bluuebunny/crossref_metadata_2025 huggingface dataset:
Characterization of adrenoceptors involved in the electrogenic chloride secretion by cultured rat epididymal epithelium
Authors: Leung A.Y.H. , Yip W.K. , Wong P.Y.D.
Published: July 2012

Short‐circuit current (SCC) technique was used to study the adrenoceptors involved in the electrogenic chloride secretion by cultured cauda epididymal epithelium of rats. Stimulation of the epithelium with noradrenaline (primarily β1‐adrenoceptor selective agonist), salbutamol (β2‐adrenoceptor selective agonist) and adrenaline (non‐selective β‐adrenoceptor agonist) led to a rise in SCC. At a low chart‐speed (2 mm min−1), the response profile to these agonists consisted of a peak followed by a sustained response considerably higher than the basal SCC.

The EC50s (doses of agonist producing 50% maximum response) of noradrenaline, salbutamol and adrenaline were 300, 115 and 10 nm respectively. Pretreating the tissues with 1 μm atenolol (β1‐selective antagonist) and 10 μm butoxamine (β2‐selective antagonist) shifted the dose‐response curves of noradrenaline (shifted EC50 = 4000 nm) and salbutamol (shifted EC50 = 1050 nm) to the right. Atenolol (1 μm) and butoxamine (10 μm) shifted the dose‐response curve of adrenaline to the right with new EC50s of 30 nm and 115 nm, respectively.

The rapidly rising phase of the SCC response to noradrenaline and adrenaline observed at low chart‐speed consisted of a brief and transient retraction followed by a rebound increase in SCC. At a high chart‐speed (1 mm s−1), the retraction and rebound phenomenon manifested as a fast initial spike which could be blocked by phentolamine (non‐specific α‐adrenoceptor antagonist) in a dose‐dependent fashion. Similar initial spikes were observed when the tissues were stimulated with phenylephrine (α1‐selective agonist) but not with isoprenaline (non‐selective β‐agonist) or forskolin (activator of adenylate cyclase). The response of the initial spike triggered by noradrenaline was dose‐dependent and the EC50 was 2000 nm.

The present study showed that the electrogenic chloride secretion by rat epididymis could be stimulated by α1, β1‐ and β2‐adrenoceptor agonists. The α1‐mediated response had a faster onset and more transient action than the β‐counterpart. It is postulated that epididymal chloride secretion might be regulated by neural (noradrenaline‐mediated) and humoral (adrenaline‐mediated) controls and that the stimulus‐secretion coupling mechanisms might involve both Ca2+ (α1‐mediated response) and adenosine 3′:5′‐cyclic monophosphate (β‐mediated response) as intracellular second messengers.
Involvement of nitric oxide pathway in the PAF‐induced relaxation of rat thoracic aorta
Authors: Moritoki Hideki , Hisayama Tetsuhiro , Takeuchi Shougo , Miyano Hiroaki , Kondoh Wataru
Published: July 2012

The mechanism of the vasorelaxant effect of platelet activating factor (PAF) on rat thoracic aorta and the effect of aging on the PAF‐induced relaxation were investigated.

PAF at concentrations causing relaxation induced marked increases in guanosine 3′:5′‐cyclic monophosphate (cyclic GMP) production, but did not induce an increase in adenosine 3′:5′‐cyclic monophosphate (cyclic AMP).

Removal of the endothelium by mechanical rubbing, and treatment with the PAF antagonists CV‐3988, CV‐6209 and FR‐900452, the nitric oxide biosynthesis inhibitor, NG‐nitro l‐arginine, the radical scavenger, haemoglobin, and the soluble guanylate cyclase inhibitor, methylene blue, inhibited PAF‐induced relaxation and abolished or attenuated PAF‐stimulated cyclic GMP production.

The relaxation was greatest in arteries from rats aged 4 weeks. With an increase in age, the response of the arteries to PAF was attenuated.

Endothelium‐dependent cyclic GMP production also decreased with increase in age of the rats.

6 These results suggest that PAF stimulates production of nitric oxide from l‐arginine by acting on the PAF receptors in the endothelium, which in turn stimulates soluble guanylate cyclase in the smooth muscle cells, and so increases production of cyclic GMP, thus relaxing the arteries. Age‐associated decrease in PAF‐induced relaxation may result from a reduction of cyclic GMP formation.
Activation of the human neutrophil 5‐lipoxygenase by leukotriene B4
Authors: McDonald Patrick P. , McColl Shaun R. , Naccache Paul H. , Borgeat Pierre
Published: July 2012

In the present study, we demonstrate that leukotriene B4 (LTB4) has the ability to activate the human neutrophil 5‐lipoxygenase (5‐LO).

Stimulation of neutrophils with 30 nm 14,15‐dideuterio‐LTB4 (D2‐LTB4) failed to induce the synthesis of LTB4 from endogenous arachidonic acid (AA), but stimulated the formation of LTB4 from 3.3 μm exogenous AA, as determined by GC‐MS analysis.

The stimulatory effect of LTB4 on 5‐LO activity was further examined with an alternative substrate; LTB4 time‐ and dose‐dependently stimulated the 5‐LO‐mediated conversion of exogenous 15(S)‐hydroperoxy‐5,8,11,13‐(Z,Z,Z,E)‐eicosatetraenoate (15‐HpETE) into 5(S), 15(S)‐dihydroxy‐6,8,11,13,‐(E,Z,Z,E)‐eicosatetraenoate (5,15‐DiHETE), with a threshold effect at 300 pm.

The ability of LTB4 to activate the 5‐LO showed structural specificity, since LTB4 was found to be 100 times more potent than ω‐hydroxy‐LTB4, and 300 times more potent than its Δ6‐trans‐12‐epi‐ isomer.

The LTB4‐induced 5‐LO activation was effectively inhibited by MK‐886 (an inhibitor of 5‐LO translocation), by pertussis toxin, and by the LTB4 receptor antagonist, LY‐223982.

