bluuebunny/crossref_metadata_2025_split huggingface dataset:
Tryptophan scanning mutagenesis in TM2 of the GABAA receptor α subunit: effects on channel gating and regulation by ethanol
Authors: Ueno Susumu , Lin Audrey , Nikolaeva Natalia , Trudell James R , Mihic S John , Harris R Adron , Harrison Neil L 
Published: January 2005

Each residue in the second transmembrane segment (TM2) of the human GABAA receptor α2 subunit was individually mutated to tryptophan. The wild‐type or mutant α2 subunits were expressed with the wild‐type human GABAA receptor β2 subunit in Xenopus oocytes, and the effects of these mutations were investigated using two‐electrode voltage‐clamp recording.
Four mutations (V257W, T262W, T265W and S270W) produced receptors which were active in the absence of agonist, and this spontaneous open channel activity was blocked by both picrotoxin and bicuculline, except in the α2(V257W)β2 mutant receptor, which was not sensitive to picrotoxin.
Six mutations (V257W, V260W, T262W, T267W, S270W and A273W) enhanced the agonist sensitivity of the receptor, by 10–100 times compared with the wild‐type α2β2 receptor. Other mutations (T261W, V263W, L269W, I271W and S272W) had little or no effect on the apparent affinity of the receptor to GABA. Eight of the tryptophan mutations (R255, T256, F258, G259, L264, T265, M266 or T268) resulted in undetectable GABA‐induced currents.
The S270W mutation eliminated potentiation of GABA by ethanol, whereas T261W markedly increased the action of ethanol. The T262W mutation produced direct activation (10% of maximal GABA response) by ethanol in the absence of GABA, while other mutations did not alter the action of ethanol significantly.
These results are consistent with a unique role for S270 in the action of ethanol within the TM2 region, and with models of GABAA receptor channel function, in which specific residues within TM2 are critical for the regulation of channel gating (S270, L264), while other residues (L269, I271 and S272) have little effect on these functions and may be non‐critical structural residues.

British Journal of Pharmacology (2000) 131, 296–302; doi:10.1038/sj.bjp.0703504
Intestinal anti‐inflammatory activity of UR‐12746, a novel 5‐ASA conjugate, on acute and chronic experimental colitis in the rat
Authors: Gálvez Julio , Garrido Margarita , Merlos Manuel , Torres Maria Isabel , Zarzuelo Antonio 
Published: January 2005

The present study was undertaken to investigate the intestinal anti‐inflammatory effects of UR‐12746 on the acute and chronic stages of a trinitrobenzene sulphonic acid (TNBS) experimental model of inflammatory bowel disease (IBD) in the rat. UR‐12746 is a novel, locally‐acting compound which combines, through an azo bond, 5‐aminosalicylic (5‐ASA) and UR‐12715, a potent platelet activating factor (PAF)‐antagonist.
UR‐12746 oral pretreatment of colitic rats (50 and 100 mg kg−1) reduced acute colonic damage when evaluated 2 days after colonic insult.
Postreatment for 4 weeks with UR‐12746 (50 and 100 mg kg−1) resulted in a faster recovery of the damaged colonic mucosa, which was macroscopically significant from the third week.
The intestinal anti‐inflammatory effect of UR‐12746 was associated with a decrease in leukocyte infiltration in the colonic mucosa, which was evidenced both biochemically, by a reduction in myeloperoxidase activity, and histologically, by a lower leukocyte count after morphometric analysis. This effect was higher than that seen with sulphasalazine, when assayed at the same doses and in the same experimental conditions.
Several mechanisms can be involved in the beneficial effects showed by UR‐12746: inhibition of leukotriene B4 synthesis in the inflamed colon, improvement of the altered colonic oxidative status, and reduction of colonic interleukin‐1β production.
The results suggest that the intestinal anti‐inflammatory activity of UR‐12746 can be attributed to the additive effects exerted by 5‐ASA and UR‐12715, the PAF antagonist compound, that are released in the colonic lumen after reduction of the azo bond by the intestinal bacteria.

