bluuebunny/crossref_metadata_2025_split huggingface dataset:
Angiotensin II responses of vascular smooth muscle cells from hypertensive rats: enhancement at the level of p42 and p44 mitogen activated protein kinase
Authors: Wilkie Neil , Ng Leong L. , Boarder Michael R. 
Published: January 2005

Stimulation of the AT1 receptor by angiotensin II (AII) gives a larger mitogenic response in vascular smooth muscle cells from spontaneously hypertensive rats (SHR) compared to those from normotensive (WKY) controls. Here we investigated whether the p42 and p44 mitogen activated protein kinase (MAPK) pathway is differentially regulated in these cells by AT1 receptors.
We showed that there is a similar level of p42 and p44 MAPK immunoreactivity in the SHR and WKY derived cells.
However, by use of an antiserum specific for the tyrosine phosphorylated form of MAPK, and an assay with a nonapeptide MAPK substrate, we showed that AII (100 nM)‐stimulated phosphorylation and activation of p42mapk and p44mapk are enhanced in the SHR derived cells.
This increased MAPK activity in SHR derived cells was also seen on protein kinase C activation with 100 nM phorbol myristate acetate (PMA). The size and time course of the response to PMA was the same as that to AII in each cell type.
The protein kinase C inhibitor Ro 31–8220 attenuated the early (2 min) phase of AII stimulation of MAPK activity and the entire stimulation caused by PMA. At longer times of AII stimulation both p42mapk and p44mapk were activated by an Ro 31–8220‐insensitive mechanism.
Agonist or PMA stimulation of MAPK activity was inhibited by the tyrosine kinase inhibitor genistein. AII stimulated tyrosine protein phosphorylation to a greater degree in SHR than WKY cells.
These results show that the MAPK response of SHR derived cells is increased over that of WKY cells by mechanisms independent of the enhanced stimulation of phospholipase C; amplification at the level of sequential protein kinase C and tyrosine kinase steps leads to the enhanced responsiveness of MAPK in the SHR derived cells.

British Journal of Pharmacology (1997) 122, 209–216; doi:10.1038/sj.bjp.0701366
Negative chronotropic actions of endothelin‐1 on rabbit sinoatrial node pacemaker cells
Authors: Tanaka Hideo , Habuchi Yoshizumi , Yamamoto Taku , Nishio Manabu , Morikawa Junichiro , Yoshimura Manabu 
Published: January 2005

The effects of endothelin‐1 (ET‐1) on sinoatrial (SA) node preparations of the rabbit heart were studied by means of whole‐cell clamp techniques.
ET‐1 at 1 nM slowed the spontaneous beating activity and rendered half of the cells quiescent. At a higher concentration of 10 nM, the slowing and cessation of spontaneous activity were accompanied by hyperpolarization.
In voltage‐clamp experiments, ET‐1 decreased the basal L‐type Ca2+ current (ICa(L)) dose‐dependently with a half‐maximal inhibitory concentration (EC50) of 0.42 nM and maximal inhibitory response (Emax) of 49.5%. The delayed rectifying K+ current (IK) was also reduced by 33.2±11.1% at 1 nM. In addition, an inwardly rectifying K+ current was activated by ET‐1 at higher concentrations (EC50=4.8 nM). These ET‐1‐induced changes in membrane currents were abolished by BQ485 (0.3 μM), a highly selective ETA receptor antagonist.
When ICa(L) was inhibited by ET‐1 (1 nM), subsequent application of 10 μM ACh showed no additional decrease in ICa(L), suggesting the involvement of cyclic AMP in the effects of ET‐1 on ICa(L). In contrast, 1 nM ET‐1 further decreased ICa(L) in the presence of 10 μM ACh, suggesting that ET‐1 activates some additional mechanism(s) which inhibit ICa(L). The ET‐1‐induced ICa(L) inhibition was abolished by protein kinase A inhibitory peptide (PKI, 20 μM) or H‐89 (5 μM). However, the ICa(L) inhibition was not affected by methylene blue (10 μM), suggesting a minor role for cyclic GMP in the effect of ET‐1 under basal conditions.
ET‐1 failed to inhibit ICa(L) when the pipette contained GDPβS (200 μM). However, incubation of the cells with pertussis toxin (PTX, 5 μg ml−1, >6 h) only reduced the ET‐1‐induced inhibition to 21.5±9.5%, whereas it abolished the inhibitory effect of ACh on ICa(L).
Intracellular perfusion of 8‐bromo cyclicAMP (8‐Br cyclicAMP, 500 μM) attenuated, but did not abolish the inhibitory effect of ET‐1 on ICa(L). This 8‐Br cyclicAMP‐resistant component (17.5±14.4%, n=20) was not affected by combined application of 8‐Br cyclicAMP with 8‐bromo cyclicGMP (500 μM), ryanodine (1 μM) or phorbol‐12‐myristate‐13‐acetate (TPA; 50 nM).
In summary, ET‐1 exerts negative chronotropic effects on the SA node via ETA‐receptors. ET‐1 inhibits both ICa(L) and IK, and increases background K+ current. The inhibition of ICa(L) by ET‐1 is mainly due to reduction of the cyclicAMP levels via PTX‐sensitive G protein, but some other mechanism(s) also seems to be operative.

