bluuebunny/crossref_metadata_2025 huggingface dataset:
Effects of chronic intravenous infusions of dopexamine and isoprenaline to rats on D1‐, β1‐ and β2‐receptor‐mediated responses
Authors: Martin S.W. , Broadley Kenneth J.
Published: July 2012

Rats received intravenous infusions of dopexamine, an agonist with selectivity for D1‐dopamine receptors and β2‐adrenoceptors (240 μg kg−1 h−1), isoprenaline, a β1‐ and β2‐adrenoceptor agonist (40 μg kg−1 h−1) or vehicle (isotonic saline at pH 2.5) for 7 days via subcutaneously implanted osmotic minipumps. Tissues were then removed for determination of functional responsivenss to β1‐adrenoceptor, β2‐adrenoceptor and dopamine D1‐receptor stimulation.

The β1‐adrenoceptor‐mediated responses of the spontaneously beating right atrium (increase in rate) and paced left atrium (increase in tension) showed significant reductions in sensitivity to isoprenaline following isoprenaline infusion. The EC50 values were significantly increased from 5.6 to 17.7 nm in right atria and from 7.4 to 27.1 nm in left atria. The maximum rate increase of right atria was also significantly less after isoprenaline infusion compared with controls (164.0 and 251.9 beats min−1, respectively) but the maximum tension responses of left atria were not significantly different. After infusion with dopexamine, however, there was no change in sensitivity of the left or right atria to β1‐adrenoceptor stimulation by isoprenaline.

β2‐Adrenoceptor‐mediated relaxation responses to isoprenaline of the pulmonary artery, constricted with noradrenaline (6 × 10−8 m), showed no significant difference in maximum response or EC50 in tissue from isoprenaline‐ or dopexamine‐infused rats compared with vehicle‐infused controls. β2‐Adrenoceptor‐mediated vasodilator responses were also examined in the kidney perfused with the thromboxane A2 analogue, U46619 (7.1 × 10−8 m) to raise perfusion pressure. As with the pulmonary artery, there was no significant difference in ED50 or maximum response to isoprenaline in kidneys from isoprenaline‐infused animals compared with vehicle controls.

The D1‐receptor‐mediated vasodilator responses to dopamine of the kidney perfused with U46619 were reduced after infusion of rats with dopexamine. The maximum fall in perfusion pressure (16.0 mmHg) and ED50 value (5.2 μg) were significantly different from those after vehicle infusion (31.5 mmHg; 1.5 μg).

These results show that functional responses mediated via β1‐adrenoceptors and D1‐receptors are reduced by intravenous infusions of isoprenaline and dopexamine, respectively. In contrast, the β2‐adrenoceptor‐mediated vasodilator responses of the pulmonary artery and kidney are not reduced by these agonists. Thus, under identical conditions, there appears to be selective down‐regulation of peripheral β1‐ and D1‐receptors, but not of β2‐adrenoceptors.
Block by capsaicin of voltage‐gated K+ currents in melanotrophs of the rat pituitary
Authors: Kehl S.J.
Published: July 2012

Whole‐cell recordings of macroscopic K+ currents were made from acutely dissociated and cultured melanotrophs isolated from the pars intermedia of the adult rat pituitary.

In acutely dissociated cells, external capsaicin reversibly decreased the amplitude both of the fast‐activating, fast‐inactivating potassium current IK(f) and the slowly‐activating, slowly‐inactivating potassium current IK(s). To simplify the investigation of the mechanism of action of capsaicin experiments were conducted on cultured melanotrophs that express only IK(s).

In control cells the activation rate and the amplitude of IK(s) increased with depolarization and the current showed very little inactivation at any voltage during pulses lasting for 100–300 ms. In capsaicin, the decrease of the current amplitude was associated with an increased rate of current decay (‘inactivation’). At a given voltage, the extent and the rate of the capsaicin‐induced inactivation was proportional to the capsaicin concentration; and, at a given concentration, the extent and rate of the inactivation increased with membrane depolarization.

The fit of the Hill equation to data derived from the steady‐state block of IK(s) evoked at 10 mV indicated an equilibrium dissociation constant (KD) of 17.4 μm (95% confidence limits 15.8–19.0) and a Hill coefficient of 1.8 (95% Cl 1.5–2.2) suggesting that at least two molecules of capsaicin must bind to the channel to block it.

Analysis of the voltage‐dependence of the steady‐state block in 100 μm capsaicin showed that half‐maximal block occurred at − 29 ± 2 mV (n = 10). Two‐pulse experiments designed to study the time‐dependence of channel block in 100 μm capsaicin indicated that the blocking kinetics were well fitted by a single exponential and that the rate of block increased with depolarization. The value for τblock at 0 mV was 24 ± 7 ms (n = 4).

Recovery from block in 100 μm capsaicin was also well fitted by a single exponential. The recovery time constant (τrecovery) was 708 ± 140 ms at − 50 mV, 70 ± 6 ms at − 70 mV and 19 ± 1.3 ms at − 90 mV (n = 4).

In 50–100 μm capsaicin, the decay of the tail current was biexponential, the values for τfast and τslow being, respectively, less than and greater than the single time constant fitted to the control tail current. Inward and outward K+ currents were equally affected by capsaicin.

Most of these effects of capsaicin on the IK(s) of melanotrophs can be accounted for by a kinetic scheme in which capsaicin binds to and blocks open K+ channels.
Effects of DAU 6215, a novel 5‐hydroxytryptamine3 (5‐HT3) antagonist on electrophysiological properties of the rat hippocampus
Authors: Passani M.B. , Pugliese A.M. , Azzurrini M. , Corradetti R.
Published: July 2012

The aim of the present study was to test the effects of DAU 6215 (endo‐N‐(8‐methyl‐8‐azabicyclo‐[3.2.1]‐octo‐3‐yl)‐2,3‐dihydro‐2‐oxo‐1H‐benzimidazole‐1‐carboxamide hydrochloride), a newly synthesized, selective 5‐hydroxytryptamine3 (5‐HT3) antagonist, on the cell membrane properties and on characterized 5‐HT‐mediated responses of pyramidal neurones in the hippocampal CA1 region.