British Journal of Pharmacology (2000) 130, 1949–1959; doi:10.1038/sj.bjp.0703505
Ligand regulation of green fluorescent protein‐tagged forms of the human β1‐ and β2‐adrenoceptors; comparisons with the unmodified receptors
Authors: McLean Alison J , Milligan Graeme 
Published: January 2005

Stable clones of HEK293 cells expressing either FLAGTM epitope‐tagged, wild type human β1‐ and β2‐adrenoceptors or C‐terminally green fluorescent protein (GFP)‐tagged forms of these receptors were established.
The binding affinity of [3H]‐dihydroalprenolol and other ligands was little affected by addition of GFP to the C‐terminal of either receptor.
Isoprenaline induced the internalisation of both β1‐adrenoceptor‐GFP and β2‐adrenoceptor‐GFP and following removal of the agonist both constructs were able to recycle to the cell surface.
The extent of internalisation of β2‐adrenoceptor‐GFP produced by isoprenaline was substantially greater than for β1‐adrenoceptor‐GFP.
C‐terminal addition of GFP slowed markedly the rate of internalization of both the β1‐adrenoceptor and the β2‐adrenoceptor in response to isoprenaline.
Sustained exposure to isoprenaline (24 h) produced substantially greater levels of downregulation of native β2‐adrenoceptor compared to β2‐adrenoceptor‐GFP although both were equally effectively removed from the plasma membrane.
Sustained exposure to isoprenaline resulted in a large fraction of β2‐adrenoceptor‐GFP becoming trapped in internal vesicles/lysosomes but not degraded.
Even after sustained exposure to isoprenaline a significant fraction of β1‐adrenoceptor‐GFP remained at the cell surface.
These results indicate that although GFP tagging of β‐adrenoceptors can provide qualitative visual patterns of agonist‐induced receptor trafficking and regulation in HEK293 cells the quantitative details vary markedly from those obtained with the unmodified receptors.

British Journal of Pharmacology (2000) 130, 1825–1832; doi:10.1038/sj.bjp.0703506
Single channel analysis of the blocking actions of BIDN and fipronil on a Drosophila melanogaster GABA receptor (RDL) stably expressed in a Drosophila cell line
Authors: Grolleau Françoise , Sattelle David B 
Published: January 2005

Single channel recordings were obtained from a Drosophila S2 cell line stably expressing the wild‐type RDLac Drosophila melanogaster homomer‐forming ionotropic GABA receptor subunit, a product of the resistance to dieldrin gene, Rdl. GABA (50 μM) was applied by pressure ejection to outside‐out patches from S2‐RDL cells at a holding potential of −60 mV. The resulting inward current was completely blocked by 100 μM picrotoxin (PTX). The unitary current‐voltage relationship was linear at negative potentials but showed slight inward rectification at potentials more positive than 0 mV. The reversal potential of the current (EGABA=−1.4 mV) was close to the calculated chloride equilibrium potential.
The single channel conductance elicited by GABA was 36 pS. A 71 pS conductance channel was also observed when the duration of the pulse, used to eject GABA, was longer than 80 ms. The mean open time distribution of the unitary events was fitted best by two exponential functions suggesting two open channel states.
When either 1 μM fipronil or 1 μM BIDN was present in the external saline, the GABA‐gated channels were completely blocked. When BIDN or fipronil was applied at a concentration close to the IC50 value for suppression of open probability (281 nM, BIDN; 240 nM, fipronil), the duration of channel openings was shortened. In addition, the blocking action of BIDN resulted in the appearance of a novel channel conductance (17 pS).
The effects of co‐application of BIDN and fipronil were examined. Co‐application of BIDN (300 nM) with various concentrations (100–1000 nM) of fipronil resulted in an additional BIDN‐induced dose‐dependent reduction of the maximum Po value.
Thus both BIDN and fipronil shorten the duration of wild‐type RDLac GABA receptor channel openings but appear to act at distinct sites.