British Journal of Pharmacology (1997) 122, 321–329; doi:10.1038/sj.bjp.0701370
Effects of increased extracellular glutamate levels on the local field potential in the brain of anaesthetized rats
Authors: Obrenovitch T. P. , Urenjak J. , Zilkha E. 
Published: January 2005

It is generally considered that glutamate‐mediated transmission can be altered from a physiological to neurotoxic action when extracellular glutamate levels become excessive subsequent to impaired uptake and/or excessive release. However, high extracellular glutamate does not consistently correlate with neuronal dysfunction and death in vivo. The purpose of this study was to examine in situ the local depolarizations, as indicated by negative shifts of the extracellular field (d.c.) potential, produced by local inhibition of high‐affinity glutamate uptake, with or without co‐application of exogenous glutamate, in three brain regions of anaesthetized rats.
Microdialysis probes incorporating an electrode were used to apply exogenous glutamate and/or its uptake inhibitor L‐trans‐pyrrolidine‐2,4‐dicarboxylate (L‐trans‐PDC), and to monitor the resulting changes in extracellular glutamate and d.c. potential at the sites of application within the cortex, striatum and hippocampus.
Perfusion of 1 to 10 mM L‐trans‐PDC markedly and concentration‐dependently increased extracellular glutamate levels (by up to 1700% of basal level in the parietal cortex). Despite their large magnitude, glutamate changes were associated with minor negative shifts of the d.c. potential (<2 mV), which were not suppressed by the N‐methyl‐D‐aspartate (NMDA)‐channel blocker, dizocilpine (MK‐801, 2 mg kg−1, i.v.), or the α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazole‐propionic acid (AMPA)/kainate‐receptor antagonist, 6‐nitro‐7‐sulphamoylbenzo(f)quinoxaline‐2,3‐dione (NBQX, 30 mg kg−1, i.p.). L‐trans‐PDC had virtually identical concentration‐dependent effects on dialysate glutamate in the hippocampus and striatum, but those induced in the cortex were around 40% larger (P<0.002). In contrast, the associated depolarizations were around twice as large in the striatum and cortex as in the hippocampus (P<0.002). Finally, co‐application of L‐trans‐PDC did not enhance the d.c. potential changes evoked by perfusion of 5 or 20 mM glutamate.
As the neurotoxic potency of glutamate agonists is considered to be linked to excessive opening of glutamate‐operated ion channels, these results challenge the notion that high extracellular glutamate levels may be the key to excitotoxicity in neurological disorders. In particular, they do not support the hypothesis that high extracellular glutamate causes the sudden negative shifts of the d.c. potential associated with ischaemia (i.e. anoxic depolarization), traumatic brain injury and spreading depression. Impaired uptake and excessive release of glutamate may well lead to excitotoxicity, but only at the synaptic level, not by spreading through the interstitial fluid.

