bluuebunny/crossref_metadata_2025 huggingface dataset:
Endothelium‐dependent and independent relaxation of the rat aorta by cyclic nucleotide phosphodiesterase inhibitors
Authors: Komas Narcisse , Lugnier Claire , Stoclet Jean‐Claude
Published: July 2012

The effects of selective inhibitors of adenosine 3′: 5′‐cyclic monophosphate (cyclic AMP) and guanosine 3′: 5′‐cyclic monophosphate (cyclic GMP) phosphodiesterases (PDEs) were investigated on PDEs isolated from the rat aorta and on relaxation of noradrenaline (1 μm) precontracted rat aortic rings, with and without functional endothelium.

Four PDE forms were isolated by DEAE‐sephacel chromatography from endothelium‐denuded rat aorta: a calmodulin‐activated PDE (PDE I) which hydrolyzed preferentially cyclic GMP, two cyclic AMP PDEs (PDE III and PDE IV) and one cyclic GMP‐specific PDE (PDE V). The latter was selectively and potently inhibited by zaprinast. The two cyclic AMP PDEs were discriminated by specific inhibitors: one was inhibited by cyclic GMP (PDE III) and by new cardiotonic agents (milrinone, CI 930, LY 195115 and SK&F 94120); the other was inhibited by denbufylline and rolipram (PDE IV). None of these drugs significantly inhibited PDE I.

The PDE III inhibitors caused endothelium‐independent relaxations of rat aortic rings with the following EC50 values (μm concentration producing 50% relaxation): LY 195115: 3.4, milrinone: 5.7, CI 930; 7.8, SK&F 94120: 14.7. Neither NG‐monomethyl‐l‐arginine (l‐NMMA, 300 μm), an inhibitor of the l‐arginine‐NO pathway, nor l‐arginine (1 mm) modified the effect of PDE III inhibitors. However, methylene blue (10 μm) an inhibitor of soluble guanylate cyclase abolished relaxation induced by PDE III inhibitors except in the case of compound CI 930.

The specific PDE IV and PDE V inhibitors both produced endothelium‐dependent relaxations which were inhibited by l‐NMMA and by methylene blue (10 μm). In the presence of l‐NMMA, relaxation was restored by subsequent addition of l‐arginine.

The relaxant effects of denbufylline and rolipram were studied in the presence of drugs stimulating either adenylate cyclase (forskolin and isoprenaline) or soluble guanylate cyclase (sodium nitroprusside, SNP), or inhibiting PDE III (milrinone). In endothelium‐denuded rings, a relaxing effect of both denbufylline and rolipram was found in the presence of milrinone (EC50 values 1.7 and 12 μm, respectively) or SNP (EC50 values 12.3 and 124 μm, respectively), but not in the presence of forskolin or isoprenaline. However in the presence of functional endothelium, relaxations produced by PDE IV inhibitors were significantly potentiated by forskolin, isoprenaline, milrinone and SNP (respective EC50 values for denbufylline: 2, 2, 0.4 and 0.7 μm and for rolipram: 7, 13, 7 and 1.2 μm).

These results indicate that the relaxant effects of inhibitors of the cyclic AMP‐specific PDE IV are markedly enhanced by cyclic GMP elevating agents and by the PDE III inhibitor milrinone. They support the hypothesis that cyclic GMP enhances cyclic AMP‐mediated relaxation, possibly through the inhibition of the cyclic GMP‐inhibited PDE III.
BAY u3405 an antagonist of thromboxane A2‐ and prostaglandin D2‐induced bronchoconstriction in the guinea‐pig
Authors: Francis H.P. , Greenham S.J. , Patel U.P. , Thompson A.M. , Gardiner P.J.
Published: July 2012

The novel thromboxane (TX) antagonist, BAY u3405, has been evaluated against bronchoconstriction induced by the TXA2 mimetic U‐46619, prostaglandin D2 (PGD2), 5‐hydroxytryptamine (5‐HT), leukotriene D4 (LTD4) and histamine in the guinea‐pig in vivo by use of a modification of the model described by Konzett & Rössler.

When given intravenously (i.v.) at 30 or 100 μg kg−1, U‐46619 caused 80% maximal bronchoconstriction in most animals. In contrast, PGD2 caused a smaller 40%‐50% maximal bronchoconstriction at the highest dose tested (300 μg kg−1, i.v.).

BAY u3405, given intravenously, orally (p.o.) or by aerosol antagonized U‐46619‐induced bronchoconstriction in a dose‐related manner. The approximate ID50 values were 600 μg kg−1, i.v., 1.7 mg kg−1 p.o. and 0.1% w/v 20 breaths by aerosol.

BAY u3405 had similar inhibitory activities against U‐46619‐induced bronchoconstriction and hypertension suggesting that it had no preferential activity on the airways.

When given intravenously BAY u3405 antagonized the bronchoconstrictor effect of intravenous PGD2 with ID50 values between 30–100 μg kg−1.

The action of BAY u3405 (10 mg kg−1, p.o.) was long lasting, causing significant inhibition of U‐46619‐induced bronchoconstriction 7 h after dosing.

At 1 mg kg−1, i.v., a dose that abolished the response to U‐46619 and PGD2, BAY u3405 had no effect on histamine‐, 5‐HT‐ or LTD4‐induced bronchoconstriction.

BAY u3405 potently and selectively antagonized U‐46619‐ or PGD2‐induced bronchoconstriction in the Konzett‐Rössler model of guinea‐pig lung function. It should therefore prove to be a useful tool for defining the role of TXA2 and PGD2 in airway diseases such as asthma.
The actions of propofol on inhibitory amino acid receptors of bovine adrenomedullary chromaffin cells and rodent central neurones
Authors: Hales Tim G. , Lambert Jeremy J.
Published: July 2012

The interaction of the intravenous general anaesthetic propofol (2,6‐diisopropylphenol) with the GABAA receptor has been investigated in voltage‐clamped bovine chromaffin cells and rat cortical neurones in cell culture. Additionally, the effects of propofol on the glycine and GABAA receptors of murine spinal neurones were determined.

