bluuebunny/crossref_metadata_2025_split huggingface dataset:
In vitro and in vivo pharmacology of a structurally novel Na+‐H+ exchange inhibitor, T‐162559
Authors: Kusumoto Keiji , Igata Hideki , Abe Akemi , Ikeda Shota , Tsuboi Ayako , Imamiya Eikoh , Fukumoto Shoji , Shiraishi Mitsuru , Watanabe Toshifumi 
Published: January 2005

We investigated the inhibitory effects of a non‐acylguanidine Na+‐H+ exchange (NHE) inhibitor, T‐162559 ((5E,7S)‐[7‐(5‐fluoro‐2‐methylphenyl)‐4‐methyl‐7,8‐dihydro‐5(6H)‐quinolinylideneamino] guanidine dimethanesulphonate), on NHE‐1, and its cardioprotective effect against ischaemia and reperfusion injury in rats and rabbits.
T‐162559 inhibited human platelet NHE‐1 in a concentration‐dependent manner, with an IC50 value of 13±3 nmol l−1, making it 16 and three times more potent than cariporide IC50: 209±75 nmol l−1, P<0.01) and eniporide (IC50: 40±11 nmol l−1, P=0.066), respectively. T‐162559 also inhibited rat NHE‐1 with an IC50 value of 14±2 nmol l−1, which was five and three times lower than that of cariporide (IC50: 75±7 nmol l−1, P<0.01) and eniporide (IC50: 44±2 nmol l−1, P<0.01), respectively.
T‐162559 inhibited, in a concentration‐dependent manner, the reduction in cardiac contractility, progression of cardiac contracture, and increase in lactate dehydrogenase release after global ischaemia and reperfusion in perfused rat hearts. The inhibitory effects of T‐162559 were observed at a lower concentration range (10 – 100 nmol l−1) than with cariporide and eniporide. T‐162559 did not alter basal cardiac contractility or coronary flow after reperfusion, suggesting that it exerts direct cardioprotective effects on the heart.
Intravenous administration of T‐162559 (0.03 and 0.1 mg kg−1) significantly inhibited the progression of myocardial infarction induced by left coronary artery occlusion and reperfusion in rabbits; the infarct size normalized by area at risk was 74±6% in the vehicle group, and 47±5% and 51±7% in the T‐162559‐0.03 mg kg−1 and T‐162559‐0.1 mg kg−1 groups (both P<0.05), respectively.
These results indicate that the new structural NHE‐1 inhibitor T‐162559 is more potent than cariporide and eniporide and possesses a cardioprotective effect against ischaemia and reperfusion injury in rat and rabbit models.

British Journal of Pharmacology (2002) 135, 1995–2003; doi:10.1038/sj.bjp.0704647
Complex regional haemodynamic effects of anandamide in conscious rats
Authors: Gardiner S M , March J E , Kemp P A , Bennett T 
Published: January 2005

Experiments were carried out in conscious, chronically instrumented, male, Sprague‐Dawley rats to delineate the regional haemodynamic effects of the putative endogenous cannabinoid, anandamide, (0.075 – 3 mg kg−1), and to dissect some of the mechanisms involved.
At all doses of anandamide, there was a significant, short‐lived increase in mean arterial blood pressure associated with vasoconstriction in renal, mesenteric and hindquarters vascular beds.
The higher doses (2.5 and 3 mg kg−1), caused an initial, marked bradycardia accompanied, in some animals, by a fall in arterial blood pressure which preceded the hypertension. In addition, after the higher doses of anandamide, the hindquarters vasoconstriction was followed by vasodilatation.
Although some of the effects described above resembled those of 5‐HT (25 μg kg−1), the bradycardia and hypotensive actions of the latter were abolished by the 5HT3‐receptor antagonist, azasetron, whereas those of anandamide were generally unaffected.
None of the cardiovascular actions of anandamide were influenced by the CB1‐receptor antagonist, AM 251, but its bradycardic effect was sensitive to atropine, and its hindquarters vasodilator action was suppressed by the β2‐adrenoceptor antagonist, ICI 118551.
The results differ, in several aspects, from those previously reported in anaesthetized animals, and underscore the important impact anaesthesia can have on responses to anandamide.

