bluuebunny/crossref_metadata_2025_split huggingface dataset:
Characterization of [125I]‐endothelin‐1 and [125I]‐BQ3020 binding to rat cerebellar endothelin receptors
Authors: Widdowson P.S. , Kirk C.N.P. 
Published: July 2012

We performed radioligand binding experiments on rat cerebellar homogenates using [125I]‐endothelin‐1 ([125I]‐ET‐1) and [125I]‐BQ3020 to examine the pharmacology of endothelin receptors in rat brain. Saturation experiments demonstrated a single population of binding sites with high affinity for both radioligands ([125I]‐ET‐1, pKd = 8.94 ± 0.17; [125I]‐BQ3020, pKd = 9.18 ± 0.14 nM; mean ± s.e.mean). However, [125I]‐BQ3020 only recognised approximately one third the number of endothelin receptors measured with [125I]‐ET‐1.
Saturation binding experiments with [125I]‐PD151242 revealed high affinity binding to a single population of ETA receptors in the cerebellar homogenates (pKd = 9.95 ± 0.14; Bmax = 30 ± 15 fmol mg−1 protein).
Competition experiments were performed with ligands that are either non‐selective or selective for endothelin receptor subtypes. The rat cerebellar endothelin receptor displayed a high affinity for endothelin‐1 (ET‐1), endothelin‐3 (ET‐3) and sarafotoxin‐S6c (STX‐6c) although the affinity for ET‐3 was slightly higher than the affinity for ET‐1 using both radioligands. The selective ETA antagonists, BQ123, BMS‐182,874 and JKC‐301 all displayed low affinities at the endothelin receptors. In contrast the selective ETB agonists, IRL1620 and [Ala1,3,11,15]ET‐1 and the selective ETB antagonist, BQ‐788 had moderate affinities at the endothelin receptor, in the low nanomolar range. The ETB agonist, BQ3020, had approximately 10 fold higher affinity than IRL1620 and [Ala1,3,11,15]ET‐1 at the rat cerebellar endothelin receptors. The non‐selective antagonists, Ro‐46,2005, Ro‐47,0203 and PD‐142,893 displayed moderate affinities at the cerebellar receptor.
Since [125I]‐BQ3020 recognises only a fraction of the [125I]‐ET‐1 binding sites, the majority of the endothelin receptors in the cerebellum cannot be classed as ETB. Although [125I]‐PD151242 was able to detect ETA receptors in the rat cerebellar homogenates, the small population of ETA receptors (2% of the total endothelin population as measured with [125I]‐ET‐1) could not account for the non‐ETB receptor population. We conclude that the rat brain cerebellar receptor has a profile similar to the ETB1 receptor as it has a high affinity for ET‐1, ET‐3, STX‐6c and was moderately sensitive to PD‐142,893. However, as the ETB ligands BQ‐788, IRL1620 and [Ala1,3,11,15]ET‐1 have only a moderate affinity for the rat cerebellar endothelin receptor and since ET‐3 has a higher affinity as compared to ET‐1, our findings suggest that the rat cerebellum contains predominately ETC receptors.
Rapid nitric oxide‐ and prostaglandin‐dependent release of calcitonin gene‐related peptide (CGRP) triggered by endotoxin in rat mesenteric arterial bed
Authors: Wang Xian , Wu Zhongxin , Tang Yueming , Fiscus Ronald R. , Han Chide 
Published: July 2012

Our objective was to determine whether endotoxin (ETX) could directly trigger the release of calcitonin gene‐related peptide (CGRP) from perivascular sensory nerves in the isolated mesenteric arterial bed (MAB) of the rat and to determine whether nitric oxide (NO) and prostaglandins (PGs) are involved.
ETX caused time‐ and concentration‐dependent release of CGRP, and as much as a 17 fold increase in CGRP levels in the perfusate at 10–15 min after the administration of ETX (50 μg ml−1).
CGRP‐like immunoreactivity in the perfusate was shown to co‐elute with synthetic rat CGRP by reverse‐phase h.p.l.c.
Pretreatment of MAB with capsaicin or ruthenium red inhibited ETX‐induced CGRP release by 90% and 71%, respectively. ETX‐evoked CGRP release was decreased by 84% during Ca2+‐free perfusion.
