bluuebunny/crossref_metadata_2025 huggingface dataset:
Ligand binding properties of putative β3‐adrenoceptors compared in brown adipose tissue and in skeletal muscle membranes
Authors: Sillence Martin N. , Moore Nikki G. , Pegg Graham G. , Lindsay Derek B.
Published: July 2012

The β‐adrenoceptor population was characterized in membrane preparations from rat brown adipose tissue (BAT) and from soleus muscle by use of the radioligand [125I]‐iodocyanopindolol ([125I]‐ICYP). In addition, atypical binding sites for [125I]‐ICYP found in both tissues were examined, and the relationship between these sites and the putative rat β3‐adrenoceptor is discussed.

It was established that BAT membranes host a mixed population of β1‐ and β2‐adrenoceptors. Of these two sites, 55% showed a high affinity for the β1‐selective ligand CGP 20712A (pK 8.5), and 45% showed a high affinity for the β2‐selective antagonist ICI 118551 (pK 8.6). Soleus muscle membranes were found to host a population of β2‐adrenoceptors, characterized by a high affinity for ICI 118551 (pK 9.1), but β1‐adrenoceptors could not be detected in this preparation. 5‐Hydroxytryptamine receptors were not detected in either preparation.

In addition to β1‐ and β2‐adrenoceptors, atypical binding sites were identified in both tissues using high concentrations of radioligand (0.5–0.6 nm) and in the presence of 1 μm (−)‐propranolol. The atypical sites were abundant, representing 80 and 81% of the total [125I]‐ICYP binding sites in BAT and soleus muscle respectively. When the pK values for 11 ligands were compared, the correlation coefficient for atypical sites in BAT and soleus muscle was 0.94.

The atypical binding sites showed a moderate affinity for (±)‐cyanopindolol (pK 7.3–7.7), poor stereoselectivity for the (+)‐ and (−)‐enantiomers of alprenolol (<10 fold), and a low affinity for β‐adrenoceptor antagonists and partial agonists in the order: (±)‐cyanopindolol>(−)‐alprenolol> (−)‐propranolol = (±)‐ICI 118551 >>(±)‐CGP20712A. The affinity of these ligands for the atypical sites reflects their behaviour in functional studies of putative β3‐adrenoceptors in rat BAT, white adipose tissue, intestine and colon.

The atypical sites labelled by [125I]‐ICYP were resistant to agonist binding, and while the order of affinity of the agonists BRL 37344 > isoprenaline > noradrenaline matches their order of potency at putative β3‐adrenoceptors, none of these compounds caused displacement of the radioligand at concentrations below 10 μm.

It is concluded that the atypical binding sites for [125I]‐ICYP found in rat BAT and soleus muscle membranes are the same, and that these sites show some relationship to the putative rat β3‐adrenoceptor identified in functional studies using antagonists. However, under the conditions used in the present study, pK values obtained for β3‐agonist binding are not useful.
Halothane inhibits the pressor effect of diphenyleneiodonium
Authors: Wang Yong‐Xiang , Pang Catherine C.Y.
Published: July 2012

We have recently found that diphenyleneiodonium (DPI), a novel inhibitor of nitric oxide (NO) synthase, causes pressor and tachycardic responses in pentobarbitone‐ but not halothane‐anaesthetized rats. The present study investigated the mechanism by which halothane suppresses the pressor response of DPI. The effects of halothane on the pressor response of DPI were also compared with those of other anaesthetic agents.

In conscious rats, i.v. bolus injections of DPI (0.025– 1.6 mg kg−1) caused dose‐dependent increases in mean arterial pressure (MAP), with ED50 of 0.07 ± 0.01 mg kg−1 and maximal rise of MAP (Emax) of 59 ± 2 mmHg. While ketamine potentiated Emax without altering the ED50 and pentobarbitone increased the ED50 without changing Emax of the pressor response to DPI, chloralose, urethane and ethanol displaced the curve to the right and potentiated Emax. In contrast, halothane (0.5–1.25%) dose‐dependently and non‐competitively reduced the pressor responses to DPI.

Intravenous bolus injection of a single dose of DPI (1.6 mg kg−1) caused immediate and large increases in plasma noradrenaline and adrenaline, as well as MAP in conscious rats. Halothane (1.25%) almost completely inhibited these increases.

The results suggest that DPI causes a pressor response in conscious rats by activating the sympathetic nervous system and halothane abolishes this pressor response by inhibiting activities of the sympathetic nervous system. The results also show that influences of anaesthetics must be taken into consideration when evaluating pressor response of vasoactive agents.
Selective reduction of N‐methyl‐D‐aspartate‐evoked responses by 1,3‐di(2‐tolyl)guanidine in mouse and rat cultured hippocampal pyramidal neurones
Authors: Fletcher Elizabeth J. , Church John , Abdel‐Hamid Khaled , MacDonald John F.
Published: July 2012

The effects of 1,3‐di(2‐tolyl)guanidine (DTG) were examined on the responses of cultured hippocampal neurones to the excitatory amino acid analogues N‐methyl‐d‐aspartate (NMDA), kainate, quisqualate and (RS)‐α‐amino‐3‐hydroxy‐5‐methylisoxazole‐4‐propionate (AMPA).

In rat hippocampal neurones loaded with the Ca2+‐sensitive dye Fura‐2, DTG (10–100 μm) produced a concentration‐dependent depression of the NMDA‐evoked rises in intracellular free calcium ([Ca2+]i), an effect that was not modified by changes in the extracellular glycine concentration. DTG (at 50 and 100 μm) also attenuated, although to a lesser extent, the rises in [Ca2+]i evoked by naturally‐derived quisqualate. In contrast, 50 and 100 μm DTG did not depress responses evoked by kainate, AMPA and synthetic, glutamate‐free (+)‐quisqualate although on occasions DTG enhanced kainate‐and AMPA‐evoked rises in [Ca2+]i.

DTG attenuated NMDA‐evoked currents recorded from mouse hippocampal neurones under whole‐cell voltage‐clamp with an IC50 (mean ± s.e.mean) of 37 ± 5 μm at a holding potential of −60 mV. The DTG block of NMDA‐evoked responses was not competitive in nature and was not dependent on the extracellular glycine or spermine concentration. The block did, however, exhibit both voltage‐, and use‐, dependency. The steady‐state current evoked by naturally‐derived quisqualate was also attenuated by DTG whereas those evoked by kainate and AMPA were not.