British Journal of Pharmacology (2000) 130, 1833–1842; doi:10.1038/sj.bjp.0703507
Serotonin inhibition of the NMDA receptor/nitric oxide/cyclic GMP pathway in human neocortex slices: involvement of 5‐HT2C and 5‐HT1A receptors
Authors: Maura Guido , Marcoli Manuela , Pepicelli Olimpia , Rosu Christian , Viola Concetta , Raiteri Maurizio 
Published: January 2005

The NMDA receptor/nitric oxide (NO)/cyclic GMP pathway and its modulation by 5‐hydroxytryptamine (5‐HT) was studied in slices of neocortical samples obtained from patients undergoing neurosurgery.
The cyclic GMP elevation produced by 100 μM NMDA was blocked by 100 μM of the NO synthase inhibitor NG‐nitro‐L‐arginine (L‐NOARG) or by 10 μM of the soluble guanylate cyclase inhibitor 1H‐[1,2,4]oxadiazolo[4,3,‐α] quinoxaline‐1‐one (ODQ).
The NMDA effect was prevented by 5‐HT or by the 5‐HT2 agonist (±)‐1‐(2,5‐dimethoxy‐4‐iodophenyl)‐2‐aminopropane ((±)‐DOI; EC50=22 nM). The (±)‐DOI inhibition was insensitive to the 5‐HT2A receptor antagonist MDL 100907 or the 5‐HT2B antagonist rauwolscine; it was largely prevented by 1 μM of the non‐selective 5‐HT2C antagonists mesulergine (5‐HT2A,B,C), ketanserin (5‐HT2A,C) or SB 200646A (5‐HT2B,C); it was completely abolished by 0.1 μM of the selective 5‐HT2C receptor antagonist SB 242084.
The NMDA‐induced cyclic GMP elevation also was potently inhibited by the selective 5‐HT2C agonist RO 60‐0175 and by the antidepressant trazodone, both added at 1 μM, in an SB 242084‐sensitive manner.
Finally, the 5‐HT1A agonist 8‐hydroxy‐2‐(di‐n‐propylamino) tetralin (8‐OH‐DPAT; 1 μM) inhibited the NMDA‐evoked cyclic GMP response, an effect blocked by the selective 5‐HT1A receptor antagonist WAY 100635.
In conclusion, the NMDA receptor/NO/cyclic GMP pathway in human neocortex slices can be potently inhibited by activation of 5‐HT2C or 5‐HT1A receptors.

British Journal of Pharmacology (2000) 130, 1853–1858; doi:10.1038/sj.bjp.0703510
Effect of genistein on native epithelial tissue from normal individuals and CF patients and on ion channels expressed in Xenopus oocytes
Authors: Mall Marcus , Wissner Andreas , Seydewitz Hans H , Hübner Martin , Kuehr Joachim , Brandis Matthias , Greger Rainer , Kunzelmann Karl 
Published: January 2005

The flavonoid genistein has been shown to activate a Cl− conductance in various cell types expressing CFTR. We examined if similar effects can be observed when genistein is applied to native ex vivo tissues from human respiratory tract and rectum. We further compared the effects when genistein was applied to oocytes of Xenopus laevis expressing CFTR.
In oocytes, both wtCFTR and ΔF508‐CFTR were activated by genistein while both cyclic AMP (KvLQT1) and Ca2+ (SK4) activated K+ channels were inhibited at high concentrations of genistein.
Biopsies from nasal polyps and rectal mucosa were obtained from normal individuals (non‐CF) and CF patients and in the presence of amiloride (10 μmol l−1; mucosal side) the effects of genistein were assessed using a perfused Ussing chamber. In non‐CF airway epithelia, genistein (50 μmol l−1; mucosal side) increased lumen negative Isc but had no additional effects on tissues pre‐stimulated with IBMX and forskolin (100 μmol l−1 and 1 μmol l−1; both sides).
In non‐CF rectal biopsies, in the presence of amiloride (10 μmol l−1; mucosal side) and indomethacin (10 μmol l−1; basolateral side), genistein increased lumen negative Isc and enabled cholinergic (carbachol; CCH, 100 μmol l−1; basolateral side) stimulation of Cl− secretion indicating activation of luminal CFTR Cl− channels. However, after stimulation with IBMX/forskolin, genistein induced opposite effects and significantly inhibited CCH activated Isc. In CF airway and intestinal tissues genistein failed to induce Cl− secretion.
Thus, genistein is able to activate luminal CFTR Cl− conductance in non‐CF tissues and mutant CFTR in oocytes. However, additional inhibitory effects on basolateral K+ conductance and missing effects in native CF tissues do not support the use for pharmacological intervention in CF.

