kg−1) had no significant effect on any of these variables.

These results suggest that reduced cardiac output following inhibition of NO synthesis is an effect of increased afterload on the heart and is reversible by nicardipine and to a lesser extent by theophylline. These findings may have potential value for those using NO synthase inhibitors to treat patients with septic shock.
Discrimination between citrulline and arginine transport in activated murine macrophages: inefficient synthesis of NO from recycling of citrulline to arginine
Authors: Baydoun Anwar R. , Bogle Richard G. , Pearson Jeremy D. , Mann Giovanni E.
Published: July 2012

The kinetics, specificity, pH‐ and Na+‐dependency of l‐citrulline transport were examined in unstimulated and lipopolysaccharide (LPS)‐activated murine macrophage J774 cells. The dependency of nitric oxide production on extracellular arginine or citrulline was investigated in cells activated with LPS (1 μg ml−1) for 24 h.

In unstimulated J774 cells, transport of citrulline was saturable (Kt = 0.16 mm and Vmax = 32 pmol μg−1 protein min−1), pH‐insensitive and partially Na+‐dependent. In contrast to arginine, transport of citrulline was unchanged in LPS‐activated (1 μg ml−1, 24 h) cells.

Kinetic inhibition experiments revealed that arginine was a relatively poor inhibitor of citrulline transport, whilst citrulline was a more potent inhibitor (Ki = 3.4 mm) of arginine transport but only in the presence of extracellular Na+. Neutral amino acids inhibited citrulline transport (Ki = 0.2–0.3 mm), but were poor inhibitors of arginine transport.

Activated J774 cells did not release nitrite in the absence of exogenous arginine. Addition of citrulline (0.01 − 10 mm), in the absence of exogenous arginine, could only partially restore the ability of cells to synthesize nitrite, which was abolished by 100 μm NG‐nitro‐l‐arginine methyl ester or NG‐iminoethyl‐l‐ornithine.

Intracellular metabolism of l‐[14C]‐citrulline to l‐[14C]‐arginine was detected in unstimulated J774 cells and was increased further in cells activated with LPS and interferon‐γ.

We conclude that J774 macrophage cells transport citrulline via a saturable but nonselective neutral carrier which is insensitive to induction by LPS. In contrast, transport of arginine via the cationic amino acid system y+ is induced in J774 cells activated with LPS.

Our findings also confirm that citrulline can be recycled to arginine in activated J774 macrophage cells. Although this pathway provides a mechanism for enhanced arginine generation required for NO production under conditions of limited arginine availability, it cannot sustain maximal rates of NO synthesis.
Effect of salmeterol on human nasal epithelial cell ciliary beating: inhibition of the ciliotoxin, pyocyanin
Authors: Kanthakumar K. , Cundell D.R. , Johnson M. , Wills P.J. , Taylor G.W. , Cole P.J. , Wilson R.
Published: July 2012

Patients with airway infection by Pseudomonas aeruginosa have impaired mucociliary clearance. Pyocyanin is a phenazine pigment produced by P. aeruginosa which is present in the sputum of colonized patients, slows human ciliary beat frequency (CBF) in vitro and slows mucociliary transport in vivo in the guinea‐pig.

We have investigated the effect of salmeterol, a long‐acting β2‐adrenoceptor agonist, on pyocyanin‐induced slowing of human CBF in vitro. Salmeterol (2 × 10−7 m) was found to reduce pyocycanin (20 μg ml−1)‐induced slowing of CBF by 53% and the fall in intracellular adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) by 26% and ATP by 29%.

Another β2‐adrenoceptor agonist, isoprenaline (2 × 10−7 m), also inhibited pyocyanin‐induced slowing of CBF by 39%.

The effects of salmeterol (30 min preincubation) persisted after washing the cells.

Propranolol (10−7 m) and the β2‐specific antagonist, ICI 118551 (10−6 m) blocked the protective effects of salmeterol completely, but atenolol (10−6 m) was less effective. These results suggested that the effects of salmeterol on pyocyanin‐induced effects were mediated primarily via the stimulation of β2‐adrenoceptors.

Pyocyanin‐induced ciliary slowing is associated with a substantial fall in intracellular cyclic AMP and ATP. Salmeterol reversed the effects of pyocyanin on cyclic AMP and ATP.

Mucociliary clearance is an important defence mechanism of the airways against bacterial infection. Salmeterol may benefit patients colonized by P. aeruginosa, not only by its bronchodilator action, but also by protecting epithelial cells from pyocyanin‐induced slowing of CBF.
Recovery after dietary vitamin E supplementation of impaired endothelial function in vitamin E‐deficient rats
Authors: Rubino Annalisa , Burnstock Geoffrey
Published: July 2012

Thoracic aortae, isolated from rats supplemented with dietary vitamin E after vitamin E deficiency, were analysed for changes in vascular reactivity.

Following 4 or 12 months of dietary vitamin E deficiency, endothelium‐dependent vasodilator responses to acetylcholine were significantly impaired. However, when animals were fed after the first 4 months of vitamin E deprivation with a vitamin E‐supplemented diet for 8 months, endothelium‐mediated responses were completely restored.

In contrast, the endothelium‐independent vasodilator or vasoconstrictor responses to sodium nitroprusside and noradrenaline, respectively were not altered either by vitamin E deficiency or supplementation.