British Journal of Pharmacology (1997) 122, 372–378; doi:10.1038/sj.bjp.0701372
Potent and voltage‐dependent block by philanthotoxin‐343 of neuronal nicotinic receptor/channels in PC12 cells
Authors: Liu Min , Nakazawa Ken , Inou Kazuhide , Ohno Yasuo 
Published: January 2005

Block by philanthotoxin‐343 (PhTX‐343), a neurotoxin from wasps, of ionic currents mediated through neuronal nicotinic acetylcholine (ACh) receptor/channels was characterized in rat phaeochromocytoma PC12 cells, by use of whole cell voltage‐clamp techniques.
In the cells held at −60 mV, PhTX‐343 at 0.1 and 1 μM inhibited an inward current activated by 100 μM ACh. The current inhibition was relieved by depolarizing steps, and augmented at negative potentials, suggesting that PhTX‐343 blocks the channel in a voltage‐dependent manner. Joro spider toxin‐3 (JSTX‐3) also exerted voltage‐dependent inhibition of ACh‐activated currents in a similar concentration range, but argiotoxin636 did not affect the currents.
Analysis of the current decay during hyperpolarizing steps indicated that the current inhibition by 100 nM PhTX‐343 develops in an order of several hundreds of milliseconds. On the other hand, the recovery from the current inhibition during depolarizing steps developed in an order of about 100 ms.
The results suggest that PhTX‐343 blocks neuronal nicotinic receptor channels in PC12 cells at concentrations lower than those required for channel block in non‐mammalian cells, and the block exhibits clear voltage‐dependence. Estimated from the voltage‐dependence, the binding site of PhTX‐343 may be located near the outer mouth of the channel.

British Journal of Pharmacology (1997) 122, 379–385; doi:10.1038/sj.bjp.0701373
Effect of acute and chronic tramadol on [3H]‐5‐HT uptake in rat cortical synaptosomes
Authors: Giusti Pietro , Buriani Alessandro , Cima Lorenzo , Lipartiti Maria 
Published: January 2005

Tramadol hydrochloride is a centrally acting opioid analgesic, the efficacy and potency of which is only five to ten times lower than that of morphine. Opioid, as well as non‐opioid mechanisms, may participate in the analgesic activity of tramadol.
[3H]‐5‐hydroxytryptamine (5‐HT) uptake in rat isolated cortical synaptosomes was studied in the presence of tramadol, desipramine, fluoxetine, methadone and morphine. Methadone and tramadol inhibited synaptosomal [3H]‐5‐HT uptake with apparent Kis of 0.27±0.04 and 0.76±0.04 μM, respectively. Morphine essentially failed to inhibit [3H]‐5‐HT uptake (Ki 0.50±0.30 M).
Methadone, morphine and tramadol were active in the hot plate test with ED50s of 3.5, 4.3 and 31 mg kg−1, respectively. At the highest tested dose (80 mg kg−1) tramadol produced only 77±5.3% of the maximal possible effect.
When [3H]‐5‐HT uptake was examined in synaptosomes prepared from rats 30 min after a single dose of morphine, methadone or tramadol, only tramadol (31 mg kg−1, s.c., equal to the ED50 in the hot plate test) and methadone (35 mg kg−1, s.c., equal to the ED90 in the hot plate test) decreased uptake.
Animals were chronically treated for 15 days with increasing doses of tramadol or methadone (5 to 40 mg kg−1 and 15 to 120 mg kg−1, s.c., respectively). Twenty‐four hours after the last drug injection, a challenge dose of methadone (35 mg kg−1, s.c.) or tramadol (31 mg kg−1, s.c.) was administered. [3H]‐5‐HT uptake was not affected in synaptosomes prepared from rats chronically‐treated with methadone, whereas chronic tramadol was still able to reduce this parameter by 42%.
Rats chronically‐treated with methadone showed a significant increase in [3H]‐5‐HT uptake (190%) 72 h after drug withdrawal. In contrast, [3H]‐5‐HT uptake in rats chronically‐treated with tramadol (110%) did not differ significantly from control animals.
These results further support the hypothesis that [3H]‐5‐HT uptake inhibition may contribute to the antinociceptive effects of tramadol. The lack of tolerance development of [3H]‐5‐HT uptake, together with the absence of behavioural alterations after chronic tramadol treatment, suggest that tramadol has an advantage over classical opioids in the treatment of pain disorders.

