bluuebunny/crossref_metadata_2025 huggingface dataset:
Regulation of the substance P‐induced contraction via the release of acetylcholine and γ‐aminobutyric acid in the guinea‐pig urinary bladder
Authors: Shirakawa Junji , Nakanishi Tateo , Taniyama Kohtaro , Kamidono Sadao , Tanaka Chikako
Published: July 2012

The action of substance P (SP) on the release of γ‐aminobutyric acid (GABA) and acetylcholine (ACh) and on contraction were studied in strips of the guinea‐pig urinary bladder. Substance P induced a dose‐dependent contraction of strips of guinea‐pig urinary bladder (EC50 =1.2 × 10−9m). This contraction was not altered by tetrodotoxin, but with a dose of 10−9m and less, there was a complete inhibition by 10−6m atropine. Contractions initiated by 3 × 10−9m SP or more were partly inhibited by atropine. The EC50 value of substance P in the presence of atropine was 7.0 × 10−9m.

Substance P induced a Ca2+‐dependent and tetrodotoxin‐resistant release of [3H]‐acetylcholine (ACh) from strips of urinary bladder preloaded with [3H]‐choline (EC50 = 4.9 × 10−10m), and this release was antagonized by [D‐Pro2,D‐Trp7′9]substance P.

Bicuculline increased the substance P‐induced contraction and the release of [3H]‐ACh from the strips.

Substance P induced a Ca2+‐dependent and tetrodotoxin‐sensitive release of [3H]‐γ‐aminobutyric acid (GABA) from strips preloaded with [3H]‐GABA (EC50 = 2.6 × 10−9m), and this release was antagonized by [D‐Pro2,D‐Trp7′9]substance P.

Therefore, substance P appears to exert excitatory effects on the contractility of urinary bladder predominantly by stimulating its own receptor located on the cholinergic nerve terminals. GABA released by substance P inhibits stimulation of the cholinergic neurone. However, the direct action of substance P on the cholinergic neurone is more potent that the indirect action via GABA release.
A comparison of haemoglobin and erythrocytes as inhibitors of smooth muscle relaxation by the NANC transmitter in the BRP and rat anococcygeus and by EDRF in the rabbit aortic strip
Authors: Gillespie John S. , Sheng Hong
Published: July 2012

The inhibitory effect of erythrocyte suspensions and haemoglobin solutions on the response of the bovine retractor penis muscle (BRP) and the rat anococcygeus to field stimulation of their non‐adrenergic non‐cholinergic (NANC) nerves has been compared. Haemoglobin 3 μm greatly reduced the relaxant response in both tissues whereas a haemoglobin‐equivalent suspension of erythrocytes was without effect.

A similar comparison of erythrocytes and haemoglobin on the response of the rabbit aortic strip to EDRF liberated by acetylcholine (ACh) showed that both reduced EDRF‐mediated relaxation, though haemoglobin was significantly more effective.

These results suggest that the NANC transmitter may not be as freely diffusible through the erythrocyte membrane as EDRF and may therefore not be nitric oxide.
The key role of sodium in the ouabain‐mediated potentiation of potassium‐evoked catecholamine release in cat adrenal glands
Authors: Abajo Francisco J. , Castro Maria Antonia S. , Sánchez‐García Pedro
Published: July 2012

The effect of [Na]o on the catecholamine release evoked by K in ouabain pretreated, isolated adrenal glands of the cat, was investigated.

Reduction of [Na]o to 70, 50 and 25 mM, with sucrose as a substitute, did not modify the spontaneous catecholamine release but progressively increased the K (17.7 mm)‐evoked secretory response.

Ouabain pretreatment (100 μm; 10 min) greatly increased the K (17.7 mm)‐evoked catecholamine secretory response in glands perfused with normal Krebs. Such an increase was still seen in glands perfused with 70 mM Na‐containing solution but disappeared when [Na]o was reduced to 25 and 10 mM.

Preperfusion of non‐ouabain treated glands with Li‐enriched Krebs, for a 40 min period, caused an increase in the K (17.7 mm)‐evoked secretory response which was dependent on [Li]o and essentially similar to that induced by ouabain, pretreatment.

Ouabain treatment (100 μm; 10 min) of glands perfused with normal Krebs evoked a long lasting catecholamine release, which reached a plateau at about 36 min and amounted to 0.68 ± 0.25 μg 2 min−1 (n = 9). Such a secretory response was dramatically increased, and its shape modified, when glands were preperfused with K (17.7 mm)‐Krebs: a peak of 3.77 ± 0.42 μg 2 min−1 (n = 7) was reached at 18 min. This response was drastically reduced in the presence of nitrendipine (1μm).

In summary, our results indicate that both [Na]o reduction or Na accumulation into the chromaffin cell by inhibition of the Na pump with ouabain, greatly enhance the secretory efficacy of small increments of [K]o, and suggest that sodium may play a role in the regulation of catecholamine release mediated by voltage‐dependent Ca channels in adrenal glands.
The interaction of hexamethonium with muscarinic receptor subtypes in vitro
Authors: Eglen R.M. , Michel A.D. , Cornett C.M. , Kunysz E.A. , Whiting R.L.
Published: July 2012

The action of hexamethonium has been studied at a range of muscarinic receptors in vitro by use of both functional and radioligand binding studies.

In functional studies, hexamethonium exhibited little or no significant (P < 0.05) antagonism of contractile responses to carbachol at muscarinic receptors in the guinea‐pig ileum, oesophageal muscularis mucosae, urinary bladder and trachea. However, antagonism was observed at muscarinic receptors in the guinea‐pig left atria mediating negative inotropic responses and the calculated pKB value was 3.80. Hexamethonium also antagonized contractile responses to carbachol in the canine saphenous vein. The pKB value at these receptors was 3.75.

