bluuebunny/crossref_metadata_2025 huggingface dataset:
Alterations in histamine receptors of guinea‐pig ileal smooth muscle produced during agonist‐induced desensitization
Authors: Horio Shuhei , Nakamura Shigeki , Ishida Yukio
Published: July 2012

The effects of prolonged treatment with histamine (10−4 m, 30 min) on desensitization at histamine H1‐receptors of guinea‐pig ileal longitudinal muscle were investigated.

This treatment did not change either the maximum amount or dissociation constant (Kd) of [3H]‐mepyramine binding to membranes of guinea‐pig ileal muscle.

In contrast, this treatment shifted the histamine inhibition curves of [3H]‐mepyramine binding to the right both in the presence and absence of 0.5 mm guanosine‐5′‐triphosphate (GTP). This rightward shift of the curves occurred rapidly in the first 10 min of exposure to histamine.

The histamine inhibition curves were analyzed with a two binding sites model. It was shown that the histamine‐induced affinity change of the receptor for the agonist occurred with the high affinity binding component (which comprise about 80% of the total), whereas no significant change occurred with the low affinity component. The GTP‐dependent decrease in the affinity of the receptor for the agonist also occurred with the high affinity component both in control and histamine‐treated preparations.

These studies suggest that histamine‐induced desensitization was caused by alteration in the affinity of the receptor for the agonist rather than reduction in the number of the receptors and that the interaction of the receptor with a guanine nucleotide regulatory protein was retained in the desensitized state.
Regional and cardiac haemodynamic effects of NG‐nitro‐l‐arginine methyl ester in conscious, Long Evans rats
Authors: Gardiner S.M. , Compton A.M. , Kemp P.A. , Bennett T.
Published: July 2012

Regional haemodynamic responses to i.v. bolus doses (0.1–10.0 mg kg−1) of NG‐nitro‐l‐arginine methyl ester (l‐NAME) were measured in conscious, Long Evans rats (n = 8) chronically instrumented with renal, mesenteric and hindquarters pulsed Doppler flow probes and intravascular catheters.

l‐NAME caused dose‐dependent pressor effects associated with renal, mesenteric and hindquarters vasoconstrictions. The mesenteric vascular bed showed earlier onset with more rapid, and greater, maximum vasoconstrictions than the renal or hindquarters vascular beds; however, the hindquarters vasoconstriction was more persistent. d‐NAME was without significant effects (n = 2).

Primed infusion of l‐arginine (100 mg kg−1 bolus followed by 100 mg kg−1 h−1 infusion), starting 10 min after an i.v. bolus injection of l‐NAME (10 mg kg−1), caused significant reversal of the pressor responses, and renal and mesenteric vasoconstrictions, but not of the hindquarters vasoconstriction. Primed infusions of l‐arginine (100 mg kg−1, 100 mg kg−1 h−1) starting 5 min after l‐NAME (1 mg kg−1) additionally caused some reversal of the hindquarters vasoconstriction, but this effect was transient.

Primed infusion of l‐arginine (100 mg kg−1, 100 mg kg−1 h−1) starting 30 min before i.v. bolus injection of l‐NAME (10 mg kg−1) caused significant attenuation of the pressor effects and the renal and mesenteric vasoconstrictions but not of the hindquarters vasoconstriction.

In a separate group of rats (n = 8) chronically instrumented with thoracic aortic electromagnetic flow probes for the measurement of cardiac haemodynamics, i.v. bolus injection of l‐NAME (10 mg kg−1) produced significant reductions in total peripheral conductance, cardiac output, stroke volume, peak thoracic aortic flow and the maximum rate of rise of aortic flow; these were coincident with the maximum pressor and vasoconstrictor effects.

These results, collectively, are consistent with l‐NAME interfering with l‐arginine‐nitric oxide pathways that have important influences on regional vascular conductances in vivo. The pressor effect resulting from l‐NAME‐induced vasoconstrictions is offset by a substantial reduction in cardiac function that may depend on direct and/or indirect effects of l‐NAME on the heart.
Reduced high‐affinity α1‐adrenoceptors in liver of senescent rats: implications of assessment at various temperatures
Authors: Borst Stephen E. , Scarpace Philip J.
Published: July 2012

We investigated the changes occurring as a result of aging in α1‐adrenoceptors in the livers of Fisher 344 female rats. For comparison, we also measured β‐adrenoceptors in this tissue. Three age groups were studied, including young adults (aged 6 months), mature adults (aged 16 months) and senescent animals (aged 25 months).

The density of α1‐receptors was measured by use of [3H]‐prazosin and was found to be reduced 39% (P > 0.01) at 25 months compared with 6 months. The percentage of α1‐receptors displaying high affinity for adrenaline was also reduced from 85.6% at 6 months to 51.6% at 25 months (P > 0.02).

In contrast, the density of β‐receptors, which was measured with [125I]‐iodocyanopindolol, was increased 104% between 6 months and 25 months. The affinity of both α1‐ and β‐adrenoceptors for antagonists was unchanged with age.

We found that receptor affinity for agonists may be measured accurately in binding studies conducted at 4°C or 25°C, but that the apparent affinity for agonist was artifactually reduced in studies conducted at 37°C. This effect is poorly reversible, in that reduced agonist‐affinity is also observed in tissue which has been incubated at 37°C and then cooled to 4°C before performing the binding studies.

It is concluded that liver α1‐adrenoceptor function is reduced and β‐adrenoceptor function increased in senescence.
Two distinct α1‐adrenoceptor subtypes involved in noradrenaline contraction of the rabbit thoracic aorta
Authors: Muramatsu Ikunobu , Kigoshi Shigeru , Oshita Masafumi
Published: July 2012

Recently, α1‐adrenoceptors in blood vessels have been classified into three subtypes (α1H, α1L and α1N). We examined which subtype (or subtypes) is involved in the noradrenaline‐induced contraction of rabbit thoracic aorta.

