bluuebunny/crossref_metadata_2025 huggingface dataset:
Frequency‐dependent inhibition of neuronal activity by lappaconitine in normal and epileptic hippocampal slices
Authors: Ameri Angela , Metzmeier Petra , Peters Thies
Published: July 2012

Extracellular recording of the stimulus‐evoked population spike in the CA1 region of rat hippocampal slices in vitro was performed in order to investigate whether lappaconitine affects neuronal excitability. Lappaconitine is a diterpene alkaloid of plants of the Aconitum genus and has analgesic properties.

The results reveal an inhibitory action of lappaconitine (10 μm) manifested in a slow attenuation of the orthodromic and antidromic population spike.

The lappaconitine‐induced inhibitory action was activity‐dependent, that is, it was potentiated when frequency of electrical stimulation was increased. In contrast, washout of the neurotoxin was accelerated when stimulation frequency was decreased.

The activity‐dependent action of lappaconitine raised the question of whether the drug is effective in suppressing the aberrant neuronal activity that occurs during an epileptic seizure. The results obtained from experiments on epileptic hippocampal slices demonstrated a selective reduction of the later spikes in the bursts with less effect on normal neuronal activity.

These data support the conclusion that lappaconitine, in addition to its antinociceptive effect, also has antiepileptic potency due to its highly activity‐dependent mode of action.
Activation by levcromakalim and metabolic inhibition of glibenclamide‐sensitive K channels in smooth muscle cells of pig proximal urethra
Authors: Teramoto Noriyoshi , Brading Alison F.
Published: July 2012

The effects of levcromakalim (BRL 38227) on ionic currents recorded from pig proximal urethra were investigated by use of tension measurement and patch clamp techniques (conventional whole‐cell configuration, nystatin perforated patch, and cell‐attached configuration).

Levcromakalim (1 μm) caused a relaxation in the resting tone. This levcromakalim‐induced relaxation was inhibited by the pretreatment with 1 μm glibenclamide.

The resting membrane potential recorded from single cells in current‐clamp mode was −36.1 ± 4.4 mV (n = 5).

Levcromakalim induced a concentration‐dependent hyperpolarization with a maximum (at ≥ 10 μm) close to the theoretical equilibrium potential of potassium (EK). The membrane hyperpolarization caused by 1 μm levcromakalim (24.7 ± 5.8 mV, n = 4) was abolished by 1 μm glibenclamide.

Levcromakalim (100 μm) caused an outward K current in whole‐cell recordings which was unaffected by iberiotoxin (300 nM) but abolished by glibenclamide (10 μm).

In cell‐attached patches, levcromakalim activated a 43 pS K channel which was inhibited by the application of glibenclamide.

The metabolic poison, cyanide (CN), also activated a 43 pS K channel which was suppressed by the application of 10 μm glibenclamide.

These results indicate that levcromakalim and metabolic inhibition activate the same 43 pS K channel in pig proximal urethra. The resultant urethral hyperpolarization might reduce the usefulness of K channel openers in the treatment of detrusor instability, but be of value in treating outflow obstruction.
Action of the calcium channel blocker lacidipine on cardiac hypertrophy and endothelin‐1 gene expression in stroke‐prone hypertensive rats
Authors: Feron Olivier , Salomone Salvatore , Godfraind Théophile
Published: July 2012

The tissue‐protective effects of calcium channel blockers in hypertension are not well dissociated from their effect on systolic blood pressure (SBP). We have previously shown that lacidipine, a dihydropyridine‐type calcium antagonist, reduced the cardiac hypertrophy and the cardiac endothelin‐1 (ET‐1) gene overexpression occurring in salt‐loaded stroke‐prone spontaneously hypertensive rats (SL‐SHRSP), an effect occurring without systolic blood pressure (SBP) change. In the present study, we have examined whether this action was dose‐related and if it could be associated with ET receptor changes. The action of lacidipine was also examined in control SHRSP and in Wistar Kyoto rats (WKY).

The daily dose of 0.3 mg kg−1 lacidipine which did not lower SBP but significantly prevented ventricle hypertrophy and cardiac preproET‐1‐mRNA expression in SL‐SHRSP was inactive in control SHRSP. With the higher dose of lacidipine (1 mg kg−1 day−1), we observed a further reduction of cardiac hypertrophy and of ET‐1 gene expression in SL‐SHRSP and a significant effect on those parameters in control SHRSP but only a small reduction of SBP in both groups.

In WKY, salt loading did not induce change in SBP or increase of cardiac ET‐1 gene expression and ventricle mass. In these normotensive rats, lacidipine (1 mg kg−1 day−1) did not modulate the basal preproET‐1‐mRNA expression and did not affect SBP or heart weight.

The maximum binding capacity (Bmax) and the dissociation constant (KD) of [125I]‐ET‐1 binding and the relative proportion of low‐ and high‐affinity binding sites for ET‐3 were not significantly affected by salt loading or lacidipine treatment in SHRSP.

These results show that lacidipine exerted a dose‐related inhibition of ventricle hypertrophy and preproET‐1‐mRNA expression in SHRSP and indicate that this effect was unrelated to SBP changes. The dose‐dependency of this inhibition suggests that salt‐induced cardiac hypertrophy could be related to ET‐1 gene overexpression. The results further show that ET receptor changes are not involved in the pathophysiological process studied here.
Further investigations into the mechanism of the antihypertensive activity of the angiotensin AT1 receptor antagonist, GR138950
Authors: Hilditch A. , Prior Helen M. , Drew G.M.
Published: July 2012

The angiotensin AT1 receptor antagonist, GR138950, produces a long‐lasting antihypertensive effect in conscious renal artery ligated hypertensive (RALH) rats but this effect does not correlate temporally with its antagonist profile against angiotensin II (AII). In the present experiments we have compared the inhibitory profiles of GR138950 and enalapril, against angiotensin I (AI), with their respective antihypertensive activities.