Administration of DAU 6215, even at concentrations several hundred fold its Ki, did not affect the cell membrane properties of pyramidal neurones, nor modify extracellularly recorded synaptic potentials, evoked by stimulating the Schaffer's collaterals.

Micromolar concentrations (15–30 μm) of 5‐HT elicited several responses in pyramidal neurones that are mediated by distinct 5‐HT receptor subtypes. DAU 6215 did not antagonize the 5‐HT1A‐induced membrane hyperpolarization and conductance increase, a response that was blocked by the selective 5‐HT1A antagonist NAN‐190 (1‐(2‐methoxyphenyl)‐4‐[4‐(2‐phtalamido)butyl‐piperazine). Similarly, DAU 6215 did not affect the membrane depolarization and decrease in amplitude of the afterhyperpolarization, elicited by the activation of putative 5‐HT4 receptors.

5‐HT increased the frequency of spontaneous postsynaptic potentials (s.p.s.ps) recorded in pyramidal neurones loaded with chloride. In agreement with previous observations, most of the s.p.s.ps were reversed GABAergic events, produced by the activation of 5‐HT3 receptors on interneurones, because they persisted in the presence of the glutamate NMDA and non NMDA antagonists, d‐aminophosphonovaleric acid (APV; 50 μm) and 6,7‐dinitroquinoxaline‐2,3‐dione (DNQX; 25 μm), and were elicited by the selective 5‐HT3 agonist, 2‐methyl‐5‐HT (2‐Me‐5‐HT, 50 μm).

The increase in frequency of s.p.s.ps induced by 5‐HT was significantly antagonized by DAU 6215 in 70% of the cases, whereas the 5‐HT3 antagonist always suppressed the effect of 2‐Me‐5‐HT, at concentrations as low as 60 nm.

The antagonistic effect of DAU 6215 was also tested on the 5‐HT3‐mediated block of induction of long‐term potentiation (LTP), elicited by a primed burst (PB) stimulation. Extracellular recordings showed that low concentrations (60 nm) of DAU 6215 suppressed the inhibitory action of 5‐HT on PB‐induced LTP, without affecting the 5‐HT1A‐induced reduction in the amplitude of the population spike.

These results provide evidence that DAU 6215 is an effective antagonist of the 5‐HT3‐mediated responses in the central nervous system and may offer a cellular correlate for the pharmacological effects of DAU 6215 as an anxiolytic and cognition enhancer.
Differential modulation of κ and μ opioid antinociception by the glycine/NMDA receptor agonist D‐serine
Authors: Hunter J.C. , Atwal P. , Woodruff G.N. , Singh L.
Published: July 2012

D‐Serine, a selective agonist for the strychnine‐insensitive glycine allosteric site associated with the NMDA receptor‐ion channel complex, was found to modulate differentially the antinociception produced by κ and μ‐opioid receptor agonists in the rat formalin test. D‐Serine (100 μg, i.c.v.) attenuated the antinociception produced by the selective κ‐opioid agonist, enadoline (0.003–0.1 mg kg−1, s.c.) against the tonic, but not acute, phase of the formalin response. Conversely, D‐serine potentiated the antinociception produced by morphine (0.3–10 mg kg−1, s.c.) against both the acute and tonic phases. These results demonstrate an important interaction between the opioid and NMDA/glycine systems in the control of nociceptive information possibly at different levels of the neuraxis.
Anti‐inflammatory and analgesic activity of the bradykinin antagonist, icatibant (Hoe 140), against an extract from Porphyromonas gingivalis
Authors: Griesbacher Thomas , Sutliff Roy L. , Lembeck Fred
Published: July 2012

Porphyromonas gingivalis is one of the bacteria likely to be related to pain in periodontitis. Several enzymes isolated from P. gingivalis have been reported to have kininogenase activity. Since kinin release could be held responsible for inflammatory symptoms and pain in periodontitis, we investigated whether the inflammatory and algesic effects of a sonic extract from P. gingivalis (PGSE) could be inhibited by the potent bradykinin B2 receptor antagonist, icatibant (Hoe 140).

In anaesthetized rats, the subplantar injection of PGSE (0.1 and 1.0 mg) caused a dose‐dependent oedema of the hind paws. The net increase of the paw volume 60 min after the injection was 23 ± 5% and 77 ± 12%, respectively. The oedema was rich in plasma proteins as determined by the Evans blue method. Pretreatment with icatibant (300 nmol kg−1, s.c.) significantly reduced the effect of 1.0 mg of PGSE whereas the effects of 0.1 mg of PGSE remained unaffected.

The subplantar injection of 1.0 mg of PGSE in unanaesthetized rats caused nociceptive behavioural responses which started about 5 min after the injection and lasted for about 10–15 min. These responses were completely prevented by pretreatment with icatibant (300 nmol kg−1, s.c.).

The present results show that the plasma extravasation induced by non‐algesic doses of a sonic extract from P. gingivalis are caused by mechanisms other than B2 kinin receptor activation whereas inflammatory effects of algesic doses are due to the action of kinins. The pain elicited by the extract is solely mediated by kinins and can be prevented by icatibant. The bradykinin antagonist could thus have a potential for a clinical use against pain associated with periodontal inflammation.
(−)‐Discretamine, a selective α1D‐adrenoceptor antagonist, isolated from Fissistigma glaucescens
Authors: Ko Feng‐Nien , Guh Jih‐Hwa , Yu Sheu‐Meei , Hou Yu‐Sheng , Wu Yang‐Chang , Teng Che‐Ming
Published: July 2012

The selectivity of (−)‐discretamine for α1‐adrenoceptor subtypes was investigated by use of functional and binding studies in rat vas deferens, spleen and aorta, and in cultured DDT1MF‐2 and A10 cells.

In prostatic portions of rat vas deferens, the competitive antagonists (−)‐discretamine, 5‐methyl‐urapidil (5‐MU) and prazosin inhibited contractions to noradrenaline (NA) with pA2 values of 6.21, 8.71 and 9.27, respectively. The irreversible antagonist, chloroethylclonidine (CEC, 100 μm) failed to affect contractions to NA while nifedipine (1 μm) blocked them almost completely.