Noradrenaline produced a concentration‐dependent contraction in the rabbit isolated thoracic aorta. Prazosin antagonized the contractions to noradrenaline, resulting in a rightward displacement of the concentration‐response curve. However, the shift was not proportional to the concentration of prazosin; Schild plots showed that the inhibition by prazosin was biphasic, implying that noradrenaline acted through two receptor populations. Two affinity constants (pKB values of 10.02 and 8.83) were determined for prazosin at these sites.

However, under continuous treatment with 1 nm prazosin, or in strips pretreated with chlorethylclonidine (CEC; an α1H inactivating agent) to remove the contribution of one receptor population, prazosin showed a single pKB or pA2 value of approximately 8.3.

Yohimbine also produced biphasic antagonism of noradrenaline‐induced contractions, resulting in two affinity constants (pKB = 6.52 and 6.17). However, a monophasic Schild plot was obtained for yohimbine either in the presence of 1 nm prazosin (pA2 = 6.08) or in strips pretreated with CEC (pA2 = 6.03).

The Schild plot for HV723 (a selective α1N‐antagonist) yielded a monophasic slope (pKB = 8.47) and the inhibition was not affected by 1 nm prazosin or CEC‐pretreatment.

[3H]‐prazosin bound to α1‐adrenoceptors of the aortic membrane preparations with two different affinities (pKD = 9.94 and 8.37). The high but not the low affinity site was completely masked by 1 nm prazosin and inactivated by pretreatment with CEC.

These results strongly suggest that noradrenaline‐induced contraction of the rabbit thoracic aorta is mediated through two distinct α1‐adrenoceptor subtypes, designated α1H and α1L.
Relative potencies for barbiturate binding to the Torpedo acetylcholine receptor
Authors: Dodson Barbara A. , Urh Rhonda R. , Miller Keith W.
Published: July 2012

The structural requirements of an allosteric barbiturate binding site on acetylcholine receptor‐rich membranes isolated from Torpedo electroplaques have been characterized by the ability of fourteen barbiturates to displace [14C]‐amobarbitone binding.

The barbiturates could be grouped into two classes with ten barbiturates producing a strong inhibition of [14C]‐amobarbitone binding (class one) and with four exerting minimal effects (class two).

Eight of the ten class one barbiturates displaced essentially all of the [14C]‐amobarbitone from its binding site, while, at their respective aqueous solubility limits, two of these barbiturates (thiopentone and dimethylbutylbarbitone (DMBB)) inhibited [14C]‐amobarbitone binding by nearly 80%. The apparent inhibition constants (KI) for the class one barbiturates ranged from 13 μm for amobarbitone to 2.8 mm for barbitone with the other eight agents lying in the range 100–600 μm, and having the rank order pentobarbitone ≅ secobarbitone > thiopentone > DMBB > butabarbitone ≅ phenobarbitone > aprobarbitone > allylbarbitone.

By contrast, the class two barbiturates had minimal effects even at close to saturating concentrations. [14C]‐amobarbitone binding was reduced slightly (< 30%) by hexobarbitone, mephobarbitone and methohexitone and was enhanced slightly (< 20%) by metharbitone.

All of the class two, but none of the class one barbiturates, were N‐methylated.
The pharmacological properties of the imidazobenzodiazepine, FG 8205, a novel partial agonist at the benzodiazepine receptor
Authors: Tricklebank M.D. , Honoré T. , Iversen S.D. , Kemp J.A. , Knight A.R. , Marshall G.R. , Rupniak N.M.J. , Singh L. , Tye S. , Watjen F. , Wong E.H.F.
Published: July 2012

The pharmacological properties of the benzodiazepine receptor ligand, FG 8205 (7‐chloro‐5,6‐dihydro‐5‐methyl‐6‐oxo‐3‐(5‐isopropyl‐1,2,4‐oxadiazol‐3‐yl)‐4H‐imidazol[1,5a][1,4]benzodiazepine) have been examined.

FG 8205 potently displaced [3H]‐flumazenil binding in rat cortical membranes with a Ki of 3.3 nm, but was inactive at 13 neurotransmitter recognition sites.

Consistent with a partial agonist profile, the affinity of FG 8205 for the benzodiazepine recognition site was increased in the presence of γ‐aminobutyric acid (GABA, 300 μm) by a degree (−log [IC50 in the presence of GABA/IC50 alone] = 0.34) significantly less than found for diazepam (0.46). FG 8205 also potentiated the inhibitory potency of the GABAA‐receptor agonist, isoguvacine, on the hippocampal CA1 population spike and, again, the maximum shift (−log dose‐ratio = 0.2) was significantly less than that seen with diazepam (0.4).

In anticonvulsant studies, the ED50 doses of FG 8205 and diazepam needed to antagonize seizures induced by pentylenetetrazol (PTZ) or by sound in audiogenic seizure prone mice were similar with values of 0.2–0.3 mg kg−1, i.p. However, even high doses of FG 8205 (50 mg kg−1) did not protect against seizures induced by electroshock.

FG 8205 released responding suppressed by footshock in a rat operant conditioned emotional response task over the dose range 0.5–50 mg kg−1 (i.p.). Similar doses of FG 8205 had a marked taming effect in cynomolgus monkeys. However, measures of sedation and ataxia (as measured by rotarod in the mouse, climbing behaviour in the rat, and by scoring arousal and co‐ordination in primates) were slight and only transiently affected by FG 8205, and FG 8205 significantly antagonized the rotarod performance deficit induced by diazepam in the mouse.

While the potentiation by FG 8205 of the response to isoguvacine in the rat hippocampal slice and the anxiolytic‐like effects of the compound in both rats and primates were reversed by the benzodiazepine receptor antagonist, flumazenil, high doses of the antagonist were able only marginally to block the protective effects of FG 8205 against seizures induced by PTZ in the mouse.