British Journal of Pharmacology (2000) 130, 1884–1892; doi:10.1038/sj.bjp.0703520
Insulin‐like effect of pinitol
Authors: Bates Sarah H , Jones Robert B , Bailey Clifford J 
Published: January 2005

D‐pinitol (3‐O‐methyl‐chiroinositol), an active principle of the traditional antidiabetic plant Bougainvillea spectabilis, is claimed to exert insulin‐like effects. This study investigates the effect of D‐pinitol on glucose homeostasis in animal models of diabetes, and on glucose transport by cultured muscle cells.
Plasma glucose concentrations were measured in normal, obese‐diabetic (ob/ob) and streptozotocin (STZ)‐diabetic mice after oral (p.o.) and intraperitoneal (i.p.) administration of D‐pinitol. Glucose transport was measured in L6 rat muscle cells by 2‐deoxyglucose (2DG) uptake.
In STZ‐diabetic mice, 100 mg kg−1 p.o. D‐pinitol acutely decreased the hyperglycaemia (by 22% at 6 h). A similar decrease in plasma glucose (by 21%) was observed after 100 mg kg−1 i.p. D‐pinitol. Insulin concentrations and the rate of insulin‐induced (1 unit kg−1 actrapid i.p.) glucose disappearance were not altered by 100 mg kg−1 p.o. D‐pinitol. Chronic administration of D‐pinitol (100 mg kg−1 i.p. twice daily for 11 days) to STZ‐diabetic mice maintained a reduction in plasma glucose concentrations from about 14 to 10 mmol l−1.
In normal non‐diabetic and severely insulin resistant ob/ob mice, 100 mg kg−1 p.o. D‐pinitol did not significantly affect plasma glucose or insulin during acute studies.
Incubation of L6 muscle cells with D‐pinitol (10−3 M) increased basal 2DG uptake by 41% after 10 min and by 34% after 4 h. The effect of D‐pinitol was inhibited by the phosphatidylinositol 3‐kinase inhibitor LY294002. D‐pinitol did not increase insulin‐stimulated 2DG uptake by L6 cells.
The data support the view that D‐pinitol can exert an insulin‐like effect to improve glycaemic control in hypoinsulinaemic STZ‐diabetic mice. D‐pinitol may act via a post‐receptor pathway of insulin action affecting glucose uptake.

British Journal of Pharmacology (2000) 130, 1944–1948; doi:10.1038/sj.bjp.0703523
The progestin levonorgestrel induces endothelium‐independent relaxation of rabbit jugular vein via inhibition of calcium entry and protein kinase C: role of cyclic AMP
Authors: Herkert Olaf , Kuhl Herbert , Busse Rudi , Schini‐Kerth Valérie B 
Published: January 2005