6 These results demonstrate that the binding of LTB4 to its cell‐surface receptor results in 5‐LO activation in a process mediated by pertussis toxin‐sensitive guanine nucleotide‐binding proteins. Our data also suggest that the underlying mechanism involves a translocation of the 5‐LO to the membrane. These findings raise the possibility that LTB4 produced by phagocytes may positively feedback on its own synthesis.
Cardiovascular effects of substituted tetrahydroisoquinolines in rats
Authors: Dong Hui , Lee Chi‐Ming , Huang Wen‐Long , Peng Si‐Xun
Published: July 2012

A series of substituted tetrahydroisoquinolines derived from the cleavage products of tetrandrine were found to inhibit [3H]‐nitrendipine binding to rat cerebral cortical membranes. Those compounds which displaced [3H]‐nitrendipine binding were also able to inhibit high KCl‐induced contraction of rat aorta in vitro.

There was a significant correlation between the ability of these tetrahydroisoquinolines to inhibit [3H]‐nitrendipine binding and KCl‐induced contraction (r = 0.99, P < 0.001).

CPU‐23 (1‐{1‐[(6‐methoxy)‐naphth‐2‐yl]}‐propyl‐2‐(1‐piperidine)‐acetyl‐6,7‐dimethoxy‐1,2,3,4,‐tetrahydroisoquinoline), one of the most potent compounds identified in this series, behaved as a simple competitive inhibitor at the [3H]‐nitrendipine binding site and reduced the apparent affinity but not the maximal number of binding sites in saturation analysis.

In contrast to nifedipine which caused hypotension and tachycardia, CPU‐23 induced both hypotension and bradycardia in a dose‐dependent manner in pentobarbitone‐anaesthetized Sprague‐Dawley rats, spontaneously hypertensive and age‐matched normotensive WKY rats.

It is suggested that CPU‐23 may exert its cardiovascular effects via interaction with the dihydropyridine binding site on the L‐type calcium channel.
Effects of cholinoceptor and 5‐hydroxytryptamine3 receptor antagonism on erythromycin‐induced canine intestinal motility disruption and emesis
Authors: Qin Xin Yu , Pilot Marie‐Anne , Thompson Hilary , Scott Mark
Published: July 2012

Erythromycin administration is associated with gastrointestinal problems, disturbed gastrointestinal motility and emesis. This study in the dog investigates the underlying mechanisms.

Intestinal myoelectrical activity and the occurrence and latency of emesis were recorded in eight conscious dogs. All drugs were administered intravenously.

Erythromycin (7 mg kg−1) increased contractions of the proximal small intestine, and caused emesis in all fasted dogs and in 5 dogs after food. Atropine (50 mg kg−1 min−1) and hexamethonium (10 mg kg−1 h−1) partially inhibited the GI motility effects but did not significantly reduce emesis.

Metoclopramide at a high dose (2 mg kg−1 h−1) reduced the incidence of emesis in the presence of increased intestinal motility, but a low dose (150 μg kg−1 h−1) was ineffective.

A 5‐hydroxytryptamine3 (5‐HT3) receptor antagonist, MDL 72222 (1 mg kg−1), reduced emesis when given alone and combined with metoclopramide (low dose). The 5‐HT4 receptor agonist BRL24924 (Renzapride, 1 mg kg−1) had no effect on emesis either alone in combination with metoclopramide.

In conclusion, erythromycin‐induced GI motility disturbances and emesis are not causally related. Whereas the increase in intestinal smooth muscle activity is possibly cholinergically mediated, emesis occurs at least in part via a 5‐hydroxytryptaminergic mechanism, but does not involve the dopamine system.
Na+/H+ exchange inhibitors reverse lactate‐induced depression in postischaemic ventricular recovery
Authors: Karmazyn Morris
Published: July 2012

By use of pharmacological approaches, the present study examined the hypothesis that the deleterious effect of lactate on postischaemic ventricular recovery may be mediated, at least in part, by enhanced activation of the Na+/H+ exchanger at the time of reperfusion.

Spontaneously beating isolated hearts of the rat were subjected to 15 min zero‐flow global ischaemia followed by 30 min reperfusion. The effects of lactate (10, 20 or 40 mm) were studied by adding it 20 min before ischaemia whereas reperfusion was carried out with lactate‐free buffer.

Pretreatment with 20 or 40 mm lactate significantly reduced postischaemic recovery of developed force to 17 ± 3% and 16 ± 4% of preischaemic values (P < 0.05) compared to a 78 ± 4% recovery in control hearts. Similarly, recovery in ventricular rate was significantly reduced to 34 ± 7.6% and 38 ± 12% with 20 and 40 mm lactate, respectively compared to 97.5 ± 6.4% recovery in control hearts. At a concentration of 10 mm, lactate was without effect on either force or ventricular rate recovery.

Coadministration of either of two Na+/H+ exchange inhibitors, amiloride (174 μm) or 5‐N,N‐hexamethylene amiloride (HMA, 1 μm) with lactate and inclusion of the two drugs during the first 5 min of reperfusion resulted in reversal of lactate‐induced inhibition of force recovery with observed recoveries of 69 ± 6.7% and 64 ± 5% with amiloride and HMA, respectively. Similarly, recovery in ventricular rate was significantly enhanced to 92 ± 10% and 89 ± 6% with amiloride and HMA, respectively compared to 38 ± 12% recovery in control hearts.

In electrically paced hearts (5 Hz) subjected to 15 min ischaemia and 30 min reperfusion, 40 mm lactate significantly reduced force recovery to 36 ± 5.5% compared to 57 ± 6% in control hearts. In the presence of amiloride or HMA, force recovery in lactate‐treated hearts was significantly increased to 68 ± 16% and 72 ± 4.7% of preischaemic values, respectively.

In spontaneously beating hearts, resting tension changes during both ischaemia and reperfusion were not statistically different between treatment groups. However, in paced hearts pretreated with 40 mm lactate the elevation in resting tension during the first 5 min of reperfusion, was significantly reduced by both amiloride and HMA.