Propofol (1.7–16.8 μm) reversibly and dose‐dependently potentiated the amplitude of membrane currents elicited by GABA (100 μm) applied locally to bovine chromaffin cells. Intracellular application of propofol (16.8 μm) was ineffective. In rat cortical neurones and murine spinal neurones, extracellular application of 8.4 μm and 1.7–16.8 μm propofol respectively produced a potentiation of GABA‐evoked currents qualitatively similar to that seen in the bovine chromaffin cell.

The potentiation by propofol (1.7 μm) was not associated with a change in the reversal potential of the GABA‐evoked whole cell current. On outside‐out membrane patches isolated from bovine chromaffin cells, propofol (1.7 μm) had little or no effect on the GABA single channel conductances, but greatly increased the probability of the GABA‐gated channel being in the conducting state.

The potentiation of GABA‐evoked whole cell currents by propofol (1.7 μm) was not influenced by the benzodiazepine antagonist flumazenil (0.3 μm). A concentration of propofol (1.7 μm) that substantially potentiated GABA currents had little effect on currents induced by the activation of the GABAA receptor by pentobarbitone (1 mm).

Bath application of propofol (8.4–252 μm), to bovine chromaffin cells voltage clamped at −60 mV, induced an inward current associated with an increase in membrane current noise on all cells sensitive to GABA. Intracellular application of propofol (16.8 μm) was ineffective in this respect. Local application of propofol (600 μm) induced whole cell currents with a reversal potential dependent upon the Cl− gradient across the cell membrane.

On outside‐out membrane patches formed from bovine chromaffin cells, propofol (30 μm) induced single channels with mean chord conductances of 29 and 12 pS. The frequency of propofol channels was greatly reduced by coapplication of 1 μm bicuculline. Under identical ionic conditions, GABA (1 μm) activated single channels with mean chord conductances of 33, 16 and 10 pS.

Bath applied propofol (0.84–16.8 μm) dose‐dependently potentiated strychnine‐sensitive currents evoked by glycine (100 μm) in murine spinal neurones.

The relevance of the present results to the general anaesthetic action of propofol is discussed.
Nitric oxide from vascular smooth muscle cells: regulation of platelet reactivity and smooth muscle cell guanylate cyclase
Authors: Mollace Vincenzo , Salvemini Daniela , Anggard Erik , Vane John
Published: July 2012

Incubation of smooth muscle cells (SMC) from bovine aorta for 3 min with human washed platelets treated with indomethacin (10 μm) promoted a cell number‐related inhibition of platelet aggregation induced by thrombin (40 mu ml−1). This inhibition was not attributable to products of the cyclooxygenase pathway for the SMC were also treated with indomethacin (10 μm).

The inhibitory activity of the SMC on platelet aggregation was enhanced by incubating the SMC with E. coli lipopolysaccharide (LPS, 0.5 μg ml−1) for a period of 9 to 24 h. This effect was attenuated when cycloheximide (10 μg ml−1) was incubated together with LPS. Cycloheximide did not prevent the inhibitory activity of the non‐treated cells.

The inhibition of platelet aggregation obtained with non‐treated or LPS‐treated SMC was potentiated by superoxide dismutase (SOD, 60 u ml−1) and ablated by oxyhaemoglobin (OxyHb, 10 μm). Preincubation of the SMC with NG‐monomethyl‐l‐arginine (l‐NMMA, 30–300 μm) for 60 min prevented their antiaggregatory activity. This effect was reversed by concurrent incubation with l‐arginine (l‐Arg, 100 μm) but not with d‐arginine (d‐Arg, 100 μm).

Exposure of the non‐treated SMC (5 × 105 cells) to stirring (1000 r.p.m., 37°C) for 10 min led to a significant increase in their levels of guanosine 3′:5′‐cyclic monophosphate (cyclic GMP) but not adenosine 3′:5′‐cyclic monophosphate (cyclic AMP). l‐NMMA (300 μm) attenuated the increase in cyclic GMP induced by stirring but did not affect the basal levels of cyclic GMP in the cells. The inhibitory activity of l‐NMMA was reversed by co‐incubation with l‐Arg (100 μm) but not d‐Arg (100 μm). l‐Arg alone had no effect on the levels of cyclic GMP. In the absence of stirring, a 10 min stimulation of the non‐treated SMC with glyceryl trinitrate (GTN, 200 μm) or atrial natriuretic factor (ANF, 10−7 m) led to an increase in the levels of cyclic GMP but not cyclic AMP. The increase in cyclic GMP promoted by GTN or ANF was not affected by l‐NMMA. The levels of cyclic GMP were higher in the LPS (0.5 μg ml−1, 18 h)‐treated cells (5 × 10−5) and stirring was more effective in increasing the levels of cyclic GMP in these cells.

These findings support the idea that non‐treated or LPS‐treated cultured SMC can produce an NO‐like factor. Production by the latter requires protein synthesis as evidenced by blockade with cycloheximide. This NO‐like factor may play a role in the auto‐regulation of smooth muscle cell reactivity through a cyclic GMP‐dependent mechanism.
Electrophysiological effects of amineptine on neurones of the rat substantia nigra pars compacta: evidence for an inhibition of the dopamine uptake system
Authors: Mercuri N.B. , Stratta F. , Calabresi P. , Bernardi G.
Published: July 2012

Intracellular recordings were made from substantia nigra pars compacta neurones of the rat maintained in vitro in order to study the effects of the tricyclic antidepressant drug, amineptine.

Amineptine hydrochloride (1–30 μm) decreased spontaneous firing and slightly hyperpolarized the membrane potential. In neurones voltage‐clamped at − 50 or − 60 mV, amineptine produced an outward membrane current. These actions were concentration‐dependent and were completely antagonized by (−)‐sulpiride.

The amineptine‐induced hyperpolarization was resistant to tetrodotoxin (1 μm) but it was abolished in 0 mm Ca2+ (plus 13 mm MgCl2) solutions.

Amineptine (300 nm‐30 μm) and cocaine (10–30 μm) increased the amplitude and duration of responses to exogenously applied dopamine. The effects of dopamine were potentiated about 5 fold by 10 μm amineptine; this potentiation persisted in calcium‐free solutions.