GR138950 (1 mg kg−1, i.a.) and enalapril (3 mg kg−1, i.a.) reduced blood pressure in RALH rats to a similar degree. Maximum reductions in blood pressure occurred approximately 5–24 h and 3–5 h after administration, respectively. The antihypertensive effect of GR138950 lasted for 24–48 h. However, the effect of enalapril lasted for only 5–24 h.

In conscious normotensive rats, inhibition of AI‐induced pressor responses was maximal 1 h after systemic administration of GR138950 and enalapril. Dose‐response curves to AI were displaced to the right, in a parallel manner, 1406 and 102 fold by GR138950 (1 mg kg−1, i.a.) and enalapril (3 mg kg−1 i.a.), respectively. The inhibitory effect of enalapril lasted for < 24 h whereas that of GR138950 lasted for up to 48 h.

Contractile responses to AI were extensively inhibited in aortae removed from either RALH rats or normotensive rats, 1 and 5 h after administration of GR138950 (1 mg kg−1, i.a.). Responses were still significantly reduced 24 h after administration but had returned to control levels after 48 h. Enalapril pretreatment (3 mg kg−1, i.a.) did not inhibit contractile responses to AI in aortae isolated from normotensive rats at any time point.

These experiments confirm that GR138950 is an effective and long‐lasting antihypertensive agent. GR138950 was a more potent and longer lasting antagonist against AI than has previously been found against AII, and the duration of its antihypertensive activity coincides better with its blockade of responses to AI. Blockade of the effects of AII generated locally within the vascular wall might play an important role in the antihypertensive profile of GR138950.
Presynaptic modulation by L‐glutamate and GABA of sympathetic co‐transmission in rat isolated vas deferens
Authors: Kwan Yiu Wa , Ngan Man‐Piu , Tsang Kay‐Yan , Lee Ha‐Man , Chu Lai‐Ah
Published: July 2012

The modulatory effects of L‐glutamate and its structural analogues, and of γ‐aminobutyric acid (GABA), on sympathetic co‐transmission were studied in the rat isolated vas deferens exposed to electrical field stimulation (EFS).

Application of exogenous L‐glutamate caused a concentration‐dependent (1 μm‐3 mM) inhibition of the rapid twitch component of the biphasic EFS contraction. However, L‐glutamate (1 μm‐3 mM) had a minimal effect on the phasic contraction induced by exogenous adenosine 5′‐triphosphate (ATP, 150 μm) and noradrenaline (50 μm). Unlike L‐glutamate, D‐glutamate had no effect on the EFS contraction.

The L‐glutamate‐induced inhibition of the EFS contractions was significantly attenuated by the glutamate decarboxylase (GAD) inhibitor 3‐mercapto‐propionic acid (150 μm) and was abolished in the presence of the GABA transaminase (GABA‐T) inhibitor, 2‐aminoethyl hydrogen sulphate (500 μm).

The L‐glutamate‐induced inhibition of the electrically evoked contraction was not affected by the adenosine A1‐receptor antagonist, 8‐cyclopentyl‐1, 3‐dipropylxanthine (DPCPX)(30 nM), reactive blue 2 (30 μm) or the GABAA receptor antagonist bicuculline (50 μm). However, the GABAB receptor antagonist 2‐hydroxysaclofen (50 μm) significantly inhibited the L‐glutamate effect.

Similar to L‐glutamate, GABA also caused a concentration‐dependent (0.1 – 100 μm) inhibition of the EFS contractions. This GABA‐induced inhibition was not affected by either the GABAA receptor antagonist bicuculline (50 μm) or reactive blue 2 (30 μm). However, a significant attenuation of the GABA‐mediated effect was recorded with the GABAB receptor antagonist 2‐hydroxysaclofen (50 μm). Contractions of the vas deferens induced by exogenous ATP and noradrenaline were not affected by GABA (0.1 – 100 μm).

The L‐glutamate analogues, N‐methyl‐D‐aspartate (NMDA) (1 μm‐1 mM) and quisqualate (Quis 0.1 μm‐0.3 mM) had no effect, whilst kainate (Kain, 1 μm‐1 mM) caused an inhibition of the EFS‐induced contractions. Effects of Kain could be abolished by the non‐NMDA receptor antagonist 6‐cyano‐7‐nitroquinoxaline‐2, 3‐dioxine (CNQX, 10 μm). NMDA, Quis and Kain had no effect on the exogenous ATP‐ or noradrenaline‐induced contractions.

It is concluded that the excitatory amino acid L‐glutamate modulates the electrically evoked vas deferens contraction through conversion to the inhibitory amino acid GABA by a specific GABA transaminase. The GABA formed may then act on GABAB receptors and cause inhibition of the contraction through a presynaptic mechanism.
Study of the mechanisms involved in adenosine‐5′‐O‐(2‐thiodiphosphate) induced relaxation of rat thoracic aorta and pancreatic vascular bed
Authors: Saïag B. , Hillaire‐Buys D. , Chapal J. , Petit P. , Pape D. , Rault B. , Allain H. , Loubatières‐Mariani M.M.
Published: July 2012

The endothelium‐dependent relaxation of blood vessels induced by P2Y‐purinoceptor activation has often been shown to involve prostacyclin and/or nitric oxide (NO) release. In this work, we have investigated the mechanisms involved in the relaxant effect of the P2Y agonist, adenosine −5′‐O‐(2‐thiodiphosphate) (ADPβS) using two complementary preparations: rat pancreatic vascular bed and aortic ring.

On the pancreatic vascular bed, ADPβS (1.5 and 15 μm) infused for 30 min induced a concentration‐dependent vasodilatation; it was progressive during the first 10 min (first period) and sustained from 10 to 30 min (second period). Indomethacin (10 μm) delayed ADPβS‐induced vasodilatation (1.5 and 15 μm) by about 6 min. Nω‐nitro‐L‐arginine methyl ester (L‐NAME) (200 μm) suppressed the relaxation for about 5 min but thereafter ADPβS at the two concentrations progressively induced an increase in the flow rate. Even the co‐administration of L‐NAME and indomethacin did not abolish the ADPβS‐induced vasorelaxation.