British Journal of Pharmacology (1997) 122, 302–306; doi:10.1038/sj.bjp.0701374
Interaction of cyclosporin derivatives with the ATPase activity of human P‐glycoprotein
Authors: Watanabe Toru , Kokubu Noriko , Charnick Steven B. , Naito Mikihiko , Tsuruo Takashi , Cohen Dalia 
Published: January 2005

P‐glycoprotein, a 170–180 kDa membrane glycoprotein that mediates multidrug resistance, hydrolyses ATP to efflux a broad spectrum of hydrophobic agents. In this study, we analysed the effects of three MDR reversing agents, verapamil, cyclosporin A and [3′‐keto‐Bmt]‐[Val*]‐cyclosporin (PSC 833), on the adenosine triphosphatase (ATPase) activity of human P‐glycoprotein.
P‐glycoprotein was immunoprecipitated with a monoclonal antibody (MRK‐16) and the P‐glycoprotein‐MRK‐16‐Protein A‐Sepharose complexes obtained were subjected to a coupled enzyme ATPase assay.
While verapamil activated the ATPase, the cyclosporin derivatives inhibited both the substrate‐stimulated and the basal P‐glycoprotein ATPase. No significant difference was observed between PSC 833 and cyclosporin A on the inhibition of basal P‐glycoprotein ATPase activity. PSC 833 was more potent than cyclosporin A for the substrate‐stimulated activity.
Kinetic analysis indicated a competitive inhibition of verapamil‐stimulated ATPase by PSC 833.
The binding of 8‐azido‐[α‐32P]‐ATP to P‐glycoprotein was not altered by the cyclosporin derivatives, verapamil, vinblastine and doxorubicin, suggesting that the modulation by these agents of P‐glycoprotein ATPase cannot be attributed to an effect on ATP binding to P‐glycoprotein.
The interaction of the cyclosporin derivatives with ATPase of P‐glycoprotein might present an alternative and/or additional mechanism of action for the modulation of P‐glycoprotein function.

British Journal of Pharmacology (1997) 122, 241–248; doi:10.1038/sj.bjp.0701377
Characterization of A2A adenosine receptors in human lymphocyte membranes by [3H]‐SCH 58261 binding
Authors: Varani Katia , Gessi Stefania , Dalpiaz Alessandro , Ongini Ennio , Andrea Borea Pier 
Published: January 2005

The present study describes for the first time the characterization of the adenosine A2A receptor in human lymphocyte membranes with the new potent and selective antagonist radioligand, [3H]‐5‐amino‐7‐(2‐phenylethyl)‐2‐(2‐furyl)‐pyrazolo [4,3‐e]‐1,2,4 triazolo [1,5‐c] pyrimidine, ([3H]‐SCH 58261). In addition, both receptor affinity and potency of reference adenosine receptor agonists and antagonists were determined in binding and adenylyl cyclase studies.
Saturation experiments revealed a single class of binding sites with Kd and Bmax values of 0.85 nM and 35 fmol mg−1 protein, respectively. A series of adenosine receptor ligands were found to compete for the binding of 0.8 nM [3H]‐SCH 58261 to human lymphocyte membranes with a rank order of potency consistent with that typically found for interactions with the A2A‐adenosine receptor. In the adenylyl cyclase assay the same compounds exhibited a rank order of potency similar to that observed in binding experiments.
Thermodynamic data indicate that [3H]‐SCH 58261 binding to human lymphocytes is entropy and enthalpy‐driven, a finding in agreement with the thermodynamic behaviour of antagonists for rat striatal A2A‐adenosine receptors.
It is concluded that in human lymphocyte membranes [3H]‐SCH 58261 directly labels binding sites showing the characteristic properties of the adenosine A2A‐receptor. The presence of A2A‐receptors in peripheral tissue such as human lymphocytes strongly suggests an important role for adenosine in modulating immune and inflammatory responses.