British Journal of Pharmacology (2002) 135, 1889–1896; doi:10.1038/sj.bjp.0704649
Mouse β3a‐ and β3b‐adrenoceptors expressed in Chinese hamster ovary cells display identical pharmacology but utilize distinct signalling pathways
Authors: Hutchinson Dana S , Bengtsson Tore , Evans Bronwyn A , Summers Roger J 
Published: January 2005

This study characterizes the mouse β3a‐adrenoceptor (AR) and the splice variant of the β3‐AR (β3b‐AR) expressed in Chinese hamster ovary cells (CHO‐K1).
Stable clones with high (∼1200), medium (∼500) or low receptor expression (∼100 fmol mg protein−1) were determined by saturation binding with [125I]‐(−)‐cyanopindolol. Competition binding studies showed no significant differences in affinity of β‐AR ligands for either receptor.
Several functional responses of each receptor were measured, namely extracellular acidification rate (EAR; cytosensor microphysiometer), cyclic AMP accumulation, and Erk1/2 phosphorylation. The β3‐AR agonists BRL37344, CL316243, GR265162X, L755507, SB251023, the non‐conventional partial β‐AR agonist CGP12177 and the β‐AR agonist (−)‐isoprenaline caused concentration‐dependent increases in EAR in cells expressing either splice variant. CL316243 caused concentration‐dependent increases in cyclic AMP accumulation and Erk1/2 phosphorylation in cells expressing either receptor.
PTX treatment increased maximum EAR and cyclic AMP responses to CL316243 in cells expressing the β3b‐AR but not in cells expressing the β3a‐AR at all levels of receptor expression.
CL316243 increased Erk1/2 phosphorylation with pEC50 values and maximum responses that were not significantly different in cells expressing either splice variant. Erk1/2 phosphorylation was insensitive to PTX or H89 (PKA inhibitor) but was inhibited by LY294002 (PI3Kγ inhibitor), PP2 (c‐Src inhibitor), genistein (tyrosine kinase inhibitor) and PD98059 (MEK inhibitor).
The adenylate cyclase activators forskolin or cholera toxin failed to increase Erk1/2 levels although both treatments markedly increased cyclic AMP accumulation in both β3a‐ or β3b‐AR transfected cells.
These results suggest that in CHO‐K1 cells, the β3b‐AR, can couple to both Gs and Gi to stimulate and inhibit cyclic AMP production respectively, while the β3a‐AR, couples solely to Gs to increase cyclic AMP levels. However, the increase in Erk1/2 phosphorylation following receptor activation is not dependent upon coupling of the receptors to Gi or the generation of cyclic AMP.

British Journal of Pharmacology (2002) 135, 1903–1914; doi:10.1038/sj.bjp.0704654
Regulation of IL‐13 synthesis in human lymphocytes: implications for asthma therapy
Authors: Pahl Andreas , Zhang Meixia , Kuss Hildegard , Szelenyi Istvan , Brune Kay 
Published: January 2005