Cocaine (10 μm) had no additional effect on the dopamine‐induced responses in the presence of amineptine (30 μm). Amineptine (10 μm) produced no detectable effects in the presence of cocaine (30 μm).

It is concluded that amineptine acts as a dopamine uptake blocker in slices of rat substantia nigra.
ATP modulates the efferent function of capsaicin‐sensitive neurones in guinea‐pig isolated atria
Authors: Rubino Annalisa , Amerini Sandra , Ledda Fabrizio , Mantelli Laura
Published: July 2012

The effect of adenosine triphosphate (ATP) and its stable analogues, α,β‐methylene‐ATP and β,γ‐methylene‐ATP, on the efferent function of capsaicin‐sensitive non‐adrenergic, non‐cholinergic (NANC) nerves was tested in guinea‐pig isolated atria.

Transmural nerve stimulation of atria isolated from reserpine‐pretreated guinea‐pigs, in the presence of 1 μm atropine and 0.3 μm CGP 20712A, induced a transient positive inotropic effect attributable to calcitonin gene‐related peptide (CGRP) release from NANC nerve endings.

ATP (1–30 μm) concentration‐dependently reduced the cardiac response to transmural nerve stimulation, without affecting the inotropic response to 10 nm exogenous CGRP. The inhibitory effect of ATP was competitively antagonized by the P1‐purinoceptor antagonist, 8‐phenyltheophylline (8‐PT, 1 μm), but was unaffected by the P2‐purinoceptor antagonist, suramin (100 μm).

β,γ‐Methylene‐ATP in the same concentration range as ATP, inhibited the cardiac response to transmural nerve stimulation. The inhibitory effect of β,γ‐methylene ATP was antagonized by 1 μm 8‐PT. The desensitizing agonist for P2‐purinoceptors, α,β‐methylene ATP did not induce any inhibitory effect either on the cardiac response to transmural nerve stimulation or on the inhibitory effect curve for ATP.

The inhibitory effect of ATP on the NANC neurotransmission was inconsistently modified in the presence of 10 μm α,β‐methylene‐adenosine diphosphate, an inhibitor of the 5′‐ectonucleotidases.

These results demonstrate that ATP modulates the efferent function of cardiac NANC nerve endings through prejunctional inhibitory receptors belonging to the P1 type. The metabolic conversion of ATP to adenosine does not seem to be a pre‐requisite for the ATP agonist activity.
Bovine polymorphonuclear leukocytes increase sensitivity to noradrenaline in isolated mesenteric arteries
Authors: De Kimpe S.J. , Van Heuven‐Nolsen D. , Nijkamp F.P.
Published: July 2012

The effects of polymorphonuclear leukocytes (PMN) on vascular function to (−)‐noradrenaline were examined in vitro. Purified bovine PMN were incubated in siliconized organ baths containing rings of bovine mesenteric arteries, after which a concentration‐effect curve in response to (−)‐noradrenaline was obtained.

PMN‐derived products induced a long lasting concentration‐dependent contraction of the blood vessels generating 24.4 ± 6.8% of the maximal tension to (−)‐noradrenaline at a cell concentration of 2.5 × 106 ml−1. The contractile response was also found in endothelium‐denuded vascular rings.

PMN present in the organ bath caused an increase in the sensitivity of vascular rings to (−)‐noradrenaline. At a cell number of 2.5 × 106 PMN ml−1 the pD2‐value for (−)‐noradrenaline was augmented 0.40 ± 0.05 (P < 0.001), while total contraction at the highest concentration (−)‐noradrenaline was not affected. This increase in sensitivity was dependent on an intact endothelium.

The increase in sensitivity to (−)‐noradrenaline by PMN was inhibited by superoxide dismutase, but not by catalase, dimethylthiourea, indomethacin or nordihydroguaiaretic acid. The non‐stimulated bovine PMN produced oxygen radicals as measured by chemiluminescence.

Simultaneous incubation of PMN and (−)‐noradrenaline with arterial rings induced an increase in the release of prostacyclin, measured by an elevated concentration of 6‐keto‐prostaglandin Fα in the supernatant.

It is concluded that PMN can increase vascular tone directly or indirectly probably via the interaction of PMN‐derived superoxide anions with endothelium‐derived relaxing factor.
Mechanism of action of MY‐1250, an active metabolite of Repirinast, in inhibiting histamine release from rat mast cells
Authors: Takei Masao , Endo Koichi , Takahashi Kazuo
Published: July 2012

When MY‐1250 (3.6 × 10−5 m) was added to mast cells, it caused a rapid increase in adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) and decrease in adenosine 5′‐triphosphate (ATP), both of which recovered to their original levels within 2 min. The accumulation of cyclic AMP was maximal at 20 s after challenge with MY‐1250. The minimum level of ATP was observed at 20 s after addition of MY‐1250.

The initial rise in [Ca2+]i and the histamine release induced by DNP‐AS antigen (40 μg ml−1) was most strongly inhibited at 20 s after incubation of the mast cells with MY‐1250.

MY‐1250 strongly and dose‐dependently inhibited the histamine release from rat mast cells induced by antigen. Moreover, MY‐1250 strongly inhibited calcium ion mobilization from the intracellular Ca2+‐store.

These results suggested that the inhibitory mechanism of MY‐1250 on the initial rise in [Ca2+]i and histamine release induced by antigen was due to the inhibition of ATP‐dependent Ca2+‐release from the intracellular Ca2+‐stores.
Comparisons of the effects of ryanodine on catecholamine secretion evoked by caffeine and acetylcholine in perfused adrenal glands of the guinea‐pig
Authors: Nakazato Yoshikazu , Hayashi Hirotaka , Teraoka Hiroki
Published: July 2012

The effect of ryanodine on catecholamine secretion induced by caffeine and muscarinic receptor activation was investigated in perfused adrenal glands of the guinea‐pig.

Caffeine (40 mm) caused only a small increase in catecholamine secretion during perfusion with standard Locke solution. Caffeine‐induced catecholamine secretion was markedly enhanced after removal of CaCl2 together with replacement of NaCl with sucrose.