On 5‐hydroxy tryptamine (5‐HT) precontracted rings mounted in isometric conditions in organ baths, we observed that ADPβS induced a concentration‐dependent relaxation of rings with a functional endothelium; this effect was stable for 25 min. The ADPβS relaxant effect was strongly inhibited by Reactive Blue 2 (30 μm) and was suppressed by pretreatment of rings with saponin (0.05 mg ml−1 for 30 min), which also abolished the acetylcholine‐induced relaxation.

ADPβS‐induced relaxation of 5‐HT precontracted rings is largely inhibited by indomethacin (100 or 10 μm) or L‐NAME (100 μm).

We conclude that: the ADPβS‐induced relaxation is endothelium‐dependent, mediated by P2Y‐purinoceptors, and at least in part linked to NO and prostacyclin release, depending on the preparation used. Furthermore, on the pancreatic vascular bed, (an)other mechanism(s) than prostacyclin and NO releases may be involved in ADPβS‐induced vasodilatation.
Modulation by nitric oxide of spontaneous motility of the rat isolated duodenum: role of tachykinins
Authors: Martinez‐Cuesta M.A. , Esplugues J.V. , Whittle B.J.R.
Published: July 2012

Incubation of proximal segments of the rat isolated duodenum with NG‐nitro‐L‐arginine (L‐NOARG; 3–100 μm) produced a concentration‐dependent increase in both resting tone and the amplitude of the spontaneous contractions. These effects were attenuated by concurrent incubation with L‐arginine (1 mM) but not D‐arginine (1 mM).

These changes in resting tone and motility induced by L‐NOARG (30 μm) were substantially reduced by concurrent incubation with tetrodotoxin (1 μm) or hexamethonium (10 μm), implicating the involvement of a local neuronal response.

The L‐NOARG‐induced increase in duodenal motility was not, however, inhibited by atropine (1 μm), guanethidine (6.4 μm) phentolamine (1 μm), or indomethacin (10 μm), indicating a non‐cholinergic, non‐adrenergic and non‐prostanoid‐mediated contractile response.

The NK1/NK2 tachykinin receptor antagonist, (D‐Pro2, D‐Trp7,9 substance P, 1–10 μm), and the NK2‐receptor antagonists, MEN 10,207 and MEN 10,376 (1–5 μm), concentration‐dependently reduced the effect of L‐NOARG (30 μm) on spontaneous duodenal motility.

The resting tone and amplitude of the spontaneous contractions was likewise increased by incubation with NG‐monomethyl‐L‐arginine (L‐NMMA; 100–1000 μm). However, incubation with L‐NMMA (100 μm) attenuated the actions of more potent L‐NOARG (30 μm) on resting motility.

Administration of E.coli endotoxin (3 mg kg−1, i.v.) to the rat 5 h prior to tissue removal, at a time of known induction of NO synthase, reduced the amplitude of spontaneous contractions of the isolated duodenum, an effect inhibited by pretreatment of the rats with dexamethasone (1 mg kg−1) 2 h prior to endotoxin challenge.

These findings indicate a role of endogenous NO in the modulation of spontaneous tone and motility in the rat duodenum. Induction of NO synthase may result in a reduction in spontaneous motility of the tissue. By contrast, inhibition of constitutive NO biosynthesis unmasks a contractile response that is neuronally mediated and involves tachykinin NK2 receptors.
Effect of treatment with cholecalciferol on the membrane potential and contractility of aortae from spontaneously hypertensive rats
Authors: Silva Eneida G. , Vianna Lucia M. , Okuyama Paulo , Paiva Therezinha B.
Published: July 2012

The diet of spontaneously hypertensive rats (SHR) and normotensive Wistar rats (NWR) was supplemented with 12.5 μg cholecalciferol per 100 g body weight daily, by gavage, for 4 weeks.

The amplitude of the contractile responses of aortic rings from SHR to potassium and adrenaline, which was smaller than in NWR aortae, was increased after treatment with cholecalciferol. No further changes were observed in the responses of NWR and SHR aortae in the presence of 100 nM apamin.

The membrane potentials of aortae from SHR, which were higher than those of aortae from NWR, decreased after treatment with cholecalciferol. Further depolarization was observed in aortic rings from NWR, but not in aortic rings from SHR, after their preincubation with 100 nM apamin.

It is concluded that cholecalciferol normalizes the membrane potential and contractility of aortae from SHR, probably through an effect on lipid composition and structure of the plasma membrane.
Effect of endothelin antagonists on the responses to prostanoid endothelium‐derived contracting factor
Authors: Moreau Pierre , Takase Hiroyuki , Lüscher Thomas F.
Published: July 2012

The effect of endothelin antagonists on endothelium‐dependent contractions was studied in conditions of stimulated endothelium‐derived contracting factor (EDCF) release and with exogenous activation of thromboxane A2‐endoperoxide receptors in the rat aorta.

The incubation of aortic rings with Nω‐nitro‐L‐arginine methyl ester (L‐NAME) led to EDCF‐mediated contraction upon stimulation with acetylcholine (24 ± 3% of KCl contraction). When vessels were preincubated with bosentan, an endothelinA‐ and endothelinB‐receptor antagonist, in addition to L‐NAME, acetylcholine‐induced contraction was reduced to 8 ± 2% (P < 0.01) of KCl contractions. PD147953, a selective endothelinA‐receptor antagonist, reduced the contraction to 14 ± 4% (P < 0.05) of KCl contractions.

Bosentan preincubation produced a significant parallel rightward shift of the contractions to U46619, a selective thromboxane A2 receptor agonist. In contrast, PD147953 failed to exhibit any inhibitory effect on U46619 contractions.