IL‐13 is an important mediator in inflammatory diseases such as asthma. IL‐13 is mainly produced by T cells. However, signalling pathways leading to induction of this cytokine are not well‐characterized. We analysed the regulation of IL‐13 in human peripheral blood mononuclear cells and CD4+ T cells.
Cyclosporine (CsA) and FK‐506 inhibited IL‐13 synthesis, when cells were stimulated by TPA/ionomycin. However, stimulation by α‐CD3/α‐CD28 led to an enhanced IL‐13 synthesis.
NF‐κB inhibitor N‐tosyl‐L‐lysine chloromethylketone (TLCK) inhibited IL‐13 synthesis more effectively after TPA/ionomycin stimulation. After α‐CD3/α‐CD28 stimulation, only 300 μM TLCK inhibited IL‐13 synthesis. Dexamethasone inhibited IL‐13 equally effective after α‐CD3/α‐CD28 and TPA/ionomycin stimulation.
p38 MAPK inhibitor SB203580 inhibited IL‐13 synthesis only partially. MEK inhibitor U0126 inhibited TPA/ionomycin induced IL‐13 synthesis very effectively, whereas α‐CD3/α‐CD28 stimulated IL‐13 induction was resistant to this drug.
These results were confirmed in purified CD4+ T cells. In difference to PBMCs α‐CD3/α‐CD28 stimulated IL‐13 synthesis was effectively inhibited by CsA, FK‐506 and U0126.
Therefore U0126 was tested in an animal model of allergic asthma. We could demonstrate for the first time that inhibition of the MEK – ERK cascade is a therapeutic option for asthma. Intraperitoneal administration of 10 mg kg−1 U0126 reduced lung eosinophilia in ovalbumin‐challenged Brown Norway rats by 44%.
These results demonstrate that different signalling pathways are involved in regulating IL‐13 synthesis in primary human T cells. Characterizing highly potent inhibitors of IL‐13 synthesis can be exploited to identify new drugs to treat immunological diseases such as asthma.

British Journal of Pharmacology (2002) 135, 1915–1926; doi:10.1038/sj.bjp.0704656
Direct visualization of ligand‐protein interactions using atomic force microscopy
Authors: Neish Calum S , Martin Ian L , Henderson Robert M , Edwardson J Michael 
Published: January 2005

Streptavidin is a 60‐kDa tetramer which binds four molecules of biotin with extremely high affinity (KA∼1014 M−1). We have used atomic force microscopy (AFM) to visualize this ligand‐protein interaction directly.
Biotin was tagged with a short (152‐basepair; 50‐nm) DNA rod and incubated with streptavidin. The resulting complexes were then imaged by AFM. The molecular volume of streptavidin calculated from the dimensions of the protein particles (105±3 nm3) was in close agreement with the value calculated from its molecular mass (114 nm3). Biotinylation increased the apparent size of streptavidin (to 133±2 nm3), concomitant with an increase in the thermal stability of the tetramer.
Images of streptavidin with one to four molecules of DNA‐biotin bound were obtained. When two ligands were bound, the angle between the DNA rods was either acute or obtuse, as expected from the relative orientations of the biotin binding sites. The ratio of acute : obtuse angles (1 : 3) was lower than the expected value (1 : 2), indicating a degree of steric hindrance in the binding of the DNA‐biotin. The slight under‐representation of higher occupancy states supported this idea.
Streptavidin with a single molecule of DNA‐biotin bound was used to tag biotinylated β‐galactosidase, a model multimeric enzyme.
The ability to image directly the binding of a ligand to its protein target by AFM provides useful information about the nature of the interaction, and about the effect of complex formation on the structure of the protein. Furthermore, the use of DNA‐biotin/streptavidin tags could potentially shed light on the architecture of multi‐subunit proteins.

British Journal of Pharmacology (2002) 135, 1943–1950; doi:10.1038/sj.bjp.0704660
Activity profile of calpains I and II in chronically infarcted rat myocardium – influence of the calpain inhibitor CAL 9961
Authors: Sandmann Steffen , Prenzel Freerk , Shaw Lee , Schauer Roland , Unger Thomas 
Published: January 2005

The calpains have been proposed to be activated following cardiac ischaemia and to contribute to myocyte damage after myocardial infarction (MI). In this study, the activity of calpains I and II in the infarcted and non‐infarcted rat myocardium and the action of the selective calpain inhibitor, CAL 9961, has been investigated.
MI was induced by permanent ligation of the left coronary artery. One, 3, 7 and 14 days post MI, the enzymes calpain I and II were separated from homogenates of the interventricular septum (IS) and left ventricular free wall (LVFW) by chromatography on DEAE‐Sepharose. The activity of the calpains was measured in sham‐operated and MI animals chronically treated with placebo or CAL 9961 (15 mg kg−1 d−1 s.c.) in a synthetic substrate assay. Treatment was started 3 days before MI induction.
Calpain I activity reached highest values in IS 14 days post MI, whereas maximum activity of calpain II was measured in LVFW 3 days post MI. In experiments in vitro, CAL 9961 completely inhibited both calpains. In vivo, chronic treatment of MI animals with CAL 9961 partially prevented the increase in calpain I activity in IS and reduced calpain II activity in LVFW to sham levels.
Our findings demonstrate that calpains I and II are activated after MI, however, both enzymes differ in their regional and temporal activation within the infarcted myocardium. Chronic inhibition of these enzymes with CAL 9961 might limit the calpain‐induced myocardial damage and preserve cardiac structural integrity post MI.