The release of CGRP evoked by ETX was enhanced by L‐arginine by 43% and inhibited by Nω‐nitro‐L‐arginine (L‐NOARG) and methylene blue by 37% and 38%, respectively. L‐Arginine reversed the effect of L‐NOARG.
Indomethacin and ibuprofen also inhibited the ETX‐induced CGRP release by 34% and 44%, respectively. No additive inhibition could be found when L‐NOARG and indomethacin were concomitantly incubated.
The data suggest that ETX triggers the release of CGRP from capsaicin‐sensitive sensory nerves innervating blood vessels. The ETX‐induced CGRP release is dependent on extracellular Ca2+ influx and involves a ruthenium red‐sensitive mechanism. Both NO and PGs appear to be involved in the ETX‐induced release of CGRP in the rat mesenteric arterial bed.
Influence of superoxide dismutase inhibition on the discrimination between NO and the nitrergic neurotransmitter in the rat gastric fundus
Authors: Lefebvre R.A. 
Published: July 2012

The influence of diethyldithiocarbamate (DETCA), that irreversibly inhibits Cu/Zn‐containing superoxide dismutase, on the inability of 6‐anilino‐5,8‐quinolinedione (LY83583), hypoxanthine/xanthine oxidase, hydroquinone and hydroxocobalamin to reduce electrically‐induced NANC relaxations in the rat gastric fundus was investigated.
Longitudinal muscle strips of the rat gastric fundus were mounted for auxotonic recording in the presence of atropine and guanethidine and tone was raised by administration of prostaglandin F2α. DETCA (3 × 10−3 m) slightly reduced the short‐lasting relaxations induced by 10−5 m exogenous nitric oxide (NO) and transmural electrical stimulation for 10 s at 4 Hz but this effect was not influenced by 1000 u ml−1 superoxide dismutase (SOD).
DETCA (3 × 10−5‐3 × 10−3 m) concentration‐dependently potentiated the inhibitory effect of LY83583 upon the electrically‐induced relaxations, although this was less pronounced than the inhibition of the NO‐induced relaxations. The inhibition of the electrically‐induced non‐adrenergic non‐cholinergic (NANC) relaxations was not reversed by SOD while that of the NO‐induced relaxations was partially reversed.
The inhibitory effect of hypoxanthine/xanthine oxidase, hydroquinone and hydroxocobalamin on the electrically‐induced NANC relaxations in the presence of DETCA (3 × 10−3 m) was not different from the inhibitory effect of DETCA alone.
It was concluded that the differentiating effect of LY83583 between exogenous NO and the endogenous nitrergic neurotransmitter is partially related to protection of the endogenous nitrergic neurotransmitter by high levels of intracellular superoxide dismutase. This mechanism does not hold for hydroquinone and hydroxocobalamin, as they still discriminate between exogenous NO and the endogenous nitrergic neurotransmitter in the presence of DETCA. The possibility that the endogenous nitrergic neurotransmitter is not free NO in the rat gastric fundus therefore remains open.
Inhibition by antioxidants of nitric oxide synthase expression in murine macrophages: role of nuclear factor kB and interferon regulatory factor 1
Authors: Hecker Markus , Preiß Christiane , Klemm Peter , Busse Rudi 
Published: July 2012

In view of the potential deleterious effects of high amounts of nitric oxide (NO) produced by the inducible isoform of NO synthase (iNOS) in inflammation, the prevention of the expression of this enzyme represents an important therapeutic goal. In cytokine‐stimulated cells, activation of nuclear factor kB (NF‐kB) is crucial for the increase in iNOS gene expression. Since NF‐kB activation appears to involve a redox‐sensitive step, we have investigated whether three structurally unrelated antioxidants, 5,7‐dihydroxyflavone (chrysin), 3,4‐dichloroisocoumarin (DCI) and N‐acetyl 5‐hydroxytryptamine (N‐acetylserotonin, NAS), affect iNOS expression in cultured RAW 264.7 monocyte/macrophages stimulated with bacterial lipopolysaccharide (LPS, 140 ng ml−1) and interferon‐γ (IFNγ, 5 u ml−1).