Thus, FG 8205 does not show the marked motor impairment characteristic of full agonists at the benzodiazepine receptor, consistent with its partial agonist profile in in vitro assay systems. Nevertheless, the compound has sufficient intrinsic activity to maintain high efficacy in anticonvulsant and anxiolytic tests.
SK&F S‐106203 inhibits leukotriene C4, leukotriene D4 and leukotriene E4 vasopressor responses in the conscious rat
Authors: Smith Edward F. , Slivjak Mark J. , Egan John W. , Eckardt R.D. , Newton J.F.
Published: July 2012

The purpose of these experiments was to investigate the effects of the selective peptidoleukotriene receptor antagonist, SK&F S‐106203, on leukotriene C4 (LTC4), LTD4 and LTE4 vasopressor responses in the conscious, normotensive rat. SK&F S‐106203 was administered as a bolus followed by a continuous infusion in order to provide information on the relationship between antagonism of leukotriene responses and steady‐state plasma concentrations.

Infusion of SK&F S‐106203 at doses of 0.2 mg kg−1 + 1 mg kg−1 h−1, 1 mg kg−1 + 3 mg kg−1 h−1 or 2 mg kg−1 + 10 mg kg−1 h−1 produced dose‐dependent steady‐state plasma drug concentrations of 1.0, 3.2 and 23.8 μg ml−1, respectively. Plasma SK&F S‐106203 concentrations appeared to increase in a linear fashion at doses of 1 and 3 mg kg−1 h−1; at the highest dose the increment in plasma drug concentrations (i.e., 7–8 fold) was greater than the increment in dose (i.e., 3 fold), suggesting saturation of the primary clearance mechanism(s) at this dose.

SK&F S‐106203 (2 mg kg−1 + 10 mg kg−1 h−1) had no effect on noradrenaline‐, vasopressin‐, isoprenaline‐, or U 46619‐induced responses.

SK&F S‐106203 produced dose‐dependent rightward shifts in the LTC4 and LTE4 dose‐response curves. Administration of SK&F S‐106203 at doses of 0.2 mg kg−1 + 1 mg kg−1 h−1, 1 mg kg−1 + 3 mg kg−1 h−1, or 2 mg kg−1 + 10 mg kg−1 h−1 produced dose‐ratios of 1.0, 3.1 and 19.9, respectively, against LTC4 responses, and dose‐ratios of 1.6, 3.8 and 9.1, respectively, against LTE4 responses.

Against LTD4 responses, SK&F S‐106203 at doses of 0.2 mg kg−1 + 1 mg kg−1 h−1, 1 mg kg−1 + 3 mg kg−1 h−1, or 2 mg kg−1 + 10 mg kg−1 h−1 produced dose‐ratios of 2.5, 2.8, and 11.4, respectively. Administration of d‐penicillamine, a non‐competitive LTD4 dipeptidase inhibitor, had no effect on LTD4 responses.

The similarity in the LTD4 dose‐ratios at the two lower infusion rates, despite increases in the plasma drug concentrations, suggests the existence of pharmacologically heterogeneous LTD4 receptors. These results indicate that SK&F S‐106203 is a potent, selective and apparently competitive antagonist of LTC4, LTD4 and LTE4 vascular responses in the intact rat.
Nedocromil sodium inhibits antigen‐induced contraction of human lung parenchymal and bronchial strips, and the release of sulphidopeptide‐leukotrienes and histamine from human lung fragments
Authors: Napier Fiona E. , Shearer Melanie A. , Temple Diana M.
Published: July 2012

The effects of nedocromil sodium on antigen‐induced release of sulphidopeptide‐leukotrienes and histamine from passively sensitized fragments of human lung, and on antigen‐induced contraction of sensitized strips of human lung parenchyma and bronchus, have been studied.

Nedocromil sodium 0.1 and 1 μm inhibited leukotriene release from fragments of human lung by 30% and 38% respectively, and histamine release by 43% for both concentrations, but 10 μm was ineffective. The lung fragments, which were passively sensitized to house dust mite, Dermataphagoides pteronyssinus, in control experiments released leukotrienes (6.58 ± 0.12 nmol equiv. leukotriene C4 per g, n = 6) and histamine (10.3% ± 1.8 of total tissue histamine, n = 5) when challenged with house dust mite extract.

Isolated strips of human lung parenchyma, passively sensitized to D. pteronyssinus, contracted when treated with house dust mite extract to a mean value of 40% of the maximal histamine response for each strip. Nedocromil sodium 0.1 and 1 μm inhibited these contractions by 50% and 70% of the control response, but 10 μm had no inhibitory effect.

Isolated rings from human bronchus, also passively sensitized to D. pteronyssinus, contracted when treated with house dust mite extract to a mean value of 86% of the maximal histamine response. Nedocromil sodium 1 μm, but not 0.1 or 10 μm, inhibited contractions by 48% of the control response.

The therapeutic effects of nedocromil sodium in allergic asthma may depend, partly, on its inhibition of antigen‐induced release of leukotrienes and histamine in human lung and its consequent inhibition of antigen‐induced contractions of parenchymal and bronchial tissue.
GABAA receptor‐mediated increase in membrane chloride conductance in rat paratracheal neurones
Authors: Allen T.G.J. , Burnstock G.
Published: July 2012

The actions of γ‐aminobutyric acid (GABA) on the intramural neurones of 14–18 day old rats were studied in situ by use of intracellular current‐ and voltage‐clamp techniques. The ionic conductance changes and the effects of various GABA‐receptor agonists and antagonists on these neurones were also investigated.

Prolonged application of GABA either by ionophoresis (10 pC‐10 nC) or superfusion (10–100 μm), evoked a biphasic membrance depolarization in over 90% of all paratracheal neurones studied. Typically, the response consisted of an initial rapid depolarization (18–45 ms) that subsequently faded over a period of 15–25 s to reveal a second smaller depolarization which was maintained for the duration of GABA application. Both components of the evoked response resulted in an increase in membrane conductance and an inward flow of current.