These data indicate that vitamin E supplementation reversed the impairment of endothelial cell function which occurs during vitamin E deficiency.
Competitive and non‐competitive effects of 5‐hydroxyindole on 5‐HT3 receptors in N1E‐115 neuroblastoma cells
Authors: Kooyman André R. , van Hooft Johannes A. , Vanderheijden Patrick M.L. , Vijverberg Henk P.M.
Published: July 2012

Effects of 5‐hydroxyindole (5‐OH‐indole) (the aromatic moiety of 5‐hydroxytryptamine (5‐HT)) on 5‐HT‐evoked ion current and the nature of these effects on 5‐HT3 receptors have been investigated in whole‐cell voltage clamp and radioligand binding experiments on cultured N1E‐115 mouse neuroblastoma cells.

The amplitude of 10 μm 5‐HT‐evoked ion currents was enhanced up to 150% of the control value by increasing concentrations up to 10 mm 5‐OH‐indole with half maximum effect of 0.8 mm. At concentrations between 10 mm and 50 mm, 5‐OH‐indole blocked the 5‐HT‐evoked ion current. Both the enhancement and the block by 5‐OH‐indole were accompanied by a marked slowing of the kinetics of decay of the 5‐HT‐evoked inward currents.

The blocking effect was surmounted when the 5‐HT concentration was raised from 10 μm to 100 μm. Conversely, the increase in amplitude and the slowing of the decay of the 5‐HT‐evoked ion current induced by 1 mm 5‐OH‐indole were not reversed by the same increase of 5‐HT concentration.

The binding of the selective antagonist [3H]‐GR65630 to 5‐HT3 receptors was displaced by 5‐OH‐indole in a concentration‐dependent manner with a pKi of 1.96. In saturation binding experiments 10 mm 5‐OH‐indole reduced the affinity of [3H]‐GR65630, whereas the total number of binding sites remained unaffected.

It is concluded that the blocking effect of high concentrations of 5‐OH‐indole is due to a competitive interaction with the antagonist recognition sites of 5‐HT3 receptors, whereas the potentiating effect of lower concentrations of 5‐OH‐indole appears to be mediated by a distinct non‐competitive interaction.
Evidence for differential roles of nitric oxide (NO) and hyperpolarization in endothelium‐dependent relaxation of pig isolated coronary artery
Authors: Kilpatrick E.V. , Cocks T.M.
Published: July 2012

The possible roles of endothelial and smooth muscle cell hyperpolarization and nitric oxide (NO) in endothelium‐dependent relaxation were examined in isolated rings of pig right coronary artery.

The effects of hyperpolarization were prevented with high K+ (30–125 mm), isotonic Krebs solutions. Functional antagonism due to high K+‐induced smooth muscle contraction was prevented with 0.3 μm nifedipine (in all treatments, for consistency). All rings were contracted with the thromboxane‐mimetic U46619, (1 − 100 nm) to bring them to an initial active force of within 30–50% of maximum contraction.

High K+ had no effects on the sensitivity (EC50) or time course of endothelium‐dependent (substance P, SP; bradykinin, BK; calcimycin, A23187) and ‐independent (sodium nitroprusside, SNP) agents. Maximum relaxations (Rmax) to SP, BK and A23187 were reduced significantly by approximately 20% but only with 125 mm K+.

In normal K+ Krebs solution (5.9 mm), NG‐nitro‐l‐arginine (l‐NOARG; 100 μm) caused 40%, 20% and no reduction in Rmax for SP, BK and SNP respectively. EC50s for SP and BK were decreased significantly by approximately two fold whereas that for SNP was increased significantly by approximately ten fold. At all high K+ concentrations (30–125 mm), l‐NOARG (100 μm) caused complete inhibition of relaxations to SP and BK but those to SNP were unaffected.

High K+ (30 mm) unmasked potent and concentration‐dependent inhibition of relaxations of SP by l‐NOARG. At 10 μm l‐NOARG, all relaxation responses to SP were abolished and at the higher concentrations of SP (1 − 10 nm) small but significant contractions were observed.

NG‐monomethyl‐l‐arginine (l‐NMMA) had similar effects on relaxations to SP in the presence of 30 mm K+ except that maximum inhibition (40%) of Rmax was achieved at 10 μm l‐NMMA and this was not increased with either 100 or 1000 μm l‐NMMA. In normal K+, l‐NMMA (1000 μm) only decreased the EC50 by approximately two fold, without affecting Rmax.

High choline+ (25, 75 and 125 mm) isotonic Krebs also had no direct effect on the relaxations to SP, but like high K+, enabled l‐NOARG (100 μm) to inhibit these responses completely. Neither charyb‐dotoxin (30 nm) nor substitution of 25 mm NaCl with 50 mm sucrose had any direct effect on relaxations to SP or on the block of relaxations to SP by l‐NOARG (100 μm).

In conclusion, most if not all of the endothelium‐dependent relaxation in the pig coronary artery in vitro is due to NO, but hyperpolarization can supplement 60%–80% of this response if NO synthesis is blocked. Multiple endothelium‐derived factors could not only explain heterogeneity of the degree of block of endothelium‐dependent relaxation responses by l‐arginine analogues, but also constitute important ‘back‐up’ mechanisms for control of arterial diameter.
Stimulation by menthol of Cl secretion via a Ca2+‐dependent mechanism in canine airway epithelium
Authors: Chiyotani A. , Tamaoki J. , Takeuchi S. , Kondo M. , Isono K. , Konno K.
Published: July 2012

To investigate the effect of menthol on airway epithelial ion transport function, we studied the bioelectrical properties of canine cultured tracheal epithelium by Ussing's short‐circuit technique in vitro.