In the absence of CaCl2 and NaCl, 50 μm ryanodine had no effect on the resting catecholamine secretion. Caffeine (40 mm) administered 15 min after treatment with ryanodine caused an increase in catecholamine secretion similar to that prior to application of ryanodine, but failed to have any effect thereafter. Combined application of ryanodine and caffeine also prevented catecholamine secretion induced by caffeine applied subsequently.

Catecholamine secretion induced by 100 μm acetylcholine (ACh) was only partially inhibited after treatment with ryanodine plus caffeine under Ca2+‐free, Na+‐deficient conditions.

Preferential influence of ryanodine on the response to caffeine was also confirmed in catecholamine secretion evoked by paired stimuli with caffeine and ACh alternately, during perfusion with either Ca2+‐free Locke or sucrose‐substituted solutions.

These results indicate that caffeine increases catecholamine secretion by mobilizing Ca2+ from intracellular Ca2+ stores through ryanodine‐sensitive mechanisms in guinea‐pig adrenal chromaffin cells. Ca2+ stores sensitive to caffeine and muscarinic receptor activation may not overlap entirely.
The role of cyclic AMP in non‐adrenergic non‐cholinergic contraction in guinea‐pig bronchi
Authors: Aikawa Takashi , Sekizawa Kiyohisa , Morikawa Masatoshi , Itabashi Shigeru , Sasaki Hidetada , Takishima Tamotsu
Published: July 2012

We investigated the role of adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) in non‐adrenergic non‐cholinergic (NANC) contraction in guinea‐pig bronchial strips.

Forskolin (3 nm to 1 μm) reduced NANC contraction induced by electrical field stimulation (EFS) in a concentration‐dependent fashion (–log EC50 was 7.22 ± 0.12 m and maximum inhibition was 100 ± 0.01%). However, forskolin (< 1 μm) did not alter the contraction induced by substance P (SP, 1 μm).

Dibutyryl cyclic AMP (1 mm) also reduced NANC contractions induced by EFS (100 ± 0.01%) without significant effect on SP (1 μm)‐induced contractions. In contrast, dibutyryl cyclic GMP (1 mm) was without effect against either NANC or SP‐induced contractions.

Both the β2‐adrenoceptor agonist, procaterol (0.1 nm to 3 nm) and theophylline (100 nm to 1 mm) concentration‐dependently reduced EFS‐induced NANC contractions without significant effect on SP (1 μm)‐induced contractions.

In contrast to forskolin, procaterol and theophylline, both sodium nitroprusside and cromakalim inhibited the EFS‐induced contractions only at those concentrations that similarly reduced the contractions induced by SP (1 μm).

These results suggest that cyclic AMP may mediate pre‐junctional inhibition of NANC contractions in guinea‐pig bronchi.
Okadaic acid inhibits activation of phospholipase C in human platelets by mimicking the actions of protein kinases A and C
Authors: Walker Trevor R. , Watson Steve P.
Published: July 2012

The effect of okadaic acid, a potent inhibitor of protein phosphatases 1 and 2A (PP1 and PP2A), on human platelets has been investigated.

Okadaic acid exerts a general increase in phosphorylation of platelet proteins but did not induce aggregation or secretion of 5‐hydroxytryptamine (5‐HT). Okadaic acid, however, did inhibit thrombin‐induced functional responses.

Maximally effective concentrations of prostacyclin, to elevate adenosine 3′:5′‐cyclic monophosphate (cyclic AMP), or phorbol dibutyrate, to activate protein kinase C, inhibited the formation of inositol phosphates by thrombin by approximately 60%. When used in combination, prostacyclin and phorbol dibutyrate reduced the levels of inositol phosphates induced by thrombin to 11%.

Okadaic acid (1 μm) decreased thrombin‐induced formation of inositol phosphates by approximately 55% and increased the inhibitory action of prostacyclin or phorbol dibutyrate. Okadaic acid had no further effect when prostacyclin and phorbol dibutyrate were used in combination.

These results suggest that protein kinases A and C act to inhibit phospholipase C by distinct mechanisms and that their action is reversed by PP1 and/or PP2A.
Characterization of P1‐purinoceptors on rat duodenum and urinary bladder
Authors: Nicholls J. , Hourani S.M.O. , Kitchen I.
Published: July 2012

The P1‐purinoceptors mediating relaxation of the rat duodenum and inhibition of contraction of the rat urinary bladder were characterized by use of adenosine and its analogues 5′‐N‐ethylcarboxamidoadenosine (NECA), N6‐cyclopentyladenosine (CPA) and 2‐p‐((carboxyethyl)phenethylamino)‐5′‐carboxamidoadenosine (CGS 21680), as well as the A1‐selective antagonist 1,3‐dipropyl‐8‐cyclopentylxanthine (DPCPX). The stable analogue of adenosine 5′‐triphosphate (ATP), adenylyl 5′‐(β,γ‐methylene)diphosphonate (AMPPCP), was also used as previous work had indicated that it has a direct action on some P1 receptors in addition to its P2‐purinoceptor activity.

In the rat duodenum, the order of potency of the adenosine agonists was NECA ≥ CPA > AMPPCP = adenosine > CGS 21680, and DPCPX antagonized CPA and AMPPCP at a concentration of 1 nm whereas equivalent antagonism of NECA and adenosine required a concentration of 1 μm. This suggests the presence of a mixture of A1 and A2 receptors in this tissue, with CPA and AMPPCP acting on the A1 and NECA and adenosine acting on the A2 receptors.

In the rat bladder, the order of potency of the adenosine agonists for inhibition of carbachol‐induced contractions was NECA ≫ adenosine > CPA = CGS 21680, and a concentration of DPCPX of 1 μm was required to antagonize responses to NECA and adenosine. This suggests the presence of A2 receptors in this tissue. ATP and AMPPCP each caused contractions which were not enhanced by DPCPX (1 μm) which suggests that in this tissue AMPPCP was acting only via P2 receptors and had no P1 agonist activity. That AMPPCP was active on the A1 receptors in the duodenum but inactive on the A2 receptors in the bladder implies that it has selectivity for the A1 subtype.

