bluuebunny/crossref_metadata_2025_split huggingface dataset:
Selectivity of [125I]‐PD 151242 for human, rat and porcine endothelin ETA receptors in the heart
Authors: Peter Markus G. , Davenport Anthony P. 
Published: July 2012

Endothelin‐1 binds with high affinity to heart where it acts as a potent positive inotropic agent. Our aim was to characterize the labelled and unlabelled ETA‐selective antagonist PD151242 in heart tissues derived from man, rat and pigs by use of radioligand binding techniques.
Binding of [125I]‐PD151242 to sections of human left ventricle was time‐dependent and reached equilibrium after 120 min at 23°C with an association rate constant of 0.0235 min−1 nm−1. The binding was reversible at 23°C with a dissociation rate constant of 0.00144 min−1.
Saturation binding assays with [125I]‐PD151242 revealed a single population of high affinity ET receptors in human left ventricle (KD = 1.07 ± 0.08 nm; Bmax = 29.8 ± 4.2 fmol mg−1 protein), porcine left ventricle (KD = 1.92 ± 0.27 nm; Bmax = 493 ± 248 fmol mg−1 protein), and rat heart (KD = 0.64 ± 0.08 nm; Bmax = 82.34.7 fmol mg−1 protein).
Unlabelled PD151242 competed with specific [125I]‐ET‐1 binding to human left ventricle tissue in a biphasic manner with high affinity binding to the ETA‐site (KD = 7.21 ± 2.83 nm) and lower affinity for the ETB‐subtype (KD = 104 ± 23 μm), indicating a greater than 10000 fold selectivity to the high affinity site.
The ETA‐selective ligand FR139317 competed for [125I]‐PD151242 binding in human left ventricle with nanomolar affinity (KD = 0.37 ± 0.10 nm), whereas the ETB‐selective compound, BQ3020, competed with only micromolar affinity (KD = 1.5 ± 0.26 μm).
The novel ETA‐selective radioligand [125I]‐PD151242 binds with high affinity to human, rat and porcine heart. In human tissue, binding was shown to be reversible and highly selective for the ETA‐subtype making [125I]‐PD 151242 a useful selective radioligand for further characterization of the ETA‐receptor in human tissue.
Presence of vasoconstrictor 5HT1‐like receptors revealed by precontraction of rabbit isolated mesenteric artery
Authors: Choppin A. , O'Connor S.E. 
Published: July 2012

A series of 5‐hydroxytryptamine (5‐HT) receptor agonists including 5‐HT, 5‐carboxamidotryptamine (5‐CT) and sumatriptan produced little or no contraction of rabbit isolated mesenteric arteries under resting tone conditions, even at concentrations up to 10−4m.
When the same agonists were retested in mesenteric artery preparations pre‐contracted with the thromboxane‐mimetic, U46619, each demonstrated concentration‐related vasoconstrictor activity. 5‐CT and 5‐HT were especially potent and effective in this model giving EC50 values of 4.3 times 10−9 m and 1.6 times 10−8 m respectively and maximum effects equivalent to those of KC1 80 mm. In preparations precontracted by U46619 (conditions retained throughout the rest of the study) the order of agonist potency was 5‐CT>5‐HT>RU 24969 = sumatriptan>8‐hydroxy‐2‐(di‐n‐propylamino)tetralin (8‐OHDPAT) > cisapride.
The vasoconstrictor effects of 5‐CT were competitively antagonized by methiothepin (pA2 8.20) but resistant to antagonism by a range of other 5‐HT receptor antagonists, i.e. pindolol (5‐HT1A/5‐HT1B), propranolol (5‐HT1B), spiperone (5‐HT2A), ondansetron (5‐HT3), ICS 205930 (5‐HT3/5‐HT4) and SDZ 205557 (5‐HT4). 5‐CT responses were slightly antagonized by a high concentration of ritanserin (5‐HT2A/5‐HT2C). Responses to 5‐HT and sumatriptan were also antagonized by methiothepin with similar affinity (pA2/pKB values ⋍ 8.0).
Metergoline and rauwolscine (10−7 −10−6m) antagonized the effects of 5‐CT in a non‐competitive fashion giving pKBapp values of 7.13 (metergoline) and 6.86 (rauwolscine).
Vasoconstrictor responses to 5‐HT were not modified in the presence of ritanserin (3 times 10−7 m) or spiperone (3 times 10−7m) and only modestly antagonized by ketanserin (10−6m) suggesting that 5‐HT2A receptors do not make a significant contribution in this model.
Hence, precontraction of rabbit mesenteric arteries reveals potent vasoconstrictor effects of 5‐HT and related agonists. Based on the agonist potency order and the antagonist studies performed, the receptor subtype responsible has the characteristics of a 5‐HT1‐like (probably 5‐HT1D) receptor. This study therefore demonstrates a particularly striking example of vasoconstrictor synergy involving 5‐HT1‐like receptors.
Differential effect of sodium ions and guanine nucleotides on the binding of thioperamide and clobenpropit to histamine H3‐receptors in rat cerebral cortical membranes
Authors: Clark E.A. , Hill S.J. 
Published: July 2012

