bluuebunny/crossref_metadata_2025 huggingface dataset:
Differential A1‐adenosine receptor reserve for inhibition of cyclic AMP accumulation and G‐protein activation in DDT1 MF‐2 cells
Authors: Baker Stephen P , Scammells Peter J , Belardinelli Luiz
Published: January 2005

The A1‐adenosine receptor (A1AdoR) reserve for N6‐cyclopentyladenosine (CPA) mediated inhibition of (−)isoprenaline stimulated cyclic AMP accumulation and stimulation of [35S]‐guanosine‐5′‐O‐(thiotriphosphate) (GTPγS) binding, a measure of guanine nucleotide binding protein (G‐protein) activation, was determined in DDT1 MF‐2 cells.

Inactivation of the A1AdoRs with the chemoreactive ligand 8‐cyclopentyl‐3‐[3‐[[4‐(fluorosulphonyl)benzoyl]oxy]propyl]‐1‐propylxanthine (FSCPX) caused a progressive rightward shift of the concentration‐response curves for CPA to inhibit cyclic AMP accumulation, with a maximum of 10 fold increase in the EC50 value. In contrast, inactivation of A1AdoR's caused only a 1.7 fold rightward shift in the CPA concentration‐response for stimulation of [35S]‐GTPγS binding.

The A1AdoR occupancy‐response relationship for CPA inhibition of cyclic AMP accumulation was hyperbolic with 43% receptor occupancy required to elicit the maximal response, i.e. a 57% A1AdoR reserve. In contrast, the A1AdoR occupancy‐response relationship for CPA mediated stimulation of [35S]‐GTPγS binding was linear indicating little or no receptor reserve for G‐protein activation. The relationship between CPA stimulation of [35S]‐GTPγS binding and cyclic AMP inhibition was also hyperbolic with 44% G‐protein activation sufficient to cause maximal inhibition.

The data suggest that the A1AdoR reserve for CPA mediated inhibition of cyclic AMP accumulation occurs at the level of G‐protein interaction with adenylyl cyclase. However, each A1AdoR appears to activate a constant fraction of the total G‐protein population suggesting signal amplification at the receptor‐G‐protein level which may also contribute to the receptor reserve for CPA.

British Journal of Pharmacology (2000) 130, 1156–1164; doi:10.1038/sj.bjp.0703405
Apoptosis in the vasculature: mechanisms and functional importance
Authors: Mallat Ziad , Tedgui Alain
Published: January 2005

Apoptotic death has now been recognized in a number of common and threatening vascular diseases, including atherosclerosis. Interest in apoptosis research relates to the fact that apoptosis, in contrast to oncosis, is a highly regulated process of cell death which raises the hope for the development of specific therapeutic strategies to alter disease progression. This review summarizes the mechanisms involved in vascular endothelial and smooth muscle cell survival/apoptosis, and the potential roles of apoptotic death in atherosclerosis and restenosis. The potential effects of modulation of apoptosis in these diseases are also discussed.

British Journal of Pharmacology (2000) 130, 947–962; doi:10.1038/sj.bjp.0703407
Cholinergic facilitation of neurotransmission to the smooth muscle of the guinea‐pig prostate gland
Authors: Lau W A K , Pennefather J N , Mitchelson F J
Published: January 2005

Functional experiments have been conducted to assess the effects of acetylcholine and carbachol, and the receptors on which they act to facilitate neurotransmission to the stromal smooth muscle of the prostate gland of the guinea‐pig.

Acetylcholine and carbachol (0.1 μM–0.1 mM) enhanced contractions evoked by trains of electrical field stimulation (20 pulses of 0.5 ms at 10 Hz every 50 s with a dial setting of 60 V) of nerve terminals within the guinea‐pig isolated prostate. In these concentrations they had negligible effects on prostatic smooth muscle tone. The facilitatory effects of acetylcholine, but not those of carbachol, were further enhanced in the presence of physostigmine (10 μM).

The facilitatory effects of carbachol were unaffected by the neuropeptide Y Y1 receptor antagonist BIBP 3226 ((R)‐N2‐(diphenylacetyl)‐N‐[(4‐hydroxyphenyl)methyl]‐argininamide) (0.3 μM, n=3) or suramin (100 μM, n=5). Prazosin (0.1 μM, n=5) and guanethidine (10 μM, n=5) alone and in combination (n=4), reduced responses to field stimulation and produced rightward shifts of the log concentration‐response curves to carbachol.

The rank orders of potency of subtype‐preferring muscarinic receptor antagonists in inhibiting the facilitatory actions of acetylcholine and carbachol were: pirenzepine > HHSiD (hexahydrosiladifenidol) > pF‐HHSiD (para‐fluoro‐hexahydrosiladifenidol) 5 himbacine, and pirenzepine > HHSiD > himbacine 5 pF‐HHSiD, respectively. These profiles suggest that muscarinic receptors of the M1‐subtype mediate the facilitatory effects of acetylcholine and carbachol on neurotransmission to the smooth muscle of the guinea‐pig prostate.

British Journal of Pharmacology (2000) 130, 1013–1020; doi:10.1038/sj.bjp.0703409
The inhibition of cholera toxin‐induced 5‐HT release by the 5‐HT3 receptor antagonist, granisetron, in the rat
Authors: Turvill J L , Connor P , Farthing M J G
Published: January 2005

The secretagogue 5‐hydroxytryptamine (5‐HT) is implicated in the pathophysiology of cholera. 5‐HT released from enterochromaffin cells after cholera toxin exposure is thought to activate non‐neuronally (5‐HT2 dependent) and neuronally (5‐HT3 dependent) mediated water and electrolyte secretion. CT‐secretion can be reduced by preventing the release of 5‐HT.

Enterochromaffin cells possess numerous receptors that, under basal conditions, modulate 5‐HT release. These include basolateral 5‐HT3 receptors, the activation of which is known to enhance 5‐HT release.

Until now, 5‐HT3 receptor antagonists (e.g. granisetron) have been thought to inhibit cholera toxin‐induced fluid secretion by blockading 5‐HT3 receptors on secretory enteric neurones. Instead we postulated that they act by inhibiting cholera toxin‐induced enterochromaffin cell degranulation.