During a 6 h incubation period neither LPS nor IFNγ alone exerted a significant effect but when combined, caused a prominent increase in nitrite formation, iNOS mRNA and protein abundance. Co‐incubation with chrysin (50 μm), DCI (50 μm) or NAS (1 mM) markedly attenuated this increase in iNOS gene expression.
DCI, but not chrysin or NAS, prevented the activation of NF‐kB in cells exposed to LPS plus IFNγ for 30 min. In contrast, all three antioxidants significantly blunted the DNA‐binding activity of interferon regulatory factor 1 (IRF‐1), which mediates the synergistic effect of IFNγ on iNOS gene expression in cells treated for 2 h with LPS plus IFNγ.
DCI thus appears to inhibit iNOS gene expression at the transcriptional level by preventing the activation of both NF‐kB and IRF‐1. The inhibitory effect of DCI on NF‐kB activation, however, does not seem to be related to its antioxidative properties, since DCI, unlike chrysin or NAS, is a potent serine protease inihbitor which stabilizes the inactive NF‐kB complex by protecting the inhibitory IkB‐α subunit from proteolytic degradation.
The virtually identical inhibitory effect of chrysin, DCI and NAS on the activation of IRF‐1 points to a redox‐sensitive step in the activation of this transcription factor, which in contrast to NF‐kB requires de novo protein synthesis.
Since iNOS gene expression in human cells and tissues usually requires the combination of several cytokines, antioxidants such as chrysin and NAS which do not interfere with the activation of NF‐kB may be of therapeutic value for selectively inhibiting the enhanced expression of this enzyme in inflammation.
Effects of cyclic GMP elevation on isoprenaline‐induced increase in cyclic AMP and relaxation in rat aortic smooth muscle: role of phosphodiesterase 3
Authors: Delpy Eric , Coste Hervé , de Gouville Anne‐Charlotte le Monnier 
Published: July 2012

In rat aortic rings precontracted with phenylephrine, the β‐adrenoceptor agonist isoprenaline (10 nM to 30 μm) produces greater relaxant effects in preparations with endothelium than in endothelium‐denuded preparations. The aim of this study was to determine the mechanisms involved in this effect and in particular investigate the possibility of a synergistic action between adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) and guanosine 3′:5′‐cyclic monophosphate (cyclic GMP).
Isoprenaline‐induced relaxation of rat aortic rings precontracted with phenylephrine was greatly reduced by the nitric oxide (NO) synthase inhibitor Nω‐nitro‐L‐arginine methyl ester (L‐NAME, 300 μm) or the soluble guanylate cyclase inhibitors methylene blue (10 μm) or 1H‐[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one (ODQ, 10 μm) but unaffected by indomethacin (10 μm), a cyclo‐oxygenase inhibitor. Similarly, in intact rings, the concentration‐response curve of forskolin (10 nM to 1 μm) was shifted to the right upon endothelium removal or treatment with methylene blue.
In endothelium‐denuded rat aortic rings, isoprenaline‐induced relaxation was potentiated by the guanylate cyclase activators atrial natriuretic factor (ANF, 1 to 10 nM) and sodium nitroprusside (SNP, 1 to 10 nM), and to a greater extent in the presence of the cyclic GMP‐specific phosphodiesterase (PDE 5) inhibitor, 1,3dimethyl‐6‐(2‐propoxy‐5‐methane sulphonylamidophenyl) pyrazolo [3,4‐d] pyrimidin‐4‐(5H)‐one (DMPPO, 30 nM). Relaxation induced by isoprenaline was also potentiated by the cyclic GMP‐inhibited PDE (PDE 3) inhibitor cilostamide (100 nM).
Intracellular cyclic nucleotide levels were measured either in rat cultured aortic smooth muscle cells or in de‐endothelialized aortic rings. In both types of preparation, isoprenaline (5 nM and 10 μm) increased cyclic AMP levels and this effect was potentiated by cilostamide (10 μm), by rolipram, a cyclic AMP‐specific PDE (PDE 4) inhibitor (10 μm) and by cyclic GMP‐elevating agents (50 nM ANF or 30 nM SNP plus 100 nM DMPPO). In isoprenaline‐stimulated conditions, the increase in cyclic AMP induced by rolipram was further potentiated by cilostamide and by cyclic GMP‐elevating agents. Cilostamide and cyclic GMP‐elevating agents did not potentiate each other, suggesting a similar mechanism of action.