In the presence of 3.2 mM hexamethonium, the pA2 value for methoctramine at atrial muscarinic receptors was reduced by approximately 10 fold (control pA2 value was 7.81 ± 0.05; pA2 value in hexamethonium was 6.73 ± 0.04). In contrast at tracheal muscarinic receptors, the pA2 values for methoctramine were unaffected in the presence of 3.2 mM hexamethonium (control pA2 = 5.58 ± 0.07; pA2 value in hexamethonium was 5.63 ± 0.12). All values quoted are mean ± s.e.mean, n = 8.

In competition radioligand binding studies, hexamethonium exhibited a higher affinity for cardiac M2 receptors (pKi = 3.68) than for cerebrocortical M1 receptors (pKi = 3.28) or for submaxillary gland M3 receptors (pKi = 2.61). At M2 receptors hexamethonium at concentrations of 0.1–10 mM, increased the half life of the dissociation rate of [3H]‐N‐methylscopolamine 1.6‐4.3 fold. This was observed at M3 receptors only at 10 mM, when the half life was increased 1.7 fold.

We conclude that hexamethonium, in addition to its well characterized nicotinic antagonist properties, can act as a weak muscarinic antagonist and differentiates between cardiac M2 receptors and glandular/smooth muscle M3 receptors. However, hexamethonium differentiates less clearly between M1 and M2 receptors. The selectivity between M2 and M3 receptors observed in the present study with hexamethonium is comparable to other M2 selective antagonists such as AF‐DX 116 and himbacine.

Caution should be exercised with regard to the inclusion of hexamethonium in functional studies of M2 muscarinic receptor subtypes at concentrations of 0.1 mM and above.
Metabolism of amines in the isolated perfused mesenteric arterial bed of the rat
Authors: Elliott Jonathan , Callingham Brian A. , Sharman Dennis F.
Published: July 2012

Semicarbazide‐sensitive amine oxidase (SSAO) activity has been demonstrated in the isolated mesenteric arterial bed of the rat in vitro by studying the metabolism of benzylamine (Bz) and tyramine (Tyr) added to the perfusing fluid.

Pretreatment of rats with (E)‐2‐(3′,4′‐dimethoxyphenyl)‐3‐fluoroallylamine (MDL72145), a potent inhibitor of SSAO in rat mesenteric blood vessels, reduced the amount of metabolites, following the addition of Bz (25 μm) or Tyr (100 μm) to the perfusing fluid, by 83% and 52% respectively. Inactivation of monoamine oxidase type A (MAO‐A) by the addition of clorgyline (10 μm) to the perfusing fluid, had little effect on the appearance of metabolites from Tyr.

The presence of 3 μm cocaine in the perfusing fluid increased the amount of metabolites produced from Tyr.

The metabolites of Tyr appearing in the perfusion fluid from control preparations were 85% p‐hydroxyphenylacetic acid and the remainder consisted of a mixture of p‐hydroxyphenylacetaldehyde and, possibly, p‐hydroxyphenylethanol.

The metabolism of Tyr by homogenates of the rat mesenteric vascular bed was carried out by SSAO (60%) and MAO‐A (40%) with very little contribution from MAO‐B. Homogenates from rats pretreated with MDL 72145 showed metabolism of Tyr by MAO‐A only.

These data indicate that SSAO is capable of metabolizing amines present in the fluid perfusing blood vessels to metabolites that are readily released. Histochemical evidence has shown that whereas MAO‐A is present in the mitochondria of smooth muscle cells and nerve endings, SSAO is located in the plasma membrane of the smooth muscle cells. This subcellular distribution may explain the differences found between metabolites released from intact vessels and the metabolism seen in homogenates. The identity of the Tyr metabolizing activity in intact vessels that is resistant to both MDL 72145 and clorgyline remains to be determined.
Spinal antinociceptive actions and naloxone reversibility of intravenous μ‐ and κ‐opioids in spinalized rats: potency mismatch with values reported for spinal administration
Authors: Parsons Chris G. , West David C. , Headley P. Max
Published: July 2012

The relative spinal effectiveness of μ‐ and κ‐opioids has been assessed by their intravenous potencies on nociceptive responses (heat and/or pinch) of single motoneurones recorded in α‐chloralose anaesthetized, spinalized rats.

The depressant actions of both μ‐ and κ‐opioids were reversed by low intravenous doses of naloxone (10 to 100 μg kg−1). When tested at a dose of 1 μg kg−1 i.V., naloxone antagonized the effects of the μ‐agonist morphine but had no effect on the κ‐opioid U‐50,488. This provides further support for the theory that the actions of μ‐ and κ‐ligands were mediated at different subclasses of opioid receptor but highlights the difficulties in using antagonists with poor receptor selectivity to differentiate between μ‐ and κ‐receptor‐mediated effects in vivo.

The molar potency ratios of fentanyl: morphine: U‐50,488: tifluadom for thermal and mechanical nociceptive responses were 620:1.0:0.74:5.7 and 520:1.0:0.56:7.7 respectively. These potency ratios, as well as the absolute potencies, agree well with those reported in several behavioural studies in which systemic administration of agonists was used in non‐thermal tests.

The agonist potency values obtained in this study contrast with those reported for local spinal administration. By this route, the potency of lipophilic opioids (e.g. fentanyl, U‐50,488 and tifluadom) relative to hydrophilic opioids (e.g. morphine) is much reduced, implying that activity of intrathecally administered opioids is more dependent on the physico‐chemical properties of the agonists used than on the relative abundance in the spinal cord of functional opioid receptors of the μ‐ and κ‐subtypes. This conclusion indicates that the results with locally applied opioids should not be used to assess spinal opioid receptor function.
On the selectivity of intravenous μ‐ and κ‐opioids between nociceptive and non‐nociceptive reflexes in the spinalized rat
Authors: Parsons Chris G. , Headley P. Max
Published: July 2012

In electrophysiological experiments in spinalized, α‐chloralose anaesthetized rats, opioids and anaesthetics were tested intravenously (i.v.) on the responses of individual motoneurones to alternating noxious (heat or pinch) and non‐noxious (tap or vibration) stimuli.