Isolated intestinal segments in anaesthetized male Wistar rats, pre‐treated with granisetron 75 μg kg−1, lidoocaine 6 mg kg−1 or saline, were instilled with a supramaximal dose of cholera toxin or saline. Net fluid movement was determined by small intestinal perfusion or gravimetry and small intestinal and luminal fluid 5‐HT levels were determined by HPLC with fluorimetric detection.

Intraluminal 5‐HT release was proportional to the reduction in tissue 5‐HT levels and to the onset of water and electrolyte secretion, suggesting that luminal 5‐HT levels reflect enterochromaffin cell activity.

Both lidocaine and granisetron inhibited fluid secretion. However, granisetron alone, and proportionately, reduced 5‐HT release.

The simultaneous inhibition of 5‐HT release and fluid secretion by granisetron suggests that 5‐HT release from enterochromaffin cells is potentiated by endogenous 5‐HT3 receptors. The accentuated 5‐HT release promotes cholera toxin‐induced fluid secretion.

British Journal of Pharmacology (2000) 130, 1031–1036; doi:10.1038/sj.bjp.0703414
Inhibition of mouse neuromuscular transmission and contractile function by okadaic acid and cantharidin
Authors: Hong S J
Published: January 2005

Phosphorylations of cellular proteins modulate biological activities. The effects of okadaic acid (0.1–10 μM) and cantharidin (1–100 μM), inhibitors of protein phosphatases, on the synaptic transmission at the mouse neuromuscular junction were explored.

Both inhibitors almost completely depressed twitch forces elicited by electrical stimulation of diaphragm muscles (the IC50s for okadaic acid and cantharidin were 1.1±0.2 and 13±1 μM, n=5, respectively) and suppressed contractures evoked by high K+ and ryanodine more than 70%. Contractures caused by cardiotoxin, which destroys the integrity of sarcolemma, were not depressed.

Both okadaic acid (10 μM) and cantharidin (100 μM) depolarized muscle membranes from ∼−80 to ∼−60 mV in a partially reversible and tetrodotoxin‐sensitive manner. The initial short‐term enhancement of twitch responses (up to ∼40%) was correlated with the inhibitors‐induced repetitive firings of muscle action potential.

Treatment with either agent resulted in nearly complete inhibitions of endplate potential (epp). The IC50s were 0.8±0.2 and 9±2 μM (n=5), respectively, for okadaic acid and cantharidin. On high frequency stimulation, the coefficient of epps was increased more than 10 fold and the extent of epp run‐down during stimulations intensified from ∼25 to ∼75%. Analyses of presynaptic quantal releases revealed decreases in epp quantal content and the immediately available vesicle pool.

The frequency of miniature epp was initially elevated up to 2 fold then suppressed down to ∼30%. The small reduction in the amplitude was antagonized when the membrane of endplate area was repolarized.

The data suggest that okadaic acid and cantharidin inhibit mobilizations of synaptic vesicles and depress Ca2+ release from sarcoplasmic reticulum and that protein phosphatases participate in the modulation of motor function.

British Journal of Pharmacology (2000) 130, 1211–1218; doi:10.1038/sj.bjp.0703418
Beneficial effects of n‐acetylcysteine on ischaemic brain injury
Authors: Cuzzocrea Salvatore , Mazzon Emanuela , Costantino Giuseppina , Serraino Ivana , Dugo Laura , Calabrò Giusy , Cucinotta Giovanni , De Sarro Angela , Caputi A P
Published: January 2005

Nitric oxide (NO), peroxynitrite, formed from NO and superoxide anion, poly (ADP‐ribole) synthetase have been implicated as mediators of neuronal damage following focal ischaemia. Here we have investigated the effects of n‐acetylcysteine (NAC) treatment in Mongolian gerbils subjected to cerebral ischaemia.

Treatment of gerbils with NAC (20 mg kg−1 30 min before reperfusion and 1, 2 and 6 h after reperfusion) reduced the formation of post‐ischaemic brain oedema, evaluated by water content.

NAC also attenuated the increase in the brain levels of malondialdehyde (MDA) and the increase in the hippocampus of myeloperoxidase (MPO) caused by cerebral ischaemia.

Positive staining for nitrotyrosine was found in the hippocampus in Mongolian gerbils subjected to cerebral ischaemia. Hippocampus tissue sections from Mongolian gerbils subjected to cerebral ischaemia also showed positive staining for poly (ADP‐ribose) synthetase (PARS). The degree of staining for nitrotyrosine and for PARS were markedly reduced in tissue sections obtained from animals that received NAC.

NAC treatment increased survival and reduced hyperactivity linked to neurodegeneration induced by cerebral ischaemia and reperfusion.

Histological observations of the pyramidal layer of CA1 showed a reduction of neuronal loss in animals that received NAC.

These results show that NAC improves brain injury induced by transient cerebral ischaemia.

British Journal of Pharmacology (2000) 130, 1219–1226; doi:10.1038/sj.bjp.0703421
Effects of riluzole on electrically evoked neurotransmitter release
Authors: Jehle T , Bauer J , Blauth E , Hummel A , Darstein M , Freiman T M , Feuerstein T J
Published: January 2005

The main purpose of the present study was to investigate the effects of the neuroprotective agent riluzole on the electrically evoked release of [3H]‐glutamate ([3H]‐Glu) in mouse neocortical slices. The reported selectivity of riluzole for excitatory amino acids was tested in release experiments with further neurotransmitters. Also distinct species, mouse, rat and man were compared.

[3H]‐Glu was formed endogenously during incubation of slices with [3H]‐glutamine ([3H]‐Gln). Released [3H]‐Glu and tissue [3H]‐Glu was separated by anion exchange chromatography. Electrically evoked [3H]‐Glu release was strongly diminished by tetrodotoxin (TTX) and Ca2+‐withdrawal.

Riluzole (100 μM) depressed the release of [3H]‐Glu up to 77% (IC50=19.5 μM). Riluzole was also able to inhibit strongly the electrically evoked release of [3H]‐acetylcholine ([3H]‐ACh) (at 100 μM by 92%, IC50=3.3 μM, and [3H]‐dopamine ([3H]‐DA) (at 32 μM by 72%, IC50=6.8 μM). However, the release of [3H]‐serotonin ([3H]‐5‐HT) was less diminished (at 100 μM by 53%, IC50=39.8 μM). Riluzole up to 100 μM did not affect [3H]‐noradrenaline ([3H]‐NA) release.