In rat spleen, the competitive antagonists (−)‐discretamine, 5‐MU and prazosin inhibited contractions to phenylephrine with pA2 values of 6.44, 7.19 and 9.45, respectively. CEC (100 μm) significantly reduced the maximum contraction to phenylephrine while nifedipine (1 μm) did not affect it.

In rat aorta, the competitive antagonists (−)‐discretamine, 5‐MU and prazosin inhibited contractions to NA with pA2 values of 7.60, 8.00 and 9.40, respectively. CEC also antagonized the contractions to NA in a competitive manner with a pA2 value of 6.10.

The specific binding of [3H]‐prazosin to DDT1MF‐2 and A10 cells was concentration‐dependent and saturated at 3–5 nM with KD values of 0.24 ± 0.02 and 0.20 ± 0.02 nM, respectively. (−)‐Discretamine, 5‐MU, CEC and prazosin inhibited specific [3H]‐prazosin binding to DDT1MF‐2 and A10 cells in a concentration‐dependent manner with IC50 values of 390.8 ± 20.6, 43.6 ± 3.9, 200.0 ± 30.0 and 0.8 ± 0.1 nM, respectively in DDT1MF‐2 cells, and 25.0 ± 3.2, 8.6 ± 1.4, 1000.0 ± 30.8 and 0.52 ± 0.03 nM, respectively in A10 cells.

Pretreatment of A10 cells with CEC (10 μm) for 30 min and then washed out thoroughly, reduced specific [3H]‐prazosin binding by 30%. The CEC‐insensitive [3H]‐prazosin binding was inhibited by (−)‐discretamine with an IC50 value of 7.0 ± 0.3 nM.

5‐MU (100 nM), CEC (1 μm) and prazosin (10 nM) markedly inhibited NA (3 μm)‐induced [3H]‐inositol monophosphate formation in DDT1MF‐2 and A10 cells, while (−)‐discretamine (100 nM) inhibited NA‐induced [3H]‐inositol monophosphate formation only in A10 cells.

In conclusion, (−)‐discretamine is a selective α1D‐adrenoceptor antagonist in vascular smooth muscle. Its selectivity among various α1‐adrenoceptor subtypes is α1A:α1B:α1D = 0.04:0.07:1.0.
Cutaneous vasodilatation induced by nitric oxide‐evoked stimulation of afferent nerves in the rat
Authors: Holzer P. , Jocič M.
Published: July 2012

The site of action at which nitric oxide (NO) may contribute to neurogenic vasodilatation in the hindpaw skin of urethane‐anaesthetized rats was examined by the use of NG‐nitro‐l‐arginine methyl ester (l‐NAME), an inhibitor of NO synthase.

Skin blood flow was measured by laser Doppler flowmetry, and neurogenic vasodilatation was evoked either by topical application of mustard oil (5%) or antidromic electrical stimulation of the saphenous nerve (antidromic vasodilatation).

l‐NAME (60 μmol kg−1, i.v.) attenuated the hyperaemia evoked by mustard oil in an enantiomer‐specific manner but failed to reduce antidromic vasodilatation and the vasodilatation due to i.v. injected calcitonin gene‐related peptide (CGRP) and substance P (0.1–1 nmol kg−1 each), two proposed mediators of neurogenic vasodilatation.

Pretreatment of rats with capsaicin (125 mg kg−1, s.c. 2 weeks beforehand), to defunctionalize afferent neurones, reduced the hyperaemic response to mustard oil and prevented l‐NAME from further decreasing the vasodilatation evoked by mustard oil.

Intraplantar infusion of sodium nitroprusside (SNP, 0.15 nmol in 1 min), a donor of NO, induced hyperaemia which was significantly diminished by the CGRP antagonist CGRP8–37 (50 nmol kg−1, i.v.) and by capsaicin pretreatment. The ability of CGRP8–37 to inhibit the vasodilator response to SNP was lost in capsaicin‐pretreated rats.

Taken together, these data indicate that NO does not play a vasorelaxant messenger role in neurogenic vasodilatation but can contribute to activation of, and/or transmitter release from, afferent nerve fibres in response to irritant chemicals.
Role of R‐type calcium channels in the response of the perfused arterial and venous mesenteric vasculature of the rat to platelet‐activating factor
Authors: Claing Audrey , Bkaily Ghassan , Berthiaume Nathalie , Sirois Pierre , Rola‐Pleszczynski Marek , D'Orléans‐Juste Pedro
Published: July 2012

The vasoactive properties of platelet‐activating factor (PAF) were studied in the arterial and venous vasculature of the rat double‐perfused mesenteric bed. Although PAF (0.01–0.3 pmol) induced a dose‐dependent vasodilatation of the arterial mesenteric vasculature, it triggered only vasoconstrictions on the venous side, with an intact endothelium as bradykinin induced a significant venodilatation.

NG‐nitro‐l‐arginine methyl ester (l‐NAME, 100 μm), a nitric oxide synthase inhibitor, markedly reduced the vasodilatation induced by PAF in the arterial mesenteric vasculature and potentiated the contractile responses of the venous side to the same agent.

The PAF antagonist, WEB‐2170, markedly reduced the response to PAF on both sides of the mesenteric vasculature. However, the IC50 of WEB‐2170 against PAF was reached at a much higher concentration (1 × 10−8 M) on the arterial side than on the venous side (5.3 × 10−11 M). Furthermore, a second antagonist of PAF receptors, SRI‐63441, although being less potent on the venous vasculature than WEB‐2170, was equipotent in antagonizing the venoconstriction and the arterial dilatation induced by PAF (IC50 of SRI‐63441, arterial side: 2.9 × 10−9 M; venous side: 3.1 × 10−9 M).

The dual L‐ and R‐calcium channel blocker, isradipine (PN 200–110), but not the L‐type calcium channel blocker, nifedipine, markedly reduced the PAF‐induced vasoactive properties on both sides of the mesenteric vasculature.