We conclude that DTG, applied at micromolar concentrations, is a selective NMDA antagonist in cultured hippocampal neurones, the block exhibiting both Mg2+‐ and phencyclidine‐like characteristics. Given the nanomolar affinity of DTG for σ binding sites it is unlikely that the antagonism observed here is mediated by σ‐receptors, but the data emphasize the potential danger of ascribing the functional consequences of DTG administration solely to σ receptor‐mediated events.
Blockade of adenosine receptors unmasks a stimulatory effect of ATP on cardiac contractility
Authors: Mantelli Laura , Amerini Sandra , Filippi Sandra , Ledda Fabrizio
Published: July 2012

The effects of ATP, α,β‐methylene ATP and β,γ‐methylene ATP on the contractile tension of guinea‐pig isolated left atria were evaluated.

ATP (1– 100 μm) produced a concentration‐dependent negative inotropic effect; this response was converted to a positive inotropic effect in the presence of the antagonist of adenosine A1 receptors, 1,3‐dipropyl‐8‐cyclopentylxanthine (DPCPX; 0.1 μm), and in the presence of 8‐phenyltheophylline (10 μm), an antagonist of A1 and A2 receptors.

The positive inotropic effect of ATP was antagonized by the P2 receptor antagonist, suramin (500 μm). Reactive blue 2 (30‐ 500 μm), a putative P2y receptor antagonist, concentration‐dependently reduced and finally abolished the effect of ATP.

In the presence of 8‐phenyltheophylline, the stable analogues of ATP, α,β‐methylene ATP and β,γ‐methylene ATP (1– 30 μm), produced a concentration‐dependent increase in atrial contractility of a lesser degree than that induced by ATP.

The results suggest that when inhibitory adenosine receptors are blocked, ATP produces a positive inotropic effect, probably mediated by P2y receptor stimulation.
Altered cardiac adrenergic neurotransmission in streptozotocin‐induced diabetic rats
Authors: Gando Satoshi , Hattori Yuichi , Kanno Morio
Published: July 2012

Functional alterations of the sympathetic neuroeffector junction of the left atria were studied in rats with streptozotocin‐induced diabetes.

Eight to 12 weeks of diabetes resulted in a marked decrease in the positive inotropic response of left atria to electrical field stimulation (EFS).

The overflow of [3H]‐noradrenaline from diabetic left atria caused by EFS was much less than that from control preparations.

The concentration‐response curves showed no change in sensitivities of the left atria to exogenous noradrenaline and tyramine in diabetic rats. The maximum positive inotropic response to these agents were similar in diabetic and control animals.

The left atrial content of noradrenaline was not significantly changed in diabetic rats. The cocaine‐sensitive uptake of [3H]‐noradrenaline was also unaltered.

Atropine enhanced the positive inotropic response and [3H]‐noradrenaline overflow induced by EFS in control left atria. Similarly, yohimbine caused an enhancement of EFS‐evoked inotropic response in control atria. However, these effects of the antagonists were not observed in diabetic left atria.

It is concluded that the decrease in the positive inotropic response of the left atria to EFS in diabetic rats is caused by an impairment of noradrenaline release from the sympathetic nerve terminals through a calcium‐dependent exocytotic mechanism. The present results also indicate that presynaptic α2‐adrenoceptors and muscarinic receptors that are linked to inhibition of the noradrenaline release during nerve stimulation may be functionally impaired in diabetic animals.
Stimulation of angiogenesis by substance P and interleukin‐1 in the rat and its inhibition by NK1 or interleukin‐1 receptor antagonists
Authors: Fan Tai‐Ping D. , Hu De‐En , Guard Steven , Gresham G. Austin , Watling Keith J.
Published: July 2012

Daily administration of 1 nmol substance P or 3 pmol recombinant human interleukin‐1 alpha (IL‐1α) caused intense neovascularization in a rat sponge model of angiogenesis. Lower doses of substance P (10 pmol) or IL‐1α (0.3 pmol) were ineffective when given alone. When combined at these low doses, substance P and IL‐1α interacted to produce an enhanced neovascular response.

By use of selective tachykinin NK1, NK2 and NK3 receptor agonists, ([Sar9,Met(O2)11]substance P, [β‐Ala8]neurokinin A(4–10), Succ‐[Asp6,MePhe8]substance P(6–11) (senktide), respectively), it was established that the activation of NK1 receptors is most likely to mediate the angiogenic response to substance P in this model.

The angiogenic activity of substance P and IL‐1α (10 pmol and 0.3 pmol day−1, respectively) was abolished by co‐administration of (i) the selective peptide NK1 receptor antagonist, L‐668,169 (1 nmol day−1), (ii) the selective non‐peptide NK1 receptor antagonists, RP 67580 and (±)‐CP‐96,345 (both at 1 nmol day−1) or (iii) the IL‐1 receptor antagonist, IL‐1ra, (50 μg day−1). In contrast, the selective NK2 receptor antagonist, L‐659,874 (1 nmol day−1) was ineffective.

The angiogenic action of substance P and IL‐1α was resistant to modification by mepyramine (1 nmol day−1) and/or cimetidine (10 nmol day−1), indomethacin (7 nmol day−1) or the platelet‐activating factor (PAF) antagonist, WEB‐2086 (22 nmol day−1), indicating that histamine, prostaglandins and PAF are not likely to be involved in this neovascular response.

The inhibition of the substance P/IL‐1 angiogenic response by selective NK1 receptor antagonists or by an IL‐1 receptor antagonist demonstrates that angiosuppression can be achieved by blocking the activity of angiogenic factors at the receptor level.
Membrane current responses to externally‐applied ATP in the longitudinal muscle of the chicken rectum
Authors: Matsuoka T. , Komori S. , Ohashi H.
Published: July 2012

Membrane current responses to ATP in enzymically‐dispersed single smooth muscle cells from the chicken rectum were investigated by the whole‐cell voltage clamp technique.