Addition of menthol (10−3 m) to the mucosal but not the submucosal solution increased the short‐circuit current (Isc) from 6.2 ± 0.9 to 14.0 ± 2.2 μA cm−2 (P < 0.001), and this effect was accompanied by increases in transepithelial potential difference and conductance. The response was dose‐dependent, with the maximal increase from the baseline value and the concentration required to produce a half‐maximal effect (EC50) being 6.4 ± 0.9 μA cm−2 (P < 0.001) and 40 μm, respectively.

Other cyclic alcohols, including menthone and cyclohexanol, had no effect on the electrical properties.

The menthol‐induced increase in Isc was not altered by pretreatment of the cells with amiloride, indomethacin, or propranolol but was abolished by diphenylamine‐2‐carboxylate, furosemide or substitution of Cl with iodide in the medium.

Menthol (10−2 m) increased cytosolic levels of free calcium ([Ca2+]i) from 98 ± 12 to 340 ± 49 nm (P < 0.01) in fura‐2‐loaded tracheal epithelium but did not affect the intracellular adenosine 3′,5′‐cyclic monophosphate content.

These results suggest that menthol stimulates Cl secretion across airway epithelium, probably through a Ca2+‐dependent mechanism, and might thus influence mucociliary transport in the respiratory tract.
Cell inward transport of l‐DOPA and 3‐O‐methyl‐l‐DOPA in rat renal tubules
Authors: Soares‐da‐Silva P. , Fernandes M.H. , Pinto‐do‐OA P.C.
Published: July 2012

The present study has determined the kinetics of the uptake of l‐3,4‐dihydroxyphenylalanine (l‐DOPA) and 3‐O‐methyl‐l‐DOPA (3‐OMDOPA) in rat renal tubules and examined the effect of 3‐OMDOPA on the inward transport of l‐DOPA and on its conversion into dopamine in kidney homogenates.

The accumulation of both l‐DOPA and 3‐OMDOPA in renal tubules was found to occur through non‐saturable and saturable mechanisms. The kinetics of the saturable component of l‐DOPA and 3‐OMDOPA uptake in renal tubules were as follows: l‐DOPA, Vmax = 11.1 nmol mg−1 protein h−1 and Km = 216 μm (n = 6); 3‐OMDOPA, Vmax = 8.1 nmol mg−1 protein h−1 and Km = 231 μm (n = 5). The diffusion constant of the non‐saturable component for the accumulation of l‐DOPA and 3‐OMDOPA was 0.0010 and 0.0014 μmol−1, respectively.

3‐OMDOPA (100 to 2000 μm) was found to produce a concentration‐dependent decrease (29% to 81% reduction) of the saturable component of the tubular uptake of l‐DOPA; the Ki value of 3‐OMDOPA for inhibition of l‐DOPA uptake was found to be 181 μm (n = 5). The accumulation of l‐DOPA obtained in experiments conducted at 4°C was not affected by 3‐OMDOPA.

In experiments conducted in kidney homogenates only l‐DOPA (10 to 5000 μm) was found to be decarboxylated. The Vmax and Km values for aromatic l‐amino acid decarboxylase determined in the absence of 3‐OMDOPA (Vmax = 14.1 nmol mg−1 protein h−1; Km = 62 μm) were not significantly different from those observed when the decarboxylation of l‐DOPA was carried out in the presence of 1000 μm 3‐OMDOPA (Vmax = 15.7 nmol mg−1 protein h−1; Km = 68 μm).

It is concluded that the tubular uptake of both l‐DOPA and 3‐OMDOPA occur through non‐saturable and saturable mechanisms; only the saturable tubular uptake of l‐DOPA was found to be inhibited by 3‐OMDOPA. It is further shown that 3‐OMDOPA neither undergoes decarboxylation into 3‐MT nor affects the decarboxylation of l‐DOPA.
Improvement of cardiovascular effects of metoprolol by replacement of common salt with a potassium‐ and magnesium‐enriched salt alternative
Authors: Mervaala Eero M.A. , Laakso Juha , Vapaatalo Heikki , Karppanen Heikki
Published: July 2012

The influence of sodium chloride (NaCl)‐enrichment of the diet (6% of the dry weight) and that of a novel sodium‐reduced, potassium‐, magnesium‐, and l‐lysine‐enriched salt alternative on the cardiovascular effects of the β1‐adrenoceptor blocking drug, metoprolol, was studied in stroke‐prone spontaneously hypertensive rats.

Increased dietary sodium chloride intake produced a marked rise in blood pressure and induced left ventricular and renal hypertrophy. By contrast, the salt alternative did not increase blood pressure and caused remarkably less cardiac and renal hypertrophy than did sodium chloride.

Metoprolol treatment at a daily dose of 250 mg kg−1 lowered blood pressure and decreased left ventricular hypertrophy index during the control diet. Sodium chloride‐enrichment blocked the antihypertensive effect of metoprolol, while a partial protective effect on left ventricular and renal hypertrophy persisted. In the presence of the salt alternative‐enrichment both at the level of 6% and 10.5% (corresponding to a NaCl level of 6%), metoprolol was fully able to exert its beneficial cardiovascular and renal effects.

Both salt supplementations, irrespective of metoprolol treatment, induced a 3 to 4 fold increase in the urinary excretion of calcium. There was a linear correlation between the urinary excretions of sodium and calcium. The urinary excretion of magnesium rose by 90% and that of potassium by 110% in the salt alternative group.

