bluuebunny/crossref_metadata_2025_split huggingface dataset:
Involvement of calyculin A inhibitable protein phosphatases in the cyclic AMP signal transduction pathway of mouse corticotroph tumour (AtT20) cells
Authors: Antaraki A , Ang K L , Antoni F A 
Published: August 2006

The role of non‐calcineurin protein phosphatases in the cyclic AMP signal transduction pathway was examined in mouse pituitary corticotroph tumour (AtT20) cells.
Blockers of protein phosphatases, calyculin A and okadaic acid, were applied in AtT20 cells depleted of rapidly mobilizable pools of intracellular calcium and activated by various cyclic AMP generating agonists. Inhibitors of cyclic nucleotide phosphodiesterases were present throughout. The accumulation of cyclic AMP was monitored by radioimmunoassay, phosphodiesterase activity in cell homogenates was measured by radiometric assay.
Neither calyculin A nor okadaic acid altered basal cyclic AMP levels but cyclic AMP formation induced by 41 amino acid residue corticotrophin releasing‐factor (CRF) was strongly inhibited (up to 80%). 1‐Norokadaone was inactive. Similar data were also obtained when isoprenaline or pituitary adenylate cyclase activating peptide1–38 were used as agonists.
Pertussis toxin did not modify the inhibition of CRF‐induced cyclic AMP production by calyculin A.
Pretreatment with calyculin A completely prevented the stimulation of cyclic AMP formation by cholera toxin even in the presence of 0.5 mM isobutylmethylxanthine (IBMX) and 0.1 mM rolipram. Cholera toxin mediated ADP‐ribosylation of the 45K and 52K molecular weight Gsα isoforms in membranes from calyculin A‐pretreated cells was enhanced to 150–200% when compared with controls.
Cholera toxin‐induced cyclic AMP was reduced by calyculin A within 10 min when calyculin A was applied after a 90 min pretreatment with cholera toxin. Under these conditions the effect of calyculin A could be blocked by the combination of 0.5 mM IBMX and 0.1 mM rolipram, but not by 0.5 mM IBMX alone.
Phosphodiesterase activity in AtT20 cell homogenates showed a significant, 2.7 fold increase after treatment with calyculin A. In control cells phosphodiesterase activity was blocked by 80% in the presence of IBMX (0.5 mM), or IBMX plus rolipram (0.1 mM). In calyculin A‐treated cells phosphodiesterase activity was also strongly inhibited by IBMX, but because of the stimulating effect of calyculin A, the activity remaining was still 55% of that found in control homogenates. This activity was reduced to 5% of control by using IBMX and rolipram in combination. Assay of phosphodiesterase in Ca2+ free conditions showed that calyculin A markedly increases the activity of rolipram sensitive (type 4) phosphodiesterase.
Taken together, blockers of protein phosphatases (PPases) impaired signal transduction through Gs‐mediated pathways and activated cyclic AMP degrading phosphodiesterase(s), indicating that PPases 1 and/or 2A are essential for agonist‐mediated regulation of cyclic AMP levels in AtT20 cells, and are thus important in maintaining the secretory phenotype of the cells.
NMDA receptor‐mediated pilocarpine‐induced seizures: characterization in freely moving rats by microdialysis
Authors: Smolders Ilse , Khan Ghous M , Manil Jacqueline , Ebinger Guy , Michotte Yvette 
Published: August 2006