Changes in functional recoveries produced by either lactate or Na+/H+ exchange inhibitors were unrelated to alterations in high energy phosphate depletion during ischaemia or to repletion of these compounds after 30 min reperfusion either in spontaneously beating or electrically paced hearts.

The results suggest that stimulated Na+/H+ exchange activation at reflow contributes, at least partially, to lactate‐induced depression of postischaemic recovery.
Effect of an intracellular calcium chelator on the regulation of electrically evoked [3H]‐noradrenaline release from rat hippocampal slices
Authors: Fredholm Bertil B. , Hu Ping‐Sheng
Published: July 2012

The electrically (3 Hz, 5 min) evoked [3H]‐noradrenaline ([3H]‐NA) release from rat hippocampal slices was reduced by prior treatment of the slices with 1,2‐bis(2‐aminophenoxy)ethane‐N,N,N′N′‐tetraacetomethylester (BAPTA/AM) in a concentration‐(10 to 500 μm) dependent manner (40% at 30 μm). Reduction of medium calcium from 1.3 to 0.5 mm caused a larger (70%) decrease. BAPTA free acid (100 mm), a non‐permeable Ca2+‐chelator had no significant effect.

Basal [3H]‐noradrenaline release was reduced by BAPTA/AM in a concentration‐dependent manner (50% at 30 μm), but reduction of external Ca2+ from 1.3 to 0.5 mm did not alter basal release.

About 10% of total [3H]‐NA in the slices was released at 3 Hz stimulation in 1.3 mm Ca2+ buffer. Addition of the α2‐adrenoceptor antagonist, idazoxan (1 μm), increased electrically evoked [3H]‐NA release to 26% but stimulated release was not altered by the adenosine A1‐receptor antagonist, 8‐cyclopentyl theophylline (8‐CPT) (1 μm).

Evoked release was reduced by the α2‐receptor agonist, UK 14,304, in a concentration‐dependent manner in the presence of 8‐CPT (1 μm). The magnitude of this effect was not altered by the treatment of slices with 30 μm BAPTA/AM.

The adenosine A1 receptor agonist, N6‐cyclohexyl adenosine (CHA) (1 μm) inhibited electrically evoked [3H]‐NA release by about 40% in the presence of idazoxan (1 μm). The effect of CHA was not significantly altered by treatment of slices with BAPTA/AM.

The N‐type Ca2+ ‐channel antagonist, ω‐conotoxin (30 nm), inhibited electrically evoked [3H]‐NA release by 30–50% and this was not altered by treatment of the slices with BAPTA/AM.

The present results show that spontaneous [3H]‐NA release is affected by reduction of intracellular Ca2+, but not by reduction of extracellular Ca2+ or by the presynaptic agonists or ω‐conotoxin. By contrast, electrically evoked release was affected more strongly by alterations of extracellular Ca2+ than by buffering intracellular Ca2+. The reduction of electrically evoked [3H]‐NA release by agonists at the adenosine A1‐receptor and α2‐adrenoceptor is probably mediated through the control of Ca2+ entry via membrane ion channels or at a low affinity Ca2+‐site governing evoked release.
Selective IK blockade as an antiarrhythmic mechanism: effects of UK66,914 on ischaemia and reperfusion arrhythmias in rat and rabbit hearts
Authors: Rees Siân A. , Curtis Michael J.
Published: July 2012

UK66,914 is a specific and selective blocker of the delayed rectifying potassium current (IK). The effectiveness of IK block as a mechanism for prevention of ischaemia‐ and reperfusion‐induced arrhythmias was tested by use of UK66,914: its actions in rat, a species deficient in cardiac IK were compared with its actions in rabbit, a species possessing functional cardiac IK. Antiarrhythmic actions in rabbit but none in rat is the only outcome possible if selective IK blockade is responsible for the antiarrhythmic actions of the drug during ischaemia and/or reperfusion.

During 30 min regional ischaemia, 0.3 and 1 μm UK66,914 had no influence on the incidence of ventricular fibrillation (VF) in rat (n = 9/group), values being 78% in controls, 100% in 0.3 μm‐treated hearts and 78% in 1.0 μm‐treated hearts (NS). UK66,914 also had no effect on reperfusion‐induced VF incidence (100% in each group), nor on the latency to onset of ischaemia‐ or reperfusion‐induced arrhythmias. In contrast, in rabbit (n = 13/group), similar concentrations of drug reduced the incidence of reperfusion‐induced VF from 77% in controls, to 38% and 31% (P < 0.05) respectively. The incidence of ischaemia‐induced arrhythmias was too low in controls to permit detection of an antiarrhythmic effect in rabbit; however no drug‐induced proarrhythmia was seen.

QT interval was not affected by UK66,914 in rat (NS), values (mean ± s.e.mean) being 114 ± 8, 121 ± 8, and 113 ± 6 ms in control, 0.3 and 1 μm groups respectively, 10 min after the onset of ischaemia. However, in the rabbit the same concentrations of UK66,914 produced concentration‐dependent widening of QT interval, values being 172 ± 14, 194 ± 9 (NS) and 233 ± 18 ms (P < 0.05), with increasing concentrations of drug, 10 min after the onset of ischaemia.

UK66,914 had no substantial effects on haemodynamics (heart rate, coronary flow or recovery of flow during reperfusion), at either concentration, in either species. Occluded zones sizes were similar with increasing concentrations of drug in rat (42 ± 4, 40 ± 2 and 40 ± 2% of total ventricular weight) and rabbit (46 ± 2, 43 ± 2 and 46 ± 3%). Thus, the species‐dependent antiarrhythmic effects of UK66,914 were not attributable to differences in these variables.

In conclusion, the rat versus rabbit data indicate that for UK66,914 to inhibit ventricular arrhythmias associated with reperfusion, IK blockade is sufficient and necessary. The rat data indicate that no antiarrhythmic effects occur with the drug during ischaemia when there is no scope for IK blockade. Whilst other mechanisms may be unlikely to be involved, they cannot be excluded completely. Together the rat and rabbit studies indicate that the antiarrhythmic effects of UK66,914 during ischaemia and reperfusion are adequately explained by IK blockade.
Differential sensitivity of antinociceptive assays to the bradykinin antagonist Hoe 140
Authors: Heapy Christopher G. , Shaw John S. , Farmer Susannah C.
Published: July 2012

The antinociceptive activity of the bradykinin (BK) BK2 receptor antagonist d‐Arg‐[Hyp3,Thi5,d‐Tic7,Oic8]BK (Hoe 140) was determined in a range of mouse abdominal constriction assays.