Our findings suggest that replacement of common salt by a potassium‐, and magnesium‐enriched salt alternative in the diet produces beneficial cardiovascular effects and improves the antihypertensive efficacy of metoprolol in stroke‐prone spontaneously hypertensive rats. Increased intake of potassium and/or magnesium and l‐lysine from the salt alternative is involved in the beneficial effects of the salt alternative. The NaCl‐induced myocardial and renal hypertrophies appear to be partially mediated by β‐adrenoceptor activation.
Evaluation of a series of novel CCKB antagonists using a functional assay in the rat central nervous system
Authors: Boden P.R. , Pinnock R.D. , Pritchard M.C. , Woodruff G.N.
Published: July 2012

Electrophysiological recordings from rat ventromedial hypothalamus (VMH) in vitro have been used to compare the effects of novel chemical entities on CCKB receptor activation in the rat central nervous system.

Twenty compounds from three different chemical series were evaluated for their ability to reduce pentagastrin‐induced increases in action potential firing rate.

All twenty compounds studied were found to be CCKB antagonists, with equilibrium constants spanning a concentration‐range of several orders of magnitude. The rank order for their ability to block pentagastrin responses correlated well with values obtained for their relative affinities for the mouse cortex CCKB binding site.

It is concluded that the VMH preparation provides a good functional correlate to binding assays in the rodent central nervous system for a structurally diverse series of CCKB antagonists.
The effects of sigma ligands and of neuropeptide Y on N‐methyl‐d‐aspartate‐induced neuronal activation of CA3 dorsal hippocampus neurones are differentially affected by pertussin toxin
Authors: Monnet François P. , Debonnel Guy , Bergeron Richard , Gronier Benjamin , de Montigny Claude
Published: July 2012

The in vivo effects of the high affinity sigma ligands 1,3‐di(2‐tolyl)guanidine (DTG), (+)‐N‐cyclopropylmethyl‐N‐methyl‐1,4‐diphenyl‐1‐ethyl‐but‐3‐en‐1‐ylamine hydrochloride (JO‐1784), (+)‐pentazocine and haloperidol, as well as of those of neuropeptide Y (NPY), on N‐methyl‐d‐aspartate (NMDA)‐ and quisqualate (Quis)‐induced neuronal activations of CA3 pyramidal neurones were assessed, using extracellular unitary recording, in control rats and in rats pretreated with a local injection of pertussis toxin (PTX), to evaluate the possible involvement of Gi/o proteins in mediating the potentiation of the neuronal response to NMDA by the activation of sigma receptors in the dorsal hippocampus.

Microiontophoretic applications as well as intravenous injections of (+)‐pentazocine potentiated selectively the NMDA response in control rats as well as in PTX‐pretreated animals. In contrast, the PTX pretreatment abolished the potentiation of the NMDA response by DTG, JO‐1784 and NPY. Moreover, microiontophoretic applications of DTG induced a reduction of NMDA‐induced neuronal activation. Neither in control nor in PTX‐treated rats, did the sigma ligands and NPY have any effect on Quis‐induced neuronal response.

In PTX‐treated rats, the potentiation of the NMDA response induced by (+)‐pentazocine was suppressed by haloperidol, whereas the reduction of the NMDA response by DTG was not affected by haloperidol.

This study provides the first in vivo functional evidence that sigma ligands and NPY modulate the NMDA response by acting on distinct receptors, differentiated by their PTX sensitivity.
Loss of preconditioning in rabbits with vascular tolerance to nitroglycerin
Authors: Szilvassy Z. , Ferdinandy P. , Bor P. , Jakab I. , Szilvassy J. , Nagy I. , Lonovics J. , Koltai M.
Published: July 2012

A preceding right ventricular overdrive pacing (VOP) of 500 b.p.m. for 5 min, markedly reduced the severity of global myocardial ischaemia produced by a subsequent 5‐min VOP in conscious rabbits. This VOP‐induced preconditioning developed in parallel with an increase in cardiac cyclic guanosine 3′: 5′‐monophosphate (cyclic GMP) content. VOP‐induced preconditioning was abolished when the animals had been made tolerant to the vasodilator effect of nitroglycerin (NG). In the heart of the NG‐tolerant rabbits, neither VOP nor preconditioning increased cyclic GMP content. This suggests that changes by NG tolerance of cyclic GMP metabolism may account for the loss of VOP‐induced preconditioning.
Comparative studies on the affinities of ATP derivatives for P2X‐purinoceptors in rat urinary bladder
Authors: Bo Xuenong , Fischer Bilha , Maillard Michel , Jacobson Kenneth A. , Burnstock Geoffrey
Published: July 2012

Radioligand binding assays have been used to determine the affinities of a series of ATP derivatives with modifications of the polyphosphate chain, adenine and ribose moieties of the ATP molecule for [3H]‐α,β‐methylene ATP ([3H]‐α,β‐MeATP) binding sites in rat urinary bladder.

The replacement of the bridging oxygen in the triphosphate chain of ATP (pIC50 = 5.58) with a methylene or imido group markedly increased the affinity (691 fold in IC50 values for β,γ‐imidoATP, 15 fold for β,γ‐methylene ATP), and the replacement of an ionized oxygen on the γ‐phosphate with a sulphur (ATPγS) also led to increased affinity (5623 fold in IC50 values).