In cells dialysed with a KCl‐rich solution under voltage clamp at a holding potential of −40 mV, ATP (10 μm) produced an inward current followed by an outward current. When the holding potential was changed to 0 mV and −80 mV, the biphasic current response to ATP was converted to an outward current alone and an inward current alone, respectively.

External application of tetraethylammonium (TEA, 5 mm), intracellular dialysis with a CsCl‐rich solution, or inclusion of EGTA (10 mm) in the pipette abolished the outward current response to ATP.

Neither depletion of Ca2+ store with caffeine (10 mm) nor block of voltage‐gated Ca2+ channels with nifedipine (10 μm) affected the biphasic current response to ATP. After removal of the extracellular Ca2+ the outward current response to ATP was abolished.

α,β‐methylene ATP (100 μm) elicited a current similar to the ATP‐induced current. In the presence of α,β‐methylene ATP (100 μm), application of ATP (100 μm) was without effect.

In CsCl‐filled cells, ATP analogues elicited an inward current and the order of potency was ATP ≒ α,β‐methylene ATP > ADP > > AMP.

Inclusion of GTP γ S (0.2 mm) or GDP β S (2 mm) in the pipette did not affect the ATP‐induced inward current in CsCl‐filled cells. The reversal potential of the ATP‐induced inward current was about 0 mV and was completely inhibited after replacement of the cations in the bath solution by Tris. The reversal potential remained almost unchanged after replacement of Na+ in the bath solution with 110 mm Ca2+, but shifted in the negative direction after replacement of Na+ or both Na+ and Ca2+ with glucosamine.

The results suggest that ATP acts on P2 purinoceptors to cause activation of cation channels with selectivity for Ca2+ over Na+. Moreover, it appears that no G‐protein‐mediated mechanism is involved and increased Ca2+ entry through the cation channels causes activation of Ca2+‐activated K+ channels.
Effects of l‐ and d‐arginine and some related esters on the cytosolic mechanisms of α‐thrombin‐induced human platelet activation
Authors: Failli Paola , Cecchi Enrica , Ruocco C. , Fazzini A. , Giotti A.
Published: July 2012

In Fura‐2 preloaded human platelets, the increase in cytosolic calcium induced by α‐thrombin was reduced by some l‐ and d‐arginine ester compounds the IC50 (μm) values of which were 7.4 for TAEE, 56.9 for BAEE, 77.6 for TAME, 560 for T(d)AME, 656.3 for l‐ArgOMe and 2206.7 for d‐ArgOMe. α‐tosyl‐l‐Arginine, l‐ and d‐arginine were inactive.

The inhibitory activity of the l‐arginine esters was not modified when platelets were pretreated with 100 μm Nω‐monomethyl‐l‐arginine.

The l‐arginine esters did not increase cyclic GMP content in platelets either in the presence or absence of indomethacin and apyrase at rest and after α‐thrombin stimulation.

The kinetic parameters of platelet Na+/H+ antiporter (amiloride‐inhibitable, evaluated after cytosolic nigericin‐induced acidification) were modified by l‐ and d‐arginine esters, while the native amino acids were ineffective.

The inhibitory effects of the l‐ and d‐arginine esters on platelet activation appear to be mainly due to their inhibitory effect on Na+/H+ antiporter.
The effects of ATP and α,β‐methylene‐ATP on cytosolic Ca2+ level and force in rat isolated aorta
Authors: Kitajima Satoshi , Ozaki Hiroshi , Karaki Hideaki
Published: July 2012

The effects of a non‐selective P2‐receptor agonist ATP and a selective P2x‐receptor agonist α,β‐methylene‐ATP on intracellular free Ca2+ level ([Ca2+]i) and force were examined in rat isolated aorta without endothelium.

Both ATP (1–1000 μm) and α,β‐methylene‐ATP (0.1–100 μm) induced transient increase followed by small sustained increase in [Ca2+]i in a concentration‐dependent manner. Compared with the force induced by a high concentration of KCl, the force induced by α,β‐methylene‐ATP was smaller and that induced by ATP was much smaller at a given [Ca2+]i.

An L‐type Ca2+ channel blocker, verapamil (10 μm), completely inhibited the high K+‐stimulated [Ca2+]i and force. Verapamil partially inhibited the transient and sustained increases in [Ca2+]i induced by 10 μm α,β‐methylene‐ATP and the sustained increase but not the transient increase induced by 1 mm ATP.

In the absence of extracellular Ca2+ (with 0.5 mm EGTA) 1 mm ATP caused transient increase in [Ca2+]i while 10 μm α,β‐methylene‐ATP was ineffective

ATP, but not α,β‐methylene‐ATP, increased the tissue adenosine 3′:5′‐cyclic monophosphate (cyclic AMP) level.

These data suggest that ATP and α,β‐methylene‐ATP increase [Ca2+]i by an activation of both L‐type and non‐L‐type Ca2+ channels. In addition, ATP, but not α,β‐methylene‐ATP, increases [Ca2+]i by a release of Ca2+ from an intracellular Ca2+ store. Possible reasons are discussed as to why the increase in [Ca2+]i due to ATP and α,β‐methylene‐ATP resulted in only a small contraction.
Desensitization of histamine H1 receptor‐mediated inositol phosphate accumulation in guinea pig cerebral cortex slices
Authors: Bristow D.R. , Banford P.C. , Bajusz I. , Vedat A. , Young J.M.
Published: July 2012

Histamine stimulated the production of [3H]‐inositol phosphates in untreated (control) guinea‐pig cerebral cortex slices with a best‐fit EC50 of 17 ± 4 μm, and a best‐fit maximum response of 385 ± 23% over basal accumulation.

Histamine pretreatment desensitized guinea‐pig cortex slices to a subsequent challenge with histamine, which was observed as a reduction in the best‐fit maximum response to 182 ± 32% over basal accumulation.

The time‐course for the histamine‐induced production of [3H]‐inositol phosphates was approximately linear over 90 min of stimulation in both control and histamine pretreated slices. The rate of production in pretreated slices was significantly slowed compared to control, such that by 90 min of histamine stimulation the desensitized slices produced 2.8 times the basal [3H]‐inositol phosphate accumulation compared to 5.3 fold the basal [3H]‐inositol phosphate accumulation in the control slices.