Between species, i.e. in mouse, rat and human neocortex, no significant differences between the effects of riluzole could be observed.

The NMDA‐receptor blocker MK‐801 (1 μM) and the AMPA/Kainate‐receptor blocker NBQX (1 μM) did neither affect the electrically evoked [3H]‐ACh release nor its inhibition by riluzole, indicating that effects of riluzole on transmitter release were neither due to modulation of ionotropic Glu receptors, nor due to indirect inhibition of Glu release through these receptors.

Taken together, riluzole inhibits the release of distinct neurotransmitters differently, but is not selective for the excitatory amino acid Glu.

British Journal of Pharmacology (2000) 130, 1227–1234; doi:10.1038/sj.bjp.0703424
Role of inflammatory mediators in lipid A analogue (ONO‐4007)‐induced vascular permeability change in mouse skin
Authors: Ishida Hiroyasu , Fujii Emiko , Irie Kaoru , Yoshioka Toshimasa , Muraki Takamura , Ogawa Ryou
Published: January 2005

Endotoxin shock is accompanied by an increase in peripheral vascular permeability. It has been postulated that most biological activities of LPS are derived from lipid A moiety. Here we examined the effect of lipid A analogue ONO‐4007 in increasing vascular permeability and the possible mediators in mouse skin by a dye leakage method.

Subcutaneous injection of ONO‐4007 (1–2 mg site−1) induced a dose‐dependent increase in vascular permeability which was evident after 120 min.

ONO‐4007‐induced dye leakage was significantly attenuated by pretreatments with anti‐tumour necrosis factor‐α (TNF‐α) and anti‐interleukin‐1α (IL‐1α) antibodies, but not with indomethacin (5 mg kg−1) or diphenhydramine (10 mg kg−1). ONO‐4007‐induced dye leakage was significantly inhibited by a pretreatment with NG‐nitro‐L‐arginine methyl ester (L‐NAME) (10 mg kg−1) but not with aminoguanidine (50 mg kg−1). In inducible nitric oxide synthase (iNOS)‐deficient mice, ONO‐4007 significantly increased the dye leakage, while ONO‐4007 dilated rat thoracic aortic rings pre‐contracted with phenylephrine, and the L‐NAME pretreatment inhibited the dilation.

Thus, TNF‐α, IL‐1α and constitutive NOSs‐derived nitric oxide but not prostaglandins or histamine play a role in ONO‐4007‐induced increase in vascular permeability. Although ONO‐4007 mimics LPS in increasing vascular permeability, mechanisms of permeability change elicited by ONO‐4007 were not identical to those of LPS.

British Journal of Pharmacology (2000) 130, 1235–1240; doi:10.1038/sj.bjp.0703425
[3H]‐8‐OH‐DPAT binding in the rat brain raphe area: involvement of 5‐HT1A and non‐5‐HT1A receptors
Authors: Assié Marie‐Bernadette , Koek Wouter
Published: January 2005

The 5‐HT1A agonist 8‐OH‐DPAT has been shown to have additional 5‐HT uptake inhibiting properties. The present work was undertaken to examine further the binding of [3H]‐8‐OH‐DPAT in the raphe area of the rat brain, a region rich in 5‐HT1A receptors and 5‐HT uptake sites.

5‐HT inhibited [3H]‐8‐OH‐DPAT binding in a biphasic manner (pKi1: 8.82±0.01, pKi2: 6.07±0.05, n=4) with the low affinity site representing 36±4% of the total population. A biphasic inhibition curve was found also with the 5‐HT1A antagonist, WAY 100635 (pKi1: 8.65±0.17, pKi2: 4.26±0.38, n=3). In the presence of 1 μM WAY 100635 to mask 5‐HT1A receptors, 5‐HT inhibited [3H]‐8‐OH‐DPAT binding in a monophasic manner (pKi: 6.04±0.07, n=3).

The affinities of various compounds for sites labelled by [3H]‐8‐OH‐DPAT in the presence of 1 μM WAY 100635 and for sites labelled by [3H]‐citalopram (a selective 5‐HT uptake inhibitor) were determined. There was a significant correlation between pKi values at 5‐HT uptake sites and at non‐5HT1A sites labelled by [3H]‐8‐OH‐DPAT (r=0.80, P<0.001, n=17), suggesting these latter sites to be 5‐HT uptake sites.

Whereas the affinities of R(+) and S(−) enantiomers of 8‐OH‐DPAT for the 5‐HT uptake site are similar, R(+)8‐OH‐DPAT has 10 times higher affinity for the non‐5‐HT1A site than S(−)8‐OH‐DPAT and was considered as an outlier in the correlation. It is suggested that [3H]‐8‐OH‐DPAT labels other, as yet unknown binding sites in the raphe.

British Journal of Pharmacology (2000) 130, 1348–1352; doi:10.1038/sj.bjp.0703426
Selective agonist of group II glutamate metabotropic receptors, LY354740, inhibits tolerance to analgesic effects of morphine in mice
Authors: Popik Piotr , Kozela Ewa , Pilc Andrzej
Published: January 2005

Antagonists of glutamate N‐methyl‐D‐aspartate (NMDA) subtype receptor inhibit the development of tolerance to the antinociceptive effects of opioids. Another way to inhibit the function of glutamate receptors is the stimulation of presynaptic metabotropic group II (mGluRII) receptors. Because LY354740 ((+)‐2‐aminobicyclo [3,1,0] hexane‐2,6‐dicarboxylic acid) is the first systemically active agonist of group II mGlu receptors, we investigated if this compound might inhibit the development of tolerance to antinociceptive effects of morphine and fentanyl.

As assessed by cumulative dose‐response approach in the tail‐flick test, administration of 10 mg kg−1 morphine bid s.c. to male Albino Swiss mice for 6 days, right‐shifted morphine dose‐response curve by ∼4 fold. In a separate group of mice, 12 injections of 0.04 mg kg−1 of fentanyl over 3 days, right‐shifted fentanyl dose‐response curve by ∼3.3 fold.