Modifications at N6, N1, and C‐8 positions on the purine base usually reduced the affinity of ATP (a decrease of 2.8 fold in IC50 values for N6‐methylATP and 8.9 fold for 8‐bromo ATP), while the attachment of an alkylthio group to the C‐2 position greatly increased the affinity for P2X‐purinoceptors (from 3.5 to 98 fold increase in IC50 values).

Replacement of the 3′‐hydroxyl group on the ribose with substituted amino or acylamino groups produced more potent P2X‐purinoceptor agonists (an increase of 447 fold in IC50 values for 3′‐deoxy‐3′‐benzylamino ATP and 28 fold for 3′‐deoxy‐3′‐(4‐hydroxyphenylpropionyl)amino ATP.

Diadenosine polyphosphates (Ap[n]A) were also shown to displace the [3H]‐α,β‐MeATP binding. The rank order of potency was Ap6A > Ap5A > Ap4A >> Ap3A >> Ap2A.

Suramin, PPADS, and reactive blue 2 could competitively displace the binding of [3H]‐α,β‐MeATP to P2X‐purinoceptors, with pIC50 values of 6.26, 5.35, and 6.22, respectively.
Muscarinic acetylcholine response in pyramidal neurones of rat cerebral cortex
Authors: Nishikawa M. , Munakata M. , Akaike N.
Published: July 2012

The effects of acetylcholine (ACh) on pyramidal neurones acutely dissociated from the rat cerebral cortex were studied in the whole‐cell mode, by use of the nystatin‐perforated patch recording configuration.

ACh induced a net inward current (IACh) accompanied by a membrane conductance decrease at a holding potential (VH) of − 40 mV. IACh increased in a concentration‐dependent manner with a half‐maximum concentration (EC50) of 8.7 × 10−7 M.

IACh mainly resulted from the suppression of the voltage‐ and time‐dependent K+ current (M‐current).

Muscarine and muscarinic agonists such as McN‐A‐343, oxotremorine and oxotremorine‐M mimicked the ACh response. The potency was in the order of oxotremorine‐M > McN‐A‐343 ≥ muscarine > oxotremorine.

Pirenzepine shifted the concentration‐response curve for ACh to the right and the corresponding Schild plot yielded a pA2 value of 7.81. Other muscarinic antagonists also reversibly blocked IACh in a concentration‐dependent manner. The inhibitory potency was in the order of atropine > 4‐DAMP > pirenzepine > AF‐DX‐116.

IACh could be induced normally even after pre‐incubation of dissociated neurones in external solution with 200 ng ml−1 pertussis toxin (PTX) for 8 h, whereas the inhibitory effect of ACh on high‐voltage‐activated Ca2+ channels was completely abolished by the PTX treatment.
[125I]‐PD 151242: a selective radioligand for human ETA receptors
Authors: Davenport Anthony P. , Kuc Rhoda E. , Fitzgerald Fiona , Maguire Janet J. , Berryman Kent , Doherty Annette M.
Published: July 2012

Our aim was to synthesize a new endothelin ETA selective radioligand, [125I]‐PD 151242 and characterize the compound in human vascular tissue. Binding of [125I]‐PD151242 to sections of human aorta was time‐dependent and reached equilibrium after 120 min at 23°C with an association rate constant of 1.26 ± 0.17 × 108 M−1 min−1 (n = 3 individuals ± s.e.mean). The binding was reversible at 23°C with an observed dissociation rate constant of 0.0025 ± 0.0006 min−1 (n = 3). Saturation binding assays using [125I]‐PD151242 revealed a single population of high affinity ET receptors (n = 3) in aorta (KD = 0.76 ± 0.17 nM; Bmax = 5.98 ± 1.56 fmol mg−1 protein), pulmonary (KD = 1.75 ± 0.20 nM; Bmax = 12.78 ± 1.39 fmol mg−1 protein) and coronary arteries (KD = 0.51 ± 0.07 nM; Bmax = 44.9 ± 1.67 fmol mg−1 protein). ETA selective ligands competed for [125I]‐PD151242 binding in aorta with nanomolar affinity (BQ123, KD = 0.41 ± 0.26 nM; FR139317, KD = 0.55 ± 0.11 nM) whereas the ETB selective compound, BQ3020, competed with micromolar affinity (KD = 1.36 ± 0.25 μm). In isolated coronary arteries, PD151242 was a functional antagonist and caused a significant, parallel rightward shift of the ET‐1 dose‐response curve with a pA2 value of 5.92 (n = 5) and a slope of unity. The high affinity and selectivity of [125I]‐PD151242 for ETA receptors will facilitate the characterization of this sub‐type in human tissues.
Behavioural and neurochemical adaptations to nicotine in rats: influence of NMDA antagonists
Authors: Shoaib M. , Benwell M.E.M. , Akbar M.T. , Stolerman I.P. , Balfour D.J.K.
Published: July 2012

The repeated co‐administration of the non‐competitive N‐methyl‐d‐aspartate (NMDA) receptor antagonist dizocilpine (0.1 and 0.3 mg kg−1, i.p.) with nicotine (0.4 mg kg−1, s.c.) attenuated the development of tolerance to the locomotor depressant effect of the nicotine in rats.

The repeated co‐administration of the competitive NMDA antagonist d‐CPPene (SDZ EAA 494; 3‐(2‐carboxypiperazin‐4‐yl)‐1‐propenyl‐1‐phosphonic acid, 2 and 8 mg kg−1, i.p.) also attenuated tolerance to the locomotor depressant effect of nicotine.