These results suggest that endothelin antagonists inhibit EDCF‐mediated contractions by blocking endothelinA receptors and that, in addition, bosentan antagonizes the direct stimulation of thromboxane A2 receptors.
Block by N6‐L‐phenylisopropyl‐adenosine of the electrophysiological and morphological correlates of hippocampal ischaemic injury in the gerbil
Authors: Domenici M.R. , de Carolis A. Scotti , Sagratella S.
Published: July 2012

The effects of the mixed A1 and A2 adenosine receptor agonist N6‐L‐phenyl‐isopropyladenosine (L‐PIA) were tested on ischaemia‐induced hippocampal neuronal injury in gerbils subjected to 5‐min bilateral carotid occlusion. For comparison, the effects of the selective A2 adenosine receptor agonist, CGS 21680 were tested.

Five‐min bilateral carotid occlusion produced within 1 week an irreversible suppression of the CA1, but not of the dentate extracellular electrical somatic responses, in 30% of gerbil hippocampal slices with respect to controls. In addition, a significant reduction occurred in the density of CA1 hippocampal pyramidal neurones but not of dentate granule cells with respect to controls.

Injection 1 h before or after bilateral carotid occlusion of L‐PIA (0.8‐1.5 mg kg−1, i.p.) but not of CGS 21680 (5 mg kg−1, i.p.), significantly prevented the irreversible disappearance of the CA1 extracellular electrical somatic responses with respect to controls. In addition, the CA1 pyramidal neuronal loss was also prevented.

The results show that activation of A1 adenosine receptors is able to prevent or block the electrophysiological and morphological correlates of hippocampal neuronal injury after global ischaemia in the gerbil, suggesting that adenosine receptor agonists might have a useful role in the treatment of neuronal functional and anatomical injury due to ischaemia.
Pharmacological characterization of desensitization in a human mGlulα‐expressing non‐neuronal cell line co‐transfected with a glutamate transporter
Authors: Desai Manisha A. , Burnett J. Paul , Mayne Nancy G. , Schoepp Darryle D.
Published: July 2012

Stimulation of phosphoinositide hydrolysis by human mGlu1α (HmGlu1α) was examined in a non‐neuronal cell line (AV12‐664) co‐expressing both HmGlu1α and a rat glutamate/aspartate transporter (GLAST).

Desensitization of HmGlu1α could be elicited by inhibition of the GLAST transporter with the glutamate uptake inhibitor, L‐trans‐pyrrolidine‐2, 4‐dicarboxylic acid (trans‐PDC). Maximal inhibition of HmGlu1α‐mediated phosphoinositide hydrolysis was induced upon 24 h pretreatment with trans‐PDC. The concentration of glutamate in the extracellular medium also rose significantly in cells pretreated with trans‐PDC. Glutamate levels increased upon incubation with trans‐PDC in a time‐dependent manner, with maximal glutamate levels attained after 24 h incubation with trans‐PDC.

The time required for desensitization of HmGlu1α by trans‐PDC was compared to the time course for desensitization elicited by the direct‐acting mGlu receptor agonists, 1‐aminocyclopentane‐1S, 3R‐dicarboxylic acid (1S, 3R‐ACPD) and (R, S)‐3, 5‐dihydroxyphenylglycine (3, 5‐DHPG). Both direct‐acting mGlu receptor agonists elicited desensitization of HmGlu1α more rapidly than did trans‐PDC, with maximal inhibition of agonist‐induced phosphoinositide hydrolysis upon 12 h pretreatment. Agonist‐induced desensitization could be fully reversed upon washout of agonist for 12 h.

Both mGlu receptor agonist‐ and trans‐PDC‐induced desensitization of HmGlu1α could be blocked by inclusion of (+)‐α‐methyl‐4‐carboxyphenylglycine (MCPG), an mGlu receptor antagonist, in the pretreatment medium.

Agonist‐stimulated phosphoinositide hydrolysis by HmGlu1α was found to parallel closely agonist‐induced desensitization of HmGlu1α. Thus, the EC50 values for 1S, 3R‐ACPD‐ and 3, 5‐DHPG‐stimulated phosphoinositide hydrolysis were similar to the EC50 values for eliciting desensitization of HmGlu1α.

These studies demonstrate desensitization of recombinant human mGlu1α receptor in a non‐neuronal cell line in which the receptor can be regulated by direct activation or by manipulation of glutamate transporter activity. Desensitization of HmGlu1α was found to be mediated by activation of the receptor since the mGlu receptor antagonist, MCPG, blocked both mGlu receptor agonist‐ and trans‐PDC‐induced desensitization of HmGlu1α. Furthermore, agonist‐induced desensitization of HmGlu1α was found to parallel receptor‐mediated stimulation of phosphoinositide hydrolysis.
The effect of linomide on the migration and the proliferation of capillary endothelial cells elicited by vascular endothelial growth factor
Authors: Parenti Astrid , Donnini Sandra , Morbidelli Lucia , Granger Harris J. , Ziche Marina
Published: July 2012

In order to assess the mechanism of action of the quinoline‐3‐carboxyamide linomide as an antiangiogenic drug, the effect of linomide was studied in vitro on postcapillary endothelial cells exposed to vascular endothelial growth factor (VEGF). Linomide did not block the spontaneous replication of endothelial cells, but significantly suppressed endothelial cell growth and migration elicited by VEGF. It is concluded that linomide appears to be an effective tool to inhibit VEGF‐dependent angiogenesis.
Extracellular site for econazole‐mediated block of Ca2+ release‐activated Ca2+ current (Icrac) in T lymphocytes
Authors: Christian Edward P. , Spence Katherine T. , Togo James A. , Dargis Pauline G. , Warawa Edward
Published: July 2012

Standard whole cell patch clamp recording techniques were used to study the pharmacological characteristics and site of econazole‐mediated inhibition of calcium release‐activated calcium current (Icrac) in the human leukaemic T cell line, Jurkat.

