bluuebunny/crossref_metadata_2025 huggingface dataset:
Effect of chronic bradykinin B2 receptor blockade on blood pressure of conscious Dahl salt‐resistant rats
Authors: Mukai Hiroya , Fitzgibbon Wayne R , Ploth David W , Margolius Harry S
Published: January 2005

In this study 3 protocols were utilized to determine the role of endogenous kinins in the resistance of the inbred Dahl (Rapp) salt‐resistant (SR/Jr) rats to high salt diet‐induced blood pressure elevation.

The bradykinin B2 receptor antagonist, Hoe 140 (D‐Arg[Hyp3, Thi5, D‐Tic7, Oic8]‐bradykinin) at doses of either 10–20 or 20–40 nmol day−1 (subcutaneously (s.c), via osmotic minipumps, for either 1 or 3 weeks during a high (8%) salt diet) effectively blocked or attenuated the hypotensive responses to 100–1000 ng of bradykinin.

In the first protocol, 5 week old SR/Jr rats treated with Hoe 140 (10–20 nmol day−1, n=9, s.c., via osmotic minipumps) for 3 weeks and concomitantly fed high (8%) NaCl diet had significantly higher conscious tail cuff blood pressures (BPc) at 1 and 3 weeks when compared with rats treated with vehicle (0.9% NaCl, n=6). The differences in BPc between the 2 groups were 13 mmHg (P<0.001) after 1 week and 8 mmHg (P<0.05) after 3 weeks of treatment.

In the second protocol, 5 week old SR/Jr rats were treated with Hoe 140 (20–40 nmol day−1, n=8, s.c., via osmotic minipumps) or vehicle (n=8) for 3 weeks. During the first week of treatment the rats were fed normal (0.8%) NaCl diet. The rats were then switched to 8% NaCl for 2 remaining weeks of the protocol. The mean BPc of Hoe 140‐treated rats was not significantly different from that of the vehicle‐treated rats when fed 0.8% NaCl diet. In contrast, rats treated with Hoe 140 and concomitantly fed high (8%) NaCl diet had significantly increased BPc (123±2 vs 111±1 mmHg, P<0.001 for the Hoe 140‐ and vehicle‐treated rats, respectively).

In the third protocol, treatment with Hoe 140 (20–40 nmol day−1, s.c., via osmotic minipumps) during high salt diet did not increase BPc in rats that were pre‐exposed to the high salt diet for 2 weeks.

At the end of 3 weeks of study, blood pressure was measured via an arterial catheter during pentobarbitone‐induced anaesthesia. Rats treated with Hoe 140 for 1 or 3 weeks had significantly lower mean arterial blood pressures than the vehicle‐treated rats.

Our findings suggest that in SR/Jr rats, kinin activation of bradykinin B2 receptors at least partially contributes to early regulatory mechanisms that resist an increase in blood pressure following exposure to a high salt diet. The mechanism underlying the decreased blood pressure during pentobarbitone anaesthesia of SR/Jr rats chronically treated with Hoe 140 has yet to be elucidated.
Responses to endothelium‐dependent agonists in subcutaneous arteries excised from hypercholesterolaemic men
Authors: Lewis Tamara V , Cooper Bridget A , Dart Anthony M , Chin‐Dusting Jaye P F
Published: January 2005

Vasomotor function of the vascular endothelium was examined in human subcutaneous arteries excised from 8 hypercholesterolaemic and 7 normolipidaemic subjects.

Left gluteal skin biopsies were performed under local anaesthesia. Subcutaneous arteries were isolated and two vessels from each subject mounted in separate myographs. A 20 ml fasting blood sample was taken at the time of the biopsy.

Hypercholesterolaemic subjects had either never been treated with lipid lowering therapy or therapy had been stopped at least two weeks before the study (n=2). At the time of the study total plasma cholesterol levels (control: 4.6±0.3 vs hypercholesterolaemic: 8.3±0.6 mmol l−1: P<0.01) were significantly elevated in hypercholesterolaemic subjects when compared with controls.