In experiment 1, LY354740 (1 and 10, but not 0.1 mg kg−1) as well as the reference compound, an uncompetitive NMDA receptor antagonist memantine (7.5 mg kg−1) inhibited the development of morphine tolerance. Neither LY354740 (10 mg kg−1) nor memantine (7.5 mg kg−1) affected the development of tolerance to fentanyl. In experiment 2, neither LY354740 (1 and 10 mg kg−1) nor memantine (7.5 mg kg−1) affected the tail‐flick antinociceptive response, or the acute antinociceptive effect of morphine.

The present results are the first to suggest that the development of antinociceptive morphine tolerance may be inhibited by metabotropic group II glutamate agonist.

British Journal of Pharmacology (2000) 130, 1425–1431; doi:10.1038/sj.bjp.0703438
Effects of combined neutral endopeptidase 24‐11 and angiotensin‐converting enzyme inhibition on femoral vascular conductance in streptozotocin‐induced diabetic rats
Authors: Arbin V , Claperon N , Fournié‐Zaluski M ‐C , Roques B P , Peyroux J
Published: January 2005

The successive effects of the angiotensin‐converting enzyme inhibitor captopril (CAP, 2 mg kg−1+1 mg kg−1 30 min−1 infusion) and the neutral endopeptidase 24‐11 inhibitor retrothiorphan (RT, 25 mg kg−1+12.5 mg kg−1 30 min−1 infusion) were studied on femoral vascular conductance (FVC) in streptozotocin‐induced diabetic (STZ‐SD) and control Sprague‐Dawley (C‐SD) rats. The role of the kinin‐nitric oxide (NO) pathway was assessed by (1) using pre‐treatments: a bradykinin (BK) B2 receptor antagonist (Hoe‐140, 300 μg kg−1), a NO‐synthase inhibitor (Nω‐nitro‐L‐arginine methyl ester, L‐NAME, 10 mg kg−1), a kininase I inhibitor (DL‐2‐mercaptomethyl‐3‐guanidinoethylthiopropanoic acid, MGTA, 10 mg kg−1+20 mg kg−1 20 min−1 infusion) and (2) comparing the effects in STZ‐induced diabetic (STZ‐BN) and control Brown‐Norway kininogen‐deficient (C‐BN) rats.

In C‐SDs, CAP and CAP+RT increased FVC similarly. In STZ‐SDs, FVC and FBF were decreased compared to C‐SDs. CAP+RT increased them more effectively than CAP alone.

In both C‐SDs and STZ‐SDs, the femoral bed vasodilatation elicited by CAP was inhibited by Hoe‐140 and L‐NAME. The FVC increase elicited by CAP+RT was not significantly reduced by Hoe‐140 but was inhibited by L‐NAME and Hoe‐140+MGTA.

In C‐BNs, the vasodilatator responses to CAP and CAP+RT were abolished and highly reduced, respectively. In STZ‐BNs, these responses were abolished.

These results show that in STZ‐SDs, CAP+RT improve FBF and FVC more effectively than CAP alone. These effects are linked to an increased activation of the kinin‐NO pathway. BK could lead to NO production by BK B2 receptor activation and another pathway in which kininase I may be involved.

British Journal of Pharmacology (2000) 130, 1297–1304; doi:10.1038/sj.bjp.0703442
Comparative effects of continuous infusion of mCPP, Ro 60‐0175 and d‐fenfluramine on food intake, water intake, body weight and locomotor activity in rats
Authors: Vickers S P , Benwell K R , Porter R H , Bickerdike M J , Kennett G A , Dourish C T
Published: January 2005

The aim of the study was to compare the effects of 14 day subcutaneous infusion of the 5‐HT2C receptor agonists, m‐chlorophenylpiperazine (mCPP, 12 mg kg−1 day−1) and Ro 60‐0175 (36 mg kg−1 day−1) and the 5‐HT releasing agent and re‐uptake inhibitor, d‐fenfluramine (6 mg kg−1 day−1), on food and water intake, body weight gain and locomotion in lean male Lister hooded rats.

Chronic infusion of all three drugs significantly reduced food intake and attenuated body weight gain. In contrast, drug infusion did not lead to significant reductions in locomotor activity in animals assessed 2 and 13 days after pump implantation.

In a subsequent 14 day study that was designed to identify possible tolerance during days 7–14, animals were given a subcutaneous infusion of mCPP (12 mg kg−1 day−1) or d‐fenfluramine (6 mg kg−1 day−1) for either 7 or 14 days. During the first 7 days both drugs significantly reduced body weight gain compared to saline‐infused controls; however, from day 7 onwards animals withdrawn from drug treatment exhibited an increase in body weight such that by day 14 they were significantly heavier than their 14‐day drug‐treated counterparts.

Both mCPP and d‐fenfluramine reduced daily food intake throughout the infusion periods. For 14‐day treated animals this hypophagia was marked during the initial week of the study but only minor during the second week. In light of the sustained drug effect on body weight, the data suggest that weight loss by 5‐HT2C receptor stimulation may be only partly dependent on changes in food consumption and that 5‐HT2C receptor agonists may have effects on thermogenesis.

These data suggest tolerance does not develop to the effects of d‐fenfluramine, mCPP and Ro 60‐0175 on rat body weight gain.

British Journal of Pharmacology (2000) 130, 1305–1314; doi:10.1038/sj.bjp.0703443
Direct effects of propylthiouracil on testosterone secretion in rat testicular interstitial cells
Authors: Chiao Yu‐Chung , Lin Ho , Wang Shyi‐Wu , Wang Paulus S
Published: January 2005

The aim of this study was to investigate the mechanism by which propylthiouracil (PTU) exerts its inhibitory effects on the production of testosterone by rat testicular interstitial cells.

The plasma testosterone concentration was decreased 60 and 120 min after an intravenous infusion of PTU (10 or 20 mg kg−1), but the concentration of plasma T4 was unaffected by the drug treatment.

Exposure of anterior pituitary tissue to PTU (3–12 mM) in vitro did not affect either basal or gonadotropin‐releasing hormone (GnRH)‐stimulated luteinizing hormone (LH) release.

PTU (3–12 mM) inhibited both the basal and the human chorionic gonadotropin (hCG, 0.05 iu ml−1)‐stimulated release of testosterone from rat testicular tissue in vitro; at the highest concentration tested (12 mM), it also inhibited the forskolin or 8‐bromo‐adenosine 3′:5′‐cyclic monophosphate (8‐Br‐cyclic AMP)‐stimulated release of testosterone.