Conflicting reports in the literature over heterogeneity (West et al., 1990) or homogeneity (Arrang et al., 1990) of histamine H3‐receptor binding sites may be attributed to the use of different incubation conditions. In the present study we have investigated the extent to which the binding of H3‐receptor ligands to rat cerebral cortical membranes can be modified by both sodium ions and guanine nucleotides.
The H3‐selective antagonist, thioperamide, discriminated between two specific binding sites for [3H]‐Nα‐methylhistamine (IC50 1 = 2.75 ± 0.87 nm, IQ50 2 = 101.6 ± 12.0 nm, % site 1 = 24 ± 2%) in 50 mm Tris HCl buffer, but showed homogeneity of binding in 50 mm Na/K phosphate buffer.
Sodium ions markedly altered the binding characteristics of thioperamide (i.e. heterogeneity was lost and IC50 value shifted towards the high affinity site). The competition curves for a second H3‐antagonist, clobenpropit and the H3‐agonist Nα‐methylhistamine however, were unaltered in the presence of sodium ions.
Guanylnucleotides displaced only 60% of specific [3H]‐Nα‐methylhistamine binding and modulated thioperamide binding in the same way as sodium ions.
These data suggest that the H3‐receptor can exist in different conformations for which thioperamide, but not Nα‐methylhistamine and clobenpropit, show differential affinity.
The potential nature of these sites, and the implications of this apparent receptor heterogeneity for H3‐receptor antagonism by thioperamide, are discussed.
Dotarizine versus flunarizine as calcium antagonists in chromaffin cells
Authors: Villarroya Mercedes , Gandía Luis , Lara Baldomero , Albillos Almudena , López Manuela G. , García Antonio G. 
Published: July 2012

Dotarizine is a novel piperazine derivative structurally related to flunarizine that is currently being evaluated in clinical trials for its antimigraine and antivertigo effects. This clinical profile may be related to its Ca2+ antagonist properties. Therefore, the actions of both compounds as calcium antagonists were compared in bovine chromaffin cells.
Dotarizine and flunarizine blocked 45Ca2+ uptake into K+ depolarized chromaffin cells (70 mm K+/0.5 mm Ca2+ for 60 s) in a concentration‐dependent manner, with IC50S of 4.8 and 6.7 μm, respectively.
Dotarizine and flunarizine also inhibited the whole‐cell Ca2+ and Ba2+ currents (Ica, IBa) in voltage‐clamped chromaffin cells, induced by depolarizing test pulses to 0 mV, during 50 ms, from a holding potential of − 80 mV. Blockade exhibited IC50S of 4 μm for dotarizine and 2.2 μm for flunarizine. Dotarizine increased the rate of inactivation of ICa and IBa; inhibition of whole‐cell currents was use‐dependent.
Transient increases of the cytosolic Ca2+ concentration, [Ca2+]i, produced by K+ stimulation (70 mm K+ for 5 s) of single fura‐2‐loaded chromaffin cells, were also inhibited by dotarizine and flunarizine with IC50s of 1.2 and 0.6 μm, respectively. Upon washout of dotarizine, the [Ca2+]i increases recovered fully after 5–10 min. In contrast, the responses remained largely inhibited 10 min after washing out flunarizine.
Catecholamine release induced by K+ stimulation (10‐s pulses of 70 mm) was inhibited by dotarizine with an IC50 of 2.6 μm and by flunarizine with an IC50 of 1.2 μm. The blocking effects of both compounds developed slowly, and was fully established after 20–30 min of superfusion. While blockade by dotarizine quickly reversed upon its washout, that of flunarizine lasted even 25 min after washing out.
Catecholamine release from electroporated chromaffin cells triggered by 10 μm Ca2+ was not affected by 10 μm dotarizine or flunarizine.
Overall, the results suggest that dotarizine behaves as a Ca2+ antagonist in bovine chromaffin cells. It exhibits a potency similar to flunarizine in blocking Ca2+ entry, Ca2+ channels, Cai2+ signals and secretion. The dotarizine effects are readily reversible suggesting that in contrast to flunarizine, it does not accumulate in cells. Dotarizine is devoid of intracellular effects on the secretory machinery. All its blocking effects on Ca2+ entry, [Ca2+]i rise and secretion are probably due to blockade of various Ca2+ channel subtypes in chromaffin cells. This blockade is use‐dependent and seems to be due to the increase by dotarizine of the rate of Ca2+ channel inactivation.
Tonic and use‐dependent block of sodium currents in isolated cardiac myocytes by bisaramil
Authors: Pugsley Michael K. , Saint David A. 
Published: July 2012