Pilocarpine administration has been used as an animal model for temporal lobe epilepsy since it produces several morphological and synaptic features in common with human complex partial seizures. Little is known about changes in extracellular neurotransmitter concentrations during the seizures provoked by pilocarpine, a non‐selective muscarinic agonist.
Focally evoked pilocarpine‐induced seizures in freely moving rats were provoked by intrahippocampal pilocarpine (10mm for 40min at a flow rate of 2μl min−1) administration via a microdialysis probe. Concomitant changes in extracellular hippocampal glutamate, γ‐aminobutyric acid (GABA) and dopamine levels were monitored and simultaneous electrocorticography was performed. The animal model was characterized by intrahippocampal perfusion with the muscarinic receptor antagonist atropine (20mm), the sodium channel blocker tetrodotoxin (1μm) and the N‐methyl‐d‐aspartate (NMDA) receptor antagonist MK‐801 (dizocilpine maleate, 100μm). The effectiveness of locally (600μm) or systemically (10mg kg−1 day−1) applied lamotrigine against the pilocarpine‐induced convulsions was evaluated.
Pilocarpine initially decreased extracellular hippocampal glutamate and GABA levels. During the subsequent pilocarpine‐induced limbic convulsions extracellular glutamate, GABA and dopamine concentrations in hippocampus were significantly increased. Atropine blocked all changes in extracellular transmitter levels during and after co‐administration of pilocarpine. All pilocarpine‐induced increases were completely prevented by simultaneous tetrodotoxin perfusion. Intrahippocampal administration of MK‐801 and lamotrigine resulted in an elevation of hippocampal dopamine levels and protected the rats from the pilocarpine‐induced seizures. Pilocarpine‐induced convulsions developed in the rats which received lamotrigine perorally.
Pilocarpine‐induced seizures are initiated via muscarinic receptors and further mediated via NMDA receptors. Sustained increases in extracellular glutamate levels after pilocarpine perfusion are related to the limbic seizures. These are arguments in favour of earlier described NMDA receptor‐mediated excitotoxicity. Hippocampal dopamine release may be functionally important in epileptogenesis and may participate in the anticonvulsant effects of MK‐801 and lamotrigine. The pilocarpine‐stimulated hippocampal GABA, glutamate and dopamine levels reflect neuronal vesicular release.
Differential effects of the neuropeptide galanin on striatal acetylcholine release in anaesthetized and awake rats
Authors: Antoniou K , Kehr J , Snitt K , Ögren S O 
Published: August 2006

In the present study the mechanisms were examined by which the neuropeptide galanin modulates the extracellular concentrations of striatal acetylcholine (ACh) in enflurane anaesthetized and in freely moving male rats by use of in vivo microdialysis and high performance liquid chromatography.
The perfusion of galanin through the microdialysis probe (0.3nmolμl−1, flow rate: 2μlmin−1) caused a statistically significant increase in the basal striatal ACh levels in anaesthetized but a decrease in awake animals. No significant effect was revealed after a low dose (0.1nmolμl−1, flow rate: 2μlmin−1) of galanin perfusion. Both the stimulating and inhibitory effects of galanin on basal ACh release were reversible.
The muscarinic antagonist scopolamine (0.1mgkg−1, subcutaneously (s.c.)) caused a significant increase in ACh release in both anaesthetized and awake animals.
The combination of galanin plus scopolamine attenuated the stimulant effect on ACh release caused by scopolamine alone in awake animals.
The putative galanin receptor antagonist M35 at 0.3nmolμl−1 but not at 0.1nmolμl−1 caused a significant reduction (20%) in ACh release, supporting the view that M35 at higher concentrations behaves as a partial agonist at the galanin receptor. When M35 (0.1nmolμl−1) was co‐infused with galanin (0.3nmolμl−1) the galanin‐evoked decrease in ACh release was completely blocked.
Taken together, these results indicate that galanin affects basal ACh release via stimulation of galanin receptors within the striatum. The mechanism involved is dependent on the anaesthesia procedure which may act via enhancement of γ‐aminobutyric acidA (GABAA) mediated transmission within striatal and/or output neurones. In addition, anaesthesia may also decrease the activity of glutamatergic striatal afferents. The results with M35 indicate that the role of galanin perfused in striatum is permissive in the normal rat. Furthermore, galanin is a potent inhibitory modulator of basal ACh release also in the striatum, as recently was shown in the ventral hippocampus in awake animals.
Differential antinociceptive effects of spinal opioids on foot withdrawal responses evoked by C fibre or Aδ nociceptor activation
Authors: Lu Y , Pirec V , Yeomans D C 
Published: August 2006

Intrathecal application of μ, δ, and κ opioids attenuate responses on several tests of animal nociception. However, the potency of these opioids differ depending on which tests were used. One factor contributing to these discrepancies is that different types of noxious stimuli activate different sets of nociceptor types, which may be differentially sensitive to opiate inhibition. To examine this hypothesis, we used a recently developed behavioural test which allows for differential assessment of nociception evoked by the activation of myelinated (Aδ) and unmyelinated C thermonociceptors.
Administration of a κ‐selective agonist was ineffective on either type of response. δ1 drugs were slightly more potent on C fibre‐mediated responses than on Aδ‐mediated responses.
Intrathecal μ and δ2 drugs were antinociceptive on both Aδ and C nociceptor‐mediated responses. However, unlike the δ1 effects, the dose‐response curves for μ and δ2 drugs were significantly more steep for Aδ than for C fibre‐mediated responses, potentially indicating differences in the mechanisms by which the drugs act on these 2 response types.
Involvement of vasoactive intestinal polypeptide in nicotine‐induced relaxation of the rat gastric fundus
Authors: Curròi Diego , Preziosi Paolo 
Published: August 2006