The 25‐OH‐cholesterol (10−7–10−5 M)‐stimulated release of pregnenolone and testosterone by the testicular interstitial cells was inhibited by PTU (12 mM, P<0.05).

The results suggest that the inhibitory actions of PTU on testosterone secretion are exerted, at least in part, at the testicular level through a mechanism which is independent of thyroid status and which involves a reduction in P450scc activity and, hence, in the conversion of cholesterol to pregnenolone.

British Journal of Pharmacology (2000) 130, 1477–1482; doi:10.1038/sj.bjp.0703444
Antidepressant drugs inhibit glucocorticoid receptor‐mediated gene transcription – a possible mechanism
Authors: Budziszewska Bogusława , Jaworska‐Feil Lucylla , Kajta Małgorzata , Lasoń Władysław
Published: January 2005

Antidepressant drugs are known to inhibit some changes evoked by glucocorticoids, as well as a hyperactivity of hypothalamic‐pituitary‐adrenal (HPA) axis, often observed in depression.

The aim of present study was to investigate effects of various antidepressant drugs on the glucocorticoid‐mediated gene transcription in fibroblast cells, stably transfected with an MMTV promoter (LMCAT cells).

The present study have shown that antidepressants (imipramine, amitriptyline, desipramine, fluoxetine, tianeptine, mianserin and moclobemide), but not cocaine, inhibit the corticosterone‐induced gene transcription in a concentration‐ and a time‐dependent manner.

Drugs which are known to augment clinical effects of medication in depressed patients (lithium chloride, amantadine, memantine), do not affect the inhibitory effects of imipramine on the glucocorticoid receptor (GR)‐mediated gene transcription.

Inhibitors of phospholipase C (PLC), protein kinase C (PKC), Ca2+/calmodulin‐dependent protein kinase (CaMK) and antagonists of the L‐type Ca2+ channel also inhibit the corticosterone‐induced gene transcription.

Inhibitors of protein kinase A (PKA) and protein kinase G (PKG) are without effect on the GR‐induced gene transcription.

Phorbol ester (an activator of PKC) attenuates the inhibitory effect of imipramine on the GR‐induced gene transcription.

Imipramine decreases binding of corticosterone‐receptor complex to DNA.

It is concluded that antidepressant drugs inhibit the corticosterone‐induced gene transcription, and that the inhibitory effect of imipramine depends partly on the PLC/PKC pathway.

British Journal of Pharmacology (2000) 130, 1385–1393; doi:10.1038/sj.bjp.0703445
Effects of dronedarone on Acetylcholine‐activated current in rabbit SAN cells
Authors: Altomare Claudia , Barbuti Andrea , Viscomi Carlo , Baruscotti Mirko , DiFrancesco Dario
Published: January 2005

The effect of the antiarrhythmic drug dronedarone on the Acetylcholine‐activated K+ current (IK(ACh)) was investigated in single cells isolated from sinoatrial node (SAN) tissue of rabbit hearts.

Externally perfused dronedarone (0.001–1 μM) caused a potent, voltage independent block of IK(ACh). Fitting of the dose response curve of IK(ACh) block yielded an IC50 value of 63 nM, a value over one order of magnitude lower than those reported for dronedarone block of other cardiac currents.

IK(ACh) block was not due to an inhibitory action of dronedarone on the muscarinic M2 receptor activation, since the drug was effective on IK(ACh) constitutively activated by intracellular perfusion with GTP‐γS.

External cell perfusion with dronedarone inhibited the activity of IK(ACh) channels recorded from cell‐attached patches by reducing the channel open probability (from 0.56 to 0.11) without modification of the single‐channel conductance.

These data suggest that dronedarone blocks IK(ACh) channels either by disrupting the G‐protein‐mediated activation or by a direct inhibitory interaction with the channel protein.

British Journal of Pharmacology (2000) 130, 1315–1320; doi:10.1038/sj.bjp.0703446
Cytoplasmic binding and disposition kinetics of diclofenac in the isolated perfused rat liver
Authors: Weiss Michael , Kuhlmann Olaf , Hung Daniel Y , Roberts Michael S
Published: January 2005

The binding kinetics of diclofenac to hepatocellular structures were evaluated in the perfused rat liver using the multiple indicator dilution technique and a stochastic model of organ transit time density.

The single‐pass, in situ rat liver preparation was perfused with buffer solution (containing 2% albumin) at 30 ml min−1. Diclofenac and [14C]‐sucrose (extracellular reference) were injected simultaneously as a bolus dose into the portal vein (six experiments in three rats). An analogous series of experiments was performed with [14C]‐diclofenac and [3H]‐sucrose.

The diclofenac outflow data were analysed using three models of intracellular distribution kinetics, assuming (1) instantaneous distribution and binding (well‐mixed model), (2) ‘slow’ binding at specific intracellular sites after instantaneous distribution throughout the cytosol (slow binding model), and (3) ‘slowing’ of cytoplasmic diffusion due to instantaneous binding (slow diffusion model).

The slow binding model provided the best description of the data. The rate constants for cellular influx and sequestration were 0.126±0.026 and 0.013±0.009 s−1, respectively. The estimated ratio of cellular initial distribution volume to extracellular volume of 2.82 indicates an almost instantaneous distribution in the cellular water space, while the corresponding ratio of 5.54 estimated for the apparent tissue distribution volume suggests a relatively high hepatocellular binding. The non‐instantaneous intracellular equilibration process was characterized by time constants of the binding and unbinding process of 53.8 and 49.5 s, respectively. The single‐pass availability of diclofenac was 86%. The results obtained with [14C]‐diclofenac and [3H]‐sucrose were not statistically different.

British Journal of Pharmacology (2000) 130, 1331–1338; doi:10.1038/sj.bjp.0703448
Lack of nitric oxide‐ and guanosine 3′:5′‐cyclic monophosphate‐dependent regulation of α‐thrombin‐induced calcium transient in endothelial cells of spontaneously hypertensive rat hearts
Authors: Failli Paola , Fazzini Alessandro , Ruocco Carlo , Mazzetti Luca , Cecchi Enrica , Giovannelli Lisa , Marra Fabio , Milani Stefano , Giotti Alberto
Published: January 2005

While the expression and/or activity of endothelial nitric oxide synthase (eNOS) has been characterized in spontaneously hypertensive (SHR) and normotensive Wistar Kyoto rat (WKY) hearts, in coronary endothelial cells (ECs) from both strains, the effect of NO on intracellular calcium concentration ([Ca2+]i) is still unknown.