The effects of bisaramil on sodium currents in rat isolated cardiac myocytes were examined by use of tight‐seal, whole‐cell patch clamp techniques. Bisaramil produced a concentration‐dependent, readily reversible reduction in peak transient sodium current. When the sodium current was evoked at 3 s intervals the estimated ED50 for bisaramil was about 11 μm.
Bisaramil (16 μm) produced a shift in the inactivation curve to hyperpolarized potentials of about 10 mV, but produced no change in the voltage‐dependence of activation.
The block of the sodium current by bisaramil showed a profound use‐dependence. A concentration of 10 μm produced a considerable block of the current with repeated stimulation. The recovery from block was biphasic, showing fast and slow components which had time constants of about 40 ms and 5 s respectively.
Bisaramil produced little tonic block of the sodium current at concentrations of 100 μm; at 300 μm it produced tonic block of around 50%, with extreme use‐dependence.
Bisaramil appeared not to interact primarily with the inactivated form of the channel, since lengthening the depolarizing pulses did not affect the degree of block produced.
Regulation of histamine‐ and UTP‐induced increases in Ins(1,4,5)P3, Ins (1,3,4,5)P4 and Ca2+ by cyclic AMP in DDT1 MF‐2 cells
Authors: Sipma Henk , Duin Marry , Hoiting Ben , den Hertog Adriaan , Nelemans Adriaan 
Published: July 2012

Stimulation of P2U‐purinoceptors with UTP or histamine H1‐receptors with histamine gave rise to the formation of inositol 1,4,5‐trisphosphate (Ins(1,4,5)P3) and inositol 1,3,4,5‐tetrakisphosphate (Ins(1,3,4,5)P4) in DDT1 MF‐2 smooth muscle cells.
Stimulation of P2U‐purinoceptors or histamine H1‐receptors caused an increase in cytoplasmic Ca2+, consisting of an initial peak, representing the release of Ca2+ from internal stores and a sustained phase representing Ca2+ influx.
The P2U‐purinoceptor‐mediated Ca2+‐entry mechanism was more sensitive to UTP than Ca2+‐mobilization (EC50: 3.3 μm ± 0.4 μm vs 55.1 μm ± 9.2 μm), in contrast to these processes activated by histamine H1‐receptors (EC50: 5.8 μm ± 0.6 μm VS 3.1 μm ± 0.5 μm).
Pre‐stimulation of cells with several adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) elevating agents, reduced the histamine H1‐receptor‐mediated formation of Ins(1,4,5)P3 and Ins(1,3,4,5)P4. Forskolin completely inhibited Ins(1,4,5)P3 formation (IC50: 158 ± 24 nm) whereas Ins(1,3,4,5)P4 formation was inhibited by only 45% (IC50: 173 ± 16 nm). The P2U‐purinoceptor‐mediated production of these inositol phosphates was not affected by cyclic AMP.
Forskolin and isoprenaline reduced the histamine‐induced increase in cytoplasmic Ca2+, as measured in Ca2+ containing medium and in nominally Ca2+‐free medium but did not change the UTP‐induced increase in cytoplasmic Ca2+.
These results clearly demonstrate that cyclic AMP differentially regulates components of the histamine induced phospholipase C signal transduction pathway. Furthermore, cyclic AMP does not affect the phospholipase C pathway activated by stimulation of P2U‐purinoceptors in DDT1 MF‐2 cells.
Inhibition of delayed rectifier K+‐current by levcromakalim in single intestinal smooth muscle cells: effects of cations and dependence on K+‐flux
Authors: McHugh D. , Beech D J. 
Published: July 2012