That CGS 21680, which has been reported to bind selectively to the high affinity A2a subclass of A2 receptors, had a very low potency on the A2 receptors in the duodenum and in the bladder suggests that these receptors are of the low affinity A2b subclass.
Dopamine release and metabolism in the rat frontal cortex, nucleus accumbens, and striatum: a comparison of acute clozapine and haloperidol
Authors: Karoum F. , Egan M.F.
Published: July 2012

The effects of the typical and typical neuroleptic agents clozapine (CLZ) (2.5–20 mg kg−1, i.p.) and haloperidol (Hal) (0.05–1.0 mg kg−1), were compared on dopamine release and metabolism in the rat prefrontal cortex (PFC), nucleus accumbens (ACC) and striatum (ST). Dopamine release was estimated by measuring the steady‐state concentration of 3‐methoxytyramine (3‐MT) and the level of 3‐MT 10 min after pargyline (3‐MT accumulation); dopamine metabolism was evaluated from the steady‐state concentrations of its acidic metabolites.

Both drugs increased 3‐MT accumulation in the PFC in a dose‐dependent manner. In contrast to Hal, CLZ failed to increase 3‐MT accumulation in the ACC or ST. The ST was the region most sensitive to Hal in terms of 3‐MT accumulation and, by inference, dopamine release.

Both CLZ and Hal dose‐dependently elevated the concentrations of 3,4‐dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) in all 3 brain regions studied. The ACC appears to be the region most sensitive to these drugs in terms of changes in the levels of HVA.

The result of the present investigations suggest measurements of 3‐MT production in the 3 brain regions analysed might be a useful and simple pharmacological tool in the search for atypical neuroleptic drugs with a selectivity of action for the cortical systems.
Effects of pinacidil on guinea‐pig isolated perfused heart with particular reference to the proarrhythmic effect
Authors: Padrini Roberto , Bova Sergio , Cargnelli Gabriella , Piovan Donatella , Ferrari Mariano
Published: July 2012

The effects of pinacidil (10, 30, 50 μm) on contractility (+ dP/dtmax), coronary perfusion pressure (cP), and ECG intervals (PR, QRS, QT) have been studied on constant‐flow perfused guinea‐pig hearts, driven at four frequencies (2.5, 3, 3.5, 4 Hz).

Pinacidil decreased +dP/dtmax, cP and the QT interval in a dose‐dependent manner, whereas the PR interval was increased. QRS duration was not modified. All these effects were independent of driving frequency. Pinacidil decreased the interval from Q‐wave to T‐wave peak (QTpeak) to a greater extent than the QT interval, thus decreasing the QTpeak/QT ratio. This effect, unlike that on QT interval, was more evident at the highest frequency of stimulation.

In 4 out of 20 hearts treated with pinacidil sustained ventricular fibrillation (VF) occurred following a short run of premature ventricular beats (R on T phenomenon).

In separate experiments, an attempt to induce VF electrically was made at drug concentrations ranging from 10 μm to 100 μm (8 experiments for each concentration). In control conditions and at the lowest concentration of pinacidil tested (10 μm) VF could never be induced; in the presence of 30 μm pinacidil VF was induced in 5 out of 8 experiments. Drug concentrations higher that 50 μm permitted the induction of VF in every case.

Although the concentrations of pinacidil producing ventricular fibrillation are 30–40 times higher than those found in patients under long term treatment with this agent, it is suggested that caution should be used in prescribing this drug, at least in patients suffering from myocardial ischaemia.
Identification of inhibitors of nitric oxide synthase that do not interact with the endothelial cell l‐arginine transporter
Authors: Bogle Richard G. , Moncada Salvador , Pearson Jeremy D. , Mann Giovanni E.
Published: July 2012

The effects of inhibitors of nitric oxide (NO) synthase and other cationic amino acids on unidirectional l‐arginine transport were studied in porcine aortic endothelial cells cultured in microwell plates or perfused in microcarrier columns. l‐Homoarginine, l‐lysine and l‐ornithine inhibited transport of l‐arginine. The NO synthase inhibitors NG‐monomethyl‐l‐arginine and NG‐iminoethyl‐l‐ornithine also reduced l‐arginine uptake, whereas NG‐nitro‐l‐arginine and its methyl‐ester had no inhibitory effect. The ability to modulate selectively endothelial cell l‐arginine transport or NO synthase activity will allow further characterization of the arginine transporter and its role in regulating NO biosynthesis.
Formoterol and salbutamol inhibit bradykinin‐ and histamine‐induced airway microvascular leakage in guinea‐pig
Authors: Advenier C. , Qian Y. , Koune J‐D. Law , Molimard M. , Candenas M‐L. , Naline E.
Published: July 2012

The effects of the β2‐adrenoceptor agonists, salbutamol and formoterol, on the increase of microvascular permeability induced by histamine or bradykinin in guinea‐pig airways have been studied in vivo. Extravasation of intravenously injected Evans blue dye was used as an index of permeability. The effects of salbutamol and formoterol on the increase in pulmonary airway resistance induced by histamine or bradykinin have also been studied.

The increase in pulmonary airway resistance induced by histamine or bradykinin was totally inhibited by salbutamol and formoterol. The ED50 of the two mediators were 0.59 ± 0.21 (n = 5) and 0.20 ± 0.14 (n = 5) μg kg−1 respectively for salbutamol, and 0.13 ± 0.12 (n = 6) and 0.02 ± 0.01 (n = 6) μg kg−1 respectively for formoterol.

Salbutamol (10 and 30 μg kg−1) and formoterol (1 and 10 μg kg−1) inhibited the increase of microvascular permeability induced by histamine (30 μg kg−1) in the guinea‐pig airways. The inhibitory effect was predominant in the trachea and the main bronchi, with a maximum inhibition of 20 to 50%. The two drugs had little or no inhibitory effect on the other structures studied, viz. nasal mucosa, larynx, proximal and distal intrapulmonary airways.

Salbutamol and formoterol (1 and 10 μg kg−1) abolished the increase in microvascular permeability induced by bradykinin (0.3 μg kg−1). This inhibitory effect of two β‐adrenoceptor stimulants was predominant in the trachea and the nasal mucosa where it was observed with 1 μg kg−1 of the β‐adrenoceptor agonists. In the main bronchi, and in the proximal and distal intrapulmonary airways, the effects of bradykinin were abolished by 10 μg kg−1 of formoterol and salbutamol.