Extracellularly applied econazole blocked Icrac in a concentration‐dependent manner (IC50 ≅ 14 μm). Block developed over a relatively slow timecourse of 30–60 s (10 μm), and only partially reversed over minutes.

Econazole dialysed from the pipette into the cytosol at concentrations ranging from 0.1 to 30 μm did not reduce Icrac, or quantitatively affect Icrac block by extracellularly applied econazole.

A less lipophilic quaternary iodide derivative of econazole was synthesized to retard absorption through the cell membrane. When applied extracellularly, this compound blocked Icrac in a concentration‐dependent manner with onset kinetics comparable to econazole.

Results with intracellularly dialysed econazole and the quaternary econazole derivative provide convergent evidence that econazole blocks Icrac via an extracellular interaction.

The inability of intracellularly applied econazole to inhibit Icrac argues against the notion that econazole inhibits capacitative Ca2+ entry pathways secondary to its known inhibitory effects on cytochrome P‐450.
Ionic mechanisms contributing to the vasorelaxant properties of iodinated contrast media: a comparison of iohexol and iodixanol in the rabbit isolated aorta
Authors: Pitman M.R. , Karlsson J.O.G. , Griffith T.M.
Published: July 2012

We have used rings of rabbit thoracic aorta to investigate the vasorelaxant properties of two different classes of non‐ionic iodinated radiographic contrast media (IRCM) and the mechanisms, underlying their mode of action. Iohexol (a triiodinated monomer) was compared with iodixanol (a hexaiodinated dimer).

Iohexol and iodixanol both relaxed phenylephrine (0.3 μm) constricted rabbit aorta in a concentration‐dependent manner that did not depend on the presence of an intact endothelium. When expressed as a function of iodine concentration, iodixanol caused significantly less relaxation than iohexol. However, the extent of relaxation was similar for both IRCM when expressed on a molar basis. Furthermore, increasing the molarity of the buffer to comparable levels with mannitol evoked only a small (∼ 15%) relaxation of phenylephrine‐induced tone.

Ouabain (10 μm) significantly inhibited both iohexol‐ and iodixanol‐induced relaxations by ∼30%. 5‐(N‐Ethyl‐N‐isopropyl)‐amiloride (EIPA, 100 nM) significantly inhibited iohexol‐induced relaxation to the same extent as ouabain, but did not alter the vasorelaxant effect of iodixanol. Co‐incubation with ouabain and EIPA had an additive effect in the case of iohexol, increasing inhibition of relaxation to ∼60%, whereas inhibition of iodixanol‐induced relaxation by the combination of ouabain plus EIPA did not differ from that of ouabain alone.

Replacing NaCl with N‐methyl‐D‐glucamine (NMDG) to lower extracellular [Na+] and thereby inhibit Na+‐Ca2+ exchange, attenuated the relaxation evoked by iohexol or by iodixanol (by ∼25%) in each case.

We conclude that iohexol‐ and iodixanol‐induced vasorelaxation in rabbit aorta is mediated through a direct action on vascular smooth muscle that is not simply a consequence of altered osmolality. It involves modulation of the Na+‐K+ ATPase and, in the case of iohexol, Na+‐H+ exchange. Both agents also appear to modulate Na+‐Ca2+ exchange, through direct and/or indirect mechanisms. This is the first study to show specific pharmacological differences between monomeric and dimeric contrast media in vascular smooth muscle.
Long Ca2+ channel opening induced by large depolarization and Bay K 8644 in smooth muscle cells isolated from guinea‐pig detrusor
Authors: Nakayama S. , Brading Alison F.
Published: July 2012

In smooth muscle cells enzymatically isolated from guinea‐pig urinary bladder, Ca2+ channel currents were recorded by conventional cell‐attached patch clamp techniques. In most recordings Bay K 8644 (2 μm) was contained in the patch pipette.

Closure of Ca2+ channels observed during the repolarizing steps was significantly slowed by preconditioning with large depolarizations (+80 and 100 mV), with or without Bay K 8644 in the pipette.

The sum of the unitary Ca2+ channel current traces obtained after large conditioning depolarizations (in the presence of Bay K 8644) showed a slowly deactivating tail current.

By use of this slow deactivating feature, the current‐voltage relationship of the unitary Ca2+ channel current was continuously measured with a ramp pulse after large depolarization. The slope conductance ranged from 22 to 30 pS, compatible with that of L‐type Ca2+ channels.

It is concluded that L‐type Ca2+ channels in guinea‐pig detrusor cells are open for much longer after large depolarizations consistent with their being two channel open states, and that Bay K 8644 prolongs the lifetime of both open states. The underlying mechanisms are discussed.
Endothelial modulation of α1‐adrenoceptor contractile responses in the tail artery of spontaneously hypertensive rats
Authors: Tabernero Antonia , Giraldo Jesús , Vivas Nuria María , Badia Albert , Vila Elisabet
Published: July 2012

Vascular contraction induced by phenylephrine was studied in tail artery rings from spontaneously hypertensive (SHR) and Wistar Kyoto rats (WKY) with particular focus on the role of endothelium. The influence of receptor reserve and the density of α1‐adrenoceptors on the possible differences observed were also analysed.

Phenylephrine (0.01–100 μm) induced concentration‐dependent vasoconstrictions. The maximum response (α, P < 0.001) was greater but the pEC50 (P < 0.05) smaller in rings from SHR than from WKY rats irrespective of the presence or absence of endothelium.

Removal of endothelial cells resulted in a decrease of the maximum contraction with no modification in the pEC50 in arteries from both WKY and SHR.

The density of α1‐adrenoceptors (Bmax) and the dissociation constant (KD) were found to be the same for preparations from SHR and WKY rats in [3H]‐prazosin binding experiments.

The apparent affinity (pKA) determined by the nested hyperbolic method and the operational model was similar in tail arteries from the two rat strains, irrespective of the presence or absence of endothelium. However, in endothelium‐denuded rings, the pKA value was enhanced when compared with intact rings, in both SHR and WKY rats.