Hoe 140 potently inhibited the response induced by i.p. injection of 10 μg BK/mouse, and 1 μg BK/mouse in mice pre‐sensitized by i.p injection of prostaglandin E2 (PGE2). The ED50 values in these assays were 1.9 and 3.7 μg kg−1 respectively. This confirms that Hoe 140 is a potent antagonist of BK in vivo.

Hoe 140 produced potent, but incomplete inhibition of the responses evoked by i.p injection of kaolin or 0.25% acetic acid. ED25 values in these assays were 2.7 and 16.1 μg kg−1, and the maximum inhibition produced was 60% and 70% respectively.

At doses up to 1 mg kg−1, Hoe 140 was completely ineffective against the abdominal constriction response induced by zymosan. In contrast, morphine, ibuprofen and indomethacin had similar potencies against zymosan, kaolin and acetic acid‐induced abdominal constriction.

Although zymosan, acetic acid and kaolin all produce qualitatively similar responses, it is appears that they achieve this by different mechanisms. The extent to which BK is involved as a mediator differs between the various types of abdominal constriction assay.
Inhibitors of nitric oxide synthetase prevent castor‐oil‐induced diarrhoea in the rat
Authors: Mascolo N. , Izzo A.A. , Barbato F. , Capasso F.
Published: July 2012

Castor oil (2 ml orally) produced copious diarrhoea in rats 3 h after its administration.

Pretreatment (intraperitoneal, i.p.) of rats with the NO synthesis inhibitors NG‐nitro‐l‐arginine methyl ester (l‐NAME, 1–25 mg kg−1) and NG‐monomethyl‐l‐arginine (l‐NMMA, 2.5–100 mg kg−1) inhibited or prevented castor‐oil‐induced diarrhoea. l‐Arginine (150–600 mg kg−1, i.p.) administered to rats pretreated with l‐NAME 10 mg kg−1, drastically reduced the antidiarrhoeal activity of l‐NAME in a dose‐related manner. d‐Arginine (900 mg kg−1) did not modify the protection by l‐NAME.

Pretreatment (i.p.) of rats with l‐NAME (2.5–25 mg kg−1) decreased the intestinal fluid accumulation and Na+ secretion induced by castor oil. l‐Arginine (600 mg kg−1) but not d‐arginine (900 mg kg−1) counteracted the inhibitory effect of l‐NAME (10 mg kg−1).

l‐NAME (10 and 25 mg kg−1) had no significant effect on the intestinal transit in normal rats or those given castor oil.

These results provide evidence that nitric oxide (NO) could play an important role in castor‐oil‐induced diarrhoea.
Renal imidazoline preferring sites and solute excretion in the rat
Authors: Allan D.R. , Penner S.B. , Smyth D.D.
Published: July 2012

Moxonidine has been found to have an approximately 600 fold greater affinity for I1 imidazoline preferring sites as compared to α2‐adrenoceptors in the rat kidney. The effects of an intrarenal infusion of moxonidine in an anaesthetized rat preparation were investigated and contrasted with the effects previously reported for α2‐adrenoceptor stimulation.

An intrarenal infusion of moxonidine (1, 3 and 10 nmol kg−1 min−1) produced an increase in urine flow rate and sodium excretion. Moxonidine increased urine volume through an increase in osmolar clearance rather than an increase in free water clearance as previously reported for α2‐adrenoceptor stimulation.

The effects of moxonidine also appeared to be unique from the effects of α2‐adrenoceptor stimulation. An imidazoline preferring site specific blocking dose of idazoxan (0.3 mg kg−1), but not an α2‐adrenoceptor specific blocking dose of rauwolscine (0.3 mg kg−1) attenuated the renal effects of moxonidine (3 nmol kg−1 min−1). Moreover, unlike α2‐adrenoceptor agonists, the effects of moxonidine were not altered by prior treatment with a V2 vasopressin receptor antagonist.

These results indicate differences between stimulation of α2‐adrenoceptors and I1 imidazoline preferring sites in the rat kidney and suggest a direct physiological function of renal imidazoline preferring sites.
Redirection of arachidonic acid metabolism by ICI D1542: effects on thrombus formation in the coronary artery of the anaesthetized dog
Authors: McAuliffe S.J.G. , Moors J.A. , Snow H.M. , Wayne M. , Jessup R.
Published: July 2012

The effects of simultaneous redirection of arachidonic acid metabolism, by inhibition of thromboxane A2 (TXA2) synthase and blockade of the platelet thromboxane A2 receptor (TP‐receptor), was examined on the rate of thrombus formation in a stenosed coronary artery with damaged endothelium in an anaesthetized dog.

Redirection of arachidonic acid metabolism was achieved by intravenous doses of ICI D1542, a selective and potent inhibitor of TXA2 synthase and the TP‐receptor.

Redirection of arachidonic acid metabolism was demonstrated in whole blood, stimulated ex vivo by collagen. The ED50 for inhibition of TXB2 production was 7.1 μg kg−1, i.v.; there were corresponding increases in the production of the eicosanoids prostaglandin D2 (PGD2), PGE2 and PGF2α.

Thrombus formation was inhibited by D1542 (ED50 0.55 μg kg−1, i.v.), but could be restarted by an intravenous infusion of adrenaline (0.2–38 μg kg−1 min−1, i.v.). In the presence of the maximum effective dose of D1542 (1 mg kg−1, i.v.) a 190 fold increase in the infusion rate of adrenaline was required to restore thrombus formation.