Displacement of [3H]‐mepyramine binding to homogenates of guinea‐pig cerebral cortex by mepyramine and histamine revealed that histamine pretreatment did not alter the apparent affinity of the H1 receptor for histamine (control Kd = 6.3 ± 0.7 μm, desensitized Kd = 7.9 ± 1.6 μm) or mepyramine (control Kd = 3.4 ± 0.8 nm, desensitized Kd = 3.4 ± 1.3 nm), nor was there any reduction in the calculated maximum number of [3H]‐mepyramine binding sites (control Bmax = 192 ± 31 fmol mg−1 protein, desensitized Bmax = 220 ± 50 fmol mg−1 protein).

The histamine‐mediated desensitization of response in guinea‐pig slices was mediated by the H1 receptor subtype, since the attentuated maximum histamine‐stimulated [3H]‐inositol phosphate accumulation could not be prevented by inclusion of an H2‐ (ranitidine) and an H3‐ (thioperamide) receptor antagonist during the pretreatment period.

The desensitized histamine‐stimulated [3H]‐inositol phosphate accumulation recovered to 90% of control levels over a period of 150 min after the removal of the conditioning dose of histamine, with a half‐time of recovery of about 95 min.
Involvement of pertussis toxin‐sensitive and ‐insensitive mechanisms in α‐adrenoceptor modulation of noradrenaline release from rat sympathetic neurones in tissue culture
Authors: Hill C.E. , Powis D.A. , Hendry I.A.
Published: July 2012

Sympathetic neurones derived from superior cervical ganglia of neonatal rats and maintained in tissue culture were used to investigate the modulation of neurotransmitter release by presynaptic receptors. Three week old cultures of neurones were loaded with [3H]‐noradrenaline to label endogenous neurotransmitter stores. Release of noradrenaline was evoked by depolarization with raised extracellular K+ in the presence of desipramine and corticosterone to prevent uptake of released catecholamine.

Potassium (55 mmol 1−1) depolarization for 30 s caused more than a four fold increase in 3H overflow from basal levels but this increase was reduced by up to 40% in the presence of exogenous noradrenaline (1 μmol 1−1). The inhibition by noradrenaline of depolarization‐evoked overflow was blocked by the α1/α2‐adrenoceptor antagonist, phentolamine. Phentolamine alone did not increase K+‐evoked 3H overflow.

The α2‐adrenoceptor antagonist, yohimbine, produced a concentration‐dependent block of the inhibition by noradrenaline of K+‐evoked overflow, while the a1‐adrenoceptor antagonist, prazosin, was without effect at concentrations up to 0.1 μmol 1−1.

The β‐adrenoceptor antagonist, propranolol, neither reduced K+‐evoked overflow nor increased the degree of inhibition caused by the addition of 1 μmol 1−1 noradrenaline.

The α2‐adrenoceptor agonist, clonidine (1 μmol 1−1) was less effective than noradrenaline at inhibiting K+‐evoked overflow, while the α1‐adrenoceptor agonist, phenylephrine (1 μmol 1−1) had no significant effect.

The L‐channel calcium blocker, nicardipine (1 μmol 1−1) significantly inhibited 3H overflow evoked by K+. In the presence of L‐channel block, however, noradrenaline still inhibited residual evoked overflow.

In the presence or absence of nicardipine, pertussis toxin pretreatment (1 nmol 1−1) reduced, but did not prevent, the effect of noradrenaline (1 μmol 1−1). Pertussis toxin alone caused a significant enhancement of K+‐evoked 3H overflow.

The data indicate that on postganglionic neurones of cultured rat sympathetic ganglia there are α2‐adrenoceptors that modulate neurotransmitter release, but no functional β‐adrenoceptors that mediate an enhancement of transmitter release. The data suggest further that in this preparation the mechanism of α2‐adrenoceptor modulation may involve pertussis toxin sensitive and insensitive G‐proteins and effects on calcium channels other than L‐type.
Calcium antagonist and antiperoxidant properties of some hindered phenols
Authors: Sgaragli G.P. , Valoti M. , Gorelli B. , Fusi F. , Palmi M. , Mantovani P.
Published: July 2012

The calcium antagonist and antioxidant activities of certain synthetic and natural phenols, related to BHA (2‐t‐butyl‐4‐methoxyphenol), were evaluated in rat ileal longitudinal muscle and in lipid peroxidation models respectively.

Compounds with a phenol or a phenol derivative moiety, with the exception of 2,2′‐dihydroxy‐3,‐3′‐di‐t‐butyl‐5,5′‐dimethoxydiphenyl (di‐BHA), inhibited in a concentration‐dependent manner the BaCl2‐induced contraction of muscle incubated in a Ca2+‐free medium. Calculated pIC50 (m) values ranged between 3.32 (probucol) and 4.96 [3,5‐di‐t‐butyl‐4‐hydroxyanisole (di‐t‐BHA)], with intermediate activity shown by khellin < gossypol < quercetin < 3‐t‐butylanisole < BHA < nordihydroguaiaretic acid (NDGA) < 2,6‐di‐t‐butyl‐4‐methylphenol (BHT) and papaverine.

The Ca2+ channel activator Bay K 8644 overcame the inhibition sustained by nifedipine, BHA and BHT, while only partially reversing that of papaverine.

BHA, BHT, nifedipine and papaverine also inhibited in a concentration‐dependent fashion CaCl2 contractions of muscle depolarized by a K+‐rich medium. This inhibition appeared to be inversely affected by the Ca2+‐concentration used.

The inhibitory effects of nifedipine, papaverine, BHA and BHT were no longer present when muscle contraction was elicited in skinned fibres by 5 μm Ca2+ or 500 μm Ba2+, suggesting a plasmalemmal involvement of target sites in spasmolysis.

Comparative antioxidant capability was assessed in two peroxyl radical scavenging assay systems. These were based either on the oxidation of linoleic acid initiated by a heat labile azo compound or on lipid peroxidation of rat liver microsomes promoted by Fe2+ ions. Across both model systems, di‐t‐BHA, NDGA, BHT, di‐BHA, BHA and quercetin ranked as the most potent inhibitors of lipid oxidation, with calculated pIC50 (m) values ranging between 7.4 and 5.7.