Full concentration‐response curves to the vasoconstrictor noradrenaline and the vasodilators acetylcholine and substance P were constructed. A single point concentration‐response to sodium nitroprusside (10 μM) was also obtained. Dilator responses were obtained in vessels pre‐constricted with a submaximal concentration of noradrenaline. Vessels were then incubated for 30 min with either L‐ or D‐arginine (10 μM) and the concentration‐response curves to the three dilator agonists repeated in the presence of the amino acid.

Maximum relaxation responses to acetylcholine (control vs hypercholesterolaemic: 83.3±6.1% vs 47.4±13.5%; P<0.05), but not to substance P or sodium nitroprusside, were dampened in the hypercholesterolaemic group when compared with controls.

Neither incubation with L‐arginine nor D‐arginine had any effect on maximum relaxation responses to acetylcholine in either the control group (pre L‐arginine vs plus L‐arginine: 83.3±6.1 vs 82.3±5.5%, pre D‐arginine vs plus D‐arginine: 98.9±1.2 vs 98.2±1.1%) or the hypercholesterolaemic group (pre L‐arginine vs plus L‐arginine: 47.4±13.5 vs 55.3±14.3%, pre D‐arginine vs plus D‐argenine: 43.3±13.6 vs 65.4±12.3%).

When results from the two study groups were pooled, the strongest predictor of maximum relaxation obtained to acetylcholine was apolipoprotein A1 (r=0.67; P=0.001).

In conclusion, relaxation responses mediated by the endothelium‐dependent agonist acetylcholine, but not by substance P, are impaired in hypercholesterolaemic patients. L‐Arginine did not improve the impaired relaxation responses to acetylcholine. We suggest that impaired endothelium‐dependent relaxation is specific to acetylcholine and not to an abnormal L‐arginine‐nitric oxide pathway in subcutaneous arteries excised from this study group.
Irreversible binding of a carbostyril‐based agonist and antagonist to the β‐adrenoceptor in DDT1 MF‐2 cells and rat aorta
Authors: Deyrup Malgorzata D , Greco Phillip G , Otero Deborah H , Dennis Donn M , Gelband Craig H , Baker Stephen P
Published: January 2005

The chemoreactive ligands 5(2‐(((1′‐(4′‐isothiocyanatophenylamino)thiocarbonyl)‐amino)‐2‐methyl‐propyl)amino‐2‐hydroxypropoxy)‐3,4‐dihydrocarbostyril (DCITC) and 8‐hydroxy‐5(2‐(((1′‐(4′‐isothiocya‐natophenylamino)thiocarbonyl)amino)‐2‐methylprop‐2‐yl)amino‐1‐hydroxyethyl)‐carbostyril (HCITC)were synthesized and shown to be potent irreversible antagonist and agonist ligands, respectively, for the β‐adrenoceptor in DDT1 MF‐2 (DDT) cells and the rat isolated aorta.

In DDT cell membranes DCITC and HCITC inhibited (−)[125I]‐iodocyanopindolol (CYP) binding to the β‐adrenoceptor with IC50 values of 1.1 and 18 nM, respectively. (−)‐Isoprenaline inhibited [125I]‐CYP binding with an IC50 of 355 nM. Pretreatment of membranes with either chemoreactive ligand produced a time‐ and concentration‐dependent decrease in the β‐adrenoceptor content, indicating irreversible receptor binding. DCITC at concentrations up to 10 μM did not stimulate cyclic AMP accumulation in DDT cells nor did it amplify forskolin‐stimulated cyclic AMP accumulation.

In the rat isolated aorta, DCITC (0.1 μM) did not affect either the phenylephrine‐mediated tissue contraction or the acetylcholine‐mediated relaxation. DCITC attenuated the maximal (−)‐isoprenaline‐mediated relaxation of a phenylephrine contracted aorta in a concentration‐dependent manner and shifted the dose‐response curves for (−)‐isoprenaline to the right. The DCITC‐induced decrease in maximal response was not reversed by extensive tissue washing. By use of the operational model of agonism, the calculated dissociation constant for (−)‐isoprenaline ws 286 nM and the estimated receptor reserve for this agonist was 23% at the maximal response.