In the presence of D1542, removal of endoperoxide metabolites by inhibition of cyclo‐oxygenase with aspirin (5 mg kg−1, i.v.) caused thrombus formation to restart, indicating the ability of the endoperoxide metabolites to inhibit thrombus formation in vivo.

These results indicate that, in the stenosed and damaged coronary artery of the dog, redirection of arachidonic acid metabolism by D1542 is more effective at preventing thrombus formation than inhibition of cyclo‐oxygenase by aspirin.
The spinal antinociceptive activity of the α2‐adrenoceptor agonist, xylazine in sheep
Authors: Kyles A.E. , Waterman A.E. , Livingston A.
Published: July 2012

The intrathecal administration of xylazine (100 μg), via a chronic indwelling, cervical intrathecal catheter, produced a marked elevation of the mechanical nociceptive thresholds in the sheep. This antinociceptive effect was abolished by the prior intrathecal administration of the α2‐adrenoceptor antagonist, idazoxan.

The intrathecal administration of the selective α2‐antagonists, idazoxan (100 μg) and RX811059 (33 μg), significantly attenuated the antinociceptive activity of intravenous xylazine, with a 60–65% reduction in the area under the antinociceptive curve. The intrathecal administration of the antagonists alone had no significant effect on nociceptive thresholds.

Examination of the distribution of tritiated idazoxan (25 μCi in 100 μl) indicated that the site of action of the drug was limited to the cervical spinal cord after intrathecal administration.

These studies demonstrate that a significant proportion of the antinociceptive effect of systemically administered xylazine is mediated by spinal α2‐adrenoceptors.
Quinapril treatment and arterial smooth muscle responses in spontaneously hypertensive rats
Authors: Arvola Pertti , Ruskoaho Heikki , Wuorela Heikki , Pekki Anu , Vapaatalo Heikki , Pörsti Ilkka
Published: July 2012

The effects of long‐term angiotensin‐converting enzyme inhibition with quinapril on arterial function were studied in spontaneously hypertensive rats, Wistar‐Kyoto rats serving as normotensive controls.

Adult hypertensive animals were treated with quinapril (10 mg kg−1 day−1) for 15 weeks, which reduced their blood pressure and the concentrations of atrial natriuretic peptide in plasma and ventricular tissue to a level comparable with that in normotensive rats.

Responses of mesenteric arterial rings in vitro were examined at the end of the study. Compared with normotensive and untreated hypertensive rats, responses to noradrenaline were attenuated in hypertensive animals on quinapril, both force of contraction and sensitivity being reduced. Quinapril also attenuated maximal contractions but not sensitivity to potassium chloride. Nifedipine less effectively inhibited vascular contractions in normotensive and quinapril‐treated than in untreated hypertensive rats.

Arterial relaxation responses by endothelium‐dependent (acetylcholine) and endothelium‐independent (sodium nitrite, isoprenaline) mechanisms were similar in normotensive and quinapril‐treated rats and more pronounced than in untreated hypertensive rats.

Cell membrane permeability to ions was evaluated by means of potassium‐free solution‐induced contractions of endothelium‐denuded denervated arterial rings. These responses were comparable in normotensive and quinapril‐treated rats and less marked than in untreated hypertensive rats.

Intracellular free calcium concentrations in platelets and lymphocytes, measured by the fluorescent indicator quin‐2, were similar in normotensive and quinapril‐treated rats and lower than in untreated hypertensive rats.

In conclusion, quinapril treatment improved relaxation responses and attenuated contractions in arterial smooth muscle of hypertensive rats. These changes may be explained by diminished cytosolic free calcium concentration, reduced cell membrane permeability, and alterations in dihydropyridine‐sensitive calcium channels following long‐term angiotensin‐converting enzyme inhibition.
Effects of dicentrine on haemodynamic, plasma lipid, lipoprotein level and vascular reactivity in hyperlipidaemic rats
Authors: Yu Sheu‐Meei , Kang Ya‐Fei , Chen Chien‐Chih , Teng Che‐Ming
Published: July 2012

The effects of dicentrine on haemodynamic, plasma lipid, lipoprotein level and vascular reactivity were investigated in Wistar‐Kyoto (WKY) and spontaneously hypertensive (SH) rats, fed a high fat‐high cholesterol diet.

In high fat‐high cholesterol (HF‐HC) diet fed WKY and SH rats, oral administration of dicentrine (5 and 10 mg kg−1, twice a day) for 4 weeks caused significant reductions in total plasma cholesterol (CE) by reducing the low density lipoprotein (LDL) fraction, and reductions in total plasma triglyceride (TG) by reducing the very low density lipoprotein (VLDL) fraction.

Dicentrine therapy was associated with increased high density lipoprotein (HDL)‐cholesterol levels; thus the ratio of total plasma cholesterol to HDL‐cholesterol was improved.

In HF‐HC diet fed conscious WKY and SH rats, oral administration of dicentrine (5 and 10 mg kg−1, twice a day) also evoked dose‐related decreases in mean arterial pressure (MAP) which were of greater magnitude in SH rats. Neither dose of dicentrine caused a significant change in heart rate (HR).

The aortic arches from SH rats fed the HF‐HC diet for 8 weeks were significantly more affected by the atherosclerotic lesions than the abdominal aortae and renal arteries of WKY and SH rats. Oral administration of dicentrine (5 and 10 mg kg−1) for 4 weeks did not diminish the atherosclerotic lesion areas in WKY and SH rats.

In aortae of the hyperlipidaemic rats, significantly attenuated EC50 values and augmented maximal responses for phenylephrine‐induced contraction were obtained. Endothelium‐dependent relaxation to acetylcholine was abolished, while endothelium‐independent relaxation to nitroprusside was well preserved. Dicentrine therapy caused significantly augmented EC50 values and attenuated maximal responses for phenylephrine‐induced contraction in hyperlipidaemic rats. However, dicentrine neither prevented the impaired relaxation to acetylcholine, nor affected the relaxation to nitroprusside during atherosclerosis progression.