Of the 32 compounds studied only 15 phenolic derivatives exhibited both antispasmogenic and antioxidant activity. Within this subgroup a linear and significant correlation was found between antispasmogenic activity and antioxidation. These bifunctional compounds were characterized by the presence of at least one hydroxyl group on the aromatic ring and a highly lipophilic area in the molecule.

Di‐t‐BHA is proposed as a lead reference compound for future synthesis of new antioxidants combining two potentially useful properties in the prevention of tissue damage after ischaemia‐reperfusion injury.
Involvement of cholecystokinin receptor types in pathways controlling oxytocin secretion
Authors: Luckman Simon M. , Hamamura Mitsuko , Antonijevic Irina , Dye Sandra , Leng Gareth
Published: July 2012

Intravenous administration of cholecystokinin (CCK) results in a transient activation of oxytocin neurones in the rat, and hence to oxytocin secretion: this activation is followed by expression of c‐fosmRNA and of Fos‐like immunoreactivity (Fos‐LI) in magnocellular oxytocin neurones. Fos‐like immunoreactivity is also induced in the regions of the brainstem that are thought to relay information from the periphery to the hypothalamus.

Administration of the selective CCKAreceptor antagonist MK‐329, but not the CCKBreceptor antagonist L‐365,260, prior to CCK injection, prevented oxytocin release as measured by radioimmunoassay and oxytocin neuronal activation as measured by electrophysiology and by the lack of induction ofc‐fosmRNA.

MK‐329 abolished the release of adrenocorticotrophic hormone (ACTH) following injection of CCK.

MK‐329 prevented the expression of Fos‐LI in the hypothalamic magnocellular nuclei and in the area postrema and dorsal vagal complex of the brainstem.

L‐365,260 had no effect on the expression of Fos‐LI in the brainstem, but attenuated that seen in the hypothalamic magnocellular nuclei.

We conclude that CCK acts on CCKAreceptors, either in the area postrema or on peripheral endings of the vagus nerve, to cause the release of hypothalamic oxytocin and ACTH. Information may be carried to the hypothalamus in part by CCK acting at CCKBreceptors.
Endothelin‐induced contraction and mediator release in human bronchus
Authors: Hay Douglas W.P. , Hubbard Walter C. , Undem Bradley J.
Published: July 2012

To elucidate the role of acetylcholine and various autacoids in endothelin‐1 (ET‐1)‐induced contraction in human bronchus, the effects of various receptor antagonists were examined. In addition, the ability of ET‐1 to stimulate the release of histamine, peptidoleukotrienes and prostanoids was determined.

ET‐1 was a potent and effective contractile agonist in human bronchus, possessing similar potency and efficacy to leukotriene D4 (LTD4); EC50 (−log m): ET‐1 = 7.76 ± 0.09, n = 7; LTD4 = 8.46 ± 0.53, n = 7; P > 0.2; maximum response (% 10 μm pre‐carbachol): ET‐1 = 103.8 ± 17.4, n = 7; LTD4 = 95.5 ± 9.3, n = 7; P > 0.6.

The cyclo‐oxygenase inhibitor, sodium meclofenamate (1 μm) or the potent and selective thromboxane receptor antagonist, SQ 29,548 (1 μm) were without significant effect on ET‐1 concentration‐response curves.

In the presence of sodium meclofenamate (1 μm), the muscarinic receptor antagonist, atropine (1 μm), the platelet activating factor (PAF) receptor antagonist, WEB 2086 (1 μm) or the combination of the H1‐histamine receptor antagonist, mepyramine (10 μm) and the leukotriene receptor antagonist, SK&F 104353 (10 μm), were without marked effect on ET‐1 concentration‐response curves. In addition, the combination of all four receptor antagonists did not antagonize ET‐1‐induced contraction.

ET‐1 (0.3 μm) did not stimulate the release of histamine or immunoreactive leukotrienes from human bronchus.

ET‐1 (0.3 μm) significantly stimulated the release of prostaglandin D2 (PGD2), 9α, 11β PGF2 (PGD2 metabolite), PGE2, 6‐keto PGF1α (PGI2 metabolite), PGF2α and thromboxane B2 (TxB2) a lower concentration, 10 nm, was without effect on prostanoid release. The production of PGD2 was increased 7.5 fold, whereas the release of the other prostanoids was stimulated only about 1.6 to 2.7 fold.

These data provide evidence that ET‐1 elicits contraction of human isolated bronchus predominantly via a direct mechanism with no significant involvement of the release of acetylcholine, leukotrienes, histamine or PAF. Although ET‐1 increased the release of several prostanoids they did not have a significant modulatory effect on the smooth muscle contraction.
The β‐adrenoceptors mediating relaxation of rat oesophageal muscularis mucosae are predominantly of the β3‐, but also of the β2‐subtype
Authors: de Boer Robert E.P. , Brouwer Frans , Zaagsma Johan
Published: July 2012

β‐Adrenoceptor‐mediated relaxation of rat oesophageal smooth muscle was investigated by studying the effects of β1‐ and β2‐selective antagonists on the relaxation induced by (−)‐isoprenaline, the β2‐selective agonists fenoterol and clenbuterol and the β3‐agonist, BRL 37344.

The highly β1‐selective antagonist CGP 20712A did not antagonize (−)‐isoprenaline‐ or BRL 37344‐induced relaxations in concentrations up to 10 μm. Only at 100 μm of CGP 20712A were clear rightward shifts of the agonist concentration‐response curves (CRCs) observed, with pA2 values of 4.70 and 4.97 against (−)‐isoprenaline and BRL 37344, respectively.

ICI 118,551, a potent and selective β2‐antagonist, at 100 nm caused moderate rightward shifts of the CRCs of (−)‐isoprenaline, fenoterol and clenbuterol; with fenoterol and clenbuterol, this was accompanied by a clear steepening of the curve. Only at the highest concentration (100 μm ICI 118,551) did the shifts to the right further increase substantially. Resulting Schild‐plots were clearly biphasic. BRL 37344‐induced relaxations were only antagonized at 100 μm ICI 118,551, yielding a pA2 value of 5.48.