Coronary microvascular ECs were isolated from SHR and WKY and characterized. Immunocytochemistry and Western blot analysis showed that eNOS was similarly expressed in ECs from both strains.

Measuring [Ca2+]i by imaging analysis of fura‐2‐loaded cells, we demonstrated that α‐thrombin (3−180 U l−1) induced a superimposable dose‐dependent calcium transient in ECs from both strains.

In WKY ECs, S‐nitroso‐N‐acetyl‐DL‐penicillamine (SNAP) dose‐dependently (10–100 μM) and 0.1 μM atrial natriuretic factor (ANF) reduced the maximum and the decay time of α‐thrombin‐induced calcium transient. The inhibitory effects of SNAP and ANF were prevented by blocking cyclic GMP‐dependent protein kinase. Non selective eNOS inhibitors prolonged the decay time of α‐thrombin‐induced calcium transient, while the selective inducible NOS inhibitor 1400 W was ineffective. SNAP (100 μM) and 0.1 μM ANF increased cyclic GMP content up to 22.9 and 42.3 fold respectively.

In SHR ECs, α‐thrombin‐induced calcium transient was not modified by SNAP, ANF or eNOS inhibition. SNAP (100 μM) and 0.1 μM ANF increased cyclic GMP content up to 9.3 and 51 fold respectively.

In WKY ECs, SNAP dose‐dependently (10–100 μM) reduced also bradykinin‐induced calcium transient, while in SHR ECs was ineffective.

We concluded that in SHR ECs, the cyclic GMP‐dependent regulation of calcium transient is lost.

British Journal of Pharmacology (2000) 130, 1468–1476; doi:10.1038/sj.bjp.0703453
Vanilloid receptors on capsaicin‐sensitive sensory nerves mediate relaxation to methanandamide in the rat isolated mesenteric arterial bed and small mesenteric arteries
Authors: Ralevic V , Kendall D A , Randall M D , Zygmunt P M , Movahed P , Högestätt E D
Published: January 2005

In the present study, the vasodilator actions of methanandamide and capsaicin in the rat isolated mesenteric arterial bed and small mesenteric arterial segments were investigated.

Methanandamide elicited concentration‐dependent relaxations of preconstricted mesenteric arterial beds (pEC50=6.0±0.1, Emax=87±3%) and arterial segments (pEC50=6.4±0.1, Emax=93±3%).

In arterial beds, in vitro capsaicin pre‐treatment blocked vasorelaxation to 1 and 3 μM methanandamide, and reduced to 12±7% vasorelaxation to 10 μM methanandamide. Methanandamide failed to relax arterial segments pre‐treated in vitro with capsaicin.

In arterial beds from rats treated as neonates with capsaicin to cause destruction of primary afferent nerves, methanandamide at 1 and 3 μM did not evoke vasorelaxation, and relaxation at 10 μM methanandamide was reduced to 26±4%.

Ruthenium red (0.1 μM), an inhibitor of vanilloid responses, attenuated vasorelaxation to methanandamide in arterial beds (pEC50=5.6±0.1, Emax=89±1%). Ruthenium red at 1 μM abolished the response to 1 μM methanandamide, and greatly attenuated relaxation at 3 and 10 μM methanandamide in arterial beds. In arterial segments, ruthenium red (0.15 μM) blocked vasorelaxation to methanandamide, but not to CGRP.

In arterial segments, the vanilloid receptor antagonist capsazepine (1 μM) inhibited, and the calcitonin gene‐related peptide (CGRP) receptor antagonist CGRP8–37 (3 μM) abolished, methanandamide‐induced relaxations. CGRP8–37, but not capsazepine, attenuated significantly relaxation to exogenous CGRP.

These data show that capsaicin and ruthenium red attenuate vasorelaxation to methanandamide in the rat isolated mesenteric arterial bed and small mesenteric arterial segments. In addition, CGRP8–37 and capsazepine antagonize responses to methanandamide in mesenteric arterial segments. In conclusion, vanilloid receptors on capsaicin‐sensitive sensory nerves play an important role in the vasorelaxant action of methanandamide in the rat isolated mesenteric arterial bed and small mesenteric arterial segments.

British Journal of Pharmacology (2000) 130, 1483–1488; doi:10.1038/sj.bjp.0703456
Characterization of [125I]‐SB‐258585 binding to human recombinant and native 5‐HT6 receptors in rat, pig and human brain tissue
Authors: Hirst Warren D , Minton Jayne A L , Bromidge Steven M , Moss Stephen F , Latter Alison J , Riley Graham , Routledge Carol , Middlemiss Derek N , Price Gary W
Published: January 2005

SB‐258585 (4‐Iodo‐N‐[4‐methoxy‐3‐(4‐methyl‐piperazin‐1‐yl)‐phenyl]‐benzenesulphonamide) is a high affinity ligand at 5‐HT6 receptors. It displays over 100 fold selectivity for the 5‐HT6 receptor over all other 5‐HT receptors tested so far. SB‐258585 has been radiolabelled, to high specific activity, for its characterization as a 5‐HT6 receptor selective radioligand.

[125I]‐SB‐258585 bound, with high affinity, to a single population of receptors in a cell line expressing human recombinant 5‐HT6 receptors. Kinetic and saturation binding experiments gave pKD values of 9.01±0.09 and 9.09±0.02, respectively.

In membranes derived from rat or pig striatum and human caudate putamen, [125I]‐SB‐258585 labelled a single site with high levels (>60%) of specific binding. Saturation analysis revealed pKD values of 8.56±0.07 for rat, 8.60±0.10 for pig and 8.90±0.02 for human. Bmax values for the tissues ranged from 173±23 and 181±25 fmol mg−1 protein in rat and pig striatum, respectively, to 215±41 fmol mg−1 protein in human caudate putamen.