British Journal of Pharmacology (1997) 122, 386–392; doi:10.1038/sj.bjp.0701378
The effect of 15‐HPETE on airway responsiveness and pulmonary cell recruitment in rabbits
Authors: Riccio M. M. , Matsumoto T. , Adcock J. J. , Douglas G. J. , Spina D. , Page C. P. 
Published: January 2005

In the present study we have investigated the effect of 15‐hydroperoxyeicosatetraenoic acid (15‐HPETE) and 15‐hydroxyeicosatetraenoic acid (15‐HETE) on airway responsiveness to inhaled histamine in rabbits in vivo.
15‐HPETE increased airway responsiveness to histamine 24 h after tracheal instillation and this was associated with a cellular infiltration consisting mainly of neutrophils, as measured by bronchoalveolar lavage. The airway hyperresponsiveness induced by 15‐HPETE was still present 72 h after tracheal instillation of 15‐HPETE, but had returned to baseline values one week post challenge. The number of neutrophils in bronchoalveolar lavage remained significantly elevated compared to pre‐challenge levels. In contrast to 15‐HPETE, the major metabolite 15‐HETE, failed to alter airway hyperresponsiveness to histamine despite the recruitment of neutrophils into the lung, suggesting that the effect of 15‐HPETE was not secondary to the generation of this metabolite nor dependent on the influx of neutrophils.
Both capsaicin and atropine but not the peripherally acting μ‐opioid receptor agonist, BW443C (H‐Tyr‐D‐Arg‐Gly‐Phe(4‐NO2)‐Pro‐NH4), attenuated 15‐HPETE‐induced hyperresponsiveness. The increased cellular infiltration induced by 15‐HPETE was only attenuated by capsaicin.
The results of the present study suggest that the release of 15‐HPETE into the airways could contribute to sensitization of afferent nerve endings analogous to the hyperalgesia induced by this mediator in skin.

British Journal of Pharmacology (1997) 122, 249–256; doi:10.1038/sj.bjp.0701379
A toxin from the spider Phoneutria nigriventer that blocks calcium channels coupled to exocytosis
Authors: Guatimosim C. , Romano‐Silva M. A. , Cruz J. S. , Beirão P. S. L. , Kalapothakis E. , Moraes‐Santos T. , Cordeiro M. N. , Diniz C. R. , Gomez M. V. , Prado M. A. M. 
Published: January 2005

The aim of the present experiments was to investigate the pharmacological action of a toxin from the spider Phoneutria nigriventer, Tx3‐3, on the function of calcium channels that control exocytosis of synaptic vesicles.
Tx3‐3, in confirmation of previous work, diminished the intracellular calcium increase induced by membrane depolarization with KCl (25 mM) in rat cerebrocortical synaptosomes. The toxin was very potent (IC50 0.9 nM) at inhibiting calcium channels that regulate calcium entry in synaptosomes. In addition, Tx3‐3 blocked the exocytosis of synaptic vesicles, as measured with the fluorescent dye FM1‐43.
Using ω‐toxins that interact selectively with distinct neuronal calcium channels, we investigated whether the target of Tx3‐3 overlaps with known channels that mediate exocytosis. The results indicate that the main population of voltage‐sensitive calcium channels altered by Tx3‐3 can also be inhibited by ω‐agatoxin IVA, an antagonist of P/Q calcium channels. ω‐conotoxin GVIA, which inhibits N type calcium channels did not decrease significantly the entry of calcium or exocytosis of synaptic vesicles in depolarized synaptosomes.
It is concluded that Tx3‐3 potently inhibits ω‐agatoxin IVA‐sensitive calcium channels, which are involved in controlling exocytosis in rat brain cortical synaptosomes.