The progestin and oestrogen component of oral contraceptives have been involved in the development of venous thromboembolic events in women. In the present study we determined the vasoactive effects of sex steroids used in oral contraceptives in isolated preconstricted rabbit jugular veins in the presence of diclofenac and examined the underlying mechanisms.
The natural hormone progesterone, the synthetic progestins levonorgestrel, 3‐keto‐desogestrel, gestodene and chlormadinone acetate, and the synthetic estrogen 17 α‐ethinyloestradiol induced concentration‐dependent relaxations of endothelium‐intact veins constricted with U46619. Levonorgestrel also inhibited constrictions evoked by either a high potassium (K+) solution or phorbol myristate acetate (PMA) in the absence and presence of extracellular calcium (Ca2+). In addition, levonorgestrel depressed contractions evoked by Ca2+ and reduced 45Ca2+ influx in depolarized veins.
Relaxations to levonorgestrel in U46619‐constricted veins were neither affected by the presence of the endothelium nor by the inhibitor of soluble guanylyl cyclase, NS2028, but were significantly improved either by the selective cyclic AMP phosphodiesterase inhibitor rolipram or in the absence of diclofenac, and decreased by the protein kinase A inhibitor, Rp‐8‐CPT‐cAMPS. Rolipram also potentiated relaxations to levonorgestrel in PMA‐constricted veins in the presence, but not in the absence of extracellular Ca2+. Levonorgestrel increased levels of cyclic AMP and inhibited PMA‐induced activation of protein kinase C in veins.
These findings indicate that levonorgestrel caused endothelium‐independent relaxations of jugular veins via inhibition of Ca2+ entry and of protein kinase C activation. In addition, the cyclic AMP effector pathway contributes to the levonorgestrel‐induced relaxation possibly by depressing Ca2+ entry.

British Journal of Pharmacology (2000) 130, 1911–1918; doi:10.1038/sj.bjp.0703524
Inhibition of sustained hypoxic vasoconstriction by Y‐27632 in isolated intrapulmonary arteries and perfused lung of the rat
Authors: Robertson Tom P , Dipp Michelle , Ward Jeremy P T , Aaronson Phil I , Evans A Mark 
Published: January 2005

We have examined the effects of Y‐27632, a specific inhibitor of Rho‐activated kinases (ROCK I and ROCK II) upon sustained hypoxic pulmonary vasoconstriction (HPV) in both rat isolated small intrapulmonary arteries (IPA) and perfused rat lungs in situ. Y‐27632 (100 nM–3 μM) was found to cause a concentration‐dependent inhibition of acute sustained HPV in rat IPA. Application of Y‐27632 (10–600 nM) in perfused rat lungs caused no change in basal perfusion pressure, but was found to inhibit HPV in a concentration‐dependent manner, resulting in complete ablation of the pressor response to hypoxia at a concentration of 600 nM. Furthermore, addition of Y‐27632 at any point during hypoxia caused a reversal of HPV in perfused rat lungs. These results suggest that activation of Rho‐associated kinase may be a pivotal step in the generation of sustained HPV.British Journal of Pharmacology (2000) 131, 5–9; doi:10.1038/sj.bjp.0703537
Prevention by dexrazoxane of down‐regulation of ryanodine receptor gene expression in anthracycline cardiomyopathy in the rat
Authors: Burke Briant E , Gambliel Hervé , Olson Richard D , Bauer Frederic K , Cusack Barry J 
Published: January 2005

Anthracyclines can cause cumulative dose‐related cardiotoxicity characterized by changes in Ca2+ metabolism, including dysfunction of the sacroplasmic reticulum (SR) and decreased expression of Ca2+‐handling proteins, such as the ryanodine receptor (RyR2). In this study, we examined the effect of dexrazoxane (ICRF‐187), an iron chelator which prevents anthracycline cardiotoxicity, on RyR2 gene expression in rats treated chronically with daunorubicin. Daunorubicin (2.5 mg kg−1 i.v. weekly for 6 weeks) produced cardiotoxicity as demonstrated by histopathologic changes. The ryanodine receptor/glyceraldehyde phosphate dehydrogenase (GAPDH) mRNA ratio was decreased by 38±3% (P<0.02) compared to values in control rats. Dexrazoxane pre‐treatment (50 mg kg−1; 1 h prior to each daunorubicin injection) prevented the decrease in RyR2/GAPDH mRNA ratio and histopathologic lesions in daunorubicin‐treated rats. This is the first report that a protective agent such as dexrazoxane can ameliorate the decreased expression of a specific gene involved in anthracycline‐induced cardiotoxicity.British Journal of Pharmacology (2000) 131, 1–4; doi:10.1038/sj.bjp.0703538
Influence of hypertension on nitric oxide synthase expression and vascular effects of lipopolysaccharide in rat mesenteric arteries
Authors: Briones Ana M , Alonso María J , Marín Jesús , Balfagón Gloria , Salaices Mercedes 
Published: January 2005