On cells that were sensitive to low doses of μ‐opioids, both fentanyl (0.5–4 μg kg−1 i.v.) and morphine (0.5 mg kg−1 i.v.) selectivity reduced reflexes to noxious stimuli to a greater degree than reflexes to non‐noxious stimuli. In contrast, on cells that were relatively resistant to these agonists, the higher doses required to reduce nociceptive reflexes (fentanyl 8 μg kg−1 i.v.; morphine 1–8 mg kg−1 i.v.) depressed non‐nociceptive reflexes to a similar degree.

A similar spectrum of selectivity was seen with U‐50,488 (0.5–16 mg kg−1 i.v.) although statistically significant selective depression of reflexes was only evident at the lowest dose tested (0.5 mg kg−1 i.v.). All effects of U‐50,488 were readily reversed by low doses of the opioid antagonist, naloxone (10–100 μg kg−1 i.v.).

The dissociative anaesthetic/PCP ligand ketamine (0.5–4 mg kg−1 i.v.) was similar in having selective actions at low doses on sensitive cells but non‐selective actions when higher doses were required. In contrast, the general anaesthetics methohexitone (4 mg kg−1 i.v.) and alphadolone/alphaxalone (1 mg kg−1 i.v.) were consistently non‐selective between reflexes to noxious and non‐noxious stimuli. α‐Chloralose (20–40 mg kg−1 i.v.) had very little effect on reflexes to any of the synaptic inputs tested.

The sensitivity of nociceptive reflexes to depression by opioids and dissociative anaesthetics thus determines the selectivity of these agents between nociceptive and non‐nociceptive reflexes. The same is not true for the general anaesthetics tested.

Taken together with our previous findings that κ‐ as well as μ‐agonists reduce thermal and mechanical nociceptive reflexes in parallel, these results indicate a general similarity of spinal μ‐ and κ‐opioid actions in the spinal cord of the rat.
Simultaneous measurement of endothelium‐derived relaxing factor by bioassay and guanylate cyclase stimulation
Authors: Kondo Kigen , Mitchell Jane Allison , de Nucci Gilberto , Vane John Robert
Published: July 2012

Endothelium‐derived relaxing factor (EDRF) released by cultured endothelial cells (EC) from bovine aortae was measured by bioassay using pre‐contracted strips of rabbit aorta and by radioimmunoassay of guanosine 3′:5′‐cyclic monophosphate (cyclic GMP) produced by stimulation of bovine lung soluble guanylate cyclase.

Bradykinin (Bk, 3 and 30 pmol) injected through a column of EC caused release of EDRF as detected by bioassay and increased cyclic GMP concentrations. Superoxide dismutase (SOD, 15 u ml−1) increased the amount of EDRF detected by the activation of soluble guanylate cyclase.

In the absence of endothelial cells, nitric oxide (NO, 1–2 μm), arachidonic acid (AA, 3–30 μm) or sodium nitröprusside (SNP, 1–100 μm) stimulated guanylate cyclase. Superoxide dismutase strongly increased the stimulation of guanylate cyclase induced by NO, but had little effect on the stimulation induced by SNP and no effect on the stimulation induced by AA.

Oxyhaemoglobin (10–300 μm) abolished the stimulation of guanylate cyclase by EDRF, NO or SNP but was much less effective as an inhibitor of AA‐induced stimulation of guanylate cyclase.

These results demonstrate that measurement of guanylate cyclase stimulation by radioimmunoassay is a viable method for detecting EDRF release, especially useful when the drugs used interfere with bioassay tissues.
Endothelium‐dependent calcium‐induced relaxation in the presence of Ca2+ ‐antagonists in canine depolarized coronary arteries
Authors: Kikkawa Kohei , Murata Sakae , Nagao Taku
Published: July 2012

We examined the mechanisms underlying Ca2+‐induced relaxation in the presence of clentiazem, a new Ca2+‐antagonist, in depolarized coronary arteries of the dog.

Ca2+ (3 × 10−5−3 × 10−3m) caused an unexpected relaxation in the presence of a high concentration of clentiazem (10−6m) in coronary, but not in mesenteric or renal arteries.

The Ca2+‐induced relaxation was also observed in the presence of established Ca2+‐antagonists such as diltiazem (3 × 10−6m), nifedipine (3 × 10−8m) and verapamil (3 × 10−6m).

The Ca2+‐induced relaxation was inhibited by removal of the endothelium, treatment with oxyhaemoglobin (1.5 × 10−6m) or methylene blue (10−5m), but not by treatment with indomethacin (5 × 10−6m).

The Ca2+‐induced relaxation was observed in an endothelium‐denuded coronary artery segment when closely apposed to an endothelium‐containing segment of coronary or mesenteric artery.

These results suggest that Ca2+‐induced relaxation in the presence of high concentrations of Ca2+‐antagonists is mediated through endothelium‐derived relaxing factor (EDRF). In addition, Ca2+‐antagonists do not affect the Ca2+‐influx necessary for the release and/or synthesis of EDRF.
The effects of L‐arginine and NG‐monomethyl L‐arginine on the response of the rat anococcygeus muscle to NANC nerve stimulation
Authors: Gillespie John S. , Liu Xiaorong , Martin William
Published: July 2012

The effect of the competitive inhibitor of L‐arginine, NG‐monomethyl L‐arginine (L‐NMMA) on the response of the rat anococcygeus muscle to non‐adrenergic, non‐cholinergic (NANC) inhibitory nerve stimulation has been examined. L‐NMMA causes a rise in muscle tone and inhibition of the response to nerve stimulation. The stereoisomer D‐NMMA is without effect. The rise in tone and inhibition of the nerve response is reversed by L‐arginine. Another analogue, L‐canavanine, which is effective against L‐arginine utilization in the macrophage, was without effect on the rat anococcygeus. These results provide indirect evidence for nitric oxide (NO) or a substance releasing NO as the transmitter of the NANC nerves in this tissue.
5‐HT1D binding sites in porcine brain can be sub‐divided by GR43175
Authors: Sumner M.J. , Humphrey P.P.A.
Published: July 2012