The amplitude of the transient inward current, recorded during the initial phase of the response, was linearly related to the membrane potential at which it was elicited and reversed symmetrically at a membrane potential of −32.7 mV. The underlying increase in conductance was largely independent of membrane potential. The equilibrium potential for the sustained inward current was −38.7 mV. Replacement of extracellular chloride with gluconate ions initially enhanced the GABA‐evoked inward current. With successive applications of GABA in low chloride, the evoked current and conductance changes declined markedly.

Muscimol superfusion (1–10 μm) or ionophoresis (10 pC‐10 nC) mimicked both the initial and late phases of the GABA‐induced conductance change and inward current. Baclofen (1–100 μm) had no effect upon either resting membrane potential or conductance in any of the cells tested.

The large transient initial phase of the GABA‐evoked inward current and depolarization were potently inhibited by picrotoxin (1–50 μm), whereas the smaller sustained inward current was largely resistant to picrotoxin.

All of the observed actions of GABA and muscimol were antagonized by bicuculline (0.1–10 μm) in an apparently competitive manner.

It is concluded that GABA acts via GABAA receptors present on the soma of paratracheal neurones to produce an increase in membrane chloride conductance. Prolonged application of GABA results in a decline in the observed current due to a combination of two processes: receptor desensitization and shifts in the chloride equilibrium potential. The possible roles for GABA in neural regulation of airway excitability are discussed.
Cerebrovascular responses to capsaicin in vitro and in situ
Authors: Edvinsson Lars , Jansen Inger , Kingman Tom A. , McCulloch James
Published: July 2012

The cerebrovascular effects of capsaicin have been examined in vitro, in feline isolated cerebral arteries (circular segments, 2–3 mm long, 300–400 μm extended diameter) and, in situ, in pial arterioles (diameter 40–200 μm) on the cortical surface of chloralose‐anaesthetized cats.

In isolated middle cerebral arteries, low concentrations of capsaicin (10−14‐10−10m) effected a concentration‐dependent relaxation of vessels precontracted with prostaglandin F2α. This relaxant response was markedly attenuated by repeated administration of capsaicin but was minimally affected by the presence of atropine, propranolol, cimetidine or spantide in the tissue bath.

In isolated middle cerebral arteries, higher concentrations of capsaicin effected a marked concentration‐dependent contraction. This contraction was not modified by 10−6 μ phentolamine or 10−6m ketanserin. A markedly reduced contraction by capsaicin was found upon the removal of calcium ions from the buffer solution. Also the calcium entry blocker nimodipine reversed the capsaicin‐induced contraction.

Subarachnoid perivascular microapplication of capsaicin around individual pial arterioles in situ elicited a biphasic response (an immediate vasoconstriction followed by a sustained vasodilatation). The maximum vasoconstriction was a 60 ± 6% reduction in diameter from base line and the maximum vasodilatation a 38 ± 7% increase in diameter. Vasodilatation occurred at lower concentrations of capsaicin (EC50, approximately 5 × 10−8 μ) than those required for vasoconstriction (EC50 3 × 10−7 μ).

Trigeminal ganglionectomy 10–16 days before the microapplication abolished the in situ vasodilator effects of capsaicin (10−6m) applied perivascularly, but was without effect on the vasoconstrictor actions of this agent.

Repeated administration of capsaicin (10−6m) around the same arteriole resulted in a progressive attenuation of the vasodilator phase of the response, with no modification of the vasoconstrictor phase.

The present study suggests that capsaicin‐induced cerebral vasodilatation is due to the release of vasoactive agents from cerebrovascular trigeminal nerve fibres, whereas the vasoconstrictor effect of capsaicin is due to a direct effect on the cerebral vasculature which is mediated via the transmembrane passage of extracellular calcium.
δ‐Opioid receptor binding sites in rodent spinal cord
Authors: Traynor J.R. , Hunter J.C , Rodriguez R.E. , Hill R.G. , Hughes J.
Published: July 2012

The δ‐opioid receptor agonist [d‐Pen2,d‐Pen5]enkephalin showed an antinociceptive effect in the mouse tail‐flick test, following intrathecal administration. This action was reversed by naloxone and by the selective δ‐opioid receptor antagonist ICI 174864.

High affinity, saturable binding of [3H]‐[d‐Pen2,d‐Pen5]enkephalin has been demonstrated in spinal cord homogenates from guinea‐pig, hamster, rat and both adult and young (18–20 g) mice. The binding was shown by autoradiography to be concentrated in the superficial laminae of the dorsal horn.

Competition studies confirmed that the binding of [3H]‐[d‐Pen2,d‐Pen5]enkephalin was to the δ‐opioid site. However, anomalies were seen with displacement assays using μ‐ligands, which may suggest some common high affinity site for δ‐ and μ‐opioid receptor agonists in the spinal cord.

The results add further evidence for a role of the δ‐opioid receptor in spinally‐mediated antinociception.
Antagonism by nifedipine of contraction and Ca2+ ‐influx evoked by ATP in guinea‐pig urinary bladder
Authors: Katsuragi T. , Usune S. , Furukawa T.
Published: July 2012

The effects of Ca2+‐antagonists, especially nifedipine, on contraction and increase of intracellular Ca2+ (Fura‐2/AM method) evoked by ATP were evaluated in a thin outer layer segment of guinea‐pig urinary bladder.

The ATP‐evoked contraction was markedly inhibited by dihydropyridine‐type Ca2+‐antagonists, such as nifedipine and nitrendipine, but not by D‐600, ω‐conotoxin and tetramethrin.

This antagonism by nifedipine of ATP‐evoked contractions was competitive from the Schild plot analysis, the pA2 value being 8.23. The reduction of ATP‐evoked contraction by nifedipine (0.1 μm) was fully reversed by administration of Bay K 8644 (0.1 μm).

ATP (100 μm) caused an increase of fluorescence brightness after loading Fura‐2/AM, which was coupled with a contraction of the bladder. Both the contraction and the elevation of intracellular Ca2+ evoked by the nucleotide were completely antagonized by nifedipine.