British Journal of Pharmacology (2002) 135, 1951–1958; doi:10.1038/sj.bjp.0704661
Xestospongin C, a novel blocker of IP3 receptor, attenuates the increase in cytosolic calcium level and degranulation that is induced by antigen in RBL‐2H3 mast cells
Authors: Oka Tatsuya , Sato Koichi , Hori Masatoshi , Ozaki Hiroshi , Karaki Hideaki 
Published: January 2005

We evaluated the role of the cross‐linking of FcεRI‐mediated inositol 1,4,5‐triphosphate (IP3) in the increase in cytosolic Ca2+ level ([Ca2+]i) using xestospongin C, a selective membrane permeable blocker of IP3 receptor, in RBL‐2H3 mast cells.
In the cells sensitized with anti‐dinitrophenol (DNP) IgE, DNP‐human serum albumin (DNP‐HSA) and thapsigargin induced degranulation of β‐hexosaminidase and a sustained increase in [Ca2+]i. Xestospongin C (3 – 10 μM) inhibited both of these changes that were induced by DNP‐HSA without changing those induced by thapsigargin.
In the absence of external Ca2+, DNP‐HSA induced a transient increase in [Ca2+]i. Xestospongin C (3 – 10 μM) inhibited this increase in [Ca2+]i.
In the cells permeabilized with β‐escin, the application of IP3 decreased Ca2+ in the endoplasmic reticulum (ER) as evaluated by mag‐fura‐2. Xestospongin C (3 – 10 μM) inhibited the effect of IP3.
After the depletion of Ca2+ stores due to stimulation with DNP‐HSA or thapsigargin, the addition of Ca2+ induced capacitative calcium entry (CCE). Xestospongin C (3 – 10 μM) inhibited the DNP‐HSA‐induced CCE, whereas it did not affect the thapsigargin‐induced CCE.
These results suggest that FcεRI‐mediated generation of IP3 contributes to Ca2+ release not only in the initial phase but also in the sustained phase of the increase in [Ca2+]i, resulting in prolonged Ca2+ depletion in the ER. The ER Ca2+ depletion may subsequently activate CCE to achieve a continuous [Ca2+]i increase, which is necessary for degranulation in the RBL‐2H3 mast cells. Xestospongin C may inhibit Ca2+ release and consequently may attenuate degranulation.

British Journal of Pharmacology (2002) 135, 1959–1966; doi:10.1038/sj.bjp.0704662
The involvement of smooth muscle P2X receptors in the prolonged vasorelaxation response to purine nucleotides in the rat mesenteric arterial bed
Authors: Ralevic V 
Published: January 2005

ATP and adenine dinucleotides can elicit three different types of vasomotor response in the rat mesenteric arterial bed; vasocontraction, rapid relaxation (which may be masked by contraction) and slow and prolonged vasorelaxation. Contraction is mediated by smooth muscle P2X receptors and rapid relaxation by endothelial P2Y receptors. The mechanism of prolonged relaxation is, however, controversial.
In the present study, bolus injection of doses of α,β‐methylene ATP (α,β‐meATP; 5 pmol – 0.5 μmol; P2X receptor agonist) in methoxamine‐preconstricted rat isolated mesenteric arterial beds, mimicked the action of ATP, causing contraction (Rmax 76±9 mmHg) followed by prolonged relaxation (78±11%; t1/2 14.6±1.5 min). KCl also elicited a biphasic response (Rmax contraction 73±8 mmHg; Rmax prolonged relaxation 70±6%; t1/2 7.7±1.9 min).
P2X receptor desensitization caused by perfusion with α,β‐meATP (10 μM) abolished contraction and prolonged relaxation to doses of α,β‐meATP (50 nmol). Rapid relaxation (32±7%; t1/2 32±2 s) was revealed, which was abolished by removal of the endothelium using distilled water.
Sodium deoxycholate treatment blocked contractile and prolonged relaxation responses to α,β‐meATP, ATP and KCl, whilst distilled water treatment had no significant effect on either phase of the biphasic responses.
These data indicate that smooth muscle P2X receptors are involved in both phases of the biphasic response (contraction followed by prolonged relaxation) to purine nucleotides in the rat isolated mesenteric arterial bed. Caution should be applied when using sodium deoxycholate to remove the endothelium because of possible damage caused by the detergent to receptors and/or the vascular smooth muscle.