We conclude that in vascular smooth muscle (VSM) cells an increase in cyclic GMP levels may inhibit PDE 3 and, thereby, cyclic AMP catabolism. Under physiological conditions of constitutive NO release, and to a greater extent in the presence of the PDE 5 inhibitor DMPPO, cyclic GMP should act synergistically with adenylate cyclase activators to relax VSM.
The effect of 5‐HT and selective 5‐HT receptor agonists and antagonists on rat dorsal vagal preganglionic neurones in vitro
Authors: Albert Anthony P. , Spyer K. Michael , Brooks Penelope A. 
Published: July 2012

Whole‐cell patch‐clamp recordings were made from 142 visually identified rat dorsal vagal preganglionic neurones (DVMs). Applications of 5‐hydroxytryptamine (5‐HT, 20 μm, 2 min) elicited a slow depolarization (8.2 ± 0.5 mV, n = 59) in 95% of the cells tested, accompanied by an increase in excitability. In (68%) of DVMs the depolarization was associated with an increase in apparent membrane resistance (Rm, 22.7 ± 2.2%). These depolarizations and increases in Rm (14.3 ± 2.6%, n = 8) were maintained in a medium which blocked synaptic transmission.
The response to 5‐HT was associated with a reversal potential (Erev) of −91 ± 1 mV at an extracellular K+ concentration ([K+]o) of 4.2 mM. This correlated well with the K+ equilibrium potential (EK = −89 mV).
The depolarizing effect of 5‐HT was attenuated by the 5‐HT2A/2C receptor antagonists, ketanserin (1 μm), LY 53,857 (1 μm) and the 5‐HT1A/2A receptor antagonist, spiperone (1 γM). The 5‐HT1A receptor antagonist, pindobind 5‐HT1A (5 μm), had no effect on the depolarizing response to 5‐HT.
The effect of 5‐HT was mimicked by the 5‐HT2A/2C receptor agonist, α‐methyl‐5‐HT (50 μm), the 5‐HT1 receptor agonist, 5‐carboxamidotryptamine (20 μm) and the putative 5‐HT4 agonist, 5‐methyoxytryptamine (50 μm). The selective 5‐HT4 receptor antagonist, GR113808, had no effect on the depolarizing effect of 5‐HT or 5‐MEOT on DVMs.
The 5‐HT3 antagonists, MDL 72222 (10 μm) and ICS‐205–930 (1 and 10 μm), partially reduced the effect of 5‐HT. The 5‐HT3 receptor agonist, 2‐methyl‐5‐HT (100–300 μm), excited a proportion of neurones tested (56%) by evoking a depolarizing and/or an increase in postsynaptic potentials (p.s.ps).
These results are consistent with direct, postsynaptic actions of 5‐HT on DVMs via 5‐HT2A receptors, being mediated, in part, by the reduction of K+ conductance.
The nitric oxide‐cyclic GMP pathway and synaptic plasticity in the rat superior cervical ganglion
Authors: Southam Eric , Charles Sarah L. , Garthwaite John 
Published: July 2012

We have investigated the possibility that nitric oxide (NO) and soluble guanylyl cyclase, an enzyme that synthesizes guanosine 3′:5′‐cyclic monophosphate (cyclic GMP) in response to NO, contributes to plasticity of synaptic transmission in the rat isolated superior cervical ganglion (SCG).
Exposure of ganglia to the NO donor, nitroprusside, caused a concentration‐dependent accumulation of cyclic GMP which was augmented in the presence of the phosphodiesterase inhibitor, 3‐isobutyl‐1‐methylxanthine. The compound, 1H‐[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one (ODQ), a selective inhibitor of soluble guanylyl cyclase, completely blocked this cyclic GMP response.