Whole‐cell voltage‐clamp recordings were made from single smooth muscle cells isolated from the longitudinal layer of the guinea‐pig small intestine.
Levcromakalim ((−)Ckm) inhibited delayed rectifier K‐current (IK(DR)) and induced a voltage‐independent K‐current (IK(‐Ckm)). Both effects were inhibited similarly by glibenclamide. In some cells, however, IK(‐ckm) could be induced without any effect on IK(DR).
Ba2+ caused a voltage‐dependent block of IK(‐Ckm). The IC50 was 0.2 mm at − 40 mV (6 cells), but at 0 mV 2 mm Ba2+ caused only a 26 ± 7% inhibition (n = 5). Ba2+ had much less effect on IK(DR), 2 mm Ba2+ having no inhibitory effect on current elicited by depolarization to − 30 mV (n = 6) or 0 mV (n = 5).
Low concentrations of Zn2+ blocked IK(‐Ckm) while having little effect on IK(DR). Zn2+ (40 μm) caused a 77 ± 1% reduction of IK(‐Ckm) at − 30 mV (n = 4) but IK(DR) was inhibited by only 10 ± 3% at the same voltage (n = 4).
Inward current amplitudes were compared in 135 mm Rb+ and 135 mm K+ bath solutions. (−)Ckm‐activated Rb+‐current was only 4% of the K+‐current, whereas delayed rectifier Rb+‐current was larger than K+‐current.
(−)Ckm did not inhibit IK(DR) if IK(‐Ckm) was blocked. In the presence of 2 mm Ba2+ or 135 mm Rb+, (−)Ckm did not induce current nor did it inhibit the delayed rectifier. When [Rb+]o was 25 mm and [K+]i was 130 mm, (−)Ckm elicited outward current and inhibited outward delayed rectifier current (at voltages positive of the reversal potential) but it did not elicit inward current or inhibit inward delayed rectifier current (at voltages negative of the reversal potential).
These experiments indicate that (−)Ckm‐activated K channels are more sensitive to inhibition by Ba2+ and Zn2+ and pass inward Rb+ current less well than delayed rectifier K channels. They also suggest that (−)Ckm does not modulate delayed rectifier K channels directly or via an intermediate protein but that the inhibitory effect of (−)Ckm on IK(DR) arises as a consequence of K+‐flux through (−)Ckm‐activated K channels.
Postsynaptic nicotinic receptor desensitized by non‐contractile Ca2+ mobilization via protein kinase‐C activation at the mouse neuromuscular junction
Authors: Kimura Ikuko , Dezaki Katsuya , Tsuneki Hiroshi , Kimura Masayasu 
Published: July 2012

Non‐contractile Ca2+ mobilization (unaccompanied by muscle contraction) was initiated by nerve stimulation in the presence of neostigmine (more than 0.03 μm) at the endplate region of mouse diaphragm muscles. In the process of nicotinic receptor desensitization, the depressant effect of non‐contractile Ca2+ on contractile Ca2+ mobilization was investigated by measurement of Ca2+‐aequorin luminescence.
When the phrenic nerve was stimulated with paired pulses having intervals of 150, 300, 600, 1000 and 2000 ms, contractile Ca2+ transients were elicited during the generation of non‐contractile Ca2+ mobilization. The amplitude of the contractile Ca2+ transients elicited by the second pulse (S2) was depressed at the shorter pulse intervals, but recovered to the initial contractile response (S1) at longer pulse intervals.
The extent of depression of S2 was enhanced by increasing the concentration of neostigmine (0.03 to 0.3 μm). When a low concentration (0.05 μm) of pancuronium, a competitive nicotinic antagonist, completely blocked non‐contractile Ca2+ mobilization, the depression of S2 was diminished.
The depression of S2 was enhanced when the peak amplitude of non‐contractile Ca2+ mobilization was raised by increasing the external Ca2+ concentration from 1.3 to 5 mm.
Staurosporine (10 nm), a protein kinase‐C inhibitor, diminished the depression of S2 despite large amounts of non‐contractile Ca2+ mobilization. The diminishing effect of staurosporine was counteracted by TPA (0.1 μm), a protein kinase‐C activator.
These findings suggest that non‐contractile Ca2+ mobilization may enhance the desensitization of the postsynaptic nicotinic receptor via activation of protein kinase‐C at the neuromuscular junction.
Characterization of α1‐adrenoceptors mediating vasoconstriction to noradrenaline and nerve stimulation in the isolated perfused mesentery of rat
Authors: Williams T.J. , Clarke D.E. 
Published: July 2012