British Journal of Pharmacology (2002) 135, 1988–1994; doi:10.1038/sj.bjp.0704663
Resolution of P‐glycoprotein and non‐P‐glycoprotein effects on drug permeability using intestinal tissues from mdr1a (−/−) mice
Authors: Stephens R H , O'Neill C A , Bennett J , Humphrey M , Henry B , Rowland M , Warhurst G 
Published: January 2005

Intestinal xenobiotic transporters are a significant barrier to the absorption of many orally administered drugs. P‐glycoprotein (PGP) is the best known, but several others, including members of the multidrug resistance‐associated protein (MRP) family, are also expressed. Definitive information on their precise effect on intestinal drug permeability is scarce due to a lack of specific inhibitors and the difficulty of studying non‐PGP activity in the presence of high PGP expression.
We have investigated the in vitro use of intestinal tissues from PGP knockout (mdr1a (−/−)) mice as a tool for dissecting the mechanisms of intestinal drug efflux. The permeability characteristics of digoxin (DIG), paclitaxel (TAX) and etoposide (ETOP) were measured in ileum from mdr1a (−/−) and wild‐type (FVB) mice mounted in Ussing chambers.
DIG and TAX exhibited marked efflux across FVB tissues (B‐A : A‐B apparent permeability (Papp) ratio 10 and 17 respectively) which was absent in mdr1a (−/−) tissues, confirming that PGP is the sole route of intestinal efflux for these compounds. The A‐B Papp of both compounds was 3 – 5 fold higher in mdr1a (−/−) than in FVB.
Polarized transport of ETOP in FVB tissues was reduced but not abolished in mdr1a (−/−) tissues. Residual ETOP efflux in mdr1a (−/−) tissues was abolished by the MRP inhibitor MK571, indicating involvement of both PGP and MRP.
MK571 abolished calcein efflux in mdr1a (−/−) tissues, while quinidine had no parallel effect in FVB tissues, suggesting involvement of MRP but not PGP.
Tissues from mdr1a (−/−) mice provide a novel approach for investigating the influence of PGP ablation on intestinal permeability and for resolving PGP and non‐PGP mechanisms that modulate drug permeability.

British Journal of Pharmacology (2002) 135, 2038–2046; doi:10.1038/sj.bjp.0704668
Bretylium or 6‐OHDA‐resistant, action potential‐evoked Ca2+ transients in varicosities of the mouse vas deferens
Authors: Jackson V Margaret , Cunnane Tom C 
Published: January 2005

Action potential‐evoked calcium transients in varicosities in mouse vas deferens were monitored using laser scanning confocal microscopy. Their significance was examined by comparison with excitatory junction potentials (EJPs) and neurogenic contractions, both indirect measurements of transmitter release. Bretylium abolished EJPs, as well as the ATP and NA‐mediated phases of contraction. However, bretylium revealed a prominent late component of contraction that was atropine‐sensitive. Bretylium abolished calcium transients in 21%, enhanced in 16% and had no effect in 63% of varicosities examined. Pre‐treatment with 6‐OHDA reduced NA levels to below detectable levels but many strings of varicosities still responded to nerve impulses with ‘normal’ calcium transients. Varicosities in which calcium transients were abolished by these agents were sympathetic. The identity of those varicosities in which calcium transients were resistant to bretylium (sympathetic but no uptake‐1 sites, parasympathetic, sensory) remains to be established.British Journal of Pharmacology (2002) 135, 1845–1850; doi:10.1038/sj.bjp.0704677
BIBN4096BS is a potent competitive antagonist of the relaxant effects of α‐CGRP on human temporal artery: comparison with CGRP(8‐37)
Authors: Verheggen Raphaela , Bumann Katja , Kaumann Alberto J. 
Published: January 2005