The pKi rank order of potency for a number of compounds, determined in competition binding assays with [125I]‐SB‐258585, at human caudate putamen membranes was: SB‐271046>SB‐258585>SB‐214111>methiothepin>clozapine>5‐Me‐OT>5‐HT>Ro 04‐6790>mianserin>ritanserin=amitriptyline>5‐CT>mesulergine. Similar profiles were obtained from pig and rat striatal membranes and recombinant 5‐HT6 receptors; data from the latter correlated well with [3H]‐LSD binding.

Thus, [125I]‐SB‐258585 is a high affinity, selective radioligand which can be used to label both recombinant and native 5‐HT6 receptors and will facilitate further characterization of this receptor subtype in animal and human tissues.

British Journal of Pharmacology (2000) 130, 1597–1605; doi:10.1038/sj.bjp.0703458
Structural factors that determine the ability of adenosine and related compounds to activate the cardiac ryanodine receptor
Authors: Chan Wei Mun , Welch William , Sitsapesan Rebecca
Published: January 2005

The effects of adenosine and adenine on the gating of native sheep cardiac ryanodine receptor (RyR) channels were investigated. By examining the mechanisms underlying channel activation and by using comparative molecular field analysis (CoMFA) we have investigated the structural features of adenine‐based ligands involved in channel activation.

In the presence of 10 μM cytosolic Ca2+, adenosine and adenine both activate the channel but only to a level approximately 10 and 20% respectively of that of ATP indicating that both are partial agonists of low efficacy.

Adenosine was able to antagonize the ATP‐induced increase in open probability (Po) as expected for a partial agonist of low efficacy at the ATP sites on the cardiac RyR.

GTP (100 μM–10 mM) had no effect on channel gating indicating that the adenine ring structure is important for agonist activity at the ATP‐sites on RyR.

CoMFA revealed an extremely strong correlation between the structural features of the five ATP analogues and the ability to increase (Po). Our model indicates that the high efficacy of ATP results primarily from the large electrostatic field established by the ionized phosphate groups. Reducing the number of phosphate groups lowers the strength of this field, leading to ligands with lower efficacy. In addition, steric interactions between the α‐phosphate and ribose moieties and the RyR are correlated with low Po.

British Journal of Pharmacology (2000) 130, 1618–1626; doi:10.1038/sj.bjp.0703459
Pharmacological characterization of metabotropic glutamate receptor‐mediated high‐affinity GTPase activity in rat cerebral cortical membranes
Authors: Nishi Nobuyuki , Odagaki Yuji , Koyama Tsukasa
Published: January 2005

Activation of heterotrimeric guanine nucleotide‐binding regulatory proteins (G‐proteins) functionally coupled to metabotropic glutamate receptors (mGluRs) was assessed by agonist‐induced high‐affinity GTPase (EC3.6.1.‐) activity in rat cerebral cortical membranes.

L‐Glutamate (1 mM) stimulated high‐affinity GTPase activity to the same extent throughout the incubation period up to 20 min, in a Mg2+‐dependent manner. The addition of 1 mM L‐glutamate augmented Vmax of the enzyme activity (1670 to 3850 pmol mg−1 protein 15 min−1) with slight increase in KM value (0.26 to 0.63 μM).

The high‐affinity GTPase activity was stimulated by the following compounds with a rank order of potency of (2S,2′R,3′R)‐2‐(2′,3′‐dicarboxycyclopropyl) glycine (DCG‐IV) >  (2S,1′S,2′S)‐2‐(carboxycyclopyropyl)glycine (L‐CCG‐I) > L‐glutamate ≥ 2R,4R‐4‐aminopyrrolidine‐2,4‐dicarboxylate [(2R,4R)‐APDC] > 1S,3R‐1‐aminocyclopentane‐1,3‐dicarboxylate [(1S,3R)‐ACPD] > (S)‐4‐carboxy‐3‐hydroxyphenylglycine [(S)‐4C3HPG] > (S)‐3‐carboxy‐4‐hydroxyphenylglycine [(S)‐3C4HPG] > ibotenate, but not by L‐(+)‐2‐amino‐4‐phosphonobutyrate (L‐AP4), (RS)‐3,5‐dihydroxyphenylglycine [(RS)‐3,5‐DHPG], quisqualate, or L‐serine‐O‐phosphate (L‐SOP), indicative of involvement of group II mGluRs, in particular mGluR2.

(2S)‐α‐Ethylglutamate (EGLU), a presumably selective antagonist against group II mGluRs, inhibited DCG‐IV‐stimulated high‐affinity GTPase activity in a competitive manner with an apparent KB of 220 μM.

L‐Glutamate‐stimulated activity was eliminated by pretreatment of the membranes with sulfhydryl alkylating agent N‐ethylmaleimide (NEM) at 30–50 μM, indicating that G‐proteins of the Gi family are involved.

These results indicate that mGluR agonist‐induced high‐affinity GTPase activity in rat cerebral cortical membranes may be used to detect the functional interaction between group II mGluRs, in particular mGluR2, and NEM‐sensitive Gi proteins.

British Journal of Pharmacology (2000) 130, 1664–1670; doi:10.1038/sj.bjp.0703464
Cytochrome P450 mono‐oxygenase gene expression and protein activity in cultures of adult cardiomyocytes of the rat
Authors: Thum Thomas , Borlak Jürgen
Published: January 2005

There are a substantial number of drugs acting either directly or indirectly on the heart, but surprisingly, little is known about the metabolic capacity of heart muscle cells.

We therefore investigated the gene expression and protein activity of cytochrome P450 isozymes in cultures of adult cardiomyocytes of the rat.

Semi‐quantitative CYP gene expression pattern suggests CYP1A1 and CYP2B1/2 to be key players in cardiomyocytes and upon treatment with Aroclor 1254 approximate 4 fold inductions could be observed for both gene families, when compared with appropriate controls.

The mRNA expression of most genes was sustained for prolonged periods of time, e.g. up to 120 h in culture and in the case of the CYP3A1 gene an approximate 10 fold induction was observed at the higher Aroclor 1254 dose level (10 μM) in 24 h old cultures.

The constitutively expressed genes, e.g. CYP2C11 and CYP2E1 are expressed throughout the entire culture period (5 days) and did not respond to Aroclor 1254 treatment.

CYP4A1 was mainly expressed in freshly isolated cardiomyocytes of control animals and its expression declined rapidly in culture.