HCITC and (−)‐isoprenaline stimulated cyclic AMP accumulation in DDT cells with pD2 values (negative logarithm to base 10 of EC50) of 7.95 and 7.97, respectively, and both mediated the same maximal stimulation. In the rat isolated aorta, HCITC produced a concentration‐dependent relaxation of the tissue with a pD2 value of 6.62, whereas the pD2 for (−)‐isoprenaline was 7.03. However, HCITC produced a greater maximal relaxation of the tissue than (−)‐isoprenaline. The HCITC‐mediated stimulation of cyclic AMP accumulation and relaxation of the isolated tissue were blocked when the β‐antagonist propranolol was added concurrently. In contrast, once the HCITC‐mediated responses were established, the addition of propranolol did not result in any attenuation indicating that HCITC is an irreversible β‐agonist.
Pharmacological properties of P2X3‐receptors present in neurones of the rat dorsal root ganglia
Authors: Rae M. G. , Rowan E. G. , Kennedy C.
Published: January 2005

The electrophysiological actions of several agonists which may differentiate between P2X1‐ and P2X3‐receptors were studied under concentration and voltage‐clamp conditions in dissociated neurones of 1–4 day old rat dorsal root ganglia.

β,γ‐Methylene‐D‐ATP (β,γ‐me‐D‐ATP) (1–300 μM), diadenosine 5′,5′′′‐P1,P5‐pentaphosphate (AP5A) (100 nM–300 μM), diadenosine 5′,5′′′‐P1,P4‐tetraphosphate (AP4A) (300 nM–300 μM) and uridine 5′‐triphosphate (UTP) (1 μM–1 mM) all activated concentration‐dependent inward currents with a latency to onset of a few ms.

The concentration‐response curves for β,γ‐me‐D‐ATP and AP5A and ATP had similar maximum values, while that for AP4A had a lower maximum. The concentration‐response curve to UTP was shallow and did not reach a maximum. β,γ‐Methylene‐L‐ATP was virtually inactive. The rank order of agonist potency was ATP>AP5A∼amp;AP4A>β,γ‐me‐D‐ATP>UTP>>β,γ‐methylene‐L‐ATP.

The inward currents were inhibited by the P2‐receptor antagonists suramin (100 μM) and pyridoxalphosphate‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS) (10 μM). PPADS also inhibited responses to ATP (800 nM) and α,β‐methylene ATP (2 μM) in a concentration‐dependent manner.

This study shows that β,γ‐me‐D‐ATP, AP5A, AP4A and UTP all act via a suramin‐ and PPADS‐sensitive P2X‐receptor to evoke rapid, transient inward currents in dissociated neurones of rat dorsal root ganglia. The very low activity of β,γ‐methylene‐L‐ATP suggests that the agonists were acting at the P2X3‐subtype to produce these effects.
Effect of the selective 5‐HT1A receptor antagonist WAY 100635 on the inhibition of e.p.s.ps produced by 5‐HT in the CA1 region of rat hippocampal slices
Authors: Maria Pugliese Anna , Passani M Beatrice , Corradetti Renato
Published: January 2005

The actions of N‐(2‐(‐4(2‐methoxyphenyl)‐1‐piperazinyl)ethyl)‐N‐(2‐pyridinyl) cyclohexane carboxamide (WAY 100635), a novel and selective 5‐hydroxytryptamine1A (5‐HT1A) antagonist, on excitatory postsynaptic potentials (e.p.s.ps) were investigated by use of intracellular recordings in pyramidal cells of the CA1 region of rat hippocampal slices.

WAY 100635 (10 nM) did not affect any of the investigated parameters of cell excitability such as membrane potential, total input resistance (Rin), firing threshold, action potential amplitude, action potential frequency adaptation, and slow afterhyperpolarization (sAHP) which follows repetitive firing of action potentials. WAY 100635 did not have any effect on either the slope or the amplitude of e.p.s.ps evoked by stimulation of the CA1 stratum radiatum.

Bath application of either 5‐hydroxytryptamine (5‐HT, 10–30 μM) or 5‐carboxamidotryptamine (5‐CT, 300 nM) hyperpolarized the membrane potential (ΔVm=−4.1±0.9 and −6.0±0.9 mV, respectively), and reduced Rin (−25±8% and −18±1%, respectively). 5‐HT blocked the action potential frequency adaptation and significantly reduced the amplitude of the sAHP that follows repetitive firing of action potentials.