These results clearly demonstrate that the BRL 37344‐induced relaxation of rat oesophageal muscularis mucosae is mediated solely through β3‐adrenoceptors, whereas (−)‐isoprenaline‐, fenoterol‐ and clenbuterol‐induced relaxations were shown to involve both β2‐ and, predominantly, β3‐adrenoceptors.
Mediation of the neuroprotective action of R‐phenylisopropyladenosine through a centrally located adenosine A1 receptor
Authors: MacGregor D.G. , Miller W.J. , Stone T.W.
Published: July 2012

Systemic injections of kainic acid, 10 mg kg−1, into adult rats resulted in lesions in the hippocampus, as assessed by peripheral benzodiazepine ligand binding. Co‐administration of clonazepam at 1 mg kg−1 or 0.2 mg kg−1 prevented major seizures associated with kainate injections, but did not alter significantly the production of hippocampal damage.

The co‐administration of the adenosine A1 agonist R‐phenylisopropyladenosine (R‐PIA, 25 μg kg−1, i.p.) abolished the lesions induced by kainic acid.

The presence of the selective A1 antagonist, 8‐cyclopentyl‐1,3‐dipropylxanthine (250 or 50 μg kg−1, i.p.) abolished the R‐PIA neuroprotective action.

The A1/A2 antagonist, 8‐(p‐sulphophenyl)theophylline (20 mg kg−1, i.p.) which cannot cross the blood brain barrier, did not alter significantly the neuroprotective action of R‐PIA, indicating that the neuroprotective action of the purine may be predominantly central.

The time course of the neuroprotection was also examined. R‐PIA was effective when administered 2 h before or after kainate administration.

The results emphasise the potential utility of systemically active adenosine A1 receptor ligands in reducing CNS gliosis induced by the activation of excitatory amino acid receptors.
Differential effects of B2 receptor antagonists upon bradykinin‐stimulated phospholipase C and D in guinea‐pig cultured tracheal smooth muscle
Authors: Pyne Susan , Pyne Nigel J.
Published: July 2012

Guinea‐pig tracheal smooth muscle cells were isolated and maintained in culture for 14–21 days prior to the study of the effect of a selective bradykinin B1 agonist and B2 antagonists upon bradykinin‐stimulated phospholipase C and D activities.

Bradykinin‐stimulated phospholipase C activity was determined by mass measurement of inositol (1,4,5)trisphosphate (Ins(1,4,5)P3) in unlabelled cells, whereas phospholipase D activity was assayed by the accumulation of [3H]‐phosphatidylbutanol ([3H]‐PtdBut) in [3H]‐palmitate‐labelled cells, which were stimulated in the presence of butan‐1‐ol (0.3%, v/v).

Bradykinin elicited the rapid and transient formation of Ins(1,4,5)P3, in a concentration‐dependent manner (log EC50 = −7.55 ± 0.1 m, n = 3). Bradykinin also rapidly activated the concentration‐dependent (log EC50 = −8.3 ± 0.4 m, n = 3) phospholipase D‐catalysed accumulation of [3H]‐PtdBut; the accumulation of [3H]‐PtdBut was sustained. These effects were not inhibited by pretreatment of the cells with indomethacin (1 μm).

The bradykinin B1 agonist, desArg9‐bradykinin (1 μm) was without effect upon phospholipase C or phospholipase D activity. Bradykinin‐stimulated (10 nm, EC40) Ins(1,4,5)P3 formation was inhibited by B2 receptor antagonists, d‐Arg‐[Hyp3,d‐Phe7]‐bradykinin (NPC 567) and d‐Arg‐[Hyp3,Thi5,8,d‐Phe7]‐bradykinin (NPC 349), with log IC50 values of −6.3 ± 0.5 m and −6.3 ± 0.4 m, respectively. However, bradykinin‐stimulated (10 nm, EC100) [3H]‐PtdBut accumulation was poorly inhibited and with low potency by each B2 receptor antagonist and bradykinin‐stimulated phospholipase D activity persisted at concentrations of antagonist that completely blocked bradykinin‐stimulated Ins(1,4,5)P3 formation (30 μm).

These observations suggest that the activation of phospholipase C by bradykinin may be mediated through a bradykinin B2 receptor population, whereas bradykinin‐stimulated phospholipase D may be activated via a distinct population of bradykinin receptors that do not appear to be either B1 or B2 receptor types, based upon pharmacological specificity. The mechanism of the activation of phospholipase D by bradykinin and the role of the putative B3 bradykinin receptor are discussed.
Antitussive effects of GABAB agonists in the cat and guinea‐pig
Authors: Bolser Donald C. , Aziz Sultan M. , DeGennaro Frances C. , Kreutner William , Egan Robert W. , Siegel Marvin I. , Chapman Richard W.
Published: July 2012

GABAB agonists inhibit neuronal processes which are important in the pathogenesis of airway disease, such as bronchospasm. Cough is a prominent symptom of pulmonary disease, but the effects of GABAB agonists on this airway reflex are unknown. Experiments were conducted to determine the antitussive effect of GABAB receptor agonists in comparison to the known antitussive agents, codeine and dextromethorphan.

Unanaesthetized guinea‐pigs were exposed to aerosols of 0.3 mm capsaicin to elicit coughing, which was detected with a microphone and counted. Cough also was produced in anaesthetized cats by mechanical stimulation of the intrathoracic trachea and was recorded from electromyograms of respiratory muscle activity.

In guinea‐pigs, the GABAB agonists baclofen and 3‐aminopropyl‐phosphinic acid (3‐APPi) produced dose‐dependent inhibition of capsaicin‐induced cough when administered by subcutaneous or inhaled routes. The potencies of baclofen and 3‐APPi compared favourably with codeine and dextromethorphan.

The GABAB antagonist, CGP 35348 (0.3– 30 mg kg−1, s.c.) inhibited the antitussive effect of baclofen (3.0 mg kg−1, s.c). However, CGP 35348 (10 mg kg−1, s.c.) had no effect on the antitussive activity of codeine (30 mg kg−1, s.c). The antitussive effect of baclofen was not influenced by the GABAA antagonist, bicuculline (3 mg kg−1, s.c.) or naloxone (0.3 mg kg−1, s.c).