The objective of this study was to investigate the α1‐adrenoceptor subtype(s) mediating vasoconstrictor responses to perfused and neuronally‐released noradrenaline (NA) in the isolated perfused mesentery preparation of rat.
Isolated mesenteric preparations (with gut attached) from male Sprague Dawley rats (250–300g) were perfused via the superior mesenteric artery with oxygenated Krebs solution at approximately 6 ml min−1. The effects of antagonists on vasoconstrictor responses to either perfused (±)‐NA or periarterial nerve stimulation (70 V, 2 ms pulse width, 10 s train) were determined.
Vasoconstrictor responses to perfused NA were antagonized by prazosin (pA2 = 9.3 ± 0.1), WB4101 (pA2 = 9.6 ± 0.1), 5‐methyl urapidil (5‐MU: pA2 = 9.0 ± 0.1), (+)‐niguldipine (insurmountable) and spiperone (pA2 = 7.7 ± 0.1). The insurmountable nature of the antagonism by (+)‐niguldipine (0.1 nm) was greatly reduced by co‐perfusion with prazosin (10 nm). Chloroethylclonidine (CEC: 100 μm for 20 min, followed by 40 min washout) caused an approximate twofold increase in the EC50 for (±)‐NA and reduced the maximum response by approximately 25%. Pre‐treatment of tissues with CEC (100 μm as above) did not significantly alter affinity estimates for prazosin (pA2 = 9.2 ± 0.1), WB4101 (pA2 = 9.3 ± 0.1) or 5‐MU (pA2 = 8.7 ± 0.2). Vasoconstrictor responses to periarterial nerve stimulation were antagonized by WB4101>5‐MU>prazosin ≫ spiperone. CEC (100 μm as above) reduced nerve‐stimulated responses by approximately 50%.
The affinity estimates for the various antagonists studied suggest that vasoconstrictor responses to both exogenous and neuronally‐released NA are mediated via the same α1‐adrenoceptor subtype. The pharmacological profile most resembles the ‘classical’ α1A‐adrenoceptor, which, in turn, appears to be a rat homologue of the cloned bovine α1C‐adrenoceptor.
Further characterization of [3H]‐CGS 21680 binding sites in the rat striatum and cortex
Authors: Kirk Ian P. , Richardson Peter J. 
Published: July 2012

The putative high affinity binding site for the adenosine A2Areceptor agonist 2‐p‐(2‐carboxyethyl)phenethyl‐amino‐5′‐N‐ ethylcarboxamidoadenosine (CGS 21680) in the rat cerebral cortex was characterized by use of a number of selective A1and A2adenosine receptor ligands, and compared to the characteristics of the more abundant striatal A2Areceptor.The binding of [3H]‐CGS 21680 to cortical membranes was performed at pH 5.5, in order to increase the amount of specific binding.Reduction of the pH from 7.4 to 5.5 increased the apparent affinity of the striatal binding side for both agonists and antagonists. The relative order of potencies of both groups of ligands were the same at both pH values, and were consistent with binding to the A2Areceptor. There was no observable change in theBmax, the values being 415 and 446 fmol mg−1protein at pH 5.5 and 7.4 respectively.The cortical binding site yielded aBmaxvalue of 117 fmol mg−1protein. The relative order of potencies of the adenosine receptor ligands observed at this binding site were not the same as those observed in the striatum, exhibiting a profile with both A1and A2characteristics.Further characterization of this cortical binding site in the presence of the A1selective antagonist 1,3‐dipropyl‐8‐cyclopentylxanthine (DPCPX) revealed a more typical A2Aprofile. This indicated that under the conditions used there were two components of [3H]‐CGS 21680 binding, approximately 20% of the A1receptor and 80% to the A2Areceptor.It is concluded that in the cerebral cortex there is a CGS 21680 binding site showing the characteristic properties of the striatal A2Areceptor, and no evidence was obtained for the existence of a novel A2A‐like binding site.
Pharmacokinetic modelling of the haemodynamic effects of the A2a adenosine receptor agonist CGS 21680C in conscious normotensive rats
Authors: Mathôt R.A.A. , Cleton A. , Soudijn W. , IJzerman A.P. , Danhof M. 
Published: July 2012