It is concluded that dicentrine decreases MAP, plasma CE, LDL‐CE, plasma TG, VLDL‐TG, vascular hyperreactivity to phenylephrine and increases HDL‐CE levels. Dicentrine may thus hold potential for the reduction of two of the major risk factors, hypertension and hyperlipidaemia, for cardiovascular disease.
Cellular mechanism of U78517F in the protection of porcine coronary artery endothelial cells from oxygen radical‐induced damage
Authors: Maeda Kaori , Kimura Masaaki , Hayashi Shigehiro
Published: July 2012

The aim of this study was to clarify the role of lipid peroxidation in cellular injury as assessed by lactate dehydrogenase (LDH) release from cultured coronary artery endothelial cells of the pig. Cells exposed to H2O2 at concentrations of 0.1 to 20 mm or to a xanthine and xanthine oxidase (X/XO) reaction mixture released LDH into the medium. Significant release from X/XO‐treated cells took place with a delay of 2 h.

Superoxide dismutase (SOD), catalase or dimethylthiourea attenuated the release of LDH from X/XO‐treated cells. Similarly the putative inhibitor of lipid peroxidation, U78517F attenuated the release of LDH by X/XO with an IC50 of 0.08 μm.

H2O2 was continuously produced by the addition of X/XO to the medium alone. However, in the presence of endothelial cells, H2O2 was eliminated at 1 h. U78517F had no effect on either process.

The oxygen radical‐induced release of LDH was associated with malondialdehyde (MDA) formation. U78517F inhibited the formation of MDA with an IC50 of 0.27 μm.

Reduction of the Ca2+ concentration in the incubation medium from 1.6 mm to 0.016 mm markedly attenuated the release of LDH from endothelial cells. Nifedipine (1 μm) did not attenuate the LDH release from the cells.

It is likely that porcine coronary artery endothelial cells can be thus injured by oxygen radicals presumably through hydroxyl radicals formed and consequent lipid peroxidation, and that the extracellular Ca2+ concentration plays an important role in the genesis of such endothelial cell damage.
Antiepileptic drug pharmacokinetics and neuropharmacokinetics in individual rats by repetitive withdrawal of blood and cerebrospinal fluid: milacemide
Authors: Semba Jun'ichi , Curzon Gerald Curzon , Patsalos Philip N.
Published: July 2012

The kinetics and metabolism of milacemide have been studied in an animal model which allows the simultaneous investigation of the temporal inter‐relationships of drugs and metabolites in blood (pharmacokinetics) and cerebrospinal fluid (CSF, neuropharmacokinetics) in individual freely moving rats.

Milacemide dose‐dependently increased CSF glycine and glycinamide (intermediary metabolite) concentrations. This confirms that milacemide is a CNS glycine prodrug.

Pretreatment with l‐deprenyl (2 mg kg−1), a specific inhibitor of monoamine oxidase type B (MAO‐B), almost completely prevented the formation of glycinamide and increased milacemide accumulation in CSF. Tmax and t1/2 were significantly increased and Cmax and AUC values were decreased for glycinamide compared to controls. Pretreatment with clorgyline (5 mg kg−1), a specific inhibitor of MAO‐type A, only moderately decreased glycinamide Cmax and AUC values.

After milacemide administration (100, 200 and 400 mg kg−1, i.p.) serum and CSF milacemide concentrations rose linearly and dose‐dependently. Serum glycinamide concentrations exhibited small dose‐dependent rises but these were not linearly related. In contrast, CSF glycinamide concentrations rose linearly and dose‐dependently with Cmax values 2.5, 3.2 and 4.1 times greater than the corresponding values for serum glycinamide after giving 100, 200 and 400 mg kg−1 respectively of milacemide.

Serum glycine concentrations were unaffected but CSF concentrations increased dose‐dependently and these were significant at the higher milacemide doses (200 and 400 mg kg−1). Animals given 400 mg kg−1 milacemide had glycine values which were still significantly elevated 7 h later.

In conclusion, serum milacemide rapidly enters and equilibrates with the CNS compartment where it is metabolised primarily by MAO‐B to glycinamide and finally to glycine. Metabolism in the peripheral compartment is negligible.
Endothelin‐3‐induced relaxation of rat thoracic aorta: a role for nitric oxide formation
Authors: Moritoki Hideki , Miyano Hiroaki , Takeuchi Shougo , Yamaguchi Mari , Hisayama Tetsuhiro , Kondoh Wataru
Published: July 2012

Endothelin‐3 (ET‐3) at concentrations below those which caused contraction (30 nm) elicited endothelium‐dependent relaxation followed by rebound contraction in rat isolated thoracic aorta.

Endothelin‐1 also relaxed the rat aorta with a similar potency.

The nitric oxide (NO) synthase inhibitor, NG‐nitro l‐arginine, the radical scavenger, haemoglobin and the soluble guanylate cyclase inhibitor, methylene blue, each inhibited the ET‐3‐induced relaxation.

The calmodulin inhibitor, calmidazolium, considerably attenuated the relaxation caused by ET‐3 without affecting that to nitroprusside.

Concentrations of ET‐3 that were necessary to induce the relaxation also caused concentration‐dependent elevation of guanosine 3′:5′‐cyclic monophosphate (cyclic GMP) levels.

NG‐nitro l‐arginine, haemoglobin, methylene blue, calmidazolium and removal of the endothelium completely abolished ET‐3‐stimulated cyclic GMP production.

These results suggest that ET‐3 triggers NO formation possibly via ETB receptors on the endothelium to activate soluble guanylate cyclase, which in turn stimulates cyclic GMP production and smooth muscle relaxation. The enzyme contributing to the NO formation may be of the calcium/calmodulin‐dependent, constitutive type.
Effects of phospholipase A2 inhibitors on coupling of α2‐adrenoceptors to inwardly rectifying potassium currents in guinea‐pig submucosal neurones
Authors: Evans R.J. , Surprenant A.
Published: July 2012

Noradrenaline hyperpolarizes guinea‐pig submucosal neurones by opening inwardly rectifying potassium channels. Intracellular recordings were made from submucosal neurones and the possible involvement of the phospholipase A2 pathway in this response was examined.