Dizocilpine (0.3 mg kg−1, i.p.) pretreatment attenuated sensitization to the locomotor stimulant effect of nicotine (0.4 mg kg−1, s.c.) and prevented sensitization of nicotine‐induced dopamine release in the nucleus accumbens. However, pretreatment with dizocilpine alone caused a modest enhancement of the behavioural response to a subsequent acute dose of nicotine.

d‐CPPene (2.0 mg kg−1, i.p.) pretreatment prevented sensitization to the nicotine‐induced dopamine release in the nucleus accumbens. There was no enhanced locomotor response that could be attributed to nicotine pretreatment when d‐CPPene was co‐administered with nicotine. However, pretreatment with d‐CPPene alone enhanced the locomotor response to an acute dose of nicotine.

The results suggest the involvement of NMDA receptors in adaptations of the behavioural and neurochemical effects of nicotine that occur as a result of repeated administration of the drug.
Inhibitory effects of capsaicin on acetylcholine‐evoked responses in rat phaeochromocytoma cells
Authors: Nakazawa Ken , Inoue Kaori , Koizumi Schuichi , Ikeda Makoto , Inoue Kazuhide
Published: July 2012

The effects of capsaicin on cellular responses evoked by acetylcholine (ACh) and those by adenosine 5′‐triphosphate (ATP) were investigated in rat phaeochromocytoma PC12 cells.

Capsaicin (1 to 30 μm) suppressed dopamine release and the intracellular Ca2+ increase evoked by 100 μm ACh. The concentration‐dependence of the ACh‐evoked release of dopamine was not shifted but the maximal response was reduced by capsaicin. Dopamine release evoked by 80 mm KC1 was also suppressed by capsaicin (3 and 30 μm), but the extent of suppression was smaller than that of the ACh‐evoked release.

Under whole‐cell voltage‐clamp, capsaicin (1 to 30 μm) reversibly inhibited the inward current activated by ACh (30 to 300 μm). The inhibition exhibited dependence on the concentration of ACh, and the current activated by a higher concentration of ACh was less inhibited. Voltage‐dependence of block by capsaicin was not observed when it was tested either by applying a ramp pulse during the current activation by ACh or by eliciting the current in cells held at various potentials.

High concentrations of capsaicin (30 to 100 μm) enhanced the inward current as well as dopamine release evoked by 30 μm ATP.

The results suggest that low concentrations of capsaicin selectively antagonize responses mediated by nicotinic receptor‐channels without affecting those mediated by purinoceptor‐coupled channels. As the antagonism by capsaicin of the ACh‐evoked responses was observed at concentrations as low as 1 μm, the influence on nicotinic receptors should be taken into account when this compound is used as a pharmacological tool to deplete neuropeptides.
Induction of nitric oxide synthase activity in phagocytic cells inhibited by tricyclodecan‐9‐yl‐xanthogenate (D609)
Authors: Tschaikowsky Klaus , Meisner Michael , Schönhuber Frank , Rügheimer Erich
Published: July 2012

The synthesis of nitric oxide (NO) by immune‐stimulated murine phagocytic cells (J774) and the modulation of this synthesis by tricyclodecan‐9‐yl‐xanthogenate (D609), a specific inhibitor of phosphatidylcholine‐specific phospholipase C (PC‐PLC), was investigated. D609 dose‐dependently suppressed production of NO, as measured by the release of nitrite and nitrate, in response to lipopolysac‐charide (LPS) and interferon‐γ (IFN‐γ) in intact cultured cells with an IC50 of approximately 20 μg ml−1. D609 at 40 μg ml−1 completely abrogated immune‐stimulated nitrite production.

The inhibitory effects of D609 on nitrite production were time‐dependent and restricted to the first 18 h post‐stimulation. D609 did not inhibit nitrite production in the cytosol of immune‐stimulated phagocytes.

These findings indicate that the xanthogenate, D609, is a potent inhibitor of the induction of NO‐synthase activity in immune‐stimulated phagocytes. Furthermore, since D609 has been demonstrated to inhibit PC‐PLC specifically, our findings suggest that the activation of this enzyme by LPS and IFN‐γ is a proximal step in the signal transduction of inducible NO‐synthase in phagocytic cells.
The effect of hyperglycaemia on function of rat isolated mesenteric resistance artery
Authors: Taylor Paul D. , Poston Lucilla
Published: July 2012

Noradrenaline sensitivity and acetylcholine‐induced relaxation were investigated in mesenteric resistance arteries from female Wistar rats (220–250 g) following exposure to isotonic supraphysiological glucose solutions (20 and 45 mm, in physiological buffer, 2h incubation).

Arteries incubated in 20 mm glucose demonstrated enhanced noradrenaline sensitivity compared with those in physiological buffer.

Profoundly impaired endothelium‐dependent relaxation to acetylcholine was observed in arteries incubated in 20 and 45 mm glucose.

Indomethacin (10 μm) normalized noradrenaline sensitivity in 20 mm glucose, but unmasked an enhanced maximum response in 20 and 45 mm glucose relative to controls.

Addition of L‐arginine (0.1 μm) prevented the abnormality of acetylcholine‐induced relaxation in the 20 mm glucose medium and significantly improved relaxation in 45 mm glucose.

The aldose reductase inhibitor, ponalrestat (10−5 m, ZENECA Pharmaceuticals), prevented impaired acetylcholine‐mediated relaxation in 20 mm glucose and significantly improved relaxation in 45 mm glucose.

Indomethacin (10 μm) improved maximum relaxation but did not alter impaired sensitivity to acetylcholine in the high glucose media (20 and 45 μm).