The effects of bradykinin, but not those of histamine, were significantly reduced (nasal mucosa, main bronchi and distal intrapulmonary airways) or abolished (trachea, proximal intrapulmonary airways) by morphine 10 mg kg−1, i.v. These results suggest that an indirect effect, through non‐adrenergic non‐cholinergic (NANC) nerves is involved in the action of bradykinin on the microvascular permeability.

In conclusion, intravenously injected β‐adrenoceptor stimulants can inhibit, partially or totally, the increase of airways microvascular permeability induced by intravenous histamine or bradykinin. However, these effects require doses that are higher than those that inhibit the increase in pulmonary airway resistance induced by these mediators. As suggested by the results obtained with morphine, the higher efficacy of β2‐adrenoceptor agonists versus bradykinin may occur through activation of presynaptic receptors of the non‐adrenergic non‐cholinergic (NANC) nerves preventing release of inflammatory neuropeptides such as substance P and neurokinin A.
Effect of treatment with vitamin D3 on the responses of the duodenum of spontaneously hypertensive rats to bradykinin and to potassium
Authors: Feres Teresa , Vianna Lucia M. , Paiva Antonio C.M. , Therezinha  , Paiva B.
Published: July 2012

The diet of spontaneously hypertensive rats (SHR) and normotensive Wistar‐Kyoto (WKY) and Wistar (NWR) rats was supplemented with either 2% calcium lactate in the drinking water or 12.5 μg vitamin D3 100 g−1 body weight daily by gavage, for 14 days.

The blood pressure of the SHR treated with either calcium or vitamin D decreased to the same levels as that of WKY and NWR.

The response to bradykinin of the SHR isolated duodenum, which is predominantly contractile, upon treatment with vitamin D (but not with calcium), became predominantly relaxant, approaching the normal behaviour of the WKY and NWR duodenum.

The relaxant responses of the SHR and WKY duodenum to potassium were smaller than those of NWR, but treatment with vitamin D increased the response in all three rat strains.

It is concluded that, besides sharing the hypotensive effect of calcium, vitamin D treatment of SHR has an effect on the duodenum smooth muscle which might be due to calmodulin‐dependent activation of calcium‐dependent potassium channels.
Characterization of histamine receptor sub‐types regulating prostacyclin release from human endothelial cells
Authors: Bull Helen A. , Courtney P.F. , Rustin M.H.A. , Dowd Pauline M.
Published: July 2012

The histamine receptor sub‐types that are involved in the initiation and maintenance of prostacyclin (PGI2) release from human endothelial cells have been investigated.

Endothelial cells cultured from umbilical vein (HUVEC) were incubated with either histamine, the selective H1‐receptor agonists, 2‐methyl histamine (2‐MeHA) or thiazolylethylamine (ThEA), the H1‐agonist/H3‐antagonist, β‐histine (β‐His), the selective H2‐agonist, dimaprit, the H2‐agonist/H3‐antagonist, impromidine, the selective H3‐agonist, (R)α‐methylhistamine ((R)α‐MeHA) and the H3‐antagonist, thioperamide.

The H1‐agonists and the H3‐agonist (R)α‐MeHA induced a concentration (100 nm‐1 mm) and time‐dependent release of PGI2 as determined by radioimmunoassay for 6‐keto‐PGF1α, but were less potent than histamine itself. The rank order of potency was the same following 30 min and 24 h incubation, i.e. histamine > ThEA > 2‐MeHA ≫ β‐His > (R)α‐MeHA.

Histamine and 2‐MeHA (1 μm‐1 mm), ThHEA (10 μm‐1mm) and (R)α‐MeHA (1 mm), but not β‐His, induced a significantly greater increase in PGI2 release after 24 h incubation than after 30 min incubation (P < 0.05).

Neither the selective H2‐agonist, dimaprit, nor the H2‐agonist/H3‐antagonist, impromidine alone induced release of PGI2.

6 The H1‐antagonist, mepyramine (10 μm), abolished release of PGI2 induced by histamine, the H1‐agonists and (R)α‐MeHA but the H2‐antagonist cimetidine (10 μm) and the H2/H3‐antagonist, burimamide (10 μm) did not significantly modulate PGI2 release.

7 Although the H3‐agonist (R)α‐MeHA induced release of PGI2, it failed to modulate PGI2 release in the presence of histamine.

8 Low concentrations of the H3‐antagonist, thioperamide (100 nm) did not modulate histamine release of PGI2 at all but after 24 h incubation, thioperamide (10−4 m) partially reduced PGI2 release in the presence of histamine.

9 These results indicate that PGI2 from HUVEC is initiated and maintained via histamine H1‐receptor occupancy. There appears to be no involvement of either H2‐ or H3‐receptors in this particular endothelial cell histaminergic response.
Participation of protein kinase C in endothelin‐1‐induced contraction in rat aorta: studies with a new tool, calphostin C
Authors: Shimamoto Hiroyuki , Shimamoto Yoriko , Kwan Chiu‐Yin , Daniel Edwin E.
Published: July 2012

Calphostin C at 10−6 m was shown to be selective and highly effective in inhibiting contractile responses of rat aortae to 12‐o‐tetradecanoylphorbol‐13‐acetate, while it had no effect on contractile responses to elevated KCl.

In the rat aorta, endothelin‐1 (ET‐1) developed a sustained tonic contraction dose‐dependently in both normal Ca2+‐containing Krebs and Ca2+‐free Krebs containing 1 mm EGTA. Calphostin C (10−6 m), a selective protein kinase C inhibitor, antagonized the maximal tensions for cumulative addition of 10−8 m ET‐1 by 13.2% in Ca2+‐containing medium and 25.8% in Ca2+‐free Krebs containing 1 mm EGTA.

In both Ca2+‐containing medium and Ca2+‐free Krebs containing 1 mm EGTA, precontraction with 10−8 m ET‐1 had no effects on the contractile response to subsequently added 10−6 m 12‐o‐tetradecanoylphorbol‐13‐acetate (TPA), an activator of protein kinase C.