There was good agreement between gene expression and translated protein activity using 7‐ethoxyresorufin and testosterone as substrates.

The data reported herein should foster the routine use of freshly isolated and cultivated cardiomyocytes for drug profiling and toxicity studies.

British Journal of Pharmacology (2000) 130, 1745–1752; doi:10.1038/sj.bjp.0703465
LU135252, an endothelinA receptor antagonist did not prevent pulmonary vascular remodelling or lung fibrosis in a rat model of myocardial infarction
Authors: Nguyen Quang Trinh , Colombo Federico , Rouleau Jean L , Dupuis Jocelyn , Calderone Angelino
Published: January 2005

The early intervention with endothelinA (ETA) receptor antagonists following coronary artery ligation has been shown to reduce the development of pulmonary hypertension, despite a lack of improvement in left ventricular function.

The present study examined the contribution of pulmonary vascular remodelling and the progression of lung fibrosis in the development of pulmonary hypertension and the subsequent role of endothelin‐1 in these processes in a rat model of myocardial infarction (MI).

The administration of 60 mg kg−1 per day of the specific ETA receptor antagonist LU135253 ((+)‐(S)‐2‐(4,6‐dimethoxy‐pyrimidin‐2‐yloxy)‐3‐methoxy‐3,3‐diphenyl‐propionic acid) 24 h following coronary artery ligation, failed to improve left ventricular contractile indices, but reduced the extent of pulmonary hypertension, as reflected by the significant decrease in right ventricular systolic pressure.

The medial wall thickness of small pulmonary arteries (50–200 μm) was significantly increased 4 weeks following MI, albeit LU135253 treatment did not ameliorate this pattern of vascular remodelling.

The steady‐state mRNA levels of collagen, fibronectin, transforming growth factor‐β1, and ‐β3 were significantly increased in the lungs of MI rats. The treatment with LU135252 did not alter this pattern of gene expression.

Thus, these data demonstrate pulmonary vascular remodelling and the increased expression of extracellular matrix proteins represent underlying mechanisms implicated in the development of pulmonary hypertension in the MI rat.

Despite the amelioration of the pulmonary hypertensive state, ETA receptor blockade was insufficient to reverse pulmonary vascular remodelling, or the development of lung fibrosis in the MI rat.

British Journal of Pharmacology (2000) 130, 1525–1530; doi:10.1038/sj.bjp.0703466
Cytotoxicity associated with induction of nitric oxide synthase in rat duodenal epithelial cells in vivo by lipopolysaccharide of Helicobacter pylori: inhibition by superoxide dismutase
Authors: Lamarque Dominique , Moran Anthony P , Szepes Zoltan , Delchier Jean‐Charles , Whittle Brendan J R
Published: January 2005

The products released by Helicobacter pylori (H. pylori) in the gastric antral and duodenal mucosa may be involved in mucosal ulceration by stimulating the local formation of cytotoxic factors such as nitric oxide (NO), superoxide or peroxynitrite.

The present study investigates the ability of purified H. pylori lipopolysaccharide (LPS) to induce nitric oxide synthase (iNOS) in rat duodenal epithelial cells following in vivo challenge and its interaction with superoxide in promoting cellular damage and apoptosis.

H. pylori LPS (0.75–3 mg kg−1 i.v. or 3–12 mg kg−1 p.o.) induced a dose–dependent expression of iNOS activity after 5 h in the duodenal epithelial cells, determined by [14C] arginine conversion to citrulline.

The epithelial cell viability, as assessed by Trypan Blue exclusion and MTT conversion, was reduced 5 h after challenge with H. pylori LPS, while the incidence of apoptosis was increased.

The iNOS activity and reduction in cell viability following H. pylori LPS challenge i.v. was inhibited by the selective iNOS inhibitor, 1400 W (0.2–5 mg kg−1 i.v.).

Concurrent administration of superoxide dismutase conjugated with polyethylene glycol (250–500 i.u. kg−1, i.v.), which did not modify the cellular iNOS activity, reduced the epithelial cell damage provoked by i.v. H. pylori LPS, and abolished the increased incidence of apoptosis.

These results suggest that expression of iNOS following challenge with H. pylori LPS provokes duodenal epithelial cell injury and apoptosis by a process involving superoxide, implicating peroxynitrite involvement. These events may contribute to the pathogenic mechanisms of H. pylori in promoting peptic ulcer disease.

British Journal of Pharmacology (2000) 130, 1531–1538; doi:10.1038/sj.bjp.0703468
Ketamine potentiates cerebrocortical damage induced by the common anaesthetic agent nitrous oxide in adult rats
Authors: Jevtovic‐Todorovic Vesna , Benshoff Nicholas , Olney John W
Published: January 2005

For general anaesthesia, patients usually receive a combination of drugs, all of which are classified as γ‐amino‐butyric acid (GABA) agonists, with two notable exceptions – ketamine and nitrous oxide (laughing gas, N2O) – which are antagonists of N‐methyl‐D‐aspartate (NMDA) glutamate receptors. At clinically relevant doses both ketamine and N2O, like other NMDA antagonists, have the potential to induce psychotomimetic reactions in humans and to cause pathomorphological changes in cerebrocortical neurons in rat brain. Because drug combinations used in clinical anaesthesia sometimes include both ketamine and N2O, we undertook experiments to evaluate whether augmented neurotoxicity results from their combined use.

Ketamine and N2O were administered alone or in combination by various dosing regimens to adult female rats for a duration of 3 h and the severity of cerebrocortical neurotoxic changes was quantified histologically. Because GABA agonists are known to protect against the psychotomimetic and neurotoxic effects of NMDA antagonists, we also evaluated whether the combined neurotoxicity of ketamine+N2O can be prevented by certain commonly used GABA agonists.

When ketamine and N2O were used in combination the neurotoxic reaction was enhanced to a degree much greater than can be explained by simple additivity. The apparent synergistic interaction was particularly striking when low doses of the agents were combined, the degree of toxic synergism at higher doses being masked by a ceiling effect. GABA agonists protected against ketamine/N2O neurotoxicity.

It is recommended that this information be taken into consideration in the selection of drugs to be used in multi‐agent protocols for general anaesthesia.

British Journal of Pharmacology (2000) 130, 1692–1698; doi:10.1038/sj.bjp.0703479