These results suggest that ATP may activate the dihydropyridine‐sensitive, voltage‐dependent Ca2+‐channels in a direct or indirect fashion and, thereby, elicit a contraction of the bladder.
Mechanism of endothelin‐induced contraction in guinea‐pig trachea: comparison with rat aorta
Authors: Hay D.W.P.
Published: July 2012

Endothelin (1nm‐0.3 μm) produced a concentration‐dependent contraction of guinea‐pig epithelium‐containing (intact) trachea (EC50 = 30.9 nm). Endothelin was a less potent agonist than leukotriene D4 (LTD4; EC50 = 0.77 nm), but was more potent than carbachol (EC50 = 0.15 μm) or substance P (EC50 = 1.4 μm). Endothelin was a more potent contractile agent in rat endothelium‐denuded aorta (EC50 = 2.1 nm) than in guinea‐pig trachea.

Endothelin‐induced contraction in guinea‐pig trachea was unaffected by mepyramine (10 μm), atropine (1 μm), SK&F 104353 (10 μm), a leukotriene receptor antgonist, or SQ 29,548 (1 μm), a thromboxane receptor antagonist. The contraction produced by 0.3 μm endothelin was potentiated by cyclo‐oxygenase inhibition with 5 μm indomethacin.

Nicardipine (0.01 or 0.1 μm) or incubation in calcium‐free medium +0.1 mm EGTA for 30 min had a relatively minor or no effect on endothelin concentration‐response curves in guinea‐pig intact trachea, but markedly inhibited responses produced by endothelin in endothelium‐denuded aorta of the rat. Increasing the EGTA concentration in calcium‐free medium to 1 mm abolished endothelin‐induced contraction in guinea‐pig trachea.

In guinea‐pig trachea, ryanodine (10 μm) produced a 2.1 fold shift to the right of endothelin concentration‐response curves and reduced the maximum response elicited by 0.3 μm endothelin.

Staurosporine (0.01 μm and 0.1 μm), a protein kinase C inhibitor, was without effect on endothelin‐ or carbachol‐induced contraction in guinea‐pig trachea, but markedly inhibited the response produced by endothelin in rat aorta.

Endothelin (3 nm‐0.3 μm) produced a concentration‐dependent stimulation of phosphatidylinositol (PI) turnover in guinea‐pig intact trachea, with an EC50 value of 45.9 nm.

Removal of the epithilium markedly potentiated endothelin‐induced contraction in guinea‐pig trachea, producing a 4.7 fold leftward shift in endothelin concentration‐response curves and an increase in the contractile response elicited by 0.3 μm endothelin.

These data indicate that endothelin is a potent agonist in guinea‐pig trachea whose response is markedly enhanced by removal of the airway epithelium. Endothelin‐induced contraction is not mediated to a marked extent by calcium influx via dihydropyridine‐sensitive calcium channels and does not involve the release of histamine, acetylcholine, leukotrienes or thromboxane. Rather, endothelin appears to produce contraction of guinea‐pig trachea via a direct action which involves stimulation of PI turnover and utilization of calcium from intracellular stores and, also, calcium influx via a pathway that is not sensitive to dihydropyridine calcium channel inhibitors. Endothelin‐induced contraction of rat aorta was more sensitive to the effects of incubation in Ca2+‐free medium, nicardipine or staurosporine, suggesting that differences exist in the relative mechanisms whereby endothelin produces contraction in different tissues.
Decreased GABAA receptor subunit mRNA concentrations following chronic lorazepam administration
Authors: Kang Inwha , Miller Lawrence G.
Published: July 2012

Chronic benzodiazepine administration has been associated with alterations in binding and function at the GABAA receptor. To evaluate effects of chronic benzodiazepine exposure on messenger RNA (mRNA) concentrations for several GABAA receptor subunits, we treated mice with lorazepam, 2 mg kg−1 daily for 1–28 days and evaluated mRNAs for the alpha1 and gamma2 subunits by Northern hybridization. In cerebral cortex, concentrations of mRNA for the alpha1 and gamma2 subunits were unchanged from vehicle or control after 1–10 days of lorazepam. However, after 14 days of treatment mRNA concentrations for both subunits decreased to approximately 50% of control values and remained decreased at 28 days. In contrast, no significant alterations were observed for either subunit mRNA in hippocampus or cerebellum over the same time course. Alterations in mRNAs in cortex occur after the development of tolerance and receptor downregulation in this model.
Inhibition of eosinophil cyclic nucleotide PDE activity and opsonised zymosan‐stimulated respiratory burst by ‘type IV’‐selective PDE inhibitors
Authors: Dent Gordon , Giembycz Mark A. , Rabe Klaus F. , Barnes Peter J.
Published: July 2012

The cyclic nucleotide phosphodiesterase (PDE) of guinea‐pig eosinophils was partially characterized and the effects of selective inhibitors of PDE isoenzymes upon opsonized zymosan (OZ)‐stimulated respiratory burst were studied.

PDE activity in eosinophil lysates appeared to be membrane‐associated, displayed substrate specificity for adenosine 3′: 5′ cyclic monophosphate (cyclic AMP) versus guanosine 3′: 5′ cyclic monophosphate (cyclic GMP) and was insensitive to cyclic GMP or Ca2+ and calmodulin.

The non‐selective PDE inhibitor, 3‐isobutyl‐1‐methylxanthine caused a concentration‐dependent inhibition of both OZ‐stimulated hydrogen peroxide (H2O2) generation and cyclic AMP hydrolysis. The type IV‐selective PDE inhibitors, rolipram and denbufylline, also inhibited H2O2 generation and cyclic AMP hydrolysis in a concentration‐dependent manner whilst SK&F 94120 and Org 9935 (type III‐selective) and zaprinast (type Ia or V‐selective) were ineffective.

Dibutyryl cyclic AMP, a cell‐permeable, non‐hydrolysable analogue of cyclic AMP, caused a concentration‐dependent inhibition of H2O2 generation stimulated by OZ. Dibutyryl cyclic GMP was ineffective.

