bluuebunny/crossref_metadata_2025 huggingface dataset:
Involvement of protein kinase C in the delayed cytoprotection following sublethal ischaemia in rabbit myocardium
Authors: Baxter G.F. , Goma F.M. , Yellon D.M.
Published: July 2012

Rabbit hearts were preconditioned with four 5 min coronary artery occlusions 24 h before 30 min coronary occlusion with 120 min reperfusion. Preconditioning significantly reduced the percentage of myocardium infarcting within the risk zone from 49.1 ±4.3% to 31.8±3.5% (P<0.05). When the protein kinase C (PKC) inhibitor, chelerythrine, was administered just before preconditioning, the delayed protection against infarction 24 h later was abolished. We conclude that the delayed cytoprotective response associated with ischaemic preconditioning of myocardium is likely to involve the early activation of one or more PKC subtypes.
P2‐purinoceptor‐mediated inhibition of noradrenaline release in rat atria
Authors: von Kügelgen Ivar , Stoffel Daniel , Starke Klaus
Published: July 2012

We looked for P2‐purinoceptors modulating noradrenaline release in rat heart atria. Segments of the atria were preincubated with [3H]‐noradrenaline and then superfused with medium containing desipramine (1 μm) and yohimbine (1 μm) and stimulated electrically, by 30 pulses/I Hz unless stated otherwise.

The adenosine Al‐receptor agonist, N6‐cyclopentyl‐adenosine (CPA; EC50 9.7 nM) and the nucleotides, ATP (EC50 6.6 μm) and adenosine‐5′‐O‐(3‐thiotriphosphate) (ATPγS; EC50 4.8 μm), decreased the evoked overflow of tritium. The adenosine A2a‐agonist, 2‐p‐(2‐carbonylethyl)‐phenethylamino‐5′‐N‐ethylcarboxamido‐adenosine (CGS‐21680; 0.03‐0.3 μm) and the P2X‐purinoceptor agonist β,γ,‐methylene‐l‐ATP (30 μm) caused no change.

The concentration‐response curve of CPA was shifted to the right by the adenosine Al‐receptor antagonist, 8‐cyclopentyl‐1,3‐dipropylxanthine (DPCPX; 3 nM; apparent pKB value 9.7) but hardly affected by the P2‐purinoceptor antagonist, cibacron blue 3GA (30 μm). In contrast, the concentration‐response curves of ATP and ATPγS were shifted to the right by DPCPX (3 nM; apparent pKB values 9.3 and 9.4, respectively) as well as by cibacron blue 3GA (30 μm; apparent pKB values 5.0 and 5.1, respectively). Combined administration of DPCPX and cibacron blue 3GA caused a much greater shift of the concentration‐response curve of ATP than either antagonist alone. The concentration‐response curve of ATP was not changed by indomethacin, atropine or the 5′‐nucleotidase blocker α,β‐methylene‐ADP.

Cibacron blue 3GA (30 μm) increased the evoked overflow of tritium by about 70%. The increase was smaller when the slices were stimulated by 9 pulses/100 Hz instead of 30 pulses/I Hz.

The results indicate that the postganglionic sympathetic axons in rat atria possess P2‐purinoceptors in addition to the known adenosine Al‐receptor. Both mediate inhibition of noradrenaline release. Some adenine nucleotides such as ATP and ATPγS act at both receptors. The presynaptic P2‐purinoceptor seems to be activated by an endogenous ligand, presumably ATP, under the condition of these experiments. This is the first evidence for presynaptic P2‐purinoceptors at cardiac postganglionic sympathetic axons.
Subcellular localization and molecular pharmacology of distinct populations of [3H]‐AMPA binding sites in rat hippocampus
Authors: Henley Jeremy M.
Published: July 2012

The subcellular distributions of [3H]‐α‐amino‐3‐hydroxy‐5‐methylisoxazolepropionate ([3H]‐AMPA) and [3H]‐kainate binding sites in rat hippocampus were investigated by cell fractionation techniques.

Two major populations of [3H]‐AMPA sites were detected with the majority of binding located intracellularly in the microsomal (P3) fraction. Most of the remaining sites were in the synaptosomal membrane fraction but some were also present in the nuclear fraction. In contrast, essentially all of the [3H]‐kainate binding sites were in the synaptosomal membrane fraction.

Saturation binding analysis yielded KD and Bmax values for [3H]‐AMPA of 147 nM and 2.6 pmol mg−1 protein respectively for the synaptosomal membrane‐associated sites and 129 nM and 5.3 pmol mg−1 protein respectively for the microsomal sites.

Both main populations of [3H]‐AMPA sites displayed the same rank order of inhibition by competitive ligands, the apparent Mr values of GluR1 subunits were equivalent, suggesting the same degree of post‐translational modification and the hydrodynamic properties of the receptor complexes were identical.

These data are consistent with the hypothesis that the movement of AMPA receptors between cellular compartments in the postsynaptic neurone could constitute one mechanism underlying long‐term potentiation in the hippocampus.
Modulatory effect of bradykinin on the release of noradrenaline from rat isolated atria
Authors: Chulak Chantal , Couture Réjean , Foucart Sylvain
Published: July 2012

We investigated the modulation by bradykinin (BK) of electrically induced noradrenaline release in rat isolated atria preincubated with [3H]‐noradrenaline.

BK (1‐100nM) enhanced significantly the stimulation‐induced outflow of radioactivity in a concentration‐dependent manner with a calculated EC50 of 0.58 nM.

Des‐Arg9‐BK (0.1‐100nM), a selective B1 receptor agonist, did not modify the stimulation‐induced outflow of radioactivity. Hoe 140 (10 nM), a selective B2 receptor antagonist, but not [Leu8]‐des‐Arg9‐BK (100 nM), a selective B1 receptor antagonist, blocked the facilitatory effect of BK.

The effect of BK was not affected by diclofenac (1 μm), a cyclo‐oxygenase inhibitor. Bisindolyl‐maleimide (1 μm), a protein kinase C inhibitor, significantly reduced the facilitatory effect of BK (10 nM), angiotensin II (0.3 μm) and phorbol dibutyrate (0.1 and 1 μm) but not of fenoterol (1 μm).