In Ca2+‐free Krebs containing 1 mm EGTA, precontraction with 10−6 m TPA potentiated the contractile response to subsequently added 10−8 m ET‐1, whereas this potentiation was abolished by pretreatment with 10−6 m calphostin C. The mechanism of the TPA‐induced potentiating effect remains to be determined.

These results suggest that the participation of protein kinase C in the 10−8 m ET‐1‐induced contraction may be 13.2% and 25.8% in the presence and absence of extracellular Ca2+, respectively, and that mechanisms other than protein kinase C may be predominantly responsible for ET‐1‐induced tonic contraction.
Capsazepine reversal of the antinociceptive action of capsaicin in vivo
Authors: Perkins M.N. , Campbell E.A.
Published: July 2012

This study was designed to investigate the antinociceptive activity of capsaicin in acute and persistent nocifensor behavioural models and to determine whether this was mediated via a specific receptor interaction, by use of the antagonist, capsazepine.

Capsaicin administered systemically in low doses produced antinociception in the knee joint hyperalgesia, rat paw pressure, rat and mouse tail flick and mouse hot plate nocifensor models with similar efficacy in all tests.

The novel competitive capsaicin antagonist, capsazepine, prevented the capsaicin‐induced antinociception when administered systemically in the range 50–100 μmol kg−1.

Capsazepine administered by itself had no antinociceptive actions.

These data describe a joint hyperalgesic model in the rat which does not rely on extreme nocifensor behavioural endpoints and it is suggested this is a useful model for investigating mechanisms in persistent pain.

6 This is the first demonstration of antagonism by capsazepine of the behavioural antinociceptive properties of capsaicin and provides further evidence that capsaicin acts to reduce nociceptive thresholds via a specific receptor.
Effects of the bradykinin antagonist, HOE 140, in experimental acute pancreatitis
Authors: Griesbacher T. , Lembeck F.
Published: July 2012

The novel bradykinin antagonist, HOE 140, completely blocked the fall in rabbit blood pressure caused, not only by i.v. bradykinin, but also by i.v. kallikrein. This shows that both the effects of exogenously administered bradykinin and those of endogenously released kinins are antagonized by HOE 140.

Acute pacreatitis was induced in rats by i.v. infusion of the cholecystokinin analogue, caerulein. This treatment resulted in massive oedema of the pancreas, increased activities of amylase and lipase in serum and a characteristic, biphasic fall in blood pressure.

HOE 140 prevented the caerulein‐induced pancreatic oedema and the second phase of hypotension whereas NPC 349, a widely used, but short‐acting, bradykinin antagonist did not show a significant inhibition. HOE 140, in contrast to its inhibitory effects on caerulein‐induced pancreatic oedema and hypotension, significantly augmented the increases in amylase and lipase activities in serum.

It is concluded that in this model of acute pancreatitis, the release of kinins induces pancreatic oedema and hypotension. Prevention by HOE 140 of the kinin‐induced oedema allows the pancreatic enzymes to leave the tissue without hindrance and thus will diminish subsequent pathological events. It is suggested that the results obtained with the highly potent and long‐acting bradykinin antagonist, HOE 140, provide a pharmacological basis for a clinical trial in acute pancreatitis.
The actions of capsaicin applied topically to the skin of the rat on C‐fibre afferents, antidromic vasodilatation and substance P levels
Authors: Lynn B. , Ye W. , Cotsell B.
Published: July 2012

Single applications of solutions of capsaicin were made to the intact skin of anaesthetized rats and the effects on cutaneous blood flow and the firing of C‐nociceptor afferents determined. Blood flow was measured by laser‐Doppler flowmetry. C‐fibre activity was recorded from filaments dissected from the saphenous nerve.

Following the application of a capsaicin solution (concentration ≥ 1 mm) to rat saphenous skin, low frequency firing occurred in C‐polymodal nociceptors that sometimes continued for > 10 min. At the some time, large increases in skin blood flow occurred exceeding 300% in some instances.

After the initial excitation, some C‐polymodal nociceptors lost their sensitivity to pressure whilst their sensitivity to heat was lost or enhanced depending on the vehicle used.

Sensitivity of C‐polymodal nociceptors to heat recovered in > 1 day following a single application of 33 mm capsaicin. Thresholds to mechanical pressure, however, were still significantly elevated by 123% on day 1, but had recovered on day 2.

Vasodilatation in response to saphenous nerve stimulation (‘antidromic vasodilatation’) was significantly reduced by 35%, 2 days after a single application of 33 mm capsaicin, but was normal at 4 days.

6 Following a single application of 33 mm capsaicin, skin substance P levels fell to only half the normal value at day 1 and remained at this level throughout the 4 day period examined.

7 It is suggested that the ability of relatively low concentrations of capsaicin to desensitize C‐fibre nociceptors may underlie the analgesic action of topical capsaicin in man.
NG‐hydroxy‐L‐arginine prevents the haemodynamic effects of nitric oxide synthesis inhibition in the anaesthetized rat
Authors: Walder Claire E. , Thiemermann Christoph , Vane John R.
Published: July 2012

We have investigated the effects of l‐hydroxy‐l‐arginine (l‐HOArg), an intermediate in the biosynthesis of nitric oxide (NO) from l‐arginine (l‐Arg), on the haemodynamic effects (systemic blood pressure and renal blood flow) of the NO synthesis inhibitor NG‐nitro‐l‐arginine methyl ester (l‐NAME) in the anaesthetized rat.

l‐Arg or l‐HOArg (3 mg kg−1 min−1), but not d‐arginine (d‐Arg) or NG‐hydroxy‐d‐arginine (d‐HOArg), elicited a slight but significant increase in total renal blood flow (RBF) of 11 ± 2% and 11 ± 1%. Since mean arterial blood pressure (MAP) did not change this dose of l‐Arg or l‐HOArg resulted in a reduced renal vascular resistance (RVR) of the same magnitude.