In rings from hypertensive rats, the operational parameter maximum possible effect (Em) was greater and the agonist efficacy (τ) was smaller than in rings from normotensive rats. When the endothelium was removed log τ and Em diminished in preparations from both rat strains.

In summary, the increased maximum responsiveness to phenylephrine in rings from SHR could be due to enhancement in Em. The log τ values indicate a deterioration in the transduction of the stimulus provided by the agonist in tail arteries from hypertensive animals. This study also suggests that the absence of endothelium modifies the α1‐adrenoceptor‐mediated vasocontriction probably by altering the transduction signalling mechanisms. The importance of analysing the degree of endothelium functionality when comparing results from different groups of rats is stated.
Cloned and transfected P2Y4 receptors: characterization of a suramin and PPADS‐insensitive response to UTP
Authors: Charlton Steven J. , Brown Colin A. , Weisman Gary A. , Turner John T. , Erb Laurie , Boarder Michael R.
Published: July 2012

The P2Y family of receptors are G protein‐coupled receptors for ATP, ADP, UTP and UDP. Recently several members of this family have been cloned, including the P2Y4, which is activated by UTP but not by ATP. In the present report, using receptors stably transfected into 1321N1 cells, we show that suramin acts as an antagonist at cloned P2Y1 and (less potently) P2Y2 receptors, but not at the cloned P2Y4 receptor. Furthermore, PPADS (pyridoxal‐phosphate‐6‐azophenyl‐2′,4′‐disulphonic acid), a potent antagonist at the P2Y1 receptor, is a relatively inneffective antagonist at the cloned P2Y4 receptor. This work moves us closer to the goal of classifying the native P2Y receptors on the basis of agonist and antagonist profiles.
Effect of intracerebroventricular and intravenous administratioi of nitric oxide donors on blood pressure and heart rate in anaesthetized rats
Authors: Nurminen Marja‐Leena , Vapaatalo Heikki
Published: July 2012

The effects of nitric oxide (NO) releasing substances, sodium nitroprusside, 3‐morpholino sydnonimine (SIN‐1) and a novel oxatriazole derivative, GEA 3162, on blood pressure and heart rate were studied after peripheral or central administration in anaesthetized normotensive Wistar rats.

Given as cumulative intravenous injections, both nitroprusside and GEA 3162 (24–188 nmol kg−1) induced short‐lasting and dose‐dependent decreases in mean arterial pressure, while SIN‐1 decreased blood pressure only slightly even after larger doses (94–3000 nmol kg−1). Heart rate increased concomitantly with the hypotensive effect of the NO‐releasing substances.

Cumulative intracerebroventricular administration of GEA 3162 (24–188 nmol kg−1) induced a dose‐dependent hypotension with slight but insignificant increases in heart rate. In contrast, intracerebroventricular nitroprusside induced little change in blood pressure, while a large dose of SIN‐1 (3000 nmol kg−1, i.c.v.) slightly increased mean arterial pressure. However, intracerebroventricular nitroprusside and SIN‐1 increased heart rate at doses that did not significantly affect blood pressure.

To determine whether the cardiovascular effects of GEA 3162 were attributable to an elevation of cyclic GMP levels, pretreatments with methylene blue, a putative guanylate cyclase inhibitor, were performed. This substance failed to attenuate the cardiovascular effects of peripherally or centrally administered GEA 3162, suggesting that the effects were independent of guanylate cyclase.

In conclusion, the centrally administered NO‐donor, GEA 3162, induced a dose‐dependent hypotensive response without significant changes in heart rate. Furthermore, intracerebroventricular injections of nitroprusside and SIN‐1 increased heart rate without affecting blood pressure. These results suggest that NO released by these drugs may affect central mechanisms involved in cardiovascular regulation independently of cyclic GMP.
Modulation by extracellular ATP of L‐type calcium channels in guinea‐pig single sinoatrial nodal cell
Authors: Qi Ai‐Dong , Kwan Yiu Wa
Published: July 2012

The effects of extracellular adenosine 5′‐triphosphate ([ATP]0) on the L‐type Ca2+ channel currents in guinea‐pig single sinoatrial nodal (SAN) cells, isolated by enzymatic dissociation, were investigated by use of whole‐cell patch‐clamp techniques.

The application of [ATP]0 (2 μm‐1 mM) produced an inhibitory effect on the L‐type Ca2+ channel current peak amplitude (10 mM Ba2+ as charge carrier) in a concentration‐dependent and reversible manner with an IC50 of 100 μm and a Hill coefficient of 1.83.

The presence of the adenosine receptor antagonists, 8‐cyclopentyl‐1,3‐dipropylxanthine (DPCPX, 0.1 μm) and 8‐phenyltheophylline (10 μm) did not affect the [ATP]O‐induced inhibition of the Ca2+ channel currents. Adenosine (100 μm) had little effect on the basal Ca2+ channel currents. Adenosine 500 μm, caused 23% inhibition of the Ca2+ channel current, which was abolished by 0.1 μm DPCPX.

The presence of the P2‐purinoceptor antagonists, suramin (1, 10 and 100 μm), reactive blue 2 (1 and 10 μm) and pyridoxal‐phosphate‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS, 50 and 100 μm) failed to affect the inhibitory action of [ATP]0 on Ca2+ channel currents.

The relative rank order of potency of different nucleotides and nucleosides, at a concentration of 100 μm, on the inhibition of the Ca2+ channel currents is as follows: adenosine 5′‐triphosphate (ATP) = α, β‐methylene‐ATP (α, βMeATP) > > 2‐methylthioATP (2‐MeSATP) ≥ adenosine 5′‐O‐(3‐thiotriphosphate) (ATPγS) > > uridine 5′‐triphosphate (UTP) = adenosine 5′‐diphosphate (ADP) > adenosine 5′‐monophosphate (AMP) ≥ adenosine.