The results suggest that BK enhances noradrenaline release via a prejunctional B2 kinin receptor in the rat atrium. The effect appears to involve protein kinase C as a second messenger.
Renal vasodilatation by dopexamine and fenoldopam due to α1‐adrenoceptor blockade
Authors: Martin S.W. , Broadley K.J.
Published: July 2012

The renal vascular responses of the rat isolated perfused kidney to the dopamine D,‐receptor agonists, dopexamine and fenoldopam, were examined.

Both kidneys were perfused in situ at constant flow rate (11ml min−1) with Krebs‐bicarbonate solution at 37°C. The perfusion pressure was monitored and to enable vasodilator responses to be measured, the resting perfusion pressure was raised by infusing noradrenaline (6 × 10−9 m).

Dose‐related vasodilator responses to bolus doses of dopexamine and fenoldopam were obtained. However, these were not antagonized by the D1‐receptor antagonist, SCH 23390, indicating that D1‐receptors were not involved.

Bolus doses of the α1‐adrenoceptor antagonist, prazosin, caused similar dose‐related vasodilator responses indicating the possibility that α1‐adrenoceptor blocking properties of dopexamine and fenoldopam were responsible for the vasodilatation.

α‐Adrenoceptor blockade by dopexamine and fenoldopam was confirmed by the parallel displacement of dose‐response curves for the vasopressor responses to noradrenaline. pA2 values were determined by Schild analysis for dopexamine, fenoldopam and prazosin antagonism of noradrenaline in the presence of neuronal (cocaine, 10−5 m) and extraneuronal uptake blockade (metanephrine, 10−5 m). The values were 6.23, 6.02 and 8.91, respectively. Schild plot slopes of unity were obtained for dopexamine and fenoldopam indicating competitive antagonism. A slope of greater than unity for prazosin may be explained by the lack of equilibrium conditions associated with bolus doses of noradrenaline, the responses of which are affected more by the high affinity antagonist, prazosin, than the two lower affinity antagonists.

This study has demonstrated that renal vasodilator responses to the D1‐receptor agonists, dopexamine and fenoldopam, are due to a brief antagonism of the α‐adrenoceptor‐mediated vasoconstriction induced by noradrenaline. This presumably masks any direct D1‐receptor‐mediated vasodilatation.
Stimulation of two vascular smooth muscle‐derived cell lines by angiotensin II: differential second messenger responses leading to mitogenesis
Authors: Morton Clare , Baines Richard , Masood Imran , Ng Leong , Boarder Michael R.
Published: July 2012

We show here that angiotensin II (All) and endothelin‐l (ET‐1) stimulate [3H]‐thymidine incorporation in a smooth muscle cell line derived from aortae of spontaneously hypertensive rats (SHR), but not in cells derived from normotensive controls (WKY). We have used the differential response of the two cell lines to investigate the relationship between second messenger systems and the mitogenic response.

All produced an increase in accumulation of inositol 1,4,5‐triphosphate which was greater in the SHR‐derived cell line than in the WKY cells.

All gave an increase in cytosolic Ca2+ in each of the cell lines, with both a larger peak (15–30 s) and plateau response (2min) in the SHR cells. ET‐1 gave an enhanced response in the SHR‐derived cells with respect to the peak but not the plateau of cytosolic Ca2+.

Phospholipase D activity was studied by monitoring the formation of [3P]‐phosphatidylbutanol in 32Pi prelabelled cells. All stimulation gave a larger phospholipase D response in SHR‐derived cells, while ET‐1 gave a larger response in WKY‐derived cells.

Stimulation of SHR‐derived cells with 100 nM All for 1 h, followed by 19 h in the absence of agonist, stimulated [3H]‐thymidine incorporation over the next 4 h. When the 1 h stimulation with All was in the presence of increasing concentrations of butanol, which diverts the product of the phospholipase D pathway, there was a loss of stimulated [3H]‐thymidine incorporation which was significant at 10 mM butanol and at 30‐50mM reached a maximum loss of 40%.

Contrasting with this there was no apparent loss of ET‐1‐stimulated thymidine incorporation when butanol was present at concentrations up to 40 mM. 7 These results suggest that phospholipase D is one of several pathways in the mitogenic response of SHR‐derived vascular smooth muscle cells to All.
Effect of illuminated nifedipine, a potent antioxidant, on intestinal and vascular smooth muscles
Authors: Bauer V. , Rekalov V.V. , Juranek I. , Gergel D. , Bohov P.
Published: July 2012

The effects of nifedipine (Nif) and its illuminated nitroso product nitrosopine (NTP) were investigated on lipid peroxidation, KC1 elevated smooth muscle tension, and ionic currents of single smooth muscle cells.

Illumination of Nif at 400–700 nm within 24–48 h changed it completely to a potent antioxidant, NTP.

Nif relaxed the KCl‐induced contractions of guinea‐pig taenia caeci and rat aorta and reduced the amplitude of the evoked inward Ca2+ current of taenia caeci cells in a concentration‐dependent manner. NTP (up to 100 μm) was ineffective in this respect. Pretreatment by NTP (10 μm) did not affect the actions of Nif.

The evidence suggests that NTP, generated by day‐light illumination from Nif, exerts antioxidant activity but is devoid of voltage‐dependent Ca2+ channel (VDC) blocking property and does not interfere with the action of Nif on the smooth muscle cell membrane VDC.
Substance P‐induced relaxation and hyperpolarization in human cerebral arteries
Authors: Petersson Jesper , Zygmunt Peter M. , Brandt Lennart , Högestätt Edward D.
Published: July 2012

Vascular effects of substance P were studied in human isolated pial arteries removed from 14 patients undergoing cerebral cortical resection.

Substance P induced a concentration‐dependent relaxation in the presence of indomethacin. No relaxation was seen in arteries where the endothelium had been removed.

Nω–nitro‐L‐arginine (l‐NOARG, 0.3 mM) abolished the relaxation in arteries from six patients. The relaxation was only partially inhibited in the remaining eight patients, the reduction of the maximum relaxation being less than 50% in each patient.

The L‐NOARG‐resistant relaxation was abolished when the external K+ concentration was raised above 30 mM.

Substance P caused a smooth muscle hyperpolarization (in the presence of L‐NOARG and indomethacin), but only when the artery showed an L‐NOARG‐resistant relaxation.