It is concluded that eosinophil respiratory burst activity induced by OZ can be regulated by intracellular cyclic AMP and that the levels of cyclic AMP are controlled exclusively by a rolipram‐ and denbufylline‐sensitive PDE isoenzyme that resembles a type IV species.
The pharmacological specificity of N‐methyl‐d‐aspartate receptors in rat cerebral cortex: correspondence between radioligand binding and electrophysiological measurements
Authors: Grimwood Sarah , Foster Alan C. , Kemp John A.
Published: July 2012

The pharmacological specificity of N‐methyl‐d‐aspartate (NMDA) receptors has been investigated in the rat cerebral cortex by use of radioligand binding and electrophysiological techniques.

A comparison was made between a functional assay (NMDA‐induced depolarizations in a rat cortical slice preparation) and NMDA‐sensitive l‐[3H]‐glutamate binding in the same brain region and species, to provide accurate affinity values for agonists and antagonists at the NMDA recognition site.

In a preparation of crude postsynaptic densities (PSD) from rat cortex, l‐[3H]‐glutamate bound with high affinity to an NMDA‐sensitive population of sites with KD (geometric mean (–s.e.mean, +s.e.mean) = 120 (114, 126) nm, Bmax (mean ± s.e.mean) = 11.4 ± 0.8 pmol mg−1 protein and Hill coefficient (mean ± s.e.mean) = 1.2 ± 0.17 (n = 3).

There was a good agreement between the relative affinities in radioligand binding and electrophysiological assays for the receptor agonists NMDA, N‐methyl‐l‐aspartate, quinolinate and trans‐2,3‐piperidine dicarboxylate, which are poor substrates of acidic amino acid transport systems. However, agonists which are good substrates for high affinity uptake systems (l‐ and d‐glutamate, l‐ and d‐aspartate, d‐aspartate‐β‐hydroxamate and l‐glutamate‐γ‐hydroxamate) were much weaker in the electrophysiological experiments.

Schild analysis of the antagonism of NMDA responses in the rat cortical slice by dl‐3‐(2‐carboxypiperazin‐4‐yl)propyl‐1‐phosphonate, d‐ and dl‐2‐amino‐5‐phosphonovalerate, d‐ and dl‐2‐amino‐ 7‐phosphonoheptanoate, d‐β‐aspartylaminomethylphosphonate, d‐γ‐glutamylglycine and d‐α‐aminoadipate (d‐AA) indicated a competitive interaction with respective pA2 values of 6.17, 5.62, 5.24, 5.28, 5.20, 5.00, 4.43 and 3.97.

In the radioligand binding experiments the same antagonists inhibited only the NMDA‐sensitive component of l‐[3H]‐glutamate binding. IC50 values showed a good correlation with the pA2 values (correlation coefficient = 0.96), with the exception of d‐AA which was more potent than anticipated in the binding experiments (IC50 = 9.8 μm).

These results confirm that NMDA‐sensitive l‐[3H]‐glutamate binding sites represent the NMDA recognition site of the NMDA receptor and provide affinity values for both agonists and antagonists in the rat cerebral cortex, agreeing well with previous estimates in this and other tissues.
Mechanical responses to catecholamines in the longitudinal muscle of guinea‐pig gastric fundus
Authors: Parekh A.B. , Syed M. Md , Tomita T.
Published: July 2012

In the longitudinal muscle of guinea‐pig gastric fundus, adrenaline and phenylephrine (1–30 μm) both produced a slow contraction preceded by a relaxation. The slow contraction was strongly inhibited by prazosin (0.1 μm), but only weakly by yohimbine (1μm), suggesting main contribution of α1‐adrenoceptors.

Most of the slow contraction was blocked by meclofenamate or indomethacin (0.1–0.3 μm). Both these drugs also inhibited spontaneously generated muscle tone. In some preparations, obtained from the apical fundus, a small contraction remained in the presence of meclofenamate.

During contraction induced by prostaglandin E2, adrenaline produced sustained relaxation and phenylephrine often transient relaxation, in the presence of meclofenamate. The transient relaxation, but not the sustained relaxation, was suppressed by prazosin.

In the presence of prostaglandin E2 (5 nm), after treating with phenoxybenzamine (30 μm) for 30 min, isoprenaline and adrenaline produced concentration‐dependent relaxation, with IC50s of 3.9 nm and 64 nm, respectively. Propranolol shifted these concentration‐response curves to the right, with apparent pA2s of 8.15 and 7.34, respectively.

It is suggested that in the fundic longitudinal muscle, adrenaline‐induced contraction is mediated mainly by an increase in endogenous prostaglandin production through activation of α1 ‐adrenoceptors and that adrenaline produces transient relaxation through α1‐adrenoceptors and sustained relaxation through β‐adrenoceptors. The β‐adrenoceptors in the longitudinal muscle are more sensitive to adrenaline and isoprenaline than those in the circular muscle.
Renal handling and effects of S(+)‐ibuprofen and R(–)‐ibuprofen in the rat isolated perfused kidney
Authors: Cox P.G.F. , Moons W.M. , Russel F.G.M. , van Ginneken C.A.M.
Published: July 2012

The renal handling and effects of S(+)‐ and R(–)‐ibuprofen have been studied in the isolated perfused kidney (IPK) of the rat.

Both ibuprofen enantiomers were extensively reabsorbed and accumulated in the kidney in a concentration‐dependent manner. No pharmacokinetic differences were observed between the two enantiomers.

3 S(+)‐ibuprofen concentrations ranging from 0.25 to 25 μg ml−1 (1.2 to 120 μm) caused a decrease in urinary flow, glomerular filtration rate (GFR) and electrolyte excretion. Urinary pH and excretion of glucose were not influenced. R(–)‐ibuprofen concentrations ranging from 2.5 to 25 μg ml−1 (12 to 120 μm) also decreased urinary flow and electrolyte excretion. This decrease, however, was less than observed with S(+)‐ibuprofen. GFR, urinary pH and glucose excretion were not affected by R(–)‐ibuprofen. Prostaglandin E2 (PGE2) concentrations of 133 ng ml−1 reversed the effects on renal function of both enantiomers.