Our results illustrate the differential vasoactive properties of PAF in the mesenteric vasculature of the rat. These vasoactive responses occur following activation of specific receptors for PAF or, alternatively, through activation of R‐type calcium channels.
Neuropeptide Y inhibits potassium‐stimulated glutamate release through Y2 receptors in rat hippocampal slices in vitro
Authors: Greber Susanne , Schwarzer Christoph , Sperk Günther
Published: July 2012

We investigated the effects of neuropeptide Y (NPY), peptide YY (PYY), NPY13–36, NPY18–36, [Leu31][Pro34]NPY and of pancreatic polypeptide Y (PPY) on calcium‐dependent, potassium‐stimulated glutamate release in superfused rat hippocampal slices.

NPY, PYY and the Y2 receptor agonist NPY13–36 equipotently inhibited the release of glutamate. The half‐maximal response was observed at about 10 nm in a dose‐dependent manner (3 to 100 nm). Maximal inhibition of 50 to 60% was obtained at 100 nm. At higher concentrations of the peptides (300 nm and 1 μm) this inhibition was partially or entirely reversed. Porcine NPY13–36 and NPY18–36, inhibited glutamate release by about 44% at 100 nm.

The specific Y1 receptor agonist, [Leu31][Pro34]NPY, caused an insignificant increase in glutamate release at 100 to 300 nm concentrations. PPY had no effect on potassium‐evoked glutamate release in hippocampal slices at concentrations of 30 nm to 1 μm.

The experiments support previous electrophysiological data. They suggest a potent inhibitory action of NPY through NPY‐Y2 receptors on the release of the excitatory amino acid glutamate in rat hippocampus. Especially under conditions of increased NPY synthesis, such as in epilepsy, this mechanism may be of pathophysiological relevance.
Facilitation by procaterol, a β‐adrenoceptor agonist, of noradrenaline release in the pithed rat independently of angiotensin II formation
Authors: Kotsonis P. , Majewski H.
Published: July 2012

The effects of the β2‐adrenoceptor agonist, procaterol, on sympathetic neuroeffector transmission were studied in the pithed adrenal demedullated rat to determine if generation of angiotensin II was involved in its effect. Pressor responses were elicited by either electrical stimulation (20 V, 2 Hz) of the entire spinal sympathetic outflow or methoxamine (0.1 mg kg−1, i.v.).

Sodium nitroprusside (3 and 5 μg−1 min−1, i.v.) produced hypotension and the pressor responses to both sympathetic nerve stimulation and methoxamine were reduced. This indicates that decreasing blood pressure in pithed rats reduces pressor responses. Procaterol (10 and 30ng kg−1 min−1, i.v.) also produced hypotension but did not alter pressor responses to sympathetic nerve stimulation. Nevertheless, procaterol (10 and 30 ng kg−1 min−1, i.v.) did reduce pressor responses to methoxamine. Together these results suggest that procaterol may have enhanced sympathetic neurotransmitter release. This was confirmed in another series of experiments where procaterol (30ngkg−1 min−1, i.v.) increased plasma noradrenaline levels during sympathetic nerve stimulation.

Captopril (5 mg kg−1, i.v.) produced hypotension and as expected reduced pressor responses to sympathetic nerve stimulation. When the hypotensive effect of captopril was abolished by concomitant vasopressin infusion (1.5‐4.5 i mu kg−1 min−1, i.v.), pressor responses to sympathetic nerve stimulation were restored to pre‐captopril levels. In this situation procaterol (10 and 30 ng kg−1 min−1, i.v.) reduced basal blood pressure and did not alter pressor responses to sympathetic nerve stimulation whereas the pressor responses were reduced by an equihypotensive infusion of sodium nitroprusside (3 and 5 μg kg−1 min−1, i.v.). The lack of reduction of pressor responses after procaterol in the presence of captopril is indirect evidence that procaterol may have enhanced noradrenaline release independently of angiotensin II.

In another series of experiments, plasma noradrenaline levels elicited by sympathetic nerve stimulation were not altered by captopril (5 mg kg−1, i.v.). In the presence of captopril (5 mg kg−1, i.v.), procaterol (30 ng kg−1 min−1, i.v.) no longer enhanced plasma noradrenaline levels during sympathetic nerve stimulation. However, since the dose of captopril is well above that required to block angiotensin converting enzyme (ACE) the effect may be non‐specific. Therefore, the selective AT1 receptor antagonist, losartan (10 mg kg−1, i.v.), was also used. Losartan (10 mg kg−1, i.v.) did not alter plasma noradrenaline levels during sympathetic nerve stimulation, and in the presence of losartan procaterol (30 ng kg−1 min−1, i.v.) enhanced plasma noradrenaline levels during sympathetic nerve stimulation. This result further suggests that β‐adrenoceptor facilitation of noradrenaline release from sympathetic nerves in the pithed rat occurs by a mechanism independent of angiotensin II generation.
Y2‐receptor‐mediated selective inhibition of slow, inhibitory postsynaptic potential in submucous neurones of guinea‐pig caecum
Authors: Cunningham Shauna M.C. , Mihara Satoshi , Lees Gordon M.
Published: July 2012

The subtype of neuropeptide Y receptor mediating the selective inhibition of the slow inhibitory postsynaptic potential (i.p.s.p.) of submucous neurones in guinea‐pig caecum was investigated by use of conventional intracellular electrophysiological recording techniques.

Neuropeptide Y (NPY) (1‐300nM) was found to depress or abolish reversibly the slow i.p.s.p. evoked by focal stimulation of internodal fibre tracts. At low concentrations (1‐30nM), a reduction in the duration of the slow i.p.s.p. was often apparent before any inhibition of the amplitude of this synaptic potential.