The results indicate that nitric oxide is an important mediator of endothelium‐dependent relaxation in human cerebral arteries. Furthermore, another endothelium‐dependent pathway, causing hyperpolarization and vasodilatation, was identified in arteries from more than half the population of patients.
Labelling with [125I]‐SB 207710 of a small 5‐HT4 receptor population in piglet right atrium: functional relevance
Authors: Kaumann Alberto J. , Lynham James A. , Brown Anthony M.
Published: July 2012

We investigated the affinity of SB 207710 for sinoatrial 5‐HT4 receptors and the density of right atrial 5‐HT4 receptors with [125I]‐SB 207710 in right atria of new‐born piglets.

SB 207710 (1–100 nM) antagonized the 5‐HT‐evoked tachycardia surmountably with a pKB of 9.8.

[125I]‐SB 207710 (5–1500 pM) labelled a small population of saturable binding sites with a pKD of 10.1 and with 5‐HT4 receptor characteristics. The density of atrial binding sites with 5‐HT4 receptor characteristics was 174 and 22 times lower respectively than those of atrial β1 and β2‐adrenoceptors, labelled with (−)−[125I]‐cyanopindolol.

We suggest that the small 5‐HT4 receptor population may in part explain why the maximal tachycardia caused by 5‐HT is smaller than that caused by catecholamines.
Characterization of an α1D‐adrenoceptor mediating the contractile response of rat aorta to noradrenaline
Authors: Kenny B.A. , Chalmers D.H. , Philpott P.C. , Naylor A.M.
Published: July 2012

The affinities of a number of α1 ‐adrenoceptor antagonists were determined by displacement of [3H]‐prazosin binding from cloned human α1A‐adrenoceptors (previously designated cloned α1c subtype), α1B α1D and rat α1D‐adrenoceptors, stably expressed in rat‐1 fibroblasts. Functional affinity estimates for these compounds were also determined from noradrenaline‐mediated contractions of rat aorta.

BMY 7378 displayed high affinity for cloned human α1D‐adrenoceptors (pKi = 8.2 ±0.10) and was selective over α1A (pAi = 6.2 ±0.10) and α1B subtypes (6.7 ± 0.11). WB 4101, benoxathian and phentolamine displayed high affinity for α1A and α1D adrenoceptors compared to the α1B subtype. Spiperone displayed high affinity and selectivity for α1B adrenoceptors (pKi 8.8 ±0.16). 5‐Methyl‐urapidil was selective for cloned α1A adrenoceptors.

Comparative binding affinities (pKi for compounds at cloned human and rat1D adrenoceptors were almost identical (r = 0.99, slope =1.08).

Prazosin, doxazosin and 5‐methyl‐urapidil were potent, competitive antagonists of noradrenaline‐mediated contractions of rat aorta (pA2 values of 9.8, 8.8 and 7.8 respectively). The selective α1D antagonist BMY 7378 was also a potent antagonist on rat aorta (pKB = 8.3±0.1) but the interaction of this compound was not consistent with competitive antagonism at a single population of receptors.

Functional affinities for compounds determined against noradrenaline‐mediated contractions of rat aorta correlated well with binding affinities at cloned α1D‐adrenoceptors (r = 0.96), but not with α1A (r = 0.61) or α1B (r = 0.46) subtypes.

Noradrenaline‐mediated contractions of rat aorta were sensitive to the alkylating effects of chlorethylclonidine (CEC). CEC (10 μm) caused a small rightward shift in the noradrenaline concentration‐response curve. CEC at 100 μm caused a further shift and suppression of the maximum response to noradrenaline.

The results of this study suggest that noradrenaline predominantly, but not exclusively, mediates contraction of rat aorta through the activation of an α1D‐adrenoceptor.
Endothelial modulation of vasoconstrictor responses to endothelin‐1 in human placental stem villi small arteries
Authors: Sabry S. , Mondon F. , Levy M. , Ferre F. , Dinh‐Xuan A.T.
Published: July 2012

The aim of this study was to assess the role of endothelial cells in the modulation of vasocontractile responses to endothelin‐1 (ET‐1) of human placental vasculature.

Isolated stem villi small arteries (diameter =170–250 μm) were obtained from healthy parturients who underwent caesarean surgery during the 39th week of pregnancy for cephalo‐pelvic disproportion. Isometric tension was measured in vascular rings mounted in a myograph system and challenged with ET‐1 (10−12 to 10−6m).

The vasocontractile response to ET‐1 was significantly (P< 0.0001) increased in endothelial‐denuded (active tension = 1156 ±214 mN mm−1) as compared with endothelial‐preserved vascular rings (active tension = 458 ± 48 mN mm−1). This difference was significantly (P<0.05) but only partly abolished by the NO synthase inhibitor NΩ‐nitro‐L‐arginine (l‐NOARG, 10−4 m).

In endothelial‐preserved rings submaximally precontracted with 5‐hydroxytryptamine (10−6 m), ET‐1 (10−12 to 10−9 m) induced dose‐dependent relaxation (maximum relaxation = 70 ±7%) at 10−9 m, which was followed, at higher doses (10−8 to 10−6 m), by a contraction. In contrast, no relaxation was seen in endothelial‐denuded rings. The relaxation in rings with endothelium was significantly (P< 0.001) reduced by l‐NOARG (10−4 m). Moreover, it was totally abolished by combined pretreatment with L‐NOARG (10−4 m) and the sulphonylurea glibenclamide (10−5 m).

In conclusion, endothelial cells modulate the vascular responses to ET‐1 through the release of NO and a substance acting on the ATP‐sensitive K+ channel of smooth muscle of stem villi small arteries from healthy parturients.
RS 39604: a potent, selective and orally active 5‐HT4 receptor antagonist
Authors: Hegde S.S. , Bonhaus D.W. , Johnson L.G. , Leung E. , Clark R.D. , Eglen R.M.
Published: July 2012

Selective antagonism of 5‐HT4 receptors may provide therapeutic benefit in certain disorders of the myocardium, alimentary and lower urinary tract. We now report on RS 39604, a novel and selective 5‐HT4 receptor antagonist and compare its pharmacological properties with those of SB 204070.