5‐HT significantly decreased the amplitude of evoked e.p.s.ps (−14±6%). This effect was greater in the presence of the GABAA receptor antagonist bicuculline (10 μM, −45±12%) and was mimicked by 5‐CT (−49±5%). Both AMPA and NMDA components of e.p.s.ps were significantly reduced in amplitude by 5–HT (−38±8%, n=6, and −29±12%, n=3, respectively; P<0.05).

WAY 100635 fully antagonized the hyperpolarization, the reduction of Rin, and the decrease in amplitude of e.p.s.ps elicited by 5‐HT, while it did not affect the action of 5‐HT on the action potential frequency adaptation. In the presence of WAY 100635, 5‐HT elicited a depolarization which was blocked by 10–30 μM RS 23597‐190, a selective 5‐HT4 receptor antagonist.

Our data demonstrate that WAY 100635 is devoid of direct effects on CA1 pyramidal cell excitability and on evoked e.p.s.ps, while it fully antagonizes the effects of 5‐HT on excitatory synaptic transmission and on hyperpolarization, without affecting the 5‐HT4 receptor‐mediated response. Since WAY 100635 selectively antagonizes 5‐HT1A receptor‐mediated actions of 5‐HT, our data also demonstrate that the inhibitory action of 5‐HT on excitatory synaptic transmission in CA1 is mediated by 5‐HT1A receptors.
Inhibition of nitrergic neurotransmission in the bovine retractor penis muscle by an oxidant stress: effects of superoxide dismutase mimetics
Authors: Mok Josephine S L , Paisley Karen , Martin William
Published: January 2005

A number of superoxide dismutase (SOD) mimetics were examined both biochemically for their ability to inhibit the superoxide‐catalyzed reduction of cytochrome c and nitro blue tetrazolium, and functionally for their ability to mimic authentic Cu/Zn SOD in restoring nitrergic neurotransmission in bovine retractor penis (BRP) muscle following its inhibition by oxidant stress.

The SOD mimetics investigated were CuSO4, MnCl2, CuDIPS (copper [II][diisopropylsalicylate]2), MnTBAP (manganese [III] tetrakis 4‐benzoic acid porphyrin), MnTMPyP (manganese [III] tetrakis 1‐methyl‐4‐pyridyl porphyrin pentachloride), tiron (4,5‐dihydroxy‐1,3‐benzene disulphonic acid), PTIYO (4‐phenyl,2,2,5,5,‐tetramethyl‐3‐imidazolin‐1‐yloxy‐3‐oxide) and tempol (4‐hydroxy‐2,2,6,6‐tetramethylpiperidine‐N‐oxyl).

The rank order of potency in inhibiting the reduction of cytochrome c was: CuSO4MnCl2CuDIPSMnTMPyP>MnTBAP>tempoltiron>PTIYO.

The requirement for EDTA (0.1 mM) prevented assessment of the activity of CuSO4, MnCl2 and CuDIPS in the assay involving inhibition of reduction of nitro blue tetrazolium. However, the rank order of potency for those agents which could be examined (MnTMPyP>MnTBAP>tirontempol>PTIYO) was essentially similar to that seen in the cytochrome c assay.

Inhibition of endogenous Cu/Zn SOD with diethyldithiocarbamate (DETCA, 3 mM, 120 min) in BRP muscle strips, followed by addition of the superoxide anion generator, LY 83583 (1 μM), resulted in almost complete abolition of nitrergic relaxation (4 Hz, 10 s).

Authentic Cu/Zn SOD (1–300 u ml−1), CuSO4 (0.1–300 μM), MnCl2 (0.1–100 μM) and MnTMPyP (10–300 μM) each restored nitrergic transmission by around 50%. However, CuDIPS (0.1–30 μM), MnTBAP (0.1–100 μM), tempol (10 μM–3 mM), PTIYO (1–300 μM) and tiron (10 μM–10 mM) all failed to restore nitrergic transmission.