We have examined the binding of 5‐carboxamidotryptamine (5‐CT) and GR43175 (3‐(2‐dimethylamino)ethyl‐N‐methyl‐1H‐indole‐5‐methane sulphonamide) to 5‐HT1D sites labelled with [3H]‐5‐hydroxytryptamine ([3H]‐5‐HT) in neonatal porcine caudate membranes. In competition studies, 5‐CT produced shallow inhibition curves (Ki 138 nM, slope 0.31), indicating binding site heterogeneity, while GR43175 interacted with a single population of binding sites (Ki 251 nM, slope 0.98), producing a maximum of only 52% inhibition of [3H]‐5‐HT binding compared to 100% for 5‐HT or 5‐CT. In the presence of excess GR43175 (10 μm), 5‐CT produced a monophasic inhibition curve with a Ki value of 800 nM for the remaining sites (slope 0.89). These preliminary data suggest that under the conditions employed, GR43175, and to a lesser extent 5‐CT, may discriminate between two sub‐populations of 5‐HT1D binding sites in porcine brain.
Mechanisms underlying the antiarrhythmic properties of β‐adrenoceptor blockade against ischaemia‐induced arrhythmias in acutely prepared rats
Authors: Paletta M.J. , Abraham S. , Beatch G.N. , Walker M.J.A.
Published: July 2012

The mechanism underlying the limited antiarrhythmic effects of β‐adrenoceptor blocking agents against occlusion‐induced arrhythmias in acutely prepared, pentobarbitone‐anaesthetized rats has been investigated.

Atenolol, ICI 111,581 and propranolol were given at low, medium and high doses calculated to shift dose‐response curves to exogenous agonists by factors of 10–30, 100–300 and 1000–3000, respectively.

Arrhythmias, blood pressure, heart rate, ECG changes and serum K+ were measured.

Antiarrhythmic activity was seen with β‐blocker treatment. This was minimal with atenolol (0.1, 1 and l0 mg kg−1) and only statistically significant with the highest dose of ICI 111,581 (5 mg kg−1), and propranolol (10 mg kg−1).

Treatment with β‐adrenoceptor blockers elevated serum potassium concentrations, as compared with saline controls, especially when measured at 30 min post‐occlusion.

Only ICI 111,581 (5 mg kg−1) and propranolol (1 and 10 mg kg−1) prolonged P‐R interval.

In order to evaluate possible mechanisms of antiarrhythmic action, attempts were made to correlate antiarrhythmic activity with β‐blockade, serum potassium concentrations, and/or with changes in the P‐R interval of the ECG.

Reductions in arrhythmias did not correlate well with presumed β‐blockade. Better correlation was obtained with elevations of serum potassium concentration, and with prolongation of P‐R interval (a presumed Class I antiarrhythmic action).

These results suggested that antiarrhythmic effects of adrenoceptor blocking agents in acutely‐prepared anaesthetized rats, subjected to occlusion of a coronary artery, are unrelated to cardiac β‐blockade. The limited antiarrhythmic effects which were observed could be attributed to elevations in serum potassium concentration (due to peripheral β‐blockade) and/or possible Class I antiarrhythmic actions.
Nicotinic excitation of rat ventral tegmental neurones in vitro studied by intracellular recording
Authors: Calabresi P. , Lacey M.G. , North R.A.
Published: July 2012

Intracellular recordings were made from presumed dopamine‐containing neurones in the ventral tegmental area (VTA) in rat brain slices.

Nicotine (10–100 μm) and acetylcholine (ACh) depolarized the neurones. The depolarization caused by ACh was typically biphasic; both components were increased by neostigmine (0.1–10 μm), but only the slower component was blocked by scopolamine (1–10 μm).

The nicotinic action of ACh, studied in the presence of neostigmine and scopolamine, persisted in the presence of tetrodotoxin (1 μm) and cobalt (2–5 mM).

ACh or carbachol (30 μm) caused inward currents in neurones voltage‐clamped near the resting potential. These currents reversed polarity at around −4mV, were blocked by hexamethonium (1–100 μm) in a voltage‐dependent manner, and showed desensitization with prolonged or repeated agonist applications.

Depolarizations caused by ACh and carbachol were reduced in slices pretreated with κ‐bungarotoxin, but were not changed by α‐bungarotoxin.

These responses to ACh and nicotine resemble those previously described on autonomic ganglion cells. The direct action on VTA neurones may contribute to the positive reinforcement associated with nicotine consumption.
Actions of anaesthetics and avermectin on GABAA chloride channels in mammalian dorsal root ganglion neurones
Authors: Robertson Brian
Published: July 2012

The γ‐aminobutyric acid (GABA)‐mimetic actions of some anaesthetics and the antehelminthic avermectin B1a were examined on freshly isolated mammalian dorsal root ganglion (DRG) neurones by use of suction electrodes and a single electrode voltage clamp.

Pentobarbitone (60 μm‐3 mM), chloralose (600 μm‐1 mM), etomidate (10–100 μm), alphaxalone (10–60 μm) and avermectin (10–60 μm) directly activated chloride channels in GABA‐sensitive DRG neurones. The agonist action was sensitive to block by bicuculline and picrotoxinin.

Steady‐state current‐voltage (I‐V) curves for the anaesthetics were either linear, or rectified in the opposite direction to steady‐state I‐V curves obtained with GABA. Current relaxations in response to voltage jumps were also of the opposite direction. An extra surge of current (‘bounce’) was commonly observed on washout of some of these agonists.

Pentobarbitone was ineffective as an agonist at alkali pH (10.4 and 9.4), but was approximately twice as effective at acid (5.4) than at normal (7.4) pH values.