In guinea‐pig striatal membranes, both RS 39604 and SB 204070 inhibited specific binding of [3H]‐GR 113808 in a concentration‐dependent manner yielding p Ki estimates of 9.1 and 10.9, respectively. RS 39604 displayed a low affinity (pKi <6.5) for 5‐HT1A, 5‐HT2C, 5‐HT3, α1c, D1? D2, M1 M2, AT1 B1 and opioid u receptors and moderate affinity for σ1 (pK1 = 6.8) and σ2 (pK1 = 7.8) sites.

In the rat isolated oesophagus, precontracted with carbachol, RS 39604 (30–300 nM) behaved as a competitive antagonist towards 5‐HT‐induced relaxation (pA2 = 9.3; Schild slope =1.0). We and others have shown previously that SB 204070 behaves as an unsurmountable antagonist in this preparation (pA2‐10.5). In the guinea‐pig isolated ileal mucosa, RS 39604 (30 nM) antagonized 5‐MeOT‐induced increase in short‐circuit current (pA2 = 9.1).

In anaesthetized vagotomized micropigs, RS 39604, administered by the i.v. or intraduodenal (i.duod.) route, produced dose‐dependent inhibition of 5‐HT‐induced tachycardia (ID50 = 4.7 μg kg−1, i.v. and 254.5 ug kg−1, i.duod). At maximal doses of 30 μg kg−1, i.v. and 6 mg kg−1, i.duod., the inhibitory effects of RS 39604 lasted for more than 6 h. In this preparation, SB 204070 was as potent as RS 39604 by the i.v. route but was inactive by the intraduodenal route at doses up to 3 mg kg−1.

In conscious mice, RS 39604, administered by the i.p. or p.o. route, produced dose‐dependent inhibition of 5‐hydroxytryptophan (5‐HTP)‐induced diarrhoea (ID50 = 81.3 ug kg−1, i.p. and 1.1 mg kg−1, p.o.). In this assay, SB 204070 was inactive by the oral route at doses up to 30 mg kg−1.

In anaesthetized guinea‐pigs, RS 39604 antagonized the contractile effect of 5‐HT in the proximal colon by producing parallel, dextral displacement of the dose‐response curve to 5‐HT. The mean dose‐ratios to 5‐HT at 0.1 mg kg−1, i.v., 1 mg kg−1, i.v. and 10 mg kg−1, i.duod. were 4.6, 30.7 and 10.8, respectively. SB 204070 behaved as an unsurmountable antagonist in this assay.

In a model of visceral pain in conscious rats, RS 39604 (0.01‐1 mg kg−1, i.v.) did not affect colorectal distension‐induced increases in arterial pressure whereas morphine (1 mg kg−1, i.v.) produced significant inhibition of the response, implying that 5‐HT4 receptors are not involved in nociception in this model.

The data suggest that RS 39604 is a high affinity and selective 5‐HT4 receptor antagonist that is orally active and long‐lasting in vivo. It is concluded that RS 39604 may be the preferable probe to use for investigating the physiological and pathophysiological role of 5‐HT4 receptors in vivo.
Blockade by sigma site ligands of N‐methyl‐D‐aspartate‐evoked responses in rat and mouse cultured hippocampal pyramidal neurones
Authors: Fletcher Elizabeth J. , Church John , Abdel‐Hamid Khaled , MacDonald John F.
Published: July 2012

The effects of a range of structurally‐dissimilar compounds which possess affinity for σ binding sites were examined on the responses of cultured hippocampal pyramidal neurones to the excitatory amino acid analogues N‐methyl‐D‐aspartate (NMDA), kainate and (RS)‐α‐ammo‐3‐hydroxy‐5‐methylisoxazole‐4‐propionic acid (AMPA).

In mouse hippocampal neurones under whole‐cell voltage‐clamp, the compounds tested reversibly attenuated NMDA‐, but not kainate‐ or AMPA‐, evoked currents with a rank order potency (IC50 values in μm): ifenprodil (0.8)>(+)‐N‐allylnormetazocine (1.1)>dextromethorphan (1.8) = haloperidol (1.9)>(+)‐pentazocine (7.2)>1S, 2R‐(−)−cis‐N‐methyl‐N‐[2‐(3, 4‐dichlorophenyl) ethyl]‐2‐(1‐pyrrolidinyl)cyclohexylamine (17) = rimcazole (18)>1,3‐di(2‐tolyl)guanidine (37)>opipramol (96)>caramiphen (110)=carbetapentane (112)> >(+)‐3‐(3‐hydroxyphenyl)‐N‐(1‐propyl)piperidine (485).

The attenuation of NMDA‐evoked responses was not mediated through interactions with the agonist, glycine (except haloperidol) or polyamine (except ifenprodil) binding sites on the NMDA receptor‐channel complex but, in the light of the voltage‐ and, in some cases, use‐dependent nature of their antagonism, an interaction with the ion channel appears to be a likely mechanism of action for many of the compounds.

Micromolar concentrations of selected σ site ligands also reduced NMDA‐evoked rises in intracellular free calcium concentration in Fura‐2‐loaded cultured hippocampal neurones of the rat with the same rank order potency as observed in the electrophysiological studies.

The data indicate that, at micromolar concentrations, the σ site ligands tested act as NMDA receptor antagonists, an action which does not appear to be mediated by high‐affinity σ binding site(s). The functional effects of micromolar concentrations of σ site ligands cannot, therefore, be attributed exclusively to interactions with high‐affinity σ binding sites.
Characterization of metabotropic glutamate receptor‐stimulated phosphoinositide hydrolysis in rat cultured cerebellar granule cells
Authors: Toms N.J. , Jane D.E. , Tse H‐W. , Roberts P.J.
Published: July 2012

The pharmacology of excitatory amino acid (EAA)‐stimulated phosphoinositide (PI) hydrolysis, monitored via [3H]‐inositol monophosphate accumulation, was investigated in primary cultures of rat cerebellar granule cells.

EAA‐stimulated PI hydrolysis peaked after 4–5 days in vitro and subsequently declined.

The agonist order of potency was found to be (EC50): L‐quisqualic acid (Quis) (2 μm)»L‐glutamate (50 μm)>(1S,3R)‐11‐aminocyclopentane‐1,3‐dicarboxylic acid ((1S,3R)‐ACPD) (102 μm). L‐Glutamate (Emax = 873% of basal activity) elicited the largest stimulation of PI hydrolysis, whereas Quis (Emax = 603%) and (1S,3R)‐ACPD (Emax = 306%) produced somewhat lower stimulations.