The ability of MnTMPyP to restore nitrergic neurotransmission may therefore provide a lead in the development of SOD mimetics as therapeutic agents in the treatment of neuropathies associated with oxidant stress.
Different actions of CCK on pancreatic and gastric growth in the rat: effect of CCKA receptor blockade
Authors: Varga Gábor , Kisfalvi Krisztina , Pelosini Iva , D'Amato Massimo , Scarpignato Carmelo
Published: January 2005

It is now well established that cholecystokinin (CCK) has a major physiological role in the regulation of pancreatic secretion and gastro‐intestinal (GI) motility. Both these actions are mediated by stimulation of CCKA‐receptors located on pancreatic acini and GI smooth muscle cells. While chronic administration of CCK‐like peptides invariably causes pancreatic hypertrophy and hyperplasia, their action on gastric growth remains controversial.

In the present investigation the action of exogenous and endogenous CCK on both pancreatic and gastric growth was studied in the same animal. In addition, the ability of dexloxiglumide, a new potent and selective CCKA‐receptor antagonist, to counteract CCK‐mediated effects was evaluated.

The amphibian peptide caerulein (1 μg kg−1 intraperitoneally three times daily) was used as a CCK agonist, while camostate (200 mg kg−1 intragastrically once daily), a synthetic protease inhibitor, was used to release endogenous CCK. They were administered to rats for seven days with or without dexloxiglumide (25 mg kg−1 subcutaneously 15 min before the stimulus). On the eighth day, animals were killed, the pancreas and stomach excised, weighed, homogenized and their protein and DNA content measured.

Both exogenous and endogenous CCK increased the weight of the pancreas as well as the total pancreatic protein and DNA content. Dexloxiglumide, which alone did not affect pancreatic size and composition, was able to counteract both caerulein‐ and camostate‐induced pancreatic changes. Neither stimuli affected gastric growth in respect of weight and composition of the oxyntic gland area and the antrum.

These results show different effects of CCK on pancreatic and gastric growth. The CCK‐induced pancreatic hypertrophy and hyperplasia are blocked by the potent and specific CCKA‐receptor antagonist, dexloxiglumide. This compound therefore represents a useful tool to investigate CCK‐receptor interactions in peripheral organs.
Pharmacological characterization of a nonpeptide bradykinin B2 receptor antagonist, FR165649, and agonist, FR190997
Authors: Asano Masayuki , Hatori Chie , Sawai Hiroe , Johki Shigeru , Inamura Noriaki , Kayakiri Hiroshi , Satoh Shigeki , Abe Yoshito , Inoue Takayuki , Sawada Yuki , Mizutani Tsuyoshi , Oku Teruo , Nakahara Kunio
Published: January 2005

The nonpeptide bradykinin (BK) B2 receptor antagonist, FR165649 (8‐[2,6‐dichloro‐3‐[N‐[(E)‐4‐(N‐ methylcarbamoyl)cinnamidoacetyl] ‐N‐methylamino] benzyloxy] ‐2 ‐ methylquinoline), and agonist, FR190997 (8‐[2,6‐dichloro‐3‐[N‐[(E)‐4‐(N‐methylcarbamoyl) cinnamidoacetyl]‐N‐methylamino]benzyloxy]‐2‐methyl‐4‐(2‐pyridylmethoxy)quinoline) have been identified. These compounds have a common chemical structure, and the 2‐pyridylmethoxy group is the only structural difference between them.

Both FR165649 and FR190997 displaced [3H]‐BK binding to B2 receptors in guinea‐pig ileum membranes, with an IC50 of 4.7×10−10 M and 1.5×10−9 M, respectively. They also displaced [3H]‐BK binding to B2 receptors in human lung fibroblast IMR‐90 cells, with an IC50 of 1.6×10−9 M and 9.8×10−10 M, respectively.