As assessed by extracellular recording, nitroprusside (100 μm) and another NO donor, S‐nitrosoglutathione (30 μm) increased the efficacy of ganglionic synaptic transmission in response to electrical stimulation of the preganglionic nerve, an effect that was reversible and which could be replicated by the cyclic GMP analogue, 8‐bromo‐cyclic GMP. Ganglionic depolarizations resulting from stimulation of nicotinic receptors with carbachol were not increased by nitroprusside. The potentiating actions of the NO donors on synaptic transmission, but not that of 8‐bromo‐cyclic GMP, were inhibited by ODQ.
Brief tetanic stimulation of the preganglionic nerve resulted in a long‐term potentiation (LTP) of synaptic transmission that was unaffected by ODQ, either in the absence or presence of the NO synthase inhibitor, NG‐nitro‐L‐arginine (L‐NOARG, 100 μm). A lack of influence of L‐NOARG was confirmed in intracellular recordings of LTP of the excitatory postsynaptic potential. Furthermore, under conditions where tetanically‐induced LTP was saturated, nitroprusside was still able to potentiate synaptic transmission, as judged from extracellular recording.
We conclude that NO is capable of potentiating ganglionic neurotransmission and this effect is mediated through the stimulation of soluble guanylyl cyclase and the accumulation of cyclic GMP. However, this potentiation is distinct from LTP of nicotinic synaptic transmission, in which neither NO nor soluble guanylyl cyclase appear to participate.
Role of a copper (I)‐dependent enzyme in the anti‐platelet action of S‐nitrosoglutathione
Authors: Gordge M.P. , Hothersall J.S. , Neild G.H. , Dutra A.A. Noronha 
Published: July 2012

S‐nitrosoglutathione (GSNO) is a potent and selective anti‐platelet agent, despite the fact that its spontaneous rate of release of nitric oxide (NO) is very slow. Our aim was to investigate the mechanism of the anti‐aggregatory action of GSNO.
The biological action of GSNO could be mediated by NO released from S‐nitrosocystylglycine, following enzymatic cleavage of GSNO by γ‐glutamyl transpeptidase. The anti‐aggregatory potency of GSNO was not, however, altered by treatment of target platelets with the γ‐glutamyl transpeptidase inhibitor acivicin (1 mM). γ‐Glutamyl transpeptidase is not, therefore, involved in mediating the action of GSNO.
The rate of breakdown of S‐nitrosoalbumin was increased from 0.19 ± 0.086 nmol min−1 to 1.52 ± 0.24 nmol min−1 (mean±s.e.mean) in the presence of cysteine (P < 0.05, n = 4). Inhibition of platelet aggregation by S‐nitrosoalbumin was also significantly increased by cysteine (P < 0.05, n = 4), suggesting that the biological activity of S‐nitrosoalbumin is mediated by exchange of NO from the protein carrier to form the unstable compound cysNO. Breakdown of GSNO showed a non‐significant acceleration in the presence of cysteine, from 0.56 ± 0.22 to 1.77 ± 0.27 nmol min−1 (mean±s.e.mean) (P = 0.064, n = 4), and its ability to inhibit platelet aggregation was not enhanced by cysteine. This indicates that the anti‐platelet action of GSNO is not dependent upon transnitrosation to form cysNO.
Platelets pretreated with the copper (I)‐specific chelator bathocuproine disulphonic acid (BCS), then resuspended in BCS‐free buffer, showed resistance to the inhibitory effect of GSNO. These findings suggest that BCS impedes the action of GSNO by binding to structures on the platelet, rather than by chelating free copper in solution.
Release of NO from GSNO was catalysed enzymatically by ultrasonicated platelet suspensions. This enzyme had an apparent Km for GSNO of 12.4 ± 2.64 μm and a Vmax of 0.21 ± 0.03 nmol min−1 per 108 platelets (mean±s.e.mean, n = 5). It was inhibited by BCS, but not by the iron chelator bathophenathroline disulphonic acid, nor by acivicin.
We conclude that the stable S‐nitrosothiol compound GSNO may exert its anti‐platelet action via enzymatic, rather than spontaneous release of NO. This is mediated by a copper‐dependent mechanism. The potency and platelet‐selectivity of GSNO may result from targeted NO release at the platelet surface.