Superoxide dismutase (SOD, 150 u ml−1) also prevented impaired acetylcholine‐induced relaxation in 20 mm glucose but not in 45 mm glucose.

Endothelium‐independent relaxation to sodium nitroprusside (SNP, 10−9‐10−5mM) was normal in 20 mm glucose but was slightly, although significantly impaired by 45 mm glucose.

Enhanced responsiveness of rat isolated mesenteric resistance arteries to noradrenaline caused by elevated glucose would appear to be mediated through abnormal cyclo‐oxygenase activity and the reduced tonic release of nitric oxide.

Hyperglycaemia may lead to abnormal endothelium‐dependent relaxation in these arteries through several mechanisms which include a role for increased free radical production, polyol pathway activation and altered L‐arginine metabolism.
The effect of non‐steroidal anti‐inflammatory drugs on the accumulation and release of interleukin‐1‐like activity by peritoneal macrophages from the mouse
Authors: Bahl A.K. , Dale M.M. , Foreman J.C.
Published: July 2012

The non‐steroidal anti‐inflammatory drugs (NSAIDs) indomethacin, 10 and 100 μm, piroxicam, 100 μm, and sodium meclofenamate, 1 and 100 μm, potentiated the lipopolysaccharide (LPS)‐stimulated release of interleukin‐1 (IL‐1)‐like activity from mouse peritoneal macrophages. Aspirin up to 100 μm was without effect. The drugs did not themselves stimulate the release of IL‐1‐like activity at the concentrations used.

LPS, 1 μg ml−1, stimulated prostaglandin E2 production by mouse peritoneal macrophages and this was totally inhibited by aspirin, 100 μm, indomethacin, 1 μm, piroxicam, 10 μm and sodium meclofenamate, 0.1 μm.

The potentiation of LPS‐stimulated release of IL‐1‐like activity produced by indomethacin, 100 μm, piroxicam, 100 μm, or sodium meclofenamate, 10 μm, was inhibited by prostaglandin E2, (PGE2) 10 ng ml−1.

Aspirin, 100 μm, indomethacin, 100nM to 10 μm, piroxicam, 1 to 100 μm, and sodium meclofenamate, 10 nm, all potentiated cell‐associated IL‐1‐like activity in LPS‐ stimulated macrophages. The drugs had no effect on cell‐associated IL‐1‐like activity by themselves.

Exogenous PGE2, 2 to 30 ng ml−1, inhibited the cell‐accumulation of IL‐1‐like activity stimulated by LPS in the presence of indomethacin, 1 μm, or sodium meclofenamate, 0.1 μm.

The 5‐lipoxygenase inhibitors BWA4C, 0.01 to 10 μm, and L‐651,392, 0.01 to 10 μm, had no effect on LPS‐stimulated released or cell‐associated IL‐1‐like activity. Over the same concentration‐ranges, neither of the 5‐lipoxygenase inhibitors affected released or cell‐associated IL‐1‐like activity in LPS‐stimulated mouse macrophages in the presence of indomethacin, 1 μm.

The synthetic diacylglycerol, DiC8, 10 to 200 μm, did not itself increase released or cell‐associated IL‐1‐like activity but in the presence of the diacylglycerol kinase inhibitor, R59022, 10 μm, DiC8 increased released and cell‐associated IL‐1‐like activity. The activity of DiC8 on released and cell‐associated IL‐1‐like activity was not increased by indomethacin, 100 μm.

NSAIDs increase LPS‐induced cell‐associated IL‐1‐like activity in mouse macrophages by inhibiting the formation of cyclo‐oxygenase products such as PGE2 but at higher concentrations the NSAIDs potentiate LPS‐induced release of IL‐1‐like activity by a mechanism independent of cyclo‐oxygenase inhibition. The potentiation of the release of IL‐1‐like activity appears not to be related to an effect of NSAIDs on either 5‐lipoxygenase or diacylglycerol metabolism.
The effects of clozapine on behavioural responses to the selective ‘D1‐like’ dopamine receptor agonist, A 68930, and to the selective ‘D2‐like’ agonist, RU 24213
Authors: Daly Siobhan A. , Waddington John L.
Published: July 2012

The influence of the atypical antipsychotic clozapine on D1 dopamine receptor‐mediated function was examined in terms of its effects on behavioural responses to the new isochroman selective D1 agonist, A 68930, and to the selective D2 agonist, RU 24213.

In rat striatal membrane preparations, radioligand binding studies with [3H]‐SCH 23390 and [3H]‐spiperone confirmed clozapine to show weak and non‐selective affinity for both D1 and D2 receptors.

Using a rapid time‐sampling behavioural check list technique, clozapine (4.0–36.0 mg kg−1) exerted only modest antagonism of RU 24213 (15.0 mg kg−1)‐induced sniffing and locomotion, and weakly released some episodes of myoclonic jerking; such antagonism with release of jerking has been shown previously to occur only during concurrent stimulation of D2 receptors and attenuation of D1 function.

Over the same dose‐range, clozapine completely blocked A68930 (0.25 mg kg−1)‐induced intense grooming but failed to influence the vacuous chewing response; this profile was similar to that demonstrated previously for selective D1 antagonists.