In guinea‐pig isolated ileum‐preparations, FR165649 had no agonistic effect on contraction and caused parallel rightward shifts of the concentration‐response curves to BK on contraction. Analysis of the data produced a nominal pA2 value of 9.2±0.1 (n=5) and a slope of 1.4±0.1 (n=5). On the other hand, FR190997 induced concentration‐dependent contraction of guinea‐pig ilea with a pD2 of 7.9±0.2 and the contraction was inhibited by a specific peptide bradykinin B2 receptor antagonist, Hoe 140 (D‐Arg‐[Hyp3, Thi5, D‐Tic7, Oic8]BK) in a non‐competitive manner.

In IMR‐90 cells, FR165649 had no agonistic effect on phosphatidyl inositol (PI) hydrolysis and caused parallel rightward shifts (approximately 200 fold shift at 10−7 M) of the concentration‐response curves to BK on PI hydrolysis. FR190997 induced concentration‐dependent PI hydrolysis in IMR‐90 cells with a pD2 of 8.4±0.1, and this effect was inhibited by Hoe 140.

These results indicate that FR165649 and FR190997 are, respectively, a potent bradykinin B2 receptor antagonist and agonist, and that the agonistic activity depends on the small part of the nonpeptide ligand. FR165649 and FR190997 may be useful tools for studying the relationship between ligands and receptors.
Interaction between superoxide anion and nitric oxide in the regulation of vascular endothelial function
Authors: Laight David W , Kaw Amar V , Carrier Martin J , Änggård Erik E
Published: January 2005

Nitric oxide (NO)‐mediated, endothelium‐dependent vasodilator function in rat aortic smooth muscle was investigated in an in vitro model of endogenous vascular superoxide anion stress, generated by pretreatment with the Cu/Zn superoxide dismutase (SOD, EC 1.15.1.1) inhibitor, diethyldithiocarbamate (DETCA).

Contraction to noradrenaline (NA, 1 nM–1 μM) in endothelium‐intact vessels was augmented after a 30 min pretreatment with DETCA (10 mM) followed by 30 min washout. This effect was abolished by NG‐nitro‐L‐arginine methyl ester (L‐NAME, 0.3 mM) and removal of the endothelium and partially reversed by exogenous Cu/Zn SOD (200 u ml−1).

Endothelium‐ and basal NO‐dependent vasorelaxation to the phosphodiesterase (PDE) type V inhibitor ONO‐1505 (4‐[2‐(2‐hydroxyethoxy)ethylamino]‐2‐(1H‐imidazol‐1‐yl)‐6‐methoxyquinazoline methanesulphonate) (0.1–10 μM) was inhibited after DETCA (10 mM) pretreatment. In addition, the ability of L‐NAME (0.3 mM) to enhance established contractile tone was effectively absent.

In contrast, DETCA pretreatment did not significantly affect vasorelaxation to acetylcholine (ACh, 1 nM–3 μM) or S‐nitroso‐N‐acetyl penicillamine (SNAP, 0.03–30 μM). However, L‐NAME (0.3 mM) unmasked an inhibitory effect of DETCA pretreatment on vasorelaxation to SNAP in endothelium‐intact vessels while markedly potentiating vasorelaxation to SNAP in control tissue.

L‐NAME (0.3 mM)‐ and exogenous catalase (200 u ml−1)‐sensitive vasorelaxation to exogenous Cu/Zn SOD (200 u ml−1) was greater after DETCA (10 mM) pretreatment in endothelium‐intact aortic rings. This difference was abolished by catalase (200 u ml−1).

In conclusion, tissue Cu/Zn SOD inhibition elicited a selective lesion in basal endothelial function in rat isolated aortic smooth muscle, consistent with the inactivation of basal NO by superoxide anion. The resulting leftward shift in nitrovasodilator reactivity, due to the loss of the tonic depression by basal NO, is likely to mask the inhibitory effect of superoxide anion on agonist‐stimulated endothelial function and nitrovasodilator‐derived NO, thereby accounting for the differential pattern of endothelial dysfunction after DETCA pretreatment.
Bladder distension and activation of the efferent function of sensory fibres: similarities with the effect of capsaicin
Authors: Lecci A , Giuliani S , Tramontana M , Santicioli P , Criscuoli M , Dion S , Maggi C A
Published: January 2005

The effects of the tachykinin NK2 receptor antagonist MEN 11420 (100 nmol kg−1, i.v.) and isoprenaline (400 nmol kg−1, i.v.) were compared in a model of distension‐induced bladder activity in isovolumetric conditions. MEN 11420 induced a relaxation of the basal tone of the urinary bladder that was dependent on the volume of the viscus: the effect was absent at low volumes (0.2 and 0.5 ml) and it was maximal at high volumes of distension (1 and 2 ml), approaching about 60% of the isoprenaline‐induced relaxation. The relaxant effect of isoprenaline was always evident at all volumes of distension.