These results suggest that some anaesthetics and avermectin are capable of ‘blocking’ GABA channels in addition to activating them.
Cardiac electrophysiological actions of captopril: lack of direct antiarrhythmic effects
Authors: Hemsworth Paul D. , Pallandi Regan T. , Campbell Terence J.
Published: July 2012

Standard microelectrode techniques were used to study the effects of captopril (1, 10 and 100 μm) on action potentials recorded from guinea‐pig ventricular cells and sinoatrial node cells.

Captopril had no effect on the maximum rate of depolarization () of ventricular action potentials in cells exposed to either normal Locke solution or ‘simulated ischaemic’ solution (K+ = 11.2 mM; pH = 6.4; Po2 < 80 mmHg), nor was there any augmentation of the normal small decline in with increasing stimulation rate (range of interstimulus intervals = 2400 ms to 300 ms).

Captopril had no effect on the duration of ventricular action potentials, nor did it alter the shortening seen on exposure to simulated ischaemia.

Captopril did not alter spontaneous sinus cycle length or any recorded parameter of sinus node action potentials.

It is concluded that any antiarrhythmic effects observed during clinical use of captopril are most unlikely to be due to direct actions of the drug on cardiac cell membrane properties.
Accumulation of extracellular calcium at the endplate of mouse diaphragm after ecothiopate in vitro
Authors: Burd P.F. , Ferry C.B. , Smith J.W.
Published: July 2012

Experiments were carried out to investigate the accumulation from the extracellular medium of 45Ca2+ by the endplate region of skeletal muscle.

Mouse diaphragm muscle was incubated in physiological saline labelled with 45Ca at 37°C for periods of up to 1.5 h.

The muscle was divided into junctional and non‐junctional portions and the Ca from the extracellular fluid accumulated at the endplate determined from the 45Ca content of the portions.

The accumulation of extracellular Ca at the endplate region of muscles incubated in physiological saline alone was nil, but there was accumulation in the presence of the anticholinesterase ecothiopate iodide 0.5 × 10−6 m (ECO). Stimulation of the phrenic nerve at 0.02 Hz caused no further increase in accumulation, but reduced the amount of spontaneous fasciculation. In tetrodotoxin (TTX) 10−6 m, the accumulation was halved, and in 3.5 mM Mg2+ the accumulation was nil. Carbachol 10−4 m resulted in an accumulation of Ca similar to that in ECO.

It is concluded that there was an accumulation of extracellular Ca following excitation of the nerve by stimulation at a low frequency and during the spontaneous fasciculations, and about half of the accumulation of extracellular Ca after ECO in the experiments was due to the postsynaptic action of ACh released non‐quantally from the nerve terminals.
Inotropic response to DPI 201‐106 in the failing human heart
Authors: Böhm Michael , Diet Frank , Kemkes Bernhard , Wankerl Michael , Erdmann Erland
Published: July 2012

The present study was designed to characterize the positive inotropic response to DPI 201‐106 in isolated papillary muscle strips obtained from heart failure patients undergoing surgery.

The positive inotropic responses to isoprenaline and milrinone and cardiac β‐adrenoceptor density were also determined.

DPI 201‐106 increased the force of contraction in papillary muscle strips from patients with moderate (NYHA II‐III) and severe (NYHA IV) heart failure, in a concentration‐dependent manner. This positive inotropic effect was more pronounced in tissues from NYHA IV patients. Furthermore, these responses were greater than those produced by milrinone or isoprenaline. The positive inotropic effects of isoprenaline and milrinone were reduced in NYHA IV compared to NYHA II‐III. Consistently, there was also a smaller density of β‐adrenoceptors in myocardium from NYHA IV than in NYHA II‐III. The positive inotropic effect of Ca2+ was similar in tissues from both groups of patients.

The positive inotropic effect of DPI 201‐106 was not antagonized by adenosine or carbachol, whereas both compounds reduced the positive inotropic effect of isoprenaline.

DPI 201‐106 did not increase the Ca2+‐sensitivity of chemically skinned ventricular fibres, whereas a significant increase of the Ca2+‐sensitivity was obtained with trifluoperazine.

It is concluded that DPI 201‐106 produces significant positive inotropic effects in tissue excised from the failing human heart. The lack of inhibition by adenosine and carbachol might contribute to its greater effectiveness in NYHA IV than NYHA II‐III and indicates that its mechanism of action is cyclic AMP‐independent. A sensitization of the contractile proteins to Ca2+ does not appear to be important for the positive inotropic action of DPI 201‐106 in the failing human heart.
Effect of hydrogen peroxide on guinea‐pig tracheal smooth muscle in vitro: role of cyclo‐oxygenase and airway epithelium
Authors: Rhoden K.J. , Barnes P.J.
Published: July 2012

Hydrogen peroxide (H2O2) (0.1 μm‐3 mM) induced variable contractions of guinea‐pig isolated trachea which were attenuated by catalase (100 u ml−1) and mannitol (15 mM) suggesting that contractions were induced by H2O2 and/or the hydroxyl anion.

Epithelial removal potentiated contractile responses of tracheal preparations to H2O2 with a leftward shift of the concentration‐response curve and an increase in the maximal response.

Indomethacin (3 μm) inhibited contractions to H2O2 of intact preparations and preparations without epithelium suggesting that contractions may be mediated by cyclo‐oxygenase products. Intact preparations (but not preparations without epithelium) contracted in response to high concentrations (>0.1 mM) of H2O2 in the presence of indomethacin suggesting that other excitatory factor(s) released by the epithelium may induce contraction.

Preincubation of intact tracheal preparations with H2O2 (1 mM) for 1 h had no effect on responses to histamine or isoprenaline.