These results suggest that [ATP]0 may play an important role in the heart beat by inhibiting the L‐type Ca2+ channel currents in single SAN cells. This inhibitory effect is not due to the formation of adenosine resulting from the enzymatic degradation of [ATP]0. Based on the relative order of inhibitory potency of different nucleotides and nucleosides on the L‐type Ca2+ channel currents and the ineffectiveness of the purinoceptor antagonists tested, a novel type of purinoceptor may be involved.
Mediation by nitric oxide of the indirect effects of adenosine on calcium current in rabbit heart pacemaker cells
Authors: Shimoni Y. , Han X. , Severson D. , Giles W.R.
Published: July 2012

Adenosine (ADO) is a potent negative chronotropic agent in the mammalian myocardium. We have used single myocytes from rabbit sino‐atrial node (SAN) to examine whether nitric oxide (NO) is a significant mediator of the effects of ADO on the pacemaker activity, or the underlying Ca2+ and K+ currents.

SAN pacemaker cells were isolated from rabbit hearts by enzymatic dispersion, and Ca2+ and K+ currents were recorded by the nystatin‐perforated patch voltage clamp method. ADO was applied in the presence of the β‐adrenoceptor agonist, isoprenaline (Iso) to mimic the adrenergic tone which the SAN is subjected to in vivo

Control experiments confirmed that isolated SAN cells responded to ADO (10–100 μm) with the expected (i) small increase in background inwardly rectifying K+ current, IK‐ADOi and (ii) pronounced decrease in L‐type Ca2+ current, ICa‐L These effects were mimicked by a selective A1 purinoceptor agonist, N6‐cyclopentyladenosine (CPA, 10 μm); and were inhibited following bath application of the antagonist, DPCPX (10 μm), which selectively blocks A1 purinoceptors. DMPX (10 μm), a blocker of A2 purinoceptor, had no effect on the actions of ADO.

A nitric oxide synthase inhibitor, L‐NMMA (100 μm), abolished the inhibitory effect of ADO on ICa‐L but did not alter activation of IK‐ADO After L‐NMMA washoff, it was possible to obtain the normal response (inhibition) of ICa‐L to ADO in the same cell.

To evaluate whether the observed effect of nitric oxide (NO) on ICa‐L was mediated by an increase in guanylyl cyclase (GC) activity and cyclic GMP formation, the guanylyl cyclase inhibitor, LY 83583 (40 μm) was applied prior to ADO. Under these conditions, the inhibitory effect of ADO on ICa‐L was abolished, but the activation of IK‐ADO was still observed.

In combination, these findings strongly suggest that in mammalian primary pacemaker tissue which is under adrenergic tone, the effects of ADO on ICa‐L are mediated by NO.
Effects of adrenomedullin and calcitonin gene‐related peptide on airway and pulmonary vascular smooth muscle in guinea‐pigs
Authors: Pinto Arthur , Sekizawa Kiyohisa , Yamaya Mutsuo , Ohrui Takashi , Jia Yu Xia , Sasaki Hidetada
Published: July 2012

The airway and pulmonary vascular effects of adrenomedullin were studied in the guinea‐pig isolated trachea, main bronchi and pulmonary artery in vitro and compared to the effects of calcitonin gene‐related peptide (CGRP).

In tracheal rings, CGRP (1 nM to 1 μm) potentiated the cholinergic contractions induced by electrical field stimulation (EFS) at 5 Hz in a concentration‐dependent manner. At a concentration of 1 μm, CGRP slightly decreased the responses to log EFS frequency, producing 50% of the maximum contraction from a control value of 0.77 ± 0.10 Hz to 0.54 ± 0.05 Hz without a significant effect on the concentration‐response curves to acetylcholine (ACh). In contrast, adrenomedullin (1 nM to 1 μm) did not alter either EFS‐induced cholinergic or ACh‐induced contractions.

In bronchial strips, CGRP (1 nM to 1 μm) slightly reduced both the non‐adrenergic non‐cholinergic (NANC) contraction induced by EFS at 10 Hz and the substance P (1 μm)‐induced contraction in a concentration‐dependent manner, whereas adrenomedullin (1 nM to 1 μm) was without effect.

Neither CGRP (1 μm) nor adrenomedullin (1 IXM) altered NANC relaxation induced by EFS at 5 Hz in tracheal rings precontracted with histamine (10 μm).

Adrenomedullin (1 nM to 1 μm) and CGRP (1 nM to 1 μm) induced a concentration‐dependent relaxation of the histamine (10 μm)‐ and prostaglandin F2α (10 μm)‐precontracted pulmonary arterial rings with intact endothelium with a similar potency.

Neither removal of the endothelium nor NG‐nitro‐L‐arginine methyl ester (100 μm) altered the vasorelaxant effects of adrenomedullin (1 nM to 1 μm) and CGRP (1 nM to 1 μm).

The putative CGRP receptor antagonist, CGRP8–37 (1 μm to 10 μm) concentration‐dependently attenuated the CGRP (3 nM to 30 nM)‐induced vasorelaxant actions, whereas it had no effect on the relaxation of vessel rings induced by adrenomedullin (3 nM to 30 nM).

These results suggest that adrenomedullin is a potent vasodilator of the pulmonary artery without any bronchomotor effect in the guinea‐pig lung, and that the vasorelaxant actions of adrenomedullin are not mediated via the activation of CGRP1 receptors.
Inhibition by propofol of [3H]‐batrachotoxinin‐A 20‐α‐benzoate binding to voltage‐dependent sodium channels in rat cortical synaptosomes
Authors: Ratnakumari L. , Hemmings H.C.
Published: July 2012

Propofol (2,6 di‐isopropylphenol), an intravenous general anaesthetic, blocks voltage‐dependent Na+ channels (Na+ channels). In this study the interaction between propofol and Na+ channels was analysed by examining its effects on neurotoxin binding to various receptor sites of the Na+ channel in rat cerebrocortical synaptosomes.