Several phenylglycine derivatives were found to be active in inhibiting 2 μm Quis‐stimulated PI hydrolysis, in order of potency (IC50): (S)‐4‐carboxy‐3‐hydroxyphenylglycine (41 μm) ≤ (S)‐4‐carboxyphenylglycine (51 μm)»(+)‐α‐methyl‐4‐carboxyphenylglycine (243 μm).

Cultured cerebellar granule cells of the rat appear to have Group I mGluR pharmacology similar to that reported for cloned mGluRl and provide an ideal system for investigating novel mGluRl ligands in a native environment.
Enhancement of the haemodynamic effects of NG‐monomethyl‐l‐arginine by transforming growth factor‐β1 in conscious, normal, but not endotoxaemic, rats
Authors: Gardiner S.M. , Kemp P.A. , March J.E. , Bennett T.
Published: July 2012

Male, Long Evans rats (350–450 g) were chronically instrumented for the measurement of regional haemodynamics, and the effects of TGF‐β1 (25 μg kg−1 i.v. bolus) were assessed during infusion of saline (n=9) or lipopolysaccharide (LPS, 150 μg kg−1 h−1; n= 12). In the same animals, responses to NG‐monomethyl‐l‐arginine (l‐NMMA 10 mg kg−1 bolus; 10 mg kg−1 h−1 infusion) were determined 18 h after administration of TGF‐β1. In a separate experiment, the effects of the endothelin antagonist, SB 209670 (10 μg kg−1 min−1) on responses to TGF‐β1 and to l‐NMMA subsequently, were determined.

In the absence of LPS, TGF‐β1 had slow‐onset bradycardic and pressor effects accompanied by mesenteric and hindquarters, but not renal, vasoconstriction. Eighteen hours after TGF‐β1, these effects had gone, but the bradycardic, pressor, and mesenteric vasoconstrictor responses to l‐NMMA were enhanced. The haemodynamic changes following TGF‐β1 and the augmentation of the subsequent responses to l‐NMMA, were inhibited by SB 209670. These results are consistent with TGF‐β1 stimulating the synthesis and release of endothelin, and an involvement of the latter in responses to l‐NMMA.

The pressor effects of TGF‐β1 were similar in LPS‐infused and saline‐infused animals, but in the former group the mesenteric vasoconstriction was enhanced and the hindquarters vasoconstriction diminished. Since, in the absence of TGF‐β1, LPS‐infused animals showed a developing hindquarters vasodilatation and mesenteric vasoconstriction, it is feasible that, in the presence of TGF‐β1 and LPS together, the haemodynamic profile represented an amalgam of the individual effects of the two interventions, rather than a specific effect of TGF‐β1 on the haemodynamic sequelae of endotoxaemia.

In the presence of LPS, haemodynamic responses to l‐NMMA were suppressed, and TGF‐β1 generally did not affect this suppression. A possible explanation of this observation is that LPS increased circulating endothelin levels, and thus resulted in desensitization to the effects of endothelin released following administration of l‐NMMA.
The ineffectiveness of the NO‐cyclic GMP signaling pathway in the atrial myocardium
Authors: Nawrath Hermann , Bäumner Dörte , Rupp Johanna , Oelert Hellmut
Published: July 2012

This study was performed to determine whether nitric oxide (NO) has direct effects on force of contraction (Fc) in atrial myocardium from rats, rabbits, guinea‐pigs, frogs, and man.

Glyceryl trinitrate, isosorbide dinitrate, 3‐morpholino‐sydnonimine hydrochloride (SIN‐1), and S‐nitroso‐N‐acetylpenicillamine (SNAP) did not signficantly reduce Fc in the various preparations investigated, either given alone or after stimulation of α‐ or β‐adrenoceptors.

SNAP did not change the time course of contractions in rat, guinea‐pig and human preparations.

8‐Bromo‐guanosine‐3′:5′‐cyclic monophosphate (8‐Br‐cyclic GMP) produced a negative inotropic effect in rat, guinea‐pig and human atrial preparations and shortened time to peak tension and relaxation time in human preparations.

High K+ (85 mmol 1−1‐induced contracture in rat heart muscle was reduced by 8‐Br‐cyclic GMP but not by SIN‐1.

N‐monomethyl‐L‐arginine (l‐NMMA), an inhibitor of NO synthase, failed to influence muscarinic effects on Fc or frequency from rat and guinea‐pig hearts.

We conclude that NO, under the experimental conditions described here, has no direct effects on the heart, although cyclic GMP may be involved in the regulation of myocardial contraction.
The involvement of ATP‐sensitive potassium channels and adenosine in the regulation of coronary flow in the isolated perfused rat heart
Authors: Randall Michael D.
Published: July 2012

The roles of adenosine 5′‐triphosphate (ATP)‐sensitive potassium channels (KATP) and endogenous adenosine in the regulation of coronary flow have been assessed in the isolated, buffer‐perfused heart of the rat.

In the presence of glibenclamide 10 μm there was a significant (P< 0.001) reduction in coronary flow from a baseline value of 8.78 ± 0.76 ml min−1 g_1 to 3.89 ± 0.59 ml min−1 g−1. This change was accompanied by a significant (P<0.01) reduction in cardiac mechanical performance as shown by the decrease in the pressure‐rate product from 21,487 ± 2,577 mmHg min−1 to 6,950 ± 1,104 mmHg min−1.

The non‐selective adenosine antagonist 8‐phenyltheophylline (10 μm) also caused a significant (P<0.001) reduction in coronary flow from a basal value of 10.4 ± 0.6 ml min−1 g_1 to 6.32 ± 0.60 ml min−1 g_1. The subsequent addition of glibenclamide, in the presence of 8‐phenyltheo‐phylline, brought about a further significant (p<0.001) reduction in coronary flow to 3.05 ± 0.55 ml min−1 g_1 and this value was similar to that in the presence of glibenclamide alone.

In hearts perfused under constant flow conditions, exogenous adenosine caused dose‐related reductions in coronary perfusion pressure described by a maximum reduction in pressure of 30.7 ± 3.9 mmHg and an ED50 of 977 ± 813 pmol. Addition of glibenclamide caused a significant (P<0.01) increase in coronary perfusion pressure of 44.7 ± 7.2 mmHg and a significant (P<0.05) rightward shift of the dose‐response curve for the depressor effects of adenosine (ED50= 13.5 ± 3.8 nmol), with a depression (P<0.05) of the maximum (16.3 ± 2.4 mmHg).