Release of CGRP during migraine may produce harmful dilatation of cranial arteries, thereby possibly causing pain. We have compared the antagonism by BIBN4096BS and CGRP(8‐37) of the relaxant effects of α‐CGRP on rings of human temporal artery.
α‐CGRP relaxed the arteries precontracted with 9 – 24 mM KCl (−logEC50=9.4) nearly as efficaciously as sodium nitroprusside (10 μM).
BIBN4096BS (0.1 – 100 nM) antagonized the effects of α‐CGRP in surmountable manner with slopes of Schild‐plots not different from unity. −LogKB values of 10.1 and 10.4 were estimated for BIBN4096BS when administered before or during the KCl‐contracture respectively.
BIBN4096BS (1 μM) did not modify the relaxant effects of papaverine and sodium nitroprusside.
CGRP(8‐37) (1 – 10 μM) antagonized the effects of α‐CGRP in a surmountable manner with slopes of Schild‐plots not different from unity. −LogKB values of 6.6 and 6.7 were estimated for CGRP(8‐37) administered before or during the KCl‐contracture respectively.
The high affinity of BIBN4096BS for CGRP receptors of human temporal artery makes it an excellent tool to explore the hypothesis of CGRP‐evoked cerebral vasodilation in migraine.

British Journal of Pharmacology (2002) 136, 120–126; doi:10.1038/sj.bjp.0704682
Cellular basis of endothelial dysfunction in small mesenteric arteries from spontaneously diabetic (db/db−/−) mice: role of decreased tetrahydrobiopterin bioavailability
Authors: Pannirselvam Malarvannan , Verma Subodh , Anderson Todd J , Triggle Chris R 
Published: January 2005

Endothelium‐dependent and ‐independent regulation of vascular tone in small mesenteric arteries (SMA) from control (db/db +/?) and diabetic (db/db −/−) mice was compared.
Phenylephrine‐induced maximum contraction, but not sensitivity, of SMA in db/db −/− compared to db/db +/? was enhanced.
Acetylcholine (ACh), but not sodium nitroprusside (SNP), ‐induced relaxation was reduced in SMA from db/db −/− compared to db/db +/?.
ACh‐induced relaxation of SMA was inhibited by a combination of Nω‐nitro‐L‐arginine and indomethacin in db/db +/?, but not in db/db −/−.
Acute incubation of SMA with tetrahydrobiopterin (BH4, 10 μM) and sepiapterin (100 μM) enhanced ACh‐induced relaxation in SMA from db/db −/−, but not from db/db +/? 2,4‐diamino‐6‐hydroxypyrimidine, an inhibitor of GTP cyclohydrolase I, (10 mM), impaired the sensitivity of SMA from db/db +/? to ACh, which was restored by co‐incubation with BH4 (10 μM).
BH4 and superoxide dismutase (SOD, 150 u ml−1), either alone or in combination, had no effect on either ACh or SNP‐induced relaxation in SMA from eNOS −/− mice.
Incubation of SMA with SOD (150 iu ml−1), catalase (200 iu ml−1) and L‐arginine (1 mM) had no effect on ACh‐induced relaxation of SMA. However, the combination of polyethylene glycol‐SOD (200 iu ml−1) and catalase (80 u ml−1) improved the sensitivity of ACh‐induced relaxation in db/db −/−, but not in db/db +/?.
These data suggest that increased production of superoxide anions and decreased availability of BH4 result in an ‘uncoupling’ of nitric oxide synthase and endothelial dysfunction in SMA from db/db −/− mice.

British Journal of Pharmacology (2002) 136, 255–263; doi:10.1038/sj.bjp.0704683