Experiments were designed to investigate the effects of the inducible nitric oxide synthase (iNOS) stimulator, lipopolysaccharide (LPS), on noradrenaline (NA) responses and on NOS activity and its expression in intact mesenteric resistance arteries (MRAs) from Wistar Kyoto (WKY) and spontaneously hypertensive (SHR) rats.
In MRAs from WKY, LPS (10 μg ml−1; 1–5 h) reduced the vasoconstrictor responses to NA (0.1–30 μM) in the presence, but not in the absence of L‐arginine (L‐Arg, 10 μM). However, in SHR arteries, LPS induced an incubation‐time dependent reduction of NA responses in the absence, as well as the presence, of L‐Arg. The LPS inhibitory effect was reduced by the non‐specific NOS inhibitor L‐NG‐nitroarginine methyl ester (L‐NAME, 100 μM) and the selective iNOS inhibitor, aminoguanidine (100 μM).
L‐NAME alone similarly shifted the concentration‐response curve to NA leftward in arteries from both strains, while aminoguanidine had no effect. L‐Arg shifted the curve to NA rightward only in SHR MRAs.
Basal activity of both iNOS and constitutive NOS (conversion of [3H]‐L‐Arg to [3H]‐L‐citrulline) was similar in arteries from both strains. After 5 h incubation with LPS, only iNOS activity in arteries from SHR was increased.
Basal iNOS protein expression was undetectable; basal endothelial (eNOS) protein expression was similar in arteries from both strains, while neuronal (nNOS) was greater in arteries from SHR. LPS induced iNOS protein expression, that was higher in arteries from SHR than in those from WKY.
These results indicate that NO production, via iNOS induction, is greater than in those from MRAs from SHR to WKY.

British Journal of Pharmacology (2000) 131, 185–194; doi:10.1038/sj.bjp.0703552
On the in vitro vasoactivity of bile acids
Authors: Ljubuncic Predrag , Said Omar , Ehrlich Yaron , Meddings Jon B , Shaffer Eldon A , Bomzon Arieh 
Published: January 2005

We compared the vasorelaxant action of nine different bile acids and correlated their vasorelaxant activity with their individual indices for hydrophobicity or lipophilicity.
Vasorelaxant activity correlated with the relative lipid solubility of bile acids with lipophilic bile acids exhibiting the greatest vasorelaxant activity with modest to no vasorelaxant activity exhibited by hydrophilic bile acids.
We also investigated whether bile acid‐induced vasorelaxation is mediated by antagonism of a prototypal contractile receptor, the α1‐adrenoceptor, by stimulation of a bile acid surface membrane receptor, by the release of endothelium‐derived relaxant factors, by promoting the generation of reactive oxygen species and increasing the extent of lipid peroxidation, or by modifying membrane fluidity.
Lipophilic bile acids induce vasorelaxation possibly by antagonizing α1‐adrenoceptors, a phenomenon that manifests itself as a lowering of the affinity of vascular α1‐adrenoceptors. Bile acid‐induced vasorelaxation was not dependent upon stimulation of a bile acid surface membrane receptor or the release of endothelium‐derived relaxant factors.
Lipophilic bile acids can also increase the extent of lipid peroxidation with a subtle reduction in the fluidity of rat vascular smooth muscle membranes not associated with loss of membrane cholesterol or phospholipid.
We have concluded that lipophilic bile acids are non‐selective vasorelaxants whose mechanism of action is a multifaceted process involving antagonism of contractile surface membrane receptors possibly effected by an increased extent of lipid peroxidation and/or membrane fluidity but occurs independent of the release of endothelial‐derived relaxant factors or stimulation of a surface membrane bile acid binding site.

British Journal of Pharmacology (2000) 131, 387–398; doi:10.1038/sj.bjp.0703554