Nicotine‐induced relaxation and release of vasoactive intestinal polypeptide (VIP)‐ and peptide histidine isoleucine (PHI)‐like immunoreactivity (LI) were measured in longitudinal muscle strips from the rat gastric fundus.
Under non‐cholinergic conditions (0.3μm atropine), nicotine (3–300 μm) produced concentration‐dependent relaxations of the 5‐hydroxytryptamine (3μm)‐precontracted strips. Under non‐adrenergic non‐cholinergic (NANC) conditions (0.3μm atropine+1μm phentolamine+1μm nadolol), relaxations induced by sub‐maximal nicotine concentrations (10 and 30μm) were significantly smaller, while that produced by the highest concentration used (300μm) was similar to that seen under non‐cholinergic conditions.
Re‐exposure to the same nicotine concentration 1h later induced smaller relaxations, indicating desensitization. The reductions seen in the second responses were proportional to the concentration used.
Under non‐cholinergic conditions, the relaxant response to 30μm nicotine was abolished by hexamethonium (100μm) and significantly reduced by tetrodotoxin (TTX, 3μm). The TTX‐resistant component was not observed under NANC conditions.
NANC relaxation induced by 30μm nicotine was significantly reduced by a specific anti‐VIP serum (approximately 35% less than that seen with normal rabbit serum).
Nicotine (30–300μm) caused significant, concentration‐dependent increases in the outflow of VIP‐ and PHI‐LI from the strips; these effects were also diminished with re‐exposure. The increases in both types of immunoreactivity evoked by nicotine (300μm) were abolished by hexamethonium (300μm), TTX (3μm) and a calcium‐free medium.
These findings indicate that VIP and possibly PHI are involved in NANC relaxation of the rat gastric fundus induced by nicotine.
Tolerance to μ‐opioid agonists in human neuroblastoma SH‐SY5Y cells as determined by changes in guanosine‐5′‐O‐(3‐[35S]‐thio)triphosphate binding
Authors: Elliott Jackie , Guo Li , Traynor John R 
Published: August 2006

The agonist action of morphine on membranes prepared from human neuroblastoma SH‐SY5Y cells was measured by an increase in the binding of the GTP analogue [35S]‐GTPγS. Morphine increased the binding of [35S]‐GTPγS to SH‐SY5Y cell membranes by 30 fmol mg−1 protein with an EC50 value of 76±10 nM.
Incubation of SH‐SY5Y cells with 10 μM morphine for 48 h caused a tolerance to morphine manifested by a 2.5 fold shift to the right in the EC50 value with a 31±6% decrease in the maximum stimulation of [35 S]‐GTPγS binding. The response caused by the partial agonist pentazocine was reduced to a greater extent.
Chronic treatment of the cells with the more efficacious μ‐ligand [D‐Ala2, MePhe4, Gly‐ol5]enkephalin (DAMGO, 10 μM) for 48 h afforded a greater effect than treatment with morphine. The maximal agonist effect of morphine was reduced to 58.9±6% of that seen in control cells while the maximal effect of DAMGO was reduced to 62.8±4%. There was a complete loss of agonist activity for pentazocine.
The development of tolerance was complete within 24 h and was blocked by naloxone and by the nonselective protein kinase inhibitor H7, but not by the putative β‐adrenoceptor kinase (β‐ARK) inhibitor suramin.
The observed tolerance effect was accompanied by a down‐regulation of μ‐opioid receptors determined by a decrease in the maximal binding capacity for the opioid antagonist [3H]‐diprenorphine of 66±4%, but with no change in binding affinity. Binding of the agonist [3H]‐DAMGO was similarly reduced.
The modulation of [35S]‐GTPγS binding in SH‐SY5Y cell membranes by opioids provides a simple method for the study of opioid tolerance at a site early in the signal transduction cascade.
mGluR‐evoked augmentation of receptor‐mediated cyclic AMP formation in neonatal and adult rat striatum
Authors: Cartmell Jayne , Schaffhauser Hervé , Wichmann Jürgen , Mutel Vincent 
Published: August 2006