Propofol (10–200 μm) exhibited concentration‐dependent inhibition of equilibrium binding of [3H]‐batrachotoxinin‐A 20‐α‐benzoate ([3H]‐BTX‐B) to receptor site 2 of the Na+ channel (mean IC50 = 26 μm; 6.5 μm free). Scatchard analysis revealed that propofol significantly increased the KD without affecting the Bmax for [3H]‐BTX‐B binding.

Kinetic studies of [3H]‐BTX‐B binding in the presence of various concentrations (25–200 μm) of propofol showed no significant changes in the association rate of [3H]‐BTX‐B. However, propofol at 200 μm significantly increased the rate of dissociation of [3H]‐BTX‐B, consistent with an indirect allosteric competitive mechanism of inhibition.

[3H]‐saxitoxin binding to receptor site 1 and [3H]‐brevetoxin‐3 binding to receptor site 5 of the Na+ channel were not inhibited by propofol (10–200 μm).

Propofol (10–100 μm) exhibited concentration‐dependent inhibition of veratridine‐evoked Na+ influx either in the absence or presence of scorpion toxin with IC50 values of 46 μm (8.8 μm free) and 44 μm (8.5 μm free), respectively.

These results suggest that propofol inhibits voltage‐dependent Na+ channels due to a preferential interaction with the inactivated state of the channel. Blockade of Na+ channels by propofol, which is known to inhibit glutamate release from synaptosomes, may contribute to its anaesthetic, anticonvulsant and neuroprotective properties.
A functional polymorphism in PER3 gene is associated with prognosis in hepatocellular carcinoma
Authors: Zhao Binyu , Lu Jianguo , Yin Jikai , Liu Hanqiang , Guo Xu , Yang Yefa , Ge Naijian , Zhu Yong , Zhang Hongxin , Xing Jinliang
Published: July 2012

Background

Previous studies have revealed that circadian genes play important roles in cell proliferation, apoptosis, cell cycle control, DNA damage response and treatment response of chemotherapy agents in cancers.

Aims

We hypothesized that the polymorphisms in circadian genes may be associated with prognosis of hepatocellular carcinoma (HCC) patients treated with transcatheter arterial chemoembolization (TACE).

Methods

Twelve functional single nucleotide polymorphisms (SNPs) in circadian negative feedback regulation genes (including
CRY1, CRY2, PER1, PER2 and
PER3) were genotyped using Sequenom iPLEX genotyping method in 337 HCC patients treated with TACE and analysed for associations with overall survival.

Results

Our data showed that one SNP rs2640908 in
PER3 gene was significantly associated with overall survival of HCC patients (P = 0.027). Patients carrying at least one variant allele of rs2640908 (WV + VV) had a significantly decreased risk of death (hazard ratio, 0.71; 95% confidence interval, 0.53–0.90), when compared with those carrying homozygous wild‐type alleles (WW). Kaplan–Meier analyses showed a significantly longer median survival time in patients with WV + VV genotypes of SNP rs2640908 than those with WW genotype (11.6 months vs. 8.1 months; log rank P = 0.030). In addition, we also observed a significant difference on the genotype distribution of SNP rs2640908 in patients with and without portal vein thrombus (P = 0.041).

Conclusions

Our study provides the first evidence that a single functional polymorphism of
PER3 gene is significantly associated with overall survival in HCC patients treated with TACE.
A Comparison of Global Conservation Prioritization Models with Spatial Spending Patterns of Conservation Nongovernmental Organizations
Authors: HOLMES GEORGE , SCHOLFIELD KATHERINE , BROCKINGTON DAN
Published: July 2012

Abstract:  In recent decades, various conservation organizations have developed models to prioritize locations for conservation. Through a survey of the spending patterns of 281 conservation nongovernmental organizations (NGOs), we examined the relation between 2 such models and spatial patterns of spending by conservation NGOs in 44 countries in sub‐Saharan Africa. We tested whether, at the country level, the proportion of a country designated as a conservation priority was correlated with where NGOs spent money. For one model (the combination of Conservation International's hotspots and High Biodiversity Wilderness Areas, which are areas of high endemism with high or low levels of vegetation loss respectively), there was no relation between the proportion of a country designated as a priority and levels of NGO spending, including by the NGO associated with the model. In the second model (Global 200), the proportion of a country designated as a priority and the amount of money spent by NGOs were significantly and positively related. Less money was spent in countries in northern and western sub‐Saharan Africa than countries in southern and eastern Africa, relative to the proportion of the country designated as a conservation priority. We suggest that on the basis of our results some NGOs consider increasing their spending on the areas designated as of conservation priority which are currently relatively underfunded, although there are economic, political, cultural, historical, biological, and practical reasons why current spending patterns may not align with priority sites.
Effects of Native and Non‐Native Vertebrate Mutualists on Plants
Authors: ASLAN CLARE E. , ZAVALETA ERIKA S. , CROLL DON , TERSHY BERNIE
Published: July 2012

Abstract:  Extinctions can leave species without mutualist partners and thus potentially reduce their fitness. In cases where non‐native species function as mutualists, mutualism disruption associated with species’ extinction may be mitigated. To assess the effectiveness of mutualist species with different origins, we conducted a meta‐analysis in which we compared the effectiveness of pollination and seed‐dispersal functions of native and non‐native vertebrates. We used data from 40 studies in which a total of 34 non‐native vertebrate mutualists in 20 geographic locations were examined. For each plant species, opportunistic non‐native vertebrate pollinators were generally less effective mutualists than native pollinators. When native mutualists had been extirpated, however, plant seed set and seedling performance appeared elevated in the presence of non‐native mutualists, although non‐native mutualists had a negative overall effect on seed germination. These results suggest native mutualists may not be easily replaced. In some systems researchers propose taxon substitution or the deliberate introduction of non‐native vertebrate mutualists to reestablish mutualist functions such as pollination and seed dispersal and to rescue native species from extinction. Our results also suggest that in places where all native mutualists are extinct, careful taxon substitution may benefit native plants at some life stages.