In conclusion, both KAtp and endogenous adenosine make major contributions towards coronary vascular tone and the regulation of coronary flow in the rat isolated heart. Furthermore, in the coronary vasculature a significant proportion of the vasodilator action of adenosine is mediated through the activation of KAtp
Mexiletine‐induced shortening of the action potential duration of ventricular muscles by activation of ATP‐sensitive K+ channels
Authors: Sato Toshiaki , Shigematsu Sakuji , Arita Makoto
Published: July 2012

A class Ib antiarrhythmic drug, mexiletine (100 μM) significantly shortened the action potential duration (APD) of guinea‐pig ventricular muscles and this effect was completely abolished in the presence of glibenclamide (50 μM), a blocker of the ATP‐sensitive K+ channel (KATP)‐ Mexiletine significantly increased the open probability of uridine diphosphate‐primed KATP channels, recorded in inside‐out patches of the ventricular cells. The results suggest that mexiletine shortens the APD of ventricular muscles, at least in part, via activation of KATP.
Improved survival and reversal of endothelial dysfunction by the 21‐aminosteroid, U‐74389G in splanchnic ischaemia‐reperfusion injury in the rat
Authors: Squadrito Franceso , Altavilla Domenica , Ammendolia Letteria , Squadrito Giovanni , Campo Giuseppe M. , Sperandeo Agostino , Canale Patrizia , Ioculano Mariapatrizia , Saitta Antonino , Caputi Achille P.
Published: July 2012

Anaesthetized rats subjected to total occlusion of the superior mesenteric artery and the coeliac trunk for 45 min developed a severe shock state (splanchnic artery occlusion, SAO shock) resulting in death within 70–90 min after release of the occlusion. Sham‐operated animals were used as controls.

Survival rate, survival time, serum tumour necrosis factor (TNF‐α), white blood cell (WBC) count, mean arterial blood pressure (MAP), plasma malonylaldehyde (MAL), myeloperoxidase activity (MPO) and the responsiveness to acetylcholine (ACh 10 nM‐10 μM) of aortic rings were investigated.

SAO shocked rats had a decreased survival rate and survival time (74 ± 10 min, while sham‐shocked rats survived more than 4 h), reduced mean arterial blood pressure, increased serum levels of TNF‐α (267 ± 13 u ml−1) and plasma levels of MAL (57 ± 7 nmol ml−1), enhanced MPO activity in the ileum (0.23 ± 0.04 u × 10−3 g−1 tissue) and in the lung (2.2 ± 0.8 u × 10−3 g−1 tissue), leukopenia and reduced responsiveness to ACh of aortic rings.

The 21‐aminosteroid U‐74389G (30 mg kg−1, i.v.) increased survival (survival time = 232 ± 15 min), lowered the serum levels of TNF‐α and the plasma levels of MAL, reduced leukopenia and MPO activity both in the ileum (0.021 ± 0.004 u × 10−3 g−1 tissue) and in the lung (0.23 ± 0.03 u × 10−3 g−1 tissue), improved MAP and restored the responsiveness to ACh of aortic rings.

Our data suggest that U‐74389G is a potent lipid peroxidation inhibitor and that it has antishock and endothelial protective actions.
Effect of diabetes and elevated glucose on nitric oxide‐mediated neurotransmission in rat anococcygeus muscle
Authors: Way Kerrie J. , Reid Julianne J.
Published: July 2012

Nitric oxide (NO)‐mediated neurotransmission is impaired in anococcygeus muscle from 8‐week streptozotocin‐induced diabetic rats. This study investigated the effects of insulin treatment, and the duration of diabetes on this impairment. In addition, the effect of in vitro exposure to elevated glucose has been investigated on NO‐mediated relaxations, in muscles from untreated rats.

Relaxant responses to field stimulation (0.5‐5 Hz, 10 s train), sodium nitroprusside (SNP; 5 and 10 nM) and NO (1 and 3 μM) were significantly impaired in anococcygeus muscles from 8‐week diabetic rats, compared to responses from control rats. Insulin treatment (5 u Lente day−1, s.c.) of diabetic rats prevented the development of this impairment.

Consistent with findings in 8‐week diabetic rats, relaxations induced by field stimulation, SNP and NO were attenuated in tissues from 2‐week and 4‐week diabetic rats compared to corresponding control responses, whereas relaxations to papaverine (3 and 10 μM) were not reduced. In contrast, diabetes of 3‐days duration did not affect relaxations to field stimulation, SNP or NO.

Incubation of anococcygeus muscles from untreated rats in medium containing elevated glucose (44.1 mM) for 6 h, significantly impaired relaxations to field stimulation compared to responses obtained after normal glucose (11.1 mM) incubation. Relaxations to SNP and to NO were not affected by 6h exposure to elevated glucose. Similarly, incubation in hyperosmolar solutions containing mannose or sucrose for 6 h, impaired relaxations to field stimulation, but not to SNP or NO.

The results indicate that the diabetes‐induced impairment of NO‐mediated neurotransmission in the rat anococcygeus muscle develops between 3 days and 2 weeks after the induction of diabetes with streptozotocin. Prevention of the impairment by insulin treatment suggests that it is specific for the diabetic state. In addition, the impairment may be related to hyperglycaemia and the consequent rise in osmolarity, since in vitro exposure to high glucose as well as to other hyperosmolar media impaired NO‐mediated relaxations to field stimulation.
Central versus peripheral site of action of the tachykinin NK1‐antagonist RP 67580 in inhibiting chemonociception
Authors: Holzer‐Petsche Ulrike , Rordorf‐Nikolić Tamara
Published: July 2012

Many studies indicate an involvement of substance P in the transmission of nociceptive stimuli, without, however, presenting any conclusive evidence as to its exact site and mode of action. The present experiments tested the involvement of substance P in the mediation of chemical nociception using the non‐peptidic specific tachykinin NK1 ‐receptor antagonist, RP 67580 (2‐[1‐imino‐2‐(2‐methoxyphenyl‐ethyl]‐7, 7diphenyl‐4‐perhydroisoindolone (3aR, 7aR)).