These inhibitory effects of NPY were mimicked by peptide YY (PYY; 0.3–100 nm), NPY13‐36 (1–300 nm) and NPY22‐36 (10‐100nM); [Leu31, Pro34]NPY ([Pro34]NPY) and bovine pancreatic polypeptide (bPP) were without pre‐ or postsynaptic effects at concentrations of up to 300 nm. The IC50 s.e. mean values for PYY, NPY, and NPY13‐36 were 2.7 ± 0.3, 7.8 ± 2.1 and 30 ± 4.8nM, respectively, and were significantly different from each other. Thus, the apparent rank order of potency was PYY>NPY>NPY13‐36>[Pro34]NPY and bPP.

In concentrations of up to 300 nm, NPY and its analogues had no depressant effects on the active and passive properties of the impaled neurone and did not affect the amplitude or duration of either cholinergic fast synaptic potentials or non‐cholinergic, slow excitatory postsynaptic potentials (e.p.s.ps). Furthermore, none of these peptides altered the amplitude or time‐course of changes in membrane potential induced by focal application of acetylcholine or noradrenaline.

It is, therefore, concluded that the selective inhibition of the slow i.p.s.p. is mediated by Y2‐receptors, located presynaptically on noradrenergic nerve terminals.
Modulation by endogenous ATP of the light‐evoked release of ACh from retinal cholinergic neurones
Authors: Neal Michael , Cunningham Jo
Published: July 2012

The retina is an area of the central nervous system that possesses intrinsic cholinergic neurones which release acetylcholine (ACh) in response to stimulation with flickering light. Using an eye‐cup preparation in anaesthetized rabbits we found that when the retina was exposed to the P2‐purinoceptor antagonist, PPADS, the light‐evoked release of ACh was strikingly increased (by over 40%). In contrast, ATP reduced the light‐evoked release of ACh by 20%. The inhibitory effect of ATP was not due to its catabolism to adenosine because it was not affected by the Aradenosine receptor antagonist, DPCPX, in combination with adenosine deaminase. The actions of both ATP and PPADS were completely blocked by strychnine. We conclude that during physiological stimulation of the retina with light, ATP is co‐released with ACh and partially inhibits ACh release by activating (with ACh) an inhibitory glycinergic feed‐back loop.
Effects of RP 67580, a tachykinin NK1 receptor antagonist, on a primary afferent‐evoked response of ventral roots in the neonatal rat spinal cord
Authors: Hosoki Rumiko , Yanagisawa Mitsuhiko , Guo Jian‐Zhong , Yoshioka Koichi , Maehara Taketoshi , Otsuka Masanori
Published: July 2012

The pharmacological characteristics of RP 67580, a non‐peptide tachykinin NK1 receptor antagonist, and its effects on a reflex response evoked by stimulation of primary afferent fibres, were examined in isolated neonatal spinal cord preparations of the rat. Potentials were recorded extracellularly from a lumbar ventral root and drugs were bath‐applied in normal artificial cerebrospinal fluid (CSF) or in the presence of tetrodotoxin (TTX).

In normal artificial CSF, RP 67580 (0.1‐0.3 μm) caused rightward shifts of the concentration‐response curves for substance P (SP), neurokinin A (NKA) and substance P methyl ester (SPOMe), an NK1selective agonist, with pA2 values of 7.25, 7.47 and 7.49, respectively.

In the presence of TTX (0.3 μm), RP 67580 also caused rightward shifts of the concentration‐response curves for SPOMe and NKA. The pA2 value of RP 67580 against SPOMe (6.75) was significantly lower than that against NKA (7.22). RP 67580 (0.3‐1 μm) did not cause a clear parallel shift of the concentration‐reponse curves for SP, and it depressed the depolarizations induced by low concentrations of SP, but slightly potentiated those induced by high concentrations of SP.

RP 67580 (1 μm) did not depress the depolarizing responses to bombesin, 1–glutamate, γ‐aminobutyric acid (GABA), thyrotropin‐releasing hormone and muscarine. RP 67580 (1 μm), however, depressed the response to acetylcholine in the presence of atropine and the response to nicotine. RP 68651 (1 μm), the enantiomer of RP 67580 devoid of activity at tachykinin NK1 receptors, also depressed the response to acetylcholine in the presence of atropine.

RP 67580 (1 μm) did not induce GABA release from the rat spinal cord.

In the neonatal gerbil spinal cord, the antagonist effects of RP 67580 (0.3‐1 μm) against SPOMe were much less potent than in the neonatal rat spinal cord.

In the rat spinal cord‐saphenous nerve preparation, electrical stimulation of the saphenous nerve at C‐fibre strength evoked a prolonged depolarization of the ipsilateral L3 ventral root (slow VRP). RP 67580 (0.1‐1 μm) depressed the saphenous nerve‐evoked slow VRP. In contrast, RP 68651 (0.3 μm) had no effect on the slow VRP.

The results of the present study indicate that RP 67580 acts as a high affinity NK1 receptor antagonist in the neonatal rat spinal cord, although it also possesses an antinicotinic action. This study further suggests the existence of a subpopulation of tachykinin NK1 receptors that are activated by NKA and SPOMe, as well as by low concentrations of SP, and are sensitive to the antagonist action of RP 67580 in the neonatal rat spinal cord. This study also provides further evidence for the involvement of SP and NKA in the slow VRP evoked by C‐fibre stimulation in the neonatal rat spinal cord.
The effects of administration of monoamine oxidase‐B inhibitors on rat striatal neurone responses to dopamine
Authors: Berry M.D. , Scarr E. , Zhu M.‐Y. , Paterson I.A. , Juorio A.V.
Published: July 2012

(−)−Deprenyl has been shown to potentiate rat striatal neurone responses to dopamine agonists at doses not altering dopamine metabolism. Since there are a number of effects of (−)−deprenyl which could result in this phenomenon, we have investigated the effects of MDL 72,145 and Ro 19–6327, whose only common effect with (−)−deprenyl is an inhibition of monoamine oxidase‐B (MAO‐B), on rat striatal neurone responses to dopamine and on striatal dopamine metabolism.

Using in vivo electrophysiology, i.p. injection of either MDL 72,145 or Ro 19–6327 was found to produce a dose‐dependent potentiation of striatal neurone responses to dopamine but not γ aminobutyric acid.