These results suggest that hydrogen peroxide generated during the inflammatory process may play a role in bronchoconstriction.
Effects of aerosolised substance P on lung resistance in guinea‐pigs: a comparison between inhibition of neutral endopeptidase and angiotensin‐converting enzyme
Authors: Lötvall Jan O. , Skoogh Bengt‐Eric , Barnes Peter J. , Chung K. Fan
Published: July 2012

We have examined in guinea‐pigs, in vivo, the effects of inhibition of neutral endopeptidase (NEP) and angiotensin‐converting enzyme (ACE) on the airway response to aerosolised substance P (SP). We aerosolised captopril (4.6 mm, 60 breaths; 210 nmol) to inhibit ACE and acetorphan (0.3, 1 and 3 mm, 60 breaths; 9 nmol, 33 nmol and 110 nmol respectively) to inhibit NEP. We also examined the effect of the highest dose of acetorphan (110 nmol) on the response to aerosolised acetylcholine (ACh).

Responsiveness to SP (or ACh) was measured as the change in lung resistance (RL) induced by nebulisation of increasing concentrations of SP (or ACh) before and after treatment with the inhibitor. PC200, defined as the provocative concentration inducing an increase in RL of 200% above baseline, was calculated for each challenge.

Administration of acetorphan before the second SP‐challenge induced a dose‐dependent decrease in PC200 for SP amounting to 1.8 (±0.3) log units after treatment with 11 nmol acetorphan. Treatment with vehicle before the second SP‐challenge or with 3 mm acetorphan before the second ACh‐challenge had no significant effect on PC200.

Treatment with captopril (21 nmol) induced only a small, nonsignificant leftward shift of PC200 to SP (0.3 ± 0.2 log units).

We conclude that a NEP‐like enzyme, but not ACE, regulates the response to aerosolised SP. We suggest that the same is true for SP released endogenously from sensory nerve endings in the airway epithelial layer.
Effects of epithelium removal on relaxation of airway smooth muscle induced by vasoactive intestinal peptide and electrical field stimulation
Authors: Farmer Stephen G. , Togo James
Published: July 2012

We have studied the effect of epithelium removal on relaxation of guinea‐pig isolated tracheal smooth muscle induced by vasoactive intestinal peptide (VIP) or stimulation of non‐adrenergic, non‐cholinergic (NANC) inhibitory nerves. Also examined were the effects of inhibitors of neutral endopeptidase (NEP) and angiotensin‐converting enzyme (ACE).

Epithelium removal produced a 3.6 ± 0.4 fold leftward shift in the VIP concentration‐response curve. The supersensitivity to VIP, following epithelium removal was abolished by phosphoramidon or thiorphan (NEP inhibitors), but unaffected by captopril (an ACE inhibitor). In intact trachea, the NEP inhibitors produced leftward shifts in the VIP curves similar to those produced by epithelium removal.

In contrast to responses to exogenous VIP, neurogenic NANC inhibitory responses to electrical field stimulation were affected neither by epithelial denudation nor by the peptidase inhibitors.

As in previous studies, epithelium removal increased tracheal sensitivity to isoprenaline. This was not altered by pretreatment with a cocktail of peptidase inhibitors. Thus, the effect of the NEP inhibitors on responses to VIP appears to be relatively specific.

These data indicate that exogenous VIP is a substrate for airway NEP, since inhibition of the enzyme potentiates the peptide. This is further evidence that the airway epithelium provides a source for the metabolism of mediators.

In guinea‐pig trachea the NEP responsible for cleaving VIP may be located largely in the epithelial layer, since NEP inhibition was without effect on sensitivity to VIP in epithelium‐denuded preparations. If VIP is a NANC inhibitory neurotransmitter in this tissue, its degradation endogenously does not appear to involve epithelial NEP.
The effects of cholecystokinin octapeptide on human isolated alimentary muscle
Authors: D'Amato M. , Stamford I.F. , Bennett A.
Published: July 2012

We studied cholecystokinin octapeptide (CCK‐OP) for its motor effects and sites of action on human isolated muscle from stomach, small intestine and colon.

CCK‐OP induced a concentration‐dependent contraction of all the longitudinal muscles and of circular muscle from the stomach and large intestine. The peptide acted directly on these muscles at a site not involving muscarinic receptors.

CCK‐OP relaxed the circular muscle of the small intestine and/or reduced the contractions to acetylcholine, by stimulating intramural postganglionic inhibitory neurones.
Antigen induces leucopenia in non‐immunised guinea‐pigs injected with platelets from actively sensitized animals
Authors: Pretolani Marina , Randon Jacques , Vargaftig B. Boris
Published: July 2012

Ovalbumin administration to animals injected with 0.5–5 × 106 washed platelets μl−1 from actively sensitized animals induced a marked decrease (maximum of around 50% at 60 min) in the number of circulating leucocytes, whereas platelet counts were unaffected. The intensity of the decrease in leucocyte counts was dependent upon the concentration of the injected platelets.

No drop in leucocyte counts was measured upon antigen challenge of guinea‐pigs injected with platelets from non‐sensitized animals.

This phenomenon was unaffected by pretreatment of the recipient animals with a platelet‐activating factor (PAF) antagonist WEB 2086 (3 mg kg−1 i.v.) but was markedly reduced (around 50% inhibition) by the anti‐allergic drug nedocromil sodium (100 mg kg−1 i.v.). By contrast, indomethacin (5 mg kg−1 i.v.) caused a significant (P < 0.01) enhancement of the antigen‐induced leucopenia, whereas the mixed cyclooxygenase and lipoxygenase inhibitor, BW 755C (10 mg kg−1 i.v.) suppressed the drop in leucocyte counts evoked by ovalbumin administration.