The non‐specific phospholipase A2 inhibitors, quinacrine (10 μm) and 4‐bromophenacyl bromide (4‐BPB, 10 μm) inhibited nerve‐evoked inhibitory synaptic potentials (i.p.s.ps) and hyperpolarizations to somatostatin and UK 14304. Quinacrine and 4‐BPB also blocked the inward rectification present in current‐voltage curves in the absence of somatostatin or UK 14304.

The more selective phospholipase A2 inhibitor, cyclosporin A (10 μm) and the lipoxygenase and cyclo‐oxygenase inhibitor, eicosatetraynoic acid (ETYA, 20 μm) and nordihydroguairetic acid (NDGA, 20 μm) did not alter i.p.s.ps or hyperpolarizations to UK 14304.

Exogenously applied arachidonic acid (1–300 μm) did not mimic the i.p.s.p. or the hyperpolarization to UK 14304.

We conclude that arachidonic acid or its eicosanoid metabolites produced by phospholipase A2 stimulation are unlikely to be involved in the receptor G‐protein coupled activation of potassium currents in submucosal neurones. The inhibition of the noradrenaline‐induced hyperpolarization by quinacrine and 4‐BPB is most likely due primarily to blockade of the basal inwardly rectifying potassium conductance present in these neurones.
Monocrotaline pyrrole‐induced changes in angiotensin‐converting enzyme activity of cultured pulmonary artery endothelial cells
Authors: Hoorn Cindy M. , Roth Robert A.
Published: July 2012

Changes in the structural and functional integrity of endothelium have been recognized as relatively early features of delayed and progressive pulmonary vascular injury caused by the pyrrolizidine alkaloid, monocrotaline (MCT). Although a number of investigators have evaluated angiotensin‐converting enzyme (ACE) activity in the lungs of rats treated with MCT, the exact nature of changes in activity of this enzyme and the role they may play in MCT pneumotoxicity remain controversial.

We examined the direct effects of monocrotaline pyrrole (MCTP), a toxic metabolite of MCT, on cultured endothelial cell ACE activity. Post‐confluent monolayers of porcine or bovine pulmonary artery endothelial cells (PECs or BECs, respectively) were treated with a single administration of MCTP at time 0; then they were examined for their ability to degrade the synthetic peptide, [3H]‐benzoyl‐Phe‐Ala‐Pro.

In PECs, which are relatively insensitive to the direct cytolytic effects of MCTP, monolayer ACE activity was unchanged initially but gradually decreased within 4 days after treatment with a high concentration of MCTP (150 μm). This decrease was transient, and PEC monolayer ACE activity returned to the control value by 10 days post treatment.

BEC monolayer ACE activity was also unchanged initially but rapidly declined within 4 days after MCTP treatment and remained depressed throughout the post treatment period. BECs were quite sensitive to the cytolytic effects of MCTP and the decline in ACE activity occurred coincident with the decrease in monolayer cellularity and appearance of marked cytotoxicity.

We conclude that high concentrations of MCTP decrease endothelial ACE activity. The decline in ACE activity is delayed and the magnitude and duration of the decrease corresponds to the degree of MCTP‐induced cytotoxicity. This suggests that altered endothelial ACE activity is unlikely to be a direct effect of MCTP on the enzyme but may reflect the delayed cell injury which results from exposure to this compound.
Competitive inhibition by procaine of carbachol‐induced stimulus‐secretion coupling in rat pancreatic acini
Authors: Ikei Nobuhiro , Busik Julia , Habara Yoshiaki , Kanno Tomio
Published: July 2012

Procaine (0.03–10 mm) inhibited carbacbol (CCh)‐induced amylase release from rat isolated pancreatic acini in a competitive manner. Kinetic analysis of the relation between CCh concentrations and the amount of amylase released in the presence of various procaine concentrations indicated that procaine caused competitive inhibition with the affinity constant (pA2) value of 5.00 ± 0.08.

Receptor binding assay confirmed that procaine (0.01–10 mm) competitively inhibited [N‐methyl‐3H]‐scopolamine chloride ([3H]‐NMS) binding to its receptor with binding affinity (pKi) of 4.63 ± 0.10.

Procaine transformed CCh‐evoked [Ca2+]i dynamics: the initial rise in [Ca2+]i followed by a gradual decay during continuous stimulation with 3 μm CCh was transformed by 0.3 mm procaine to the oscillatory [Ca2+]i dynamics, which resembled the response to 0.3 μm CCh in the absence of procaine. The initial phase of [Ca2+]i oscillation corresponded to the initial phase of CCh‐induced amylase release in isolated perfused acini.

Procaine (0.3–3 mm) did not inhibit the secretory response to cholecystokinin octapeptide (CCK‐8) in isolated incubated acini. A higher concentration of procaine (10 mm) caused weak but significant inhibition of the response to only limited concentrations of CCK‐8, 30 and 100 pm. Procaine lower than 10 mm was ineffective on [125I]‐BH‐CCK‐8 binding, although procaine (10 mm) caused weak but significant inhibition of the binding.
Salmeterol, a long‐acting β2‐adrenoceptor agonist mediating cyclic AMP accumulation in a neuronal cell line
Authors: McCrea Karen E. , Hill Stephen J.
Published: July 2012

The accumulation of cyclic AMP stimulated by salmeterol, a long‐acting β2‐adrenoceptor agonist and by isoprenaline, a non‐selective β‐adrenoceptor agonist have been compared in the B50 neuroblastoma cell line.

Salmeterol produced a concentration‐dependent increase in the accumulation of total [3H]‐cyclic AMP in B50 cells yielding an EC50 value of 37 nm which was lower than that obtained with isoprenaline (294 nm). The maximum response to salmeterol was only 46% of that obtained with isoprenaline.

The β2‐adrenoceptor antagonist, ICI 118551, inhibited the responses to both salmeterol (apparent KD 2.2 nm) and isoprenaline (apparent KD 1.6 nm). However, the β1‐adrenoceptor antagonist, atenolol, produced no significant effect at concentrations up to 100 μm.