In the cat, baclofen (0.3–3.0 mg kg−1, i.v.) decreased mechanically‐induced cough in a dose‐dependent manner. In this model, baclofen (ED50 = 0.63 mg kg−1) was less potent than either codeine or dextromethorphan. The antitussive effect of baclofen in the cat was antagonized by the GABAB antagonists, CGP 35348 (10 mg kg−1, i.v.) and 3‐aminopropylphosphonic acid (3 mg kg−1, i.v.).

We show that baclofen and 3‐APPi have antitussive effects in the guinea‐pig and cat and these effects are mediated by GABAB receptors.
Increases in intracellular calcium via activation of an endogenous P2‐purinoceptor in cultured CHO‐K1 cells
Authors: Iredale Philip A. , Hill Stephen J.
Published: July 2012

Increases in intracellular calcium ([Ca2+]i) were measured in Chinese hamster cultured ovary cells (clone, CHO‐K1), by use of the fluorescent, calcium‐sensitive dye, fura‐2.

Addition of both ATP and UTP elicited rapid increases in [Ca2+]i due to mobilization from intracellular stores and calcium entry across the plasma membrane.

Omission of calcium from the extracellular medium and pre‐incubation with the inorganic calcium channel blocker, nickel (Ni2+) prevented the calcium entry components of the responses.

Investigation of the concentration‐response relationships of various analogues of ATP suggests the presence of a purinoceptor which cannot be characterized as P2X or P2Y. In addition, there appears to be a sub‐population of P2Y‐purinoceptors which do not cross‐react with the ‘nucleotide’ receptor population.

Cross‐desensitization and additivity experiments suggest that both ATP and UTP activate the same receptor.

Pre‐incubation with the tumour‐promoting agent, β‐phorbol‐12,13 dibutyrate (PDBu), caused a reduction in the increases in [Ca2+]i, suggesting a role for protein kinase C in feedback inhibition of purinoceptor responses in this cell line.

In summary, we present evidence for the existence of an endogenous P2U‐purinoceptor (or ‘nucleotide receptor’) which is linked to increases in [Ca2+]i in CHO‐K1 cells.
Zeneca ZD7114 acts as an antagonist at β3‐adrenoceptors in rat isolated ileum
Authors: Growcott James W. , Holloway Brian , Green Michelle , Wilson Campbell
Published: July 2012

The relaxant effects of Zeneca ZD7114, BRL37344 (putative β3‐adrenoceptor agonists) and various phenylethylamine‐based agonists were studied in isolated ileum of the rat where tone was increased with carbachol (0.5 μm). Agonist‐induced relaxation was measured under equilibrium conditions with α‐, β1‐ and β2‐adrenoceptors inhibited.

Relaxant responses were obtained to isoprenaline, noradrenaline, and BRL37344, although, the efficacy of this latter agent was significantly lower than that of isoprenaline. Salbutamol caused weak relaxation (<20%) at high concentrations (10 μm) and ZD7114 was without significant relaxant effect even at high concentrations (10 μm).

Relaxant responses to isoprenaline and BRL37344 were weakly antagonized by high concentrations of (±)‐propranolol (10 and 100 μm) yielding pKB values of 5.7 with isoprenaline as the agonist and 5.5 with BRL37344 as the agonist.

The non‐selective β‐adrenoceptor antagonist, (±)‐alprenolol (1–100 μm) caused competitive antagonism of the relaxant responses to isoprenaline (pA2 value = 6.5). A similar pKB value was obtained when BRL37344 was used as the agonist (6.4).

Relaxant effects of isoprenaline and BRL37344 were also antagonized by ZD7114 (1–100 μm) yielding pA2 and pKB values of 6.3 and 6.7 respectively.

The low potencies of (±)‐propranolol and (±)‐alprenolol as antagonists of the relaxant responses to isoprenaline and BRL37344 indicate that both the agonists and antagonists employed in the current study may interact with β3‐adrenoceptors in the rat isolated ileum. Contrary to the previous findings in guinea‐pig ileum, where BRL37344 and ZD7114 were full agonists, in the current study, BRL37344 was a partial agonist and ZD7114 an antagonist at the β3‐adrenoceptor in rat ileum.
Inhibition by rapamycin of leukocyte migration and bronchial hyperreactivity induced by injection of Sephadex beads to guinea‐pigs
Authors: de Francischi Janetti Nogueira , Conroy Dolores M. , Maghni Karim , Sirois Pierre
Published: July 2012

The effect of rapamycin (0.001 to 5 mg kg−1) on the increased leukocyte counts in bronchoalveolar lavage (BAL) fluid and hyperreactivity of isolated bronchial strips to histamine and acetylcholine (ACh) was studied following the intravenous injection of Sephadex beads to guinea‐pigs.

The intramuscular (i.m.) injection of rapamycin (0.012 to 5 mg kg−1) dose‐dependently inhibited the increase in leukocyte counts in BAL fluid. Rapamycin (5 mg kg−l) reduced the numbers of eosinophils neutrophils, macrophages and lymphocytes in BAL fluid by 64, 55, 19 and 50% respectively. In addition, rapamycin (0.012 to 5 mg kg−1) significantly inhibited the Sephadex‐induced hyperreactivity of bronchial tissue to both histamine and ACh.

At a dose of 0.001 mg kg−1, rapamycin did not significantly reduce leukocyte infiltration or bronchial hyperreactivity.

Cyclosporin (5 mg kg−1) significantly reduced both lymphocyte and eosinophil numbers in BAL fluid of Sephadex‐injected guinea‐pigs whereas dexamethasone (1 mg kg−1) significantly reduced lymphocyte numbers. Neither drug affected the bronchial hyperreactivity to histamine and ACh.

It is concluded that the new immunosuppressive drug, rapamycin, is a potent inhibitor of leukocyte migration and bronchial hyperreactivity observed following the intravenous injection of Sephadex beads to guinea‐pigs. Rapamycin also appears to be more effective than cyclosporin or dexamethasone in reducing leukocyte counts and bronchial hyperreactivity in this model.