Neurochemical investigations revealed that this occurred at doses (0.25‐1 mg kg−1) which, while not affecting levels of dopamine or its metabolites, 3,4‐dihydroxyphenylacetic acid or homo vanillic acid, did cause a significant, dose‐dependent, elevation in striatal levels of the putative neuromodulator, 2‐phenylethylamine (PE).

Inhibition of PE synthesis by i.p. injection of the aromatic 1‐amino acid decarboxylase inhibitor, NSD 1015, produced a reversal of the effects of MDL 72,145 and Ro 19–6327.

Neurochemical analysis revealed this to occur at a dose of NSD 1015 (10 mg kg−1) selective for reduction of elevated PE levels.

These results suggest that PE can act as a neuromodulator of dopaminergic responses and that MAO‐B inhibitors may potentiate neuronal responses to dopamine via the indirect mechanism of elevation of PE following MAO‐B inhibition.
Antithrombotic actions of argatroban in rat models of venous, ‘mixed’ and arterial thrombosis, and its effects on the tail transection bleeding time
Authors: Berry Christopher N. , Girard Denise , Lochot Sylvette , Lecoffre Catherine
Published: July 2012

The antithrombotic action of argatroban, a synthetic thrombin inhibitor, was studied in three models of thrombosis in the rat, and in the tail transection bleeding time test. Heparin was studied as a reference anticoagulant.

In the model of venous thrombosis induced by thromboplastin followed by stasis of the abdominal vena cava, argatroban had an ED50 of 125 μg kg−1, when administered as an i.v. bolus 5 min prior to the thromboplastin injection: the ED50 of heparin was 42 μg kg−1, where ED50 is the dose which reduces the weight of the thrombus by 50% compared with that of the control animals. When the two compounds were administered by continuous i.v. infusion, argatroban (ED50= 1.5 μg kg1 min−1) had the same potency as heparin (ED50= 1.2 μg kg1 min1).

Argatroban was active in the arterio‐venous shunt model with an ED50 of 0.6mg kg−1 when the compound was given as a bolus. The ED50 of herapin was 0.04 mg kg−1 under the same conditions. The two compounds had ED50 values of 6 μg kg1 min−1(argatroban) and 3 μg kg1 min−1(heparin), when administered by continuous i.v. infusion.

When tested against occlusive arterial thrombus formation by electrical stimulation of the left carotid artery, both compounds given as either an i.v. bolus or a continuous infusion led to dose‐dependent increases in the duration of post‐lesion vessel patency. Heparin bolus was more active than argatroban on a weight basis, in that 2mg kg−1 gave a similar increase in the time to occlusion as 8mg kg−1argatroban. As in the other models, when given as continuous infusions, argatroban (111% increase in time to occlusion at 20 μg kg−1, min−1) had similar activity to that of heparin (180% increase at 25 μg kg−1 min−1) on a weight basis. Hoever, the antithrombotic effects of argatroban were accompanied by only moderate changes in the coagulation parameters (thrombin time and activated partial thromboplastin time, APTT), whereas, even at a subthreshold dose of heparin (12.5 μg kg−1 min−1), both the thrombin time and the APTT were greater than 150 s.

Infusions of both compounds caused dose‐dependent increases in the tail transection bleeding time, with the dose of argatroban that doubles the bleeding time (11 μg kg−1 min−1) being five times greater than that of heparin (ED100= 2.2 μg kg−1 min−1).

These data show that, when administered as an intravenous infusion, argatroban is a potent antithrombotic agent in rat models of venous ‘mixed’ and arterial thrombosis, this effect can be obtained with a lower degree of systemic anticoagulation than with heparin in the arterial model, and argatroban has a lower haemorrhagic potential than that of heparin.
The binding of 1,3‐[3H]‐dipropyl‐8‐cyclopentylxanthine to adenosine Ai receptors in rat smooth muscle preparations
Authors: Peachey J.A. , Hourani S.M.O. , Kitchen I.
Published: July 2012

The binding of 1,3‐[3H]‐dipropyl‐8‐cyclopentylxanthine ([3H]‐DPCPX), an antagonist radioligand selective for adenosine A1 receptors, was studied in rat duodenum, colon muscularis mucosae and longitudinal muscle, urinary bladder and vasa deferentia.

[3H]‐DPCPX bound with high affinity to a single site in all membrane preparations studied with the exception of the rat urinary bladder in which no specific binding was detected. The affinity (Kd) of the binding site for [3H]‐DPCPX was similar in all membrane preparations, the colon longitudinal muscle (1.18 ± 0.47 nm), colon muscularis mucosae (0.84 ± 0.15 nm), duodenum (1.59 ± 0.18 nm) and vasa deferentia (0.93 ± 0.17 nm). The density of [3H]‐DPCPX binding sites was similar in the duodenum (38.8 ± 4fmol mg−1 protein), muscularis mucosae (43 ± 3.5 fmol mg−1 protein) and vasa deferentia (43.3 ± 12.2 fmol mg−1 protein), but in the longitudinal muscle 6–7 fold more binding sites (295 ± 70 fmol mg−1 protein) were identified.

Inhibition studies using DPCPX (0.1–100 nm), N6‐cyclopentyladenosine (CPA) (0.1–100 nm), 5′‐N‐ethylcarboxamidoadenosine (NECA) (2nM‐10 μm) and (R)‐N6‐phenylisopropyladenosine (R‐PIA) (1 nm‐1 μm) to displace the binding of [3H]‐DPCPX at a concentration around the Kd value (1 nm), demonstrated an order of potency of displacement in all tissues of DPCPX ≥ CPA > R‐ PIA > NECA. This potency order is characteristic of an A1 receptor, indicating that [3H]‐DPCPX binds to adenosine A1 receptors in the rat duodenum, colon and vasa deferentia. Two site analysis revealed that the agonists bind to both a high and low affinity state of the receptor.