Very high S(+)‐ and R(–)‐ibuprofen concentrations (> 400 μg ml−1) resulted in an increase in urinary flow and fractional excretion of sodium, chloride, potassium, glucose and calcium.

It is concluded that the pharmacokinetic behaviour of ibuprofen in the kidney is not stereoselective. Relatively high concentrations of both enantiomers increased the urinary flow and electrolyte excretion in a nonstereoselective manner. Lower concentrations of S(+)‐ibuprofen decreased urinary flow and electrolyte excretion. The pharmacologically inactive R(–)‐ibuprofen was also able to affect renal function in a similar way, but at different concentrations. These effects on renal function are probably caused by inhibition of PGE2 synthesis.
Modulation of the structure‐binding relationships of antagonists for muscarinic acetylcholine receptor subtypes
Authors: Pedder E. Kristina , Eveleigh Phike , Poyner David , Hulme Edward C. , Birdsall Nigel J.M.
Published: July 2012

Membranes from rat cerebral cortex, myocardium and extraorbital lacrimal gland were used as sources of M1 M2 and M3 muscarinic acetylcholine receptors respectively and the affinities of seven antagonists for the three subtypes were examined under different experimental conditions.

The affinities for the membrane‐bound receptors were measured at different ionic strengths and temperatures and compared with those determined on the receptor solubilised in the neutral detergent digitonin or the zwitterionic detergent, CHAPSO.

The range of measured affinity constants of a given antagonist for a specific subtype varied from 2 (atropine at M1 receptors) to 1000 (AF‐DX 116 at M2 receptors).

As a consequence of these changes in affinity, which were dependent on the drug, the subtype and the experimental conditions, both the structure‐binding relationships of a given subtype can be markedly changed as well as the selectivity of a drug for the different subtypes. For example it is possible to change the relative affinities of AF‐DX 116 and gallamine at membrane‐bound M1 receptors from 50:1 to 1:60.

Experimental conditions for the observation of high selectivity of pirenzepine, AF‐DX 116, gallamine and hexahydrosiladiphenidol for the three subtypes are given.

When the receptors are removed from their membrane environment by solubilisation in detergent, antagonist affinities are changed but the subtypes still retain different structure‐binding relationships.

In general, AF‐DX 116 and the allosteric antagonist, gallamine, behave differently from the other antagonists, suggesting that they bind in different ways to muscarinic receptors.

Careful attention should therefore be paid to the experimental conditions in binding assays used to assess the affinities and selectivities of new muscarinic antagonists in order to avoid misleading results.

The ability to produce enhanced or attenuated affinities and selectivities of antagonists, resulting from the induction of different conformations of the receptor by a variety of physical, chemical or molecular biological perturbations may lead to a better understanding of the structural basis of drug receptor interactions.
Nitric oxide is the mediator of ATP‐induced dilatation of the rabbit hepatic arterial vascular bed
Authors: Mathie R.T. , Ralevic V. , Alexander B. , Burnstock G.
Published: July 2012

Livers of 10 New Zealand White rabbits were perfused in vitro with Krebs‐Bülbring buffer via the hepatic artery (HA) and portal vein (PV) at constant flows of 23 ± 1 and 77 + 1 ml min−1 100 g−1 respectively. The tone of the preparation was raised with noradrenaline (concentration: 10 μm).

Dose‐response curves for the vasodilatation produced by adenosine 5′‐triphosphate (ATP), acetylcholine (ACh), adenosine, and sodium nitroprusside (SNP) were obtained following injection into the HA supply. Injections were then repeated in the presence of the l‐arginine to nitric oxide pathway inhibitors N‐monomethyl‐l‐arginine (l‐NMMA, n = 6) and N‐nitro‐l‐arginine methyl ester (l‐NAME, n = 4) at concentrations of 30 μm and 100 μm for each inhibitor.

Both l‐NMMA and l‐NAME antagonized the responses to ATP and ACh; l‐NAME was 2–3 times more potent than l‐NMMA as an inhibitor of these endothelium‐dependent vasodilatations. Neither l‐NMMA nor l‐NAME attenuated responses of the endothelium‐independent vasodilators, adenosine and SNP.

These results indicate that nitric oxide is the mediator of ATP‐induced vasodilatation in the HA vascular bed of the rabbit and that the receptor responsible for the release of nitric oxide, the P2y‐purinoceptor, is located predominantly on the endothelium.
Tissue selectivity and spasmogen selectivity of relaxant drugs in airway and pulmonary vascular smooth muscle contracted by PGF2α or endothelin
Authors: O'Donnell Stella R. , Wanstall Janet C. , Kay Christine S. , Zeng Xiang‐ping
Published: July 2012

The spasmolytic effects of smooth muscle relaxant drugs with different mechanisms of action have been examined on isolated preparations of guinea‐pig trachea and rat pulmonary artery. The preparations were contracted with concentrations of prostaglandin F2α (PGF2α) or endothelin selected to give approximately 80% of the agonist maximum response on each tissue. These concentrations also caused similar levels of tone (% of tissue maximum contraction) on each tissue.

With endothelin as the spasmogen, the potassium channel opening drug, pinacidil, was more potent on trachea (– log IC50 5.49) than on pulmonary artery (4.39), i.e. was airway‐vascular selective, whereas with PGF2α as the spasmogen it was more potent on pulmonary artery (6.01) than on trachea (5.27), i.e. was vascular‐airway selective.

With endothelin as the spasmogen, fenoterol was also airway‐vascular selective (8.35 on trachea; little effect on pulmonary artery), nitroprusside was vascular‐airway selective (7.50 on pulmonary artery; 5.99 on trachea) and forskolin was non‐selective (6.69 on trachea; 6.70 on pulmonary artery). Thus, the airway‐vascular selectivity of the relaxant drugs varied with the drug.