The effects of selective agonists at group I, II and III metabotropic glutamate receptors (mGluRs) on adenosine A2 receptor‐mediated cyclic AMP formation were compared in cross‐chopped slices of adult and neonatal (8 days old) rat striatum, in the presence of 1 u ml−1 adenosine deaminase.
The group II selective agonist, (2S,1R,2R,3R)‐2‐(2,3‐dicarboxycyclopropyl)glycine (DCG‐IV), elicited a potentiation of 5′‐N‐ethylcarboxamidoadenosine (NECA)‐stimulated cyclic AMP production with similar potencies in adult (EC50 value 122±35 nM) and neonatal (EC50 value 285±6 nM) brain. In contrast, the group I selective agonist (S)‐dihydroxyphenylglycine ((S)‐DHPG) augmented the NECA cyclic AMP response in neonatal striatum (EC50 value 9±1 μM), but at a concentration of 100 μM, (S)‐DHPG failed to affect the NECA response in adult striatal slices.
The potentiation evoked by (S)‐DHPG was specific for group I mGluRs as (2S,3S,4S,)‐2‐methyl‐2‐(carboxycyclopropyl)glycine (MCCG), a group II antagonist, was ineffective on the (S)‐DHPG (100 μM) response at a concentration (500 μM) which reversed a similar augmentation elicited by DCG‐IV (300 nM). Furthermore, a protein kinase C inhibitor (Ro 31‐8220, 10 μM) markedly reversed the effect of (S)‐DHPG without affecting the response to DCG‐IV.
The mGluR agonist (2S,3S,4S,)‐α‐(carboxycyclopropyl)glycine (L‐CCG‐I), elicited a greater potentiation of NECA‐stimulated cyclic AMP production in neonatal striatum in comparison with that observed in adult rat brain. Moreover, EC50 values obtained from adult and neonatal striatum were 2±1 μM and 9±1 μM, respectively. These differences in potency might reflect co‐activation of both group I and group II mGluRs by L‐CCG‐I in neonatal striatum.
Distinct patterns of mGluR expression in various brain areas might account for previous conflicting data on the nature of the mGluR able to evoke such potentiated responses.
Electrically‐evoked dopamine and acetylcholine release from rat striatal slices perfused without magnesium: regulation by glutamate acting on NMDA receptors
Authors: Jin Shaoyu , Fredholm Bertil B 
Published: August 2006

Rat striatal slices, preincubated with [3H]‐dopamine and [14C]‐choline, were continuously superfused and electrically stimulated. Electrically evoked release of [3H]‐dopamine and [14C]‐acetylcholine (ACh) was not significantly changed by elimination of Mg2+ from superfusion buffer, but the basal release of [3H]‐dopamine was doubled.
Kynurenic acid (100–800 μM) caused, in the absence but not presence of Mg2+, a concentration‐dependent decrease in the evoked release of these two transmitters. The addition of glycine reversed the inhibition of the evoked release of both transmitters caused by kynurenic acid (400 μM) in a concentration‐dependent manner. In addition, glycine increased the evoked release of [3H]‐dopamine via a site inhibitable by strychnine (1 μM).
Another two antagonists at N‐methyl‐D‐aspartate (NMDA) receptors, 2‐amino‐5‐phosphonovaleric acid and dizocilpine, also decreased significantly the evoked release of the two transmitters in a concentration‐dependent manner in the absence, but not presence of Mg2+. By contrast, an antagonist of non‐NMDA receptors, 6‐cyano‐7‐nitroquinoxaline‐2,3‐dione (10 μM) significantly decreased the evoked release of the two transmitters in the presence, but not in the absence of Mg2+.
Electrical field stimulation evoked release of endogenous adenosine, and this release tended to be higher in the absence of Mg2+. However, the addition of a selective adenosine A1 receptor antagonist 8‐cyclopentyl‐1,3‐dipropylxanthine (200 nM) did not influence the evoked release of the two transmitters, showing that the released adenosine is of little importance in controlling ACh and dopamine release from striatal slices. Non‐NMDA receptors may play a similar role when Mg2+ ions are present.
The results indicate that NMDA receptors activated in the absence of Mg2+ participate in the electrically‐evoked release of [3H]‐dopamine and [14C]‐ACh from the striatum.
Interactions between loreclezole, chlormethiazole and pentobarbitone at GABAA receptors: functional and binding studies
Authors: Zhong Yu , Simmonds Michael A 
Published: August 2006