Bolus injections of l‐NAME, at 0.3 or 1 mg kg−1 i.v., produced a significant fall in RBF of 11 ± 2% and 32 ± 5% and an increase in MAP of 7 ± 3 mmHg and 22 ± 5 mmHg, respectively. Consequently, RVR was elevated by 21 ± 5% and 52 ± 10%.

l‐Arg or l‐HOArg (3 mg kg−1 min−1) reduced the l‐NAME‐induced (0.3 or 1 mg kg−1) falls in RBF and increases in RVR by more than 65%. Neither d‐Arg nor d‐HOArg (3 mg kg−1 min−1) had any significant effect on the changes in RBF or RVR induced by l‐NAME.

l‐Arg or l‐HOArg (3 mg kg−1 min−1) attenuated the pressor effect of l‐NAME (3 mg kg−1) by 73% and 64%, respectively, while neither the d‐isomer of arginine nor hydroxyarginine had any effect.

6 These results demonstrate that l‐HOArg antagonizes the haemodynamic effects of NO‐biosynthesis inhibition in vivo, thus supporting the hypothesis that l‐HOArg is an intermediate in the formation of NO from l‐Arg.
L‐689,660, a novel cholinomimetic with functional selectivity for M1 and M3 muscarinic receptors
Authors: Hargreaves R.J. , McKnight A.T. , Scholey K. , Newberry N.R. , Street L.J. , Hutson P.H. , Semark J.E. , Harley E.A. , Patel S. , Freedman S.B.
Published: July 2012

L‐689,660, 1‐azabicyclo[2.2.2]octane, 3‐(6‐chloropyrazinyl)maleate, a novel cholinomimetic, demonstrated high affinity binding (pKD (apparent) 7.42) at rat cerebral cortex muscarinic receptors. L‐689,660 had a low ratio (34) of pKD (apparent) values for the displacement of binding of the antagonist [3H]‐N‐methylscopolamine ([3H]‐NMS) compared with the displacement of the agonist [3H]‐oxotremorine‐M ([3H]‐Oxo‐m), in rat cerebral cortex. Low NMS/Oxo‐M ratios have been shown previously to be a characteristic of compounds that are low efficacy partial agonists with respect to stimulation of phosphatidyl inositol turnover in the cerebral cortex.

L‐689,660 showed no muscarinic receptor subtype selectivity in radioligand binding assays but showed functional selectivity in pharmacological assays. At M1 muscarinic receptors in the rat superior cervical ganglion, L‐689,660 was a potent (pEC50 7.3 ± 0.2) full agonist in comparison with (±)‐muscarine. At M3 receptors in the guinea‐pig ileum myenteric plexus‐longitudinal muscle or in trachea, L‐689,660 was again a potent agonist (pEC50 7.5 ± 0.2 and 7.7 ± 0.3 respectively) but had a lower maximum response than carbachol. In contrast L‐689,660 was an antagonist at M2 receptors in guinea‐pig atria (pA2 7.2 (95% confidence limits 7, 7.4)) and at muscarinic autoreceptors in rat hippocampal slices.

The putative M1‐selective muscarinic agonist, AF102B (cis‐2‐methylspiro‐(1,3‐oxathiolane 5,3′)‐quinuclidine hydrochloride) was found to have a profile similar to L‐689,660 but had up to 100 times less affinity in binding and functional assays. RS‐86 (2‐ethyl‐8‐methyl‐2,8‐diazospiro[4,5]decan 1,3‐dione hydrochloride) also had lower affinity than L‐689,660, and had no binding selectivity for muscarinic receptor subtypes. RS‐86 had a higher NMS/Oxo‐M ratio than L‐689,660 and was a full agonist at M1, M2 and M3 receptors in the functional pharmacological assays.

The functional selectivity of L‐689,660 in muscarinic pharmacological assays is consistent with the effects of a low efficacy partial agonist in tissues with different effective receptor reserves.
The role of endothelial cells in the relaxations induced by 13‐hydroxy‐ and 13‐hydroperoxylinoleic acid in canine arteries
Authors: De Meyer Guido R.Y. , Bult Hidde , Verbeuren Tony J. , Herman Arnold G.
Published: July 2012

One of the major fatty acids in the arterial wall is linoleic acid. It has been shown that its 13‐hydroxy metabolite (13‐HODE) is generated in significant amounts by cultured endothelial cells. The aim of the present study was to investigate the relaxations to 13‐HODE and its hydroperoxyprecursor (13‐HPODE) and to examine the role of the endothelial cells.

Ring segments of canine circumflex and splenic artery were mounted in organ chambers for isometric tension recording. During contractions induced by prostaglandin F2α or noradrenaline, 13‐HODE and 13‐HPODE evoked dose‐dependent relaxations. Removal of the endothelial cells reduced the relaxations to 13‐HODE, but had no effect on those elicited by 13‐HPODE.

Indomethacin and meclofenamate (0.3 μm to 30 μm) blocked the relaxations evoked by 13‐HODE and 13‐HPODE in endothelium‐denuded rings. In segments with endothelium, both cyclo‐oxygenase inhibitors again abolished the relaxations to 13‐HODE, but only diminished those to 13‐HPODE.

Prostacyclin biosynthesis, as measured by radioimmunoassay, increased upon incubation with 13‐HODE and 13‐HPODE (10 μm). Bioassay of the release of nitric oxide (NO) indicated that NO was not involved in the relaxations elicited by either metabolite. Moreover, l‐NG‐nitroarginine (100 μm), a specific inhibitor of NO synthesis, did not influence the relaxations to 13‐HODE and 13‐HPODE. The responses to 13‐HPODE were also not altered by superoxide dismutase.

In the splenic artery 13‐HPODE and 13‐HODE induced contractions above 3 μm which were blocked by the thromboxane receptor antagonist, daltroban. In the circumflex artery contractile responses to high concentrations of 13‐HODE could be observed only after inhibition of cyclo‐oxygenase.

6 We conclude that the vasodilatation induced by 13‐HODE and 13‐HPODE was due to stimulation of prostacyclin biosynthesis both in the endothelium and smooth muscle cells or other subendothelial structures. An additional, unidentified intermediate, which was neither NO nor a cyclo‐oxygenase product nor superoxide anion, contributed to the relaxations to 13‐HPODE in arteries with endothelium.