Salmeterol (1 μm) changed the concentration‐response curve of isoprenaline in the manner of a partial agonist interacting with a full agonist. The KD of salmeterol obtained from the interaction was 55.6 nm.

Whereas salmeterol has a slow onset of action in airway smooth muscle compared to other β2‐adrenoceptor agonists, in B50 monolayers both salmeterol and isoprenaline produced a rapid increase in cyclic AMP accumulation (t1/2 1.1 min and 0.4 min respectively).

Despite the existence of cyclic AMP efflux mechanisms that exist in this cell line it was possible to investigate the duration of agonist action by measuring intracellular levels of the second messenger. Replacement of drug‐containing medium with fresh buffer led to a rapid reduction in intracellular levels of cyclic AMP in isoprenaline‐stimulated cells whereas cyclic AMP accumulation was sustained for much longer periods in salmeterol‐stimulated cells. However, the persistent action of salmeterol could be reversed by the addition of a β2‐selective antagonist.

These results confirm that salmeterol has a high affinity, but low efficacy (relative to isoprenaline) for β2‐adrenoceptors coupled to cyclic AMP accumulation and that the drug persists at its site of action for long periods in the B50 neuronal cell line.
Bradykinin‐induced airflow obstruction and airway plasma exudation: effects of drugs that inhibit acetylcholine, thromboxane A2 or leukotrienes
Authors: Kawikova Ivana , Arakawa Hirokazu , Löfdahl Claes‐Göran , Skoogh Bengt‐Eric , Lötvall Jan
Published: July 2012

The mechanisms behind bradykinin‐induced effects in the airways are considered to be largely indirect. The role of cholinergic nerves and eicosanoids, and their relationship in these mechanisms were investigated in guinea‐pigs.

The role of cholinergic nerves was studied in animals given atropine (1 mg kg−1, i.v.), hexamethonium (2 mg kg−1, i.v.), or vagotomized. To study the role of eicosanoids, animals were pretreated with a thromboxane A2 (TxA2) receptor antagonist (ICI 192,605; 10−6 mol kg−1, i.v.) or with a leukotriene (LT) receptor C4/D4/E4 antagonist (ICI 198,615; 10−6 mol kg−1, i.v.).

After pretreatment with a drug, bradykinin (150 nmol) was instilled into the tracheal lumen. We measured both airway insufflation pressure (Pi), to assess airway narrowing, and the content of Evans blue dye in airway tissue, to assess plasma exudation.

Bradykinin instillation into the trachea caused an increase in Pi and extravasation of Evans blue dye. The increase in Pi was significantly attenuated by atropine or the TxA2 receptor antagonist, but not by hexamethonium, vagotomy or the LT receptor antagonist.

The bradykinin‐induced exudation of Evans blue dye was significantly attenuated in the intrapulmonary airways by the TxA2 receptor antagonist, but not by atropine, hexamethonium, cervical vagotomy or the LT receptor antagonist.

A thromboxane‐mimetic, U‐46619 (20 nmol kg−1, i.v. or 10 nmol intratracheally), caused both an increase in Pi and extravasation of Evans blue dye at all airway levels. Atropine pretreatment slightly attenuated the peak Pi after the intratracheal administration of U‐46619, but not after i.v. administration.

We conclude that peripheral cholinergic nerves are involved in bradykinin‐induced airflow obstruction but not plasma exudation, and that TxA2 is involved in both airflow obstruction and airway plasma exudation induced by bradykinin given via the airway route. TxA2‐induced airflow obstruction is mediated only to a minor degree, via the release of acetylcholine in the airways.
An electrophysiological investigation of the properties of 5‐HT3 receptors of rabbit nodose ganglion neurones in culture
Authors: Peters John A. , Malone Hilary M. , Lambert Jeremy J.
Published: July 2012

The biophysical and pharmacological properties of 5‐hydroxytryptamine (5‐HT)‐evoked currents in rabbit nodose ganglion neurones in culture have been determined by use of the whole‐cell and outside‐out membrane patch recording modes of the patch‐clamp technique.

In 49% of cells investigated the bath application of 10−5 m 5‐HT at negative holding potentials elicited an inward current. The whole‐cell response to 5‐HT reversed in sign (E5‐HT) at approximately − 2 mV and exhibited inward rectification.

The influence of various ion substitutions upon E5‐HT established that the 5‐HT‐evoked current is mainly mediated by a mixed Na+, K+ cation conductance with little or no contribution from Cl− ions. The omission of Ca2+ and Mg2+ from the extracellular solution enhanced the amplitude of the 5‐HT‐induced current.

On isolated outside‐out membrane patches, the bath application of 10−6 m 5‐HT induced single channel currents with a chord conductance of approximately 17 pS at − 70 mV and an average slope conductance of 19 pS over the range − 100 to − 40 mV. The 5‐HT‐induced single channels exhibited modest inward rectification and were reduced in frequency, but not amplitude, by the 5‐HT3 receptor antagonist metoclopromide (10−6 m).

The bath application of 5‐HT (3 × 10−7 − 3 × 10−5 m) to whole cells voltage clamped at − 60 mV produced dose‐dependent inward currents which were mimicked by 2‐methyl‐5‐HT and 1‐phenylbiguanide with equipotent molar ratios, relative to 5‐HT, of 2.5 and 32 respectively.

Whole‐cell inward currents produced by the local pressure application of 5‐HT (10−5 m) were unaffected by 10−6 m methysergide, 10−6 m ketanserin or 10−6 m citalopram, but were concentration‐dependently antagonized by the selective 5‐HT3 receptor antagonists tropisetron (IC50 = 4.6 × 10−11 m) ondansetron (IC50 = 5.7 × 10−11 m), and bemesetron (IC50 = 3.3 × 10−10 m). The response to 5‐HT was also blocked by the non‐selective antagonists metoclopramide (IC50 = 1.2 × 10−8 m), cocaine (IC50 = 8.3 × 10−8 m) and (+)‐tubocurarine (IC50 = 1.6 × 10−7 m).