These results indicate that platelets from actively sensitized guinea‐pigs transferred to normal animals still responds to the specific antigen with the activation of circulating leucocytes. This phenomenon appears to be independent of the production of PAF and of cyclo‐oxygenase metabolites. By contrast, the production of the metabolites of the lipoxygenase pathway by platelets could account for the marked leucopenia observed following the immunological stimulation.
Evidence that agonist and antagonist enantiomers of the dihydropyridine PN 202–791 act at different sites on the voltage‐dependent calcium channel of vascular muscle
Authors: Hughes A.D. , Hering S. , Bolton T.B.
Published: July 2012

S(+)‐PN 202–791, a calcium channel agonist, and its optical isomer R(−)‐PN 202–791, a calcium channel antagonist, respectively increased and decreased inward current carried by barium through voltage‐dependent calcium channels in isolated ear artery cells of the rabbit in a concentration‐dependent manner. The EC50 or IC50 derived from the concentration‐response relationship was unaltered in the presence of the other enantiomer indicating that no competitive antagonism exists between these enantiomers and suggesting that the actions of these two enantiomers involve two separate binding sites.
Interactions of spironolactone with (+)‐[3H]‐isradipine and (−)‐[3H]‐desmethoxyverapamil binding sites in vascular smooth muscle
Authors: Mironneau J. , Sayet I. , Rakotoarisoa L. , Dacquet C. , Mironneau C.
Published: July 2012

Spironolactone partially inhibited the specific binding of (+)‐[3H]‐isradipine and (−)‐[3H]‐desmethoxyverapamil to vascular smooth muscle membranes. It is suggested that spironolactone interacts at a binding site of the calcium channel complex and allosterically modulates ligand binding at receptor sites in the channel.
Potassium‐stimulated release of nitric oxide from cerebellar slices
Authors: Dickie B.G.M. , Lewis M.J. , Davies J.A.
Published: July 2012

This study shows that nitric oxide release can be evoked from intact neuronal tissue. Stimulation of rat cerebellar slices for 4 min with 50 mM potassium significantly increased the release of nitric oxide and glutamate and was associated with a rise in the guanosine 3′:5′‐cyclic monophosphate (cyclic GMP) content of the tissue. Nitric oxide levels in the perfusate increased from 160 to 262 parts per billion. It is suggested that an increased release of endogenous glutamate in the slice, induced by exposure to potassium, may mediate the release of nitric oxide.
Pharmacological estimation of agonist affinity: detection of errors that may be caused by the operation of receptor isomerisation or ternary complex mechanisms
Authors: Leff P. , Harper D. , Dainty I.A. , Dougall I.G.
Published: July 2012

Recent theoretical studies have questioned the pharmacological estimation of agonist affinity. They showed that when receptor isomerisation or ternary complex mechanisms operate, the receptor inactivation method can substantially overestimate affinity, whereas methods for partial agonist analysis are more accurate. We previously suggested that the operation of such mechanisms and therefore the presence of errors could be detected by analysing the same partial agonist by the receptor inactivation and comparative methods. This paper describes the practical application of this test.

The ternary complex mechanism was simulated for a partial agonist under various conditions relating receptor (R) and transducer (T) concentrations, one of which also corresponds to the receptor isomerisation mechanism. The theoretical data so generated were then analysed by the inactivation and comparative methods to quantify the magnitude of error of affinity estimation that could occur.

This analysis showed that for a partial agonist with approximately 85% of the activity of a full agonist, the inactivation method could produce an affinity (pKA) estimate up to 0.7 log10 units higher than that produced by the comparative method. This difference would occur when the total receptor concentration ([R0]) is less than or equal to the total transducer concentration ([T0]). It also showed that the over‐estimation of affinity by the inactivation method was accompanied by drastic overestimation of Em, the maximal effect parameter.

The test was then exemplified using the muscarinic receptor system in the guinea‐pig isolated left atrial preparation, where there is evidence that a ternary complex mechanism operates. The test agonist was pilocarpine, which produced on average 83% of the activity of the full agonist, carbachol. Pilocarpine was analysed in comparison with carbachol and by receptor inactivation in the same tissue resulting in small and statistically insignificant differences in Em (96.7% and 97.3% respectively) and pKA (5.03 and 4.95 respectively).

In conclusion, in this experimental system, there was no evidence for the errors in agonist affinity estimation predicted by theory. Although this conclusion only applies to this system and application of the test to others is necessary to establish the generality of the present results, further examination of the theoretical basis for the predicted errors is required.
Pharmacological evidence for the existence of a local renin‐angiotensin system in porcine interlobar renal arteries
Authors: Müller‐Schweinitzer Else , Olea‐Baza Itziar
Published: July 2012

Changes in tension in response to cumulative additons of angiotensins (i.e., angiotensinogen, angiotensin I and angiotensin II), bradykinin and acetylcholine were monitored isometrically on ring preparations from porcine interlobar renal arteries.

Angiotensins consistently elicited contractile responses, whereas both bradykinin and acetylcholine produced relaxation of the arterial rings when active tone was induced by prostaglandin F2α.

Contractile responses to angiotensin II could be completely blocked by the combined action of the cyclo‐oxygenase inhibitor, indomethacin (1 μm) and the lipoxygenase inhibitor, nordihydroguaiaretic acid (NDGA, 10 μm).

Relaxant responses to bradykinin were unchanged during blockade of thromboxane A2 synthesis by dazoxiben (30μm) and proved to be largely resistant to blockade by indomethacin (1 μm) and the prostaglandin I2 (prostacyclin) synthesis inhibitor, tranylcypromine (40 μm).

The angiotensin receptor blocker, saralasin (10 and 100 nm) antagonized responses to angiotensinogen, angiotensin I and angiotensin II effectively and with similar potency. Enalaprilic acid, the active metabolite of the converting enzyme inhibitor enalapril (300 nm), attenuated responses to angiotensin I but failed to inhibit those to angiotensinogen up to 1 μm. The serine protease kallikrein (0.001 to 1 u ml−1) produced a dose‐dependent shift to the left of the concentration‐response curve for angiotensinogen.

It is suggested that the porcine interlobar renal artery possesses a local renin‐angiotensin system with activatable angiotensin II forming enzyme(s) within the vessel wall.
Alterations in regional blood flow in rats following sensitization to the nematode Nippostrongylus brasiliensis: effects of PAF antagonists
Authors: Mathison Ronald , Rimmer Carol , Davison Joseph S. , Wallace John L. , Befus A. Dean
Published: July 2012

Changes in tissue and organ blood flow associated with sensitization of rats to the nematode parasite, Nippostrongylus brasiliensis, were studied 30 to 35 days after infection, a time when very few worms remain in the animal.