On pulmonary artery, pinacidil, nitroprusside and forskolin were all more potent against PGF2α than against endothelin, i.e. 42, 4 and 7 fold respectively. On trachea, these drugs were equipotent against PGF2α and endothelin.

The results suggest that, in pulmonary artery, but not in trachea, the relative contribution of protein kinase C activation and calcium influx to the maintenance of tonic contractions to endothelin and PGF2α may be different. If protein kinase C activation should be the predominant mechanism for endothelin in pulmonary artery, then it may be more difficult to reverse this with relaxant drugs that lower intracellular calcium.

The study indicates that the airway‐vascular selectivity of relaxant drugs can be spasmogen‐dependent as well as dependent on the mechanism of action of the relaxant drug. Thus, relaxant drugs, whether of interest for their airway or vascular effects, should be tested against a full range of spasmogens of likely pathophysiological importance.
Pharmacokinetic characterization of phosphatidylserine liposomes in the rat
Authors: Palatini Pietro , Viola Giampietro , Bigon Emilio , Menegus Anna Maria , Bruni Alessandro
Published: July 2012

The plasma decay, tissue uptake and biotransformation of radiolabelled phosphatidylserine (PS) liposomes have been investigated in rats following bolus i.v. injection (2 mg kg−1).

PS plasma concentration showed a biexponential decay with half‐lives of 0.85 and 40 min. The following interpretation of the biphasic decay is proposed: (1) The rapid initial decline is due to the irreversible uptake of PS liposomes by the mononuclear phagocyte system, as demonstrated by the almost exclusive accumulation of PS in liver and spleen. (2) The slow decay phase reflects the elimination of that fraction of PS that has been incorporated into high density plasma lipoproteins (HDL). A kinetic model has been developed to describe these phenomena and a good agreement has been observed between experimental data and theoretical values.

Evidence has been obtained that a large fraction of PS is hydrolyzed at the injection site, probably by phospholipase A2 and other hydrolytic enzymes released by platelets. Hydrolysis at the injection site has also been observed following intraperitoneal and intramuscular injections.

As shown by the comparative analysis of the biotransformation products found in tissues after administration of either [3H]‐glycerol‐PS or [14C]‐serine‐PS, parenterally administered PS follows two distinct metabolic pathways: (1) decarboxylation to phosphatidylethanolamine and (2) extensive hydrolytic degradation with release of the individual components of the molecule. These pathways probably reflect the two main mechanisms of PS uptake, incorporation into the plasma membrane and internalization by endocytosis, respectively.
Secretion of growth hormone elicited by intravenous desipramine in the conscious, unrestrained rat
Authors: Grealy M. , O'Donnell J.M.
Published: July 2012

Acute intravenous administration of either clonidine (Clon) (50 μg kg−1) or desipramine (DMI) (5 mg kg−1) elicited a pulse of growth hormone (GH) and corticosterone secretion in conscious, unrestrained rats.

The responses to DMI were similar to those with Clon, except that the GH pulse following DMI was delayed and was not dose‐dependent.

The GH response to DMI was inhibited by prior administration of idazoxan (1 mg kg−1) or yohimbine (0.5 mg kg−1), but not by atropine (10 μg kg−1), sulpiride (5 mg kg−1) or prazosin (1 mg kg−1).

The corticosterone secretion following DMI was not altered by prior atropine, sulpiride or prazosin, but was augmented by idazoxan (1 mg kg−1).

GH secretion was not influenced by atropine, sulpiride, prazosin or idazoxan given alone. Idazoxan or yohimbine given alone elicited significant secretion of corticosterone.

It is concluded that i.v. DMI caused an activation through indirect mechanisms of α2‐adrenoceptors specifically involved in hypothalamic‐pituitary regulation of GH release and also a distinct, independent and transient generalized activation of the pituitary‐adrenal axis.
Evidence for regulatory control of iron uptake from ferric maltol across the small intestine of the rat
Authors: Barrand Margery A. , Callingham Brian A.
Published: July 2012

59Fe absorption from the novel iron compound, ferric maltol, was studied in rats pretreated twice daily for two weeks with non‐radioactive ferric maltol in oral doses containing 7 mg elemental iron. Tissue accumulation of 59Fe 2 h after administration of radioactive ferric maltol into the stomach was significantly lower in iron pretreated animals than in saline‐treated controls.

59Fe uptake from ferric maltol into isolated fragments of ileum and of duodenum was of similar magnitude in control animals but in iron‐treated animals, duodenal uptake was significantly lower than that of the ileum.

Absorption of 59Fe was also investigated in anaesthetized rats after intestinal perfusion with saline (controls) or with 5 mm chenodeoxycholate to render the intestines more permeable.

Changes in permeability of the small intestine were monitored by estimating the amount of [14C]‐mannitol absorbed and fluid secreted with reference to the non‐absorbable [3H]‐inulin in the perfusate effluents.

Despite the increased permeability of the intestines after bile salt treatment, there was little difference from control in the tissue accumulation of 59Fe from ferric maltol 2 h after intraduodenal administration. However 59Fe absorption from ferrous sulphate was significantly increased after bile salt treatment.

Gel filtration profiles of plasma made 5 and 60 min after intraduodenal administration of [59Fe]‐ferric [3H]‐maltol demonstrated that metal and ligand do not enter the circulation as the complex even when intestinal permeability is increased.

Uptake of 59Fe was investigated in isolated fragments of rat small intestine after saline or bile salt perfusion. Although 59Fe uptake from ferric maltol was somewhat greater in the bile salt‐treated intestinal fragments, saturable kinetics were still observed. By contrast, 59Fe uptake from ferrous sulphate: ascorbate was greatly enhanced by bile salt pretreatment and a very large diffusional component of uptake was evident.

It is concluded that iron uptake from ferric maltol may well be under endogenous regulatory control even in damaged intestines, so it is unlikely that this novel iron compound can bring about iron overload when administered orally.