Effects of γ‐aminobutyric acid (GABA) agonists and GABA uptake inhibitors on pharmacosensitive and pharmacoresistant epileptiform activity in vitro
Authors: Pfeiffer Michael , Draguhn Andreas , Meierkord Hartmut , Heinemann Uwe 
Published: July 2012

Lowering of the extracellular Mg2+‐concentration induces various patterns of epileptiform activity in combined rat entorhinal cortex‐hippocampal brain slices. After a prolonged period of exposure to Mg2+‐free medium seizure‐like events in the entorhinal cortex change to a state of late recurrent discharges which cannot be blocked by clinically available antiepileptic drugs. This late epileptiform activity thus represents a useful model to test the effects of new anticonvulsant substances.
A mechanism possibly underlying the development of sustained seizure‐like activity is the loss of synaptically released γ‐aminobutyric acid (GABA). Drugs which increase the amount of GABA available in presynaptic endings might thus be useful in the treatment of these therapeutically complicated forms of epilepsy.
Therefore, we studied the effects of various substances increasing GABA‐mediated inhibition on early and late forms of epileptiform activity. GABA and the GABAA receptor agonist muscimol blocked both the pharmacosensitive discharges in the hippocampus and entorhinal cortex as well as the late recurrent discharges in the medial entorhinal cortex. The GABAB receptor agonist baclofen blocked the recurrent short discharges very potently, but did not consistently block seizure‐like events and late recurrent discharges in the entorhinal cortex.
GABA uptake blockers showed a differential potency to block the various discharge patterns. Whereas nipecotic acid and β‐alanine suppressed all forms of epileptiform activity albeit at high concentrations (1–5 mM), tiagabine was much more potent in blocking the hippocampal recurrent short discharges and the seizure‐like events in the medial entorhinal cortex, but could not block the late recurrent discharges.
Our data support the idea that prolonged neuronal overactivity might result in a loss of synaptically available GABA. Selective block of uptake into glia cells or substitution of the transmitter may therefore be an efficient strategy for the treatment of severe prolonged epileptic discharges whereas block of neuronal GABA uptake fails to counteract synchronized discharges in this situation.
Influence of levosimendan, pimobendan, and milrinone on the regional distribution of cardiac output in anaesthetized dogs
Authors: Pagel Paul S. , Hettrick Douglas A. , Warltier David C. 
Published: July 2012

The distribution of cardiac output during administration of levosimendan, a new myofilament calcium sensitizer, is unknown. We examined and compared the effects of levosimendan, pimobendan, and milrinone on regional tissue perfusion by use of the radioactive microsphere technique in barbiturate‐anaesthetized dogs.
Haemodynamics and regional blood flow were determined before and during infusions of levosimendan (0.75, 1.5, and 3.0 μg kg−1 min−1), pimobendan (10, 20, and 40 μg kg−1 min−1), or milrinone (1.0, 2.0, and 4.0 μg kg−1 min−1).
All three drugs caused similar increases in heart rate, cardiac output, and left ventricular + dP/dt and decreases in end‐diastolic pressure and systemic vascular resistance. No changes in subendocardial, midmyocardial, and subepicardial blood flow occurred during administration of levosimendan. However, a redistribution of blood flow from subendocardium to subepicardium was observed. Pimobendan increased midmyocardial and subepicardial blood flow and reduced the endo/epi ratio to a greater degree than levosimendan. Milrinone did not affect myocardial perfusion.
Levosimendan increased blood flow to the renal medulla and decreased renal medullary and cortical vascular resistance. Levosimendan increased blood flow to the small intestine and liver and reduced vascular resistance in these organs. Pimobendan increased hepatic blood flow to a greater degree than levosimendan but did not alter small intestinal perfusion. All three drugs decreased splenic blood flow to similar degrees. Levosimendan and pimobendan reduced cerebral vascular resistance. Levosimendan and milrinone reduced skeletal muscle vascular resistance.
The results indicate that levosimendan, pimobendan, and milrinone cause subtlety different alterations in regional tissue perfusion while producing similar haemodynamic effects.