Neither active nor passive sensitization modified heart rate, mean arterial blood pressure, cardiac output or total peripheral resistance. Passive sensitization and administration of non‐immune sera did not modify blood flow to any of the tissues studied.

Active sensitization increased hepatic arterial blood flow, but decreased blood flow to the stomach, duodenum, jejunum and the submandibular glands. These effects cannot be attributed to residual nematode infections as treatment with the anthelmintic, thiabendazole, did not alter blood flow relative to untreated, actively sensitized rats.

The effects of active sensitization on blood flow were probably due to an action of platelet‐activating factor (PAF) since treatment of actively sensitized animals with the selective antagonists, WEB‐2086 and BN 52021, reversed the decrease in flow seen to the intestinal regions. The PAF antagonists increased blood flow to the kidneys and the trachea of sensitized animals.

These results suggest that the PAF released from undetermined sources in nematode‐sensitized rats, produces altered blood flow, primarily to the stomach and proximal small bowel.
Dihydropyridines alter adenosine sensitivity in the rat hippocampal slice
Authors: Bartrup J.T. , Stone T.W.
Published: July 2012

The effects of adenosine and a range of adenosine analogues, which are resistant to uptake processes, were studied in the presence of dihydropyridines and verapamil on the population spike potential recorded from the CA1 area of the hippocampal slice.

Nifedipine and Bay K 8644, a calcium channel antagonist and activator respectively, enhanced the inhibitory action of adenosine in a concentration‐dependent manner. This was in contrast to their effect on adenosine analogues where the inhibition of the population potential was significantly attenuated. Similiar interactions between the adenosine compounds and the dihydropyridines were also displayed in studies on spontaneous epileptiform activity in the CA3 region.

This effect of nifedipine and Bay K 8644 was not shown by the dihydropyridines, nimodipine or nitrendipine, or by the phenylalkylamine, verapamil.

Addition of the adenosine uptake blocker dipyridamole reversed the action of nifedipine on adenosine, so that inhibition by adenosine was now attenuated by nifedipine in a similar manner to that observed with the adenosine analogues.

These results can be explained with reference to binding studies that show displacement of adenosine analogues from the adenosine receptor by dihydropyridines. An action at the adenosine uptake site by the dihydropyridines explains the enhancement of adenosine inhibition.

The possible sites for this interaction are discussed.
Endothelium modulation of the effects of nitroglycerin on blood vessels from dogs with pacing‐induced heart failure
Authors: Forster Christine , Main Jeffrey S. , Armstrong Paul W.
Published: July 2012

The relaxant actions of nitroglycerin (previously considered to be an endothelium‐independent relaxing agent) and acetylcholine (an endothelium‐dependent relaxing agent) were compared on 4 vascular preparations (dorsal pedal artery, saphenous vein, left anterior descending coronary artery and circumflex coronary artery) from dogs with and without pacing‐induced congestive heart failure (CHF).

Responses of the coronary arteries to acetylcholine were unaltered in endothelium‐intact rings from dogs with and without heart failure. Similarly no such changes were observed in the peripheral vessels. The maximum relaxation produced by acetylcholine was always greater in the coronary vessels compared to the peripheral vessels.

Before heart failure, the coronary vessels were more sensitive and reactive to nitroglycerin compared to the peripheral vessels.

Removal of the endothelium in both the control (dogs without CHF) and experimental (dogs with CHF) rings enhanced the relaxant effects of nitroglycerin, such that the EC50 for nitroglycerin became significantly lower in all denuded rings, with the exception of the saphenous vein and the left anterior descending coronary artery, before the development of CHF.

When CHF was maximally developed, vascular sensitivity to nitroglycerin was increased in peripheral vessels with an intact endothelium, but not in the coronary vessels.

These findings indicate that relaxation produced by nitroglycerin cannot be considered as entirely endothelium‐independent but should be considered endothelium‐modulated.
Phentolamine and yohimbine inhibit ATP‐sensitive K+ channels in mouse pancreatic β‐cells
Authors: Plant T.D. , Henquin J.C.
Published: July 2012

The effects of phentolamine and yohimbine on adenosine 5‐triphosphate (ATP)‐sensitive K+ channels were studied in normal mouse β‐cells.

In the presence of 3 mM glucose, many ATP‐sensitive K+ channels are open in the β‐cell membrane. Under these conditions, phentolamine inhibited 86Rb efflux from the islets. This inhibition was faster with 100 than with 20 μm phentolamine but its steady‐state magnitude was similar with both concentrations. Yohimbine (20–100 μm) also inhibited the efflux rate but was not as potent as phentolamine.

In the presence of 6 mM glucose, most ATP‐sensitive K+ channels are closed in the β‐cell membrane. Their opening by 100 μm diazoxide caused a marked acceleration of 86Rb efflux from the islets. This acceleration was almost entirely prevented by 20 μm phentolamine. It was barely affected by 20 μm yohimbine and reduced by 50% by 100 μm yohimbine.

ATP‐sensitive K+ currents were studied in single β‐cells by the whole cell patch‐clamp technique. Phentolamine (20–100μm) caused a progressive but almost complete and irreversible inhibition of the current. The effects of yohimbine were faster but smaller; the inhibition was still incomplete with 100 μm yohimbine.

The increase in ATP‐sensitive K+ current produced by 100 μm diazoxide was prevented by 100 μm phentolamine but only partially attenuated by 100 μm yohimbine.

It is concluded that phentolamine inhibits ATP‐sensitive K+ channels in pancreatic β‐cells. This novel effect of phentolamine resembles that of hypoglycaemic sulphonylureas. It may account for previously unexplained effects of the drug. These observations also call for reinterpretation of many studies in which phentolamine was used as an allegedly specific blocker of α‐adrenoceptors.