The aim of the present investigation was to determine the relationship between the blood concentration and haemodynamic effects of the adenosine A2a receptor agonist, CGS 21680C (the sodium salt of 2‐p‐(2‐carboxyethyl)phenylethylamino‐5′‐N‐ethylcarboxamidoadenosine) in conscious normotensive rats.
Chronically cannulated rats were randomly assigned to three groups which received 300, 1000 or 3000 μg kg−1 (0.56, 1.9 or 5.6 μmol kg−1) of CGS 21680C intravenously over 15min. The mean arterial blood pressure (MAP) and heart rate (HR) were monitored continuously during the experiment and serial arterial blood samples were taken for analysis of drug concentration. The ratio MAP/HR was also calculated, which may reflect changes in total peripheral resistance on the assumption that no changes in stroke volume occur.
For each individual rat the reduction in mean arterial pressure was related to the blood concentration according to the sigmoidal Emax model. The concentration‐effect relationships were consistent for the different treatment groups. The potency based on free drug concentrations (EC50,u) was 5.8 ng ml−1 (11 nm) (mean ± s.e.; n = 19) and correlated well with the reported adenosine A2a receptor affinity (Ki 19 nm). In comparison with the reduction in blood pressure, CGS 21680C exhibited a greater potency for the reduction of the ratio MAP/HR.
It is concluded that estimates can be obtained for the potency and intrinsic activity of adenosine A2a receptor agonists in vivo by pharmacokinetic‐pharmacodynamic analysis of mean arterial pressure data in a rat model. In future studies, total peripheral resistance may also be useful as a pharmacodynamic parameter for A2a activation, provided that possible changes of the stroke volume are also assessed.
Characterization of endothelin receptors in rat renal artery in vitro
Authors: Clark Kenneth L. , Pierre Lisa 
Published: July 2012

The aim of this study was to investigate the function and characteristics of endothelin receptors in rat main branch renal artery in vitro.
Endothelin(ET)‐1 (mean EC50 = 9.8 nm) was approximately 12 fold more potent than ET‐3 (mean EC50 = 120 nm) as a contractile agonist and produced a greater maximum response. In contrast, neither of the ETB receptor‐selective agonists, alanine1,13,11,15] ET‐1 nor sarafotoxin S6c, (0.1 nm‐1 μm), induced any contractile effect, or any relaxant effect in endothelium‐intact preparations pre‐contracted with the thromboxane A2 mimetic, U‐46619. Sarafotoxin S6c (30 nm) also failed to induce any further contraction in tissues pre‐contracted with an EC50 concentration of ET‐1.
The ETA receptor‐selective antagonist, BQ123, behaved as a weak and variable antagonist of the contractile effects of ET‐1 (mean pA2 estimates in the range 5.8–6.3). In contrast, BQ123 antagonized ET‐3 with a potency (mean pA2 = 7.6) consistent with its affinity for ETA receptors. Co‐incubation of BQ123 (3 μm) with the putative ETB receptor‐selective antagonist, IRL1038 (10 μm), produced no greater antagonism of ET‐1 responses than was induced by BQ123 (3 μm) alone.
In conclusion, ETB receptors do not appear to be present in rat main branch renal artery. The contractile effects of ET‐3 in this tissue seem to be mediated by ETA receptors. While ETA receptors partly mediate the contractile effects of ET‐1, these data raise the possibility that a population of novel BQ123‐insensitive endothelin receptors may also contribute to this response.