British Journal of Pharmacology (1997) 122, 591–597; doi:10.1038/sj.bjp.0701381
Effect of chronic treatment with the GABA transaminase inhibitors γ‐vinyl GABA and ethanolamine O‐sulphate on the in vitro GABA release from rat hippocampus
Authors: Qume M. , Fowler L. J. 
Published: January 2005

The effects of 2, 8 and 21 day oral treatment with the specific γ‐aminobutyric acid transaminase (GABA‐T) inhibitors γ‐vinyl GABA (GVG) and ethanolamine O‐sulphate (EOS) on brain GABA levels, GABA‐T activity, and basal and stimulated GABA release from rat cross‐chopped brain hippocampal slices was investigated.
Treatment with GABA‐T inhibitors lead to a reduction in brain GABA‐T activity by 65–80% compared with control values, with a concomitant increase in brain GABA content of 40–100%.
Basal hippocampal GABA release was increased to 250–450% of control levels following inhibition of GABA‐T activity. No Ca2+ dependence was observed in either control or treated tissues.
GVG and EOS administration led to a significant elevation in the potassium stimulated release of GABA from cross‐chopped hippocampal slices compared with that of controls. Although stimulated GABA release from control tissues was decreased in the presence of a low Ca2+ medium, GVG and EOS treatment abolished this Ca2+ dependency.
GABA compartmentalization, Na+ and Cl− coupled GABA uptake carriers and glial release may provide explanations for the loss of the Ca2+ dependency of stimulated GABA release observed following GVG and EOS treatment.
Administration of GABA‐T inhibitors led to increases in both basal and stimulated hippocampal GABA release. However, it is not clear which is the most important factor in the anticonvulsant activity of these drugs, the increased GABA content ‘leaking’ out of neurones and glia leading to widespread inhibition, or the increase in stimulated GABA release which may occur following depolarization caused by an epileptic discharge.

British Journal of Pharmacology (1997) 122, 539–545; doi:10.1038/sj.bjp.0701383
Role of ATP in fast excitatory synaptic potentials in locus coeruleus neurones of the rat
Authors: Nieber Karen , Poelchen Wolfgang , Illes Peter 
Published: January 2005

Intracellular recordings were made in a pontine slice preparation of the rat brain containing the nucleus locus coeruleus (LC). The pressure application of α,β‐methylene ATP (α,β‐meATP) caused reproducible depolarizations which were depressed by suramin (30 μM) and abolished by suramin (100 μM). Pyridoxal‐phosphate‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS; 10, 30 μM) also concentration‐dependently inhibited the α,β‐meATP‐induced depolarization, although with a much slower time‐course than suramin. Almost complete inhibition developed with 30 μM PPADS. Reactive blue 2 (30 μM) did not alter the effect of α,β‐meATP, while reactive blue 2 (100 μM) slightly depressed it.
Pressure‐applied (S)‐α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA) also depolarized LC neurones. Kynurenic acid (500 μM) depressed and 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (CNQX; 50 μM) abolished the response to AMPA. Suramin (100 μM) potentiated the AMPA effect.
Pressure‐applied noradrenaline hyperpolarized LC neurones. Suramin (100 μM) did not alter the effect of noradrenaline.
Focal electrical stimulation evoked biphasic synaptic potentials consisting of a fast depolarization (p.s.p.) followed by a slow hyperpolarization (i.p.s.p.). A mixture of D(−)‐2‐amino‐5‐phosphonopentanoic acid (AP‐5; 50 μM), CNQX (50 μM) and picrotoxin (100 μM) depressed both the p.s.p. and the i.p.s.p. Under these conditions suramin (100 μM) markedly inhibited the p.s.p., but did not alter the i.p.s.p. In the combined presence of AP‐5 (50 μM), CNQX (50 μM), picrotoxin (100 μM), strychnine (0.1 μM), tropisetron (0.5 μM) and hexamethonium (100 μM), a high concentration of suramin (300 μM) almost abolished the p.s.p. without changing the i.p.s.p.
In the presence of kynurenic acid (500 μM) and picrotoxin (100 μM), PPADS (30 μM) depressed the p.s.p. Moreover, the application of suramin (100 μM) to the PPADS (30 μM)‐containing medium failed to cause any further inhibition. Neither PPADS (30 μM) nor suramin (100 μM) altered the i.p.s.p.
It was concluded that the cell somata of LC neurones are endowed with excitatory P2‐purinoceptors. ATP may be released either as the sole transmitter from purinergic neurones terminating at the LC or as a co‐transmitter of noradrenaline from recurrent axon collaterals or dendrites of the LC neurones themselves.

British Journal of Pharmacology (1997) 122, 423–430; doi:10.1038/sj.bjp.0701386