Mean arterial pressure (MAP) and intragastric pressure (IGP) were measured in anaesthetized rats. The reflex changes of these parameters in response to i.p. or s.c. injections of hydrochloric acid or capsaicin were taken to indicate nociception.

Intravenous administration of RP 67580 up to 5 mg kg−1 had little influence on the reflex changes in MAP or IGP in response to hydrochloric acid or capsaicin. In contrast, the sensitization of rats to i.p. capsaicin by preinjection of prostaglandin E2 was significantly reduced by 1 mg kg−1 RP 67580.

Intrathecal injection of 5 μg RP 67580 inhibited the reflex changes of MAP and IGP in response to i.p. or s.c. capsaicin whereas the inactive enantiomer RP 68651 was ineffective.

The results indicate that spinal NK1 ‐receptors are involved in the acute transmission of chemically induced pain, while such receptors in the periphery take part in the sensitization by prostaglandin E2. The rather minor ability of i.v. RP 67580 to inhibit the acute nociceptive reflex is attributed to an insufficient penetration of the blood‐brain‐barrier.
Factors involved in the relaxation of female pig urethra evoked by electrical field stimulation
Authors: Werkström Viktoria , Persson Katarina , Ny Lars , Bridgewater Melissa , Brading Alison F. , Andersson Karl‐Erik
Published: July 2012

Non‐adrenergic, non‐cholinergic (NANC) relaxations induced by electrical field stimulation (EFS) were studied in pig isolated urethra. The mechanism for relaxation was characterized by measurement of cyclic nucleotides and by study of involvement of different subsets of voltage‐operated calcium channels (VOCCs).

EFS evoked frequency‐dependent and tetrodotoxin‐sensitive relaxations in the presence of propranolol (1 JIM), phentolamine (1 μM) and scopolamine (1 μIM). At low frequencies (<:12Hz), relaxations were rapid, whereas at high (> 12 Hz) frequencies distinct biphasic relaxations were evoked. The latter consisted of a rapidly developing first phase followed by a more long‐lasting second phase.

Treatment with the NO‐synthesis inhibitor NG‐nitro‐L‐arginine (L‐NOARG; 0.3 mM) inhibited relaxations at low frequencies of stimulation. At high frequencies (>12 Hz) only the first relaxation phase was affected.

Measurement of cyclic nucleotides in preparations subjected to continuous nerve‐stimulation, revealed an increase in guanosine 3′:5′‐cyclic monophosphate (cyclic GMP) levels from 1.3 ± 0.3 to 3.0±0.4 pmol mg−1 protein (P<0.01). In the presence of L‐NOARG, there was a significant decrease in cyclic GMP content to control. However, there was no increase in cyclic GMP content in response to EFS. Levels of cyclic AMP remained unchanged following EFS.

Treatment with the N‐type VOCC‐inhibitor, ω‐conotoxin GVIA (0.1 μMs) reduced NO‐dependent relaxations, the effect being most pronounced at low frequencies (1‐4 Hz) of stimulation. The NO‐independent second phase of the relaxation, studied in the presence of L‐NOARG (0.3 mM) at 16–30 Hz, was however markedly reduced or abolished by ω‐conotoxin GVIA. ω‐Conotoxin MVIIC (1 μM) or ω‐agatoxin IVA (30 nM) had no effect on electrically evoked relaxations.

These results suggest that NANC‐nerve derived urethral relaxation in the pig consists of two apparently independent components. One is mediated by NO and associated with an increase in cyclic GMP content. The other mediator is unknown and produces relaxations not associated with changes in levels of cyclic nucleotides. The release of this mediator seems to involve the N‐type VOCC, since the relaxation was markedly reduced or abolished by ω‐conotoxin GVIA.
pH‐dependent transport of procainamide in cultured renal epithelial monolayers of OK cells: consistent with nonionic diffusion
Authors: Dudley Adam J. , Brown Colin D.A.
Published: July 2012

Previous studies suggest that procainamide is a substrate for organic cation/proton antiport. In order to study the coupling between procainamide flux and proton flux in greater detail we investigated the effects of extracellular procainamide addition upon intracellular pH in cultured monolayers of renal OK cells. Intracellular pH was monitored by use of BCECF as a probe.

Apical addition of procainamide (10 mM) caused a significant alkalinisation of intracellular pH. Basolateral addition of procainamide was equally effective in raising intracellular pH. A similar alkalinisation was found in two other renal cell Unes: MDCK strain 1 and LLCPK1.

In contrast, both tetraethylammonium and N‐methylnicotinamide, archetypal substrates for organic cation/proton antiport were without effect upon intracellular pH.

At physiological pH values, procainamide exists as a neutral weak base (B) and its conjugate weak acid (BH+). To test which species of procainamide was responsible for the alkalinisation, experiments in which [B] was kept constant whilst [BH+] was varied from 1.15 mM to 7.25 mM were performed. The results suggested that the neutral weak base (B) was the permeant species.

Procainamide efflux from procainamide‐loaded cell monolayers resulted in a significant acidification of intracellular pH. As with procainamide uptake, this result could be ascribed to the movement of neutral weak base.

These effects of procainamide upon intracellular pH are consistent with nonionic diffusion of procainamide rather than an interaction of procainamide with the organic cation/proton antiporter. In addition, the results suggest that organic cation/proton antiport is not highly expressed in OK cells.
Lens opacification by antioestrogens: tamoxifen vs ICI 182, 780
Authors: Zhang Jin Jun , Jacob Tim J.C. , Hardy Simon P. , Higgins Christopher F. , Valverde Miguel A.
Published: July 2012

The antioestrogen, tamoxifen, blocks volume‐regulated chloride channels and reduces transparency in bovine lenses maintained in vitro. In contrast to tamoxifen, the sterodial antioestrogen, ICI 182780, did not block volume‐regulated chloride currents in three cultured cell lines and required 10 fold higher concentration to induce significant opacification of bovine lenses maintained in vitro. These data suggest that ocular toxic side effects will be minimized by use of the steroidal (ICI 182780) rather than nonsteroidal antioestrogens (tamoxifen).
Interaction of human adrenomedullin 13–52 with calcitonin gene‐related peptide receptors in the microvasculature of the rat and hamster
Authors: Hall Judith M. , Siney Lynn , Lippton Howard , Hyman Albert , Kang‐Chang Jaw , Brain Susan D.
Published: July 2012

Adrenomedullin (ADM), a recently discovered circulating hypotensive peptide, shares limited sequence homology with the sensory nerve‐derived vasodilator, calcitonin gene‐related peptide (CGRP). This study compared the vasodilator effect of sequence 13–52 of human adrenomedullin (ADM13–52) with that of human α CGRP (CGRP), in the microvasculature of the hamster cheek pouch and rat skin in vivo.