Interations were investigated between loreclezole, chlormethiazole and pentobarbitone as potentiators of depolarization responses mediated by γ‐aminobutyric acidA (GABAA) receptors on afferent nerve terminals in the rat cuneate nucleus in vitro. These drugs were also compared as modulators of [3H]‐flunitrazepam (FNZ) binding to synaptic membranes prepared from rat whole brain homogenate.
In rat cuneate nucleus slices, the drugs shifted muscimol log dose–response lines to the left in an approximately parallel fashion with the result that 200 μM chlormethiazole potentiated muscimol responses by 0.567±0.037 log unit (mean±s.e.mean, n=4) while loreclezole gave a maximal potentiation at 10 μM of only 0.121±0.037 (n=6) log unit and 0.071±0.039 (n=22) at 50 μM.
While 50 μM chlormethiazole and 30 μM pentobarbitone showed no significant interactions between each other when potentiating muscimol responses in combination, 50 μM loreclezole in combination with either chlormethiazole or pentobarbitone attenuated their potentiating effects, possibly by inducing desensitization of GABAA receptors.
In the [3H]‐FNZ binding studies on well‐washed membranes, loreclezole enhanced binding to a maximum of 47.3±2.83% of control (mean±s.e.mean, n=3) at 300 μM. Scatchard analysis revealed no change in Bmax but a decrease in KD for [3H]‐FNZ from 3.9±0.29 nM to 2.7±0.10 nM (mean±s.e.mean, n=4) in the presence of 100 μM loreclezole. In contrast, 100 μM chlormethiazole caused no potentiation. A small component of the enhancement by loreclezole could be blocked by 100 μM bicuculline and could also be blocked by 100 μM chlormethiazole. It seems likely that the effects on [3H]‐FNZ binding are due predominantly to direct actions of the drugs on the GABAA receptor and are separate from the GABA‐potentiating effects.
The results indicate distinctly different profiles of action for loreclezole, chlormethiazole and pentobarbitone on GABAA receptors.
Effect of oxytocin as a partial agonist at vasoconstrictor vasopressin receptors on the human isolated uterine artery
Authors: Jovanović Aleksandar , Jovanović Sofija , Tulić Ivan , Grbović Leposava 
Published: August 2006

The effect of oxytocin on endothelium‐intact and endothelium‐denuded segments of the human uterine artery rings was investigated.
In both types of preparation oxytocin induced contraction of human uterine artery with similar potency and efficacy (pEC50 values: 6.95±0.05 vs 7.06±0.01; maximal response values: 61±4.1% vs 63±5.1% for arteries with and without endothelium, respectively).
In contrast, human uterine arteries, both intact and denuded of endothelium, did not respond to the addition of the selective oxytocin receptor agonist, [Thr4, Gly7]oxytocin (10 nM–1 μM).
The vasopressin receptor antagonists, [d(CH2)5Tyr(Me)]AVP (10–100 nM) and [d(CH2)5,D‐Ile2,Ile4]AVP (300 nM–3 μM) produced parallel rightward shifts of the curves for oxytocin. The Schild plots constrained to a slope of unity gave the following −log KB values: [d(CH2)5Tyr(Me)] AVP vs [d(CH2)5,D‐Ile2,Ile4] AVP 9.24 vs 6.91 and 9.26 vs 6.84 for human uterine artery with intact and those denuded of endothelium, respectively. In contrast, in both types of preparations the oxytocin receptor antagonist, [d(CH2)5Tyr(OMe), 2Orn8]vasotocin (1 μM), did not significantly affect oxytocin‐induced contractions.
The calculated pKA values for oxytocin itself also did not differ between preparations: 6.56 and 6.43 for human uterine artery with and without endothelium, respectively. In both types of preparations, the receptor reserve (KA/EC50) was close to unity (intact vs denuded: 3.9 vs 3.0).
It is concluded that, in human uterine artery, oxytocin induces contractions that are not modulated by the endothelium. It is likely that oxytocin acts as a partial agonist on human uterine artery, regardless of the endothelial condition. On the basis of differential antagonists affinity and affinity of oxytocin itself, it is probable that receptors involved in oxytocin‐induced contraction in human uterine arteries belong to the V1A vasopressin receptors.