Our results suggest that inflammatory mechanisms which are inhibited by rapamycin may be important in the induction of Sephadex‐induced hyperreactivity.
Different muscarinic receptor subtypes mediating the phasic activity and basal tone of pig isolated intravesical ureter
Authors: Hernández M. , Simonsen U. , Prieto D. , Rivera L. , García P. , Ordaz E. , García‐Sacristán A.
Published: July 2012

We have studied the effects of muscarinic cholinoceptor agonists and specific antagonists on both phasic activity and basal tone of the isolated intravesical ureter of the pig by means of isometric techniques in vitro.

Acetylcholine in the presence and absence of physostigmine increased both phasic activity and basal tone of ureteral strips in a concentration‐dependent manner. Moreover carbachol, methacholine and oxotremorine‐M increased both contractile parameters while bethanechol and McN‐A‐343 evoked only increases in tone without affecting the frequency of the phasic contractions.

The nicotinic receptor blocker, hexamethonium (10−6–10−4 m), failed to modify the contractions evoked by a single dose of carbachol (10−5 m), whilst the muscarinic antagonist, atropine inhibited both phasic and tonic responses.

The muscarinic M1 (pirenzepine), M2 (AF‐DX 116 and methoctramine), M3 (4‐DAMP, HHSiD and p‐F‐HHSiD), and putative M4 receptor (tropicamide) antagonists significantly reversed increases in both frequency of phasic activity and baseline tone induced by a submaximal dose of carbachol (10−5 m). The pIC50 values for inhibition of the induced phasic activity were: atropine (10.16) > 4‐DAMP (9.12) > HHSiD (8.22) = methoctramine (7.98) = p‐F‐HHSiD (7.88 > tropicamide (7.62) = pirenzepine (7.53) = AF‐DX 116 (7.45) and for inhibition of basal tone were: atropine (10.73) > 4‐DAMP (9.32) > HHSiD (8.65) = pirenzepine (8.43) = p‐F‐HHSiD (8.38) > methoctramine (7.79) > tropicamide (7.53) > AF‐DX 116 (7.04).

The antagonist profile indicates that an M1 receptor mediates the tonic response while the phasic activity could involve either both M2 and M3 or an M4 muscarinic receptor. These results suggest that different muscarinic receptor subtypes mediate the phasic and tonic contractile activity induced by a submaximal concentration of carbachol in the porcine intravesical ureter.
Dual contractile effects of ATP released by field stimulation revealed by effects of α,β‐methylene ATP and suramin in rat tail artery
Authors: Bao Jian‐Xin , Stjärne Lennart
Published: July 2012

The field stimulation‐induced release of endogenous ATP and noradrenaline (NA) and contractile response in rat isolated tail artery were examined. The release of ATP was studied by extracellular electrophysiological recording and that of NA by a novel voltammetrical technique. The effects of the P2‐purinceptor antagonist, suramin, on these parameters were compared with those of α,β‐methylene ATP, a P2X‐purinoceptor desensitizing agent.

Neither α,β‐methylene ATP (10 μm) nor suramin (100−500 μm) had significant effects on the extracellularly recorded nerve terminal action potential but both abolished the ATP‐induced excitatory junction current caused by stimulation at 0.1 Hz. Neither agent affected significantly the voltammetrically measured release of NA induced by 10 or 100 pulses at 20 Hz.

Combined blockade of both postjunctional α1‐ and α2‐adrenoceptors by prazosin and yohimbine (both 0.1 μm) profoundly depressed the contractile response to 10 pulses at 20 Hz. The small and fast residual contraction in the presence of these agents was abolished by α,β‐methylene ATP (10 μm) and inhibited by suramin in a concentration‐dependent manner (10–500 μm; IC50 75 μm) and was hence probably caused by ATP or a related nucleotide.

When added first, α,β‐methylene ATP (10 μm) or suramin (100–500 μm) delayed the onset and enhanced the amplitude of the neurogenic contraction. This enhanced response was abolished by further addition of prazosin and yohimbine (both 0.1 μm).

The K+ channel blocker, tetraethylammonium (10 mm), dramatically enhanced the contractile response to 100 pulses at 1 Hz and caused it to become diphasic. Addition of α,β‐methylene ATP (10 μm) or suramin (100–500 μm) abolished the large initial twitch component of this contraction and depressed the tonic phase.

Like α,β‐methylene ATP, suramin (500 μm) had no effect on the contraction caused by exogenous NA (10 nm − 100 μm) or KCl (60 mm); both agents almost abolished the contraction caused by ATP (100 μm).

In conclusion, (i) the contractile response of rat tail artery to electrical field stimulation is mediated by both ATP and NA, and is thus an expression of ATP‐NA co‐transmission, (ii) the released ATP exerts two opposite effects via ‘P2X‐like’ purinoceptors, triggering the initial rapid phase of the neurogenic contraction and restricting the NA‐mediated component of the contraction; and (iii) the source and possible physiological role of the ATP which causes the inhibitory effect are unknown at present.
Mechanism of rat uterine smooth muscle contraction induced by endothelin‐1
Authors: Tsunoda Hajime , Miyauchi Takashi , Fujita Kayo , Kubo Takeshi , Goto Katsutoshi
Published: July 2012

Endothelin (ET)‐1 has been demonstrated to cause contraction of uterine smooth muscle. We investigated the role of ET receptor subtypes (ETA and ETB receptors) in ET‐1‐induced contraction of rat uterine smooth muscle by using the ETA receptor antagonist BQ‐123 and the ETB receptor agonist BQ‐3020.

ET‐1 caused a contraction with superimposed oscillations of the rat isolated uterus suspended in Krebs‐Ringer solution; both the amplitude of contraction as well as the oscillation frequency increased in a dose‐dependent manner (10−11–10−7 m).

BQ‐123 (10−6 m) markedly shifted the dose‐response curve of ET‐1 for both contractile effects and oscillation frequency to the right.

BQ‐3020 (10−11–3 × 10−7 m) did not cause uterine contraction; neither did it affect the dose‐response curve of ET‐1 for either the contractile effect or the increase in oscillation frequency. Thus, stimulation of ETB receptors is not involved in these responses.

The present findings suggest that ET‐1‐induced contractile responses and the increase in oscillation frequency in rat uterine smooth muscle is mediated through ETA receptors, and that ETB receptors, play no role in these responses.