Single arterioles (20–40 μm diameter) in the hamster cheek pouch were visualised by intravital microscopy and video recording, and measured by image analysis. Both ADM13–52 (1 pmol–0.4 nmol) and CGRP (0.1 pmol–1 nmol) evoked dose‐related increases in the diameter of preconstricted arterioles (n = 6). ADM13–52 (ED50 14 pmol) was 20 fold less active than CGRP (ED50 0.71 pmol). The kinetics of onset and decline of vasodilator responses to both peptides were similar, with vasodilator responses to both peptides reaching a maximum at ca. 2 min, and reversing after 10–15 min (n = 5–7). The submaximal increase in blood flow evoked by ADM13–52 was significantly inhibited (P < 0.05; n = 6) by the CGRP1 receptor antagonist, CGRP8–37, at a dose (300 nmol kg−1, i.v.) that we have previously shown to inhibit significantly equivalent vasodilator responses to CGRP in this preparation.

In experiments measuring changes in local blood flow in rat skin by a 133xenon clearance technique, intradermal injection of both ADM13–52 (3–300 pmol) and CGRP (0.1–30 pmol) evoked dose‐related increases in local blood flow. ADM13–52 (ED50 27 pmol) was 17 fold less potent than CGRP (ED50 1.6 pmol) (n = 6). The submaximal increase in blood flow evoked by both peptides was significantly inhibited (P < 0.02; n = 5) by CGRP837 (100 nmol kg−1, i.v.).

We conclude that ADM13–52 is a potent vasodilator in the microvasculature of the hamster and rat in vivo. It mediates its vasodilator effect by arteriolar dilatation and this effect is due, at least in part, to the stimulation of CGRP1 receptors.
On the inhibition of prostanoid formation by SK&F 96365, a blocker of receptor‐operated calcium entry
Authors: Leis Hans J. , Zach Doris , Huber Evelyn , Ziermann Laszlo , Gleispach Helmut , Windischhofer W.
Published: July 2012

The proposed blocker of receptor‐operated calcium channels, SK&F 96365 was shown to inhibit formation of prostaglandin E2 in two osteoblast‐like cell lines, MC3T3‐E1 and UMR‐106 in a dose‐dependent manner at an IC50 of 3–4 μm. Inhibition was observed with various stimuli (arachidonic acid, bradykinin and calcium ionophore A23187).

This effect was also observed in human platelets, where SK&F 96365 dose‐dependently blocked thromboxane biosynthesis and formation of 12‐hydroxy‐heptadecatrienoic acid after stimulation with arachidonic acid (IC50 = 4 μm).

The compound had no effect on 12‐hydroxy‐eicosatetraenoic acid production by human platelets. Additionally, linoleic acid oxidation by soybean 15‐lipoxidase was not impaired by SK&F 96365. The results thus provide evidence for cyclo‐oxygenase inhibition by SK&F 96365 at concentrations used to block receptor‐operated calcium influx.
Cardiovascular effects of the calcium sensitizer, levosimendan, in heart failure induced by rapid pacing in the presence of aortic constriction
Authors: Udvary Éva , Papp Julius Gy , Végh Ágnes
Published: July 2012

The haemodynamic effects of a novel cardiotonic drug, levosimendan, which has both calcium‐sensitizing and phosphodiesterase III (PDE III) inhibitory properties, were studied in conscious dogs in which heart failure had been induced by prolonged cardiac pacing in the presence of aortic constriction. These effects were compared with those in sham‐operated dogs with essentially normal cardiac function.

Eighteen mongrel dogs were instrumented for the measurement of left ventricular pressure (LVSP, LVEDP) and contractile function (dP/dt; dP/dt/P). In twelve dogs a balloon catheter, positioned in the thoracic aorta, was inflated producing an approximate 60% reduction in effective aortic diameter. Twenty min later rapid ventricular pacing (240 beats mean−1) was commenced and maintained for 48 h by means of a bipolar pacing electrode introduced into the right ventricle. This electrode served also for recording changes in the endocardial electrogram in the absence of pacing. Six of these dogs were used to evaluate the haemodynamic changes of pacing‐induced heart failure; a further six of these dogs the haemodynamic changes elicited by levosimendan under these conditions. Six sham‐operated dogs (group 2) served as controls.

In six dogs (group 1) the haemodynamic alterations were assessed after the development of heart failure. In the presence of aortic constriction, 48 h continuous rapid cardiac pacing resulted in a marked deterioration in left ventricular function which remained stable for at least 48 h after cessation of pacing. Thus, there was a marked reduction in LVSP (15%), + dP/dtmax (35%), ‐dP/dtmax (36%) and also in dP/dt/P (29%), whereas LVEDP was increased considerably (from 6.4 ± 1.4 to 20.0 ± 2.2 mmHg). A marked elevation occurred in endocardial ST‐segment (138%), lasting for 20 min.

Levosimendan was administered intravenously in doses of 0.005, 0.01 and 0.03 μmol kg−1 to 2 groups of conscious dogs. In the sham‐operated dogs (group 2), only the higher dose (0.03 μmol kg−1) produced significant increases in LVSP (19%), + dP/dtmax (37%), and in dP/dt/P (32%). In dogs with heart failure (group 3) doses of 0.005, 0.01 and 0.03 μmol kg−1 levosimendan resulted in an improvement in + dP/dtmax (26%, 38% and 49%), ‐dP/dtmax (20%, 25% and 38%) and in dP/dt/P (19%, 34% and 50%) and reduction in the elevated LVEDP (from 20 ± 2.2 mmHg to 16 ±1.0, 10 ± 1.3 and 9± 1.0 mmHg, respectively).

Levosimendan proved to be a potent cardiotonic drug at the doses used, and was approximately three times more effective under conditions of impaired left ventricular function than in normal hearts.