The existence of A1 binding sites in the rat vasa deferentia, colon muscularis mucosae and duodenum, and their absence in the urinary bladder, is consistent with previous functional studies. However, in contrast to the findings of the [3H]‐DPCPX binding assay, no functional response mediated by adenosine A1 receptors could be detected by measuring contractile or relaxant responses to CPA in the colon longitudinal muscle. The functional significance of the binding sites in this tissue has therefore yet to be determined.
Effect of type A and B monoamine oxidase selective inhibition by Ro 41–1049 and Ro 19–6327 on dopamine outflow in rat kidney slices
Authors: Pestana M. , Soares‐da‐Silva P.
Published: July 2012

The influence of pargyline and of selective inhibitors of type A and B monoamine oxidase (MAO), Ro 41–1049 and Ro 19–6327 respectively, on the outflow of dopamine and 3,4‐dihydroxyphenylacetic acid (DOPAC) in slices of rat renal cortex loaded with exogenous L‐3,4‐dihydroxyphenylalanine (L‐DOPA) was examined. Dopamine and DOPAC in the tissues and in the effluent were assayed by means of h.p.l.c. with electrochemical detection.

The levels of newly‐formed dopamine and DOPAC in the perifusate decreased progressively with time. In control conditions, DOPAC/dopamine ratios in the perifusate were 3 to 5 fold those in the tissue and were found to increase progressively with time. The addition of pargyline (100 μm), produced a marked decrease in the outflow levels of DOPAC (45 to 54% reduction) and significantly increased the levels of dopamine in the effluent (102 to 158% increase); DOPAC/dopamine ratios in the perifusate remained stable throughout the perifusion and were similar to those found in the tissues. The addition of the MAO‐A inhibitor Ro 41–1049 to the perifusion fluid also significantly decreased DOPAC outflow (41% to 54% reduction) and increased dopamine outflow (19% to 80% increase). In the presence of Ro 41–1049 DOPAC/dopamine ratios in the perifusate were lower (P < 0.01) than in controls; in contrast with the effect of pargyline, this ratio was found to increase (P < 0.01) throughout the perifusion period. Ro 19–6327 did not reduce the outflow of DOPAC, but significantly increased (by 40–60%) that of dopamine. In the presence of Ro 19–6237, the proportion of DOPAC to dopamine in the perifusate was similar to that of controls and significantly increased throughout the perifusion; however, this increase was less than that observed in the control group.

When benserazide (50 μm) was added to the perifusion fluid, the levels of both dopamine and DOPAC in the effluent were similar to those observed in the absence of benserazide. However, in the presence of benserazide, DOPAC/dopamine ratios in the perifusate did not increase with time. In conditions of decarboxylase inhibition, the effects of pargyline, Ro 41–1049 and Ro 19–6327 on dopamine and DOPAC outflow were less pronounced than in experiments conducted in the absence of benserazide.

In conclusion, the results presented here show that the fraction of newly‐formed dopamine which leaves the compartment where the synthesis has occurred is a constant source for deamination into DOPAC. The results provide evidence favouring the view that MAO‐A is the main form of the enzyme involved in this process; however, the data described here suggest that dopamine would also have access to MAO‐B.
Peripheral and central sites of action of GABA‐B agonists to inhibit the cough reflex in the cat and guinea pig
Authors: Bolser Donald C. , DeGennaro Frances C. , O'Reilly Sandra , Chapman Richard W. , Kreutner William , Egan Robert W. , Hey John A.
Published: July 2012

The GABA‐B receptor agonists baclofen and 3‐aminopropylphosphinic acid (3‐APPi) have antitussive activity in the cat and guinea pig. The purpose of this study was to investigate the sites of action of these GABA‐B receptor agonists to inhibit the cough reflex.

Single intracerebroventricular (i.e.v.) cannulas were placed in the lateral ventricles of anaesthetized guinea pigs. Approximately 1 week later, the animals were exposed to aerosols of capsaicin (0.3 μm) to elicit coughing. Coughs were detected with a microphone and counted.

Cough was produced in anaesthetized cats by mechanical stimulation of the intrathoracic trachea and was recorded from electromyograms of respiratory muscle activity. Cannulas were placed for intravenous (i.v.) or, in separate groups of animals, intravertebral arterial (i.a.) administration of baclofen, 3‐APPi, the centrally active antitussive drug codeine or the peripherally active antitussive drug BW443c. Dose‐response relationships for i.v. and i.a. administration of each drug were generated to determine a ratio of i.v. ED50 to i.a. ED50, known as the effective dose ratio (EDR). The EDR will be 20 or greater for a centrally acting drug.

In the guinea pig, baclofen (3 mg kg−1, s.c.) and 3‐APPi (10 mg kg−1, s.c.) inhibited capsaicin‐induced cough by 50% and 35% respectively. The antitussive activity of baclofen was completely blocked by i.e.v. administration of the GABA‐B receptor antagonist CGP 35348 (10 μg). Conversely, the antitussive effect of 3‐APPi was unaffected by i.e.v. CGP 35348. However, systemic administration of CGP 35348 (30 mg kg−1, s.c.) completely blocked the antitussive activity of 3‐APPi (10 mg kg−1, s.c). In separate experiments baclofen alone (1 μg, i.c.v.) inhibited capsaicin‐induced cough by 78%. 3‐APPi (10 and 100 μg, i.c.v.) had no effect on capsaicin‐induced cough in the guinea pig.

In the cat, potencies (ED50) of the standards and GABA‐B agonists by the i.v. route were: codeine (0.34 mg kg−1), BW443C (0.17 mg kg−1), baclofen (0.63 mg kg−1) and 3‐APPi (2.3 mg kg−1). Potencies of these drugs by the i.a. route were: codeine, 0.013 mg kg−1; BW443C, 0.06 mg kg−1; baclofen, 0.016 mg kg−1; and 3‐APPi, 0.87 mg kg−1. The EDRs for each drug were: codeine, 26; BW443C, 3; baclofen, 39; and 3‐APPi, 3.