On the basis of complete blockade of typical D1 agonist‐induced grooming and weak release of atypical jerking to D2 agonism in the face of modest reduction in typical D2‐stimulated behaviours, clozapine apears to exert some preferential but not selective attenuation of D1 receptor‐mediated function. Clozapine may attenuate activity through a classical D1 receptor at a level beyond the recognition site, for which it has little affinity, or by way of new, putative ‘Dplike’ site(s) that subserve distinct elements of dopaminergic behaviour.
The sensitivity of nicotinic synapses in bullfrog sympathetic ganglia to α‐bungarotoxin and neuronal‐bungarotoxin
Authors: Shen Wei‐Xing , Jobling Phillip , Horn John P.
Published: July 2012

The sensitivity of nicotinic synapses to α‐bungarotoxin (α‐Bgt) and neuronal‐bungarotoxin (n‐Bgt) was measured in the B and C cell systems of bullfrog paravertebral sympathetic ganglia 9 and 10 by recording extracellular compound postganglionic action potentials from the rami communicantes.

High concentrations (10 μm) of α‐Bgt applied for up to 8 h had no effect upon synaptic transmission in either the B or C cell system. Ganglia pretreated with collagenase were also insensitive to α‐Bgt. In control experiments on isolated sartorius muscle preparations, nerve‐evoked twitches were fully blocked by 30‐100nM α‐Bgt.

Nicotinic transmission in the B and C cell systems was reversibly blocked by 30–300 nm n‐Bgt. Block appeared within 25–45 min of exposure to toxin and reversed fully with a half‐time of 40‐80min. This was indistinguishable from washout times after block by 100 μm (+)‐tubocurarine.

The results demonstrate close parallels between the bungarotoxin sensitivity of neuronal nicotinic receptors mediating ganglionic transmission in functional subclasses of bullfrog sympathetic neurones and the bungarotoxin sensitivity which has been reported for autonomic neurones in avian and mammalian preparations.
Role of K+ channels in the vasodilator response to bradykinin in the rat heart
Authors: Fulton David , McGiff John C. , Quilley John
Published: July 2012

The role of K+ channels in the nitric oxide (NO)‐independent coronary vasodilator efffect of bradykinin was examined in the Langendorff heart preparation in which nitroarginine was used to inhibit NO synthesis and elevate perfusion pressure; cyclo‐oxygenase was inhibited with indomethacin.

The K+ channel inhibitors, tetraethylammonium, procaine and charybdotoxin, but not glibenc‐lamide, further increased perfusion pressure suggesting a role for K+ channels, other than ATP‐sensitive K+ channels, in the regulation of coronary vascular tone under the experimental conditions adopted here.

The non‐specific K+ channel inhibitors, tetraethylammonium and procaine, reduced vasodilator responses to bradykinin and cromakalim but not those to nitroprusside in the perfused heart treated with nitroarginine and indomethacin.

Glibenclamide, an inhibitor of ATP‐sensitive K+ channels, reduced vasodilator responses to cromakalim but did not affect those to bradykinin or nitroprusside.

Charybdotoxin, an antagonist of Ca2+‐activated K+ channels, inhibited responses to bradykinin but did not affect those to cromakalim or nitroprusside.

Nifedipine inhibited vasodilator responses to bradykinin and cromakalim without affecting those to nitroprusside.

Inhibition of cytochrome P450 with clotrimazole reduced responses to bradykinin but did not modify those to cromakalim or nitroprusside.

These results suggest that bradykinin utilizes a Ca2+‐activated K+ channel to produce vasodilatation in the rat heart.
Modulatory effect of neuropeptide Y on acetylcholine‐induced oedema and vasoconstriction in isolated perfused lungs of rabbit
Authors: Delaunois Annie , Gustin Pascal , Dessy‐Doize Cécile , Ansay Michel
Published: July 2012

The modulatory role of neuropeptide Y (NPY) on pulmonary oedema induced by acetylcholine and capsaicin was investigated. The effects of NPY on the haemodynamic response to acetylcholine, phenylephrine and substance P were also investigated.

Isolated, ventilated, exsanguinated lungs of the rabbit were perfused with a constant flow of recirculating blood‐free perfusate. The double/arterial/venous occlusion method was used to partition the total pressure gradient (ΔPt) into four components: the arterial gradient (ΔPa), the pre‐ and post‐capillary gradients (respectively ΔPa' and ΔPv') and the venous pressure gradient (ΔPv). Endothelial permeability was evaluated by measuring the capillary filtration coefficient (Kf,c).

Acetylcholine (10−8 m to 10−4 m) and substance P (SP, 10−10 m to 10−6 m) induced a concentration‐dependent increase in the Kf,c. Capsaicin (10−4m) and 5‐hydroxytryptamine (5‐HT) (10−4m) also increased this parameter. NPY (10−8m) completely inhibited the effects of acetylcholine and capsaicin on the Kf,c, without preventing the effects of substance P and 5‐HT.

Acetylcholine induced concentration‐dependent vasoconstriction in the precapillary segment. The effect was inhibited by NPY and aspirin, an inhibitor of cyclo‐oxygenase, while ketanserin, a 5‐HT2 receptor antagonist, and SR140333, a new NK1 antagonist, had no protective effect. Phenylephrine increased ΔPa at high concentration, an effect also inhibited by NPY and aspirin. Substance P had no significant haemodynamic effect. When injected together with NPY, substance P (10−6m) induced a significant increase in the total pressure gradient.

It was concluded that NPY can protect the lung against acetylcholine‐ and capsaicin‐induced oedema via a prejunctional modulatory effect on the C‐fibres. NPY also inhibits acetylcholine‐evoked precapillary and phenylephrine‐induced arterial vasoconstriction, probably by interfering with cyclo‐oxygenase products synthesis.