We conclude that in both the cat and guinea pig baclofen inhibits cough by a central site of action, while 3‐APPi inhibits cough by a peripheral site of action.
Modulation of morphine antinociception in the mouse by endogenous nitric oxide
Authors: Brignola Giovanni , Calignano Antonio , Di Rosa Massimo
Published: July 2012

L‐Arginine (100–1000 mg kg−1) administered orally (p.o.) or intraperitoneally (i.p.), but not intracerebroventricularly (i.c.v., 0.08 mg per mouse), reduced the antinociceptive effect of morphine (0.5–10 mg kg−1 s.c.) assessed in mice using three different tests: hot plate, tail‐flick and acetic acid‐induced writhing. D‐Arginine (up to 1000 mg kg−1 p.o. or i.p.) was ineffective.

NG‐Monomethyl‐L‐arginine (L‐NMMA, 5–50 mg kg−1 i.p.) and NG‐nitro‐L‐arginine methyl ester (L‐NAME, 5‐ 30 mg kg−1 i.p.), but not NG‐nitro‐D‐arginine methyl ester (D‐NAME, 30 mg kg−1 i.p.), reversed in all assays the effect of L‐arginine on morphine‐induced antinociception.

Morphine (10 mg kg−1 s.c), L‐arginine (1000 mg kg−1 p.o.) or L‐NAME (30 mg kg−1 i.p.), either alone or in combination, did not produce changes in locomotor activity or sensorimotor performance of animals.

These results suggest that the L‐arginine‐nitric oxide pathway plays a modulating role in the morphine‐sensitive nociceptive processes.
Antinociceptive and toxic effects of (+)‐epibatidine oxalate attributable to nicotinic agonist activity
Authors: Rupniak N.M.J. , Patel S. , Marwood R. , Webb J. , Traynor J.R. , Elliott J. , Freedman S.B. , Fletcher S.R. , Hill R.G.
Published: July 2012

Epibatidine is an analgesic substance, isolated from the skin of the poisonous frog Epipedobates tricolor, for which the mechanism of action was previously unknown.

The IC50 of synthetic (+)‐epibatidine oxalate (the naturally occurring isomer) for [3H]‐nicotine binding to rat whole‐brain membranes was 0.1 nm. The (−)−isomer also exhibited high affinity (IC50 = 0.2 nm).

(+)‐ and (−)−Epibatidine exhibited much lower affinity for displacement of the muscarinic ligand [3H]‐N‐methylscopolamine binding to rat cortical membranes (Kapp − 6.9μm and 16.0 μm respectively). The (+)‐enantiomer of epibatidine had an antagonist/agonist (NMS/oxo‐M) binding ratio of 4.2 This is consistent with a muscarinic antagonist profile.

(+)‐Epibatidine oxalate (10 μm) did not cause significant (>30%) displacement of radioligand binding to opioid, excitatory amino acid, benzodiazepine, 5‐HT, dopamine, adrenaline or peptide receptors.

(+)‐ and (−)−Epibatidine (5–20 μg kg−1 s.c.) doubled response latency in the mouse hot‐plate test. Antinociception and behavioural depression induced by (+)‐epibatidine (5 μg kg−1) was fully blocked by the nicotinic antagonists mecamylamine (2 mg kg−1 s.c.) or dihydro‐β‐erythroidine (2 mg kg−1 s.c). The muscarinic antagonist scopolamine (0.4 and 10 mg kg−1 s.c.) caused partial reversal of antinociception induced by (+)‐epibatidine in mice, but not in rats.

These findings demonstrate that (+)‐epibatidine oxalate salt is a highly selective and potent nicotinic analgesic agent.
Ion channel modulation by NS 1619, the putative BKCa channel opener, in vascular smooth muscle
Authors: Edwards G. , Niederste‐Hollenberg A. , Schneider J. , Noack Th. , Weston A.H.
Published: July 2012

The effects of NS 1619, the putative BKCa channel opener, were investigated on rat intact portal veins and on single smooth muscle cells enzymatically separated from the same tissue.

Under whole‐cell patch clamp conditions with K‐rich pipettes, exposure of single cells held at −10 mV to NS 1619 (10–33 μm) induced a noisy, outward current which reached a maximum (33 μm NS 1619; mean 35.8 ± 17 pA, n = 8) within about 6min.

On stepping to test potentials (range − 50 to + 50 mV) from a holding potential of − 10 mV, the NS 1619‐induced noisy current exhibited time‐dependent activation and marked outward rectification.

The stimulation of outward currents by NS 1619 at − 10 mV was independent of the presence of Ca2+ in the bath or pipette solutions but was antagonized by either charybdotoxin (250 nm) or penitrem A (100 nm) in the bath solution.

Stationary fluctuation analysis of the noisy current induced by NS 1619 at − 10 mV yielded a value of 70 ± 8 pS (n = 4) (under the quasi‐physiological conditions of the experiment) for the unitary conductance of the channel involved.

At − 10 mV, NS 1619 (10–33 μm) rapidly inhibited spontaneous transient outward currents.

With a holding potential of − 90 mV, NS 1619 (10–33 μm) produced a reduction of outward currents evoked by depolarizing steps to + 50 mV, an effect associated with marked inhibition of the delayed rectifier current, IK(V).

NS 1619 (3–100 μm) produced a concentration‐dependent inhibition of spontaneous activity in rat portal vein characterized by a reduction in the amplitude and duration of the tension waves. This inhibition was slightly potentiated in the presence of either charybdotoxin (250 nm) or penitrem A (1 μm). NS 1619 also totally inhibited contractions of rat aorta induced by KC1 (both 20 mm and 80 μm).

Under whole‐cell recording conditions and using Cs‐rich pipettes, Ca‐currents evoked in portal vein cells by stepping from a holding potential of − 90 mV to test potentials in the range − 30 to + 50 mV were totally inhibited in the presence of 33 μm NS 1619.

NS 1619 (33 μm) inhibited the induction of IK(ATP) by levcromakalim (10 μm).

It is concluded that NS 1619 activates the large conductance, Ca2+‐sensitive channel, BKCa and over the same concentration range it inhibits both Kv and L‐type Ca‐channels. The observed NS 1619‐induced mechanical inhibition in rat portal vein and aorta seems most likely to be due to the observed inhibition of Ca‐currents.