Tetrodotoxin (1–100 μM, intravesically applied) abolished distension‐evoked micturition contractions, but did not prevent the relaxant effect of MEN 11420‐ or isoprenaline on the bladder tone.

The cyclo‐oxygenase inhibitor S‐ketoprofen (0.5 μmol kg−1, i.v.) produced a marked decrease of the bladder tone and a concomitant reduction of bladder motility at 1 ml volume of distension. At 2 ml of distension, S‐ketoprofen still decreased the minimal pressure but had no significant effect on other parameters of vesical motility. In S‐ketoprofen‐pretreated rats, the relaxant effect of MEN 11420 was significant at 2 but not at 1 ml of distension, and that of isoprenaline was reduced by 50% at both 1 and 2 ml.

Ruthenium red (10 μmol kg−1, i.v.) had no effect at a low volume of distension (0.2 ml) or at highest volume (2 ml) but decreased the basal tone and the frequency of bladder contractions at 1 ml of distension. In ruthenium red‐pretreated rats, MEN 11420 failed to decrease bladder tone at 1 ml, whereas at 2 ml the effect of MEN 11420 was not different from that observed in controls (43 vs 60% of isoprenaline‐induced relaxation, respectively).

At both 1 and 2 ml of distension, capsaicin pretreatment (164 μmol kg−1, s.c. 5 days before) reduced the frequency of micturition contractions but had no effect on the bladder tone. Capsaicin pretreatment prevented the relaxant effect of MEN 11420 on the bladder tone both at 1 and at 2 ml of distension.

It is concluded that the release of tachykinins from capsaicin‐sensitive afferent nerves induced by bladder distension is resistant to tetrodotoxin and to prostaglandin synthesis inhibition. Tachykinins modulate the vesical tone by acting through NK2 receptors.
Effects of tachykinin NK1 receptor antagonists on vagal hyperreactivity and neuronal M2 muscarinic receptor function in antigen challenged guinea‐pigs
Authors: Costello Richard W , Fryer Allison D , Belmonte Kristen E , Jacoby David B
Published: January 2005

The role of tachykinin NK1 receptors in the recruitment of eosinophils to airway nerves, loss of inhibitory neuronal M2 muscarinic receptor function and the development of vagal hyperreactivity was tested in antigen‐challenged guinea‐pigs.

In anaesthetized guinea‐pigs, the muscarinic agonist, pilocarpine (1–100 μg kg−1, i.v), inhibited vagally induced bronchoconstriction, in control, but not in antigen‐challenged guinea‐pigs 24 h after antigen challenge. This indicates normal function of neuronal M2 muscarinic receptors in controls and loss of neuronal M2 receptor function in challenged guinea‐pigs. Pretreatment of sensitized guinea‐pigs with the NK1 receptor antagonists CP99994 (4 mg kg−1, i.p.), SR140333 (1 mg kg−1, s.c.) or CP96345 (15 mg kg−1, i.p.) before antigen challenge, prevented M2 receptor dysfunction.

Neither administration of the NK1 antagonists after antigen challenge, nor pretreatment with an NK2 receptor antagonist, MEN10376 (5 μmol kg−1, i.p.), before antigen challenge, prevented M2 receptor dysfunction.

Electrical stimulation of the vagus nerves caused a frequency‐dependent (2–15 Hz, 10 V, 0.2 ms for 5 s) bronchoconstriction that was significantly increased following antigen challenge. Pretreatment with the NK1 receptor antagonists CP99994 or SR140333 before challenge prevented this increase.

Histamine (1–20 nmol kg−1, i.v.) caused a dose‐dependent bronchoconstriction, which was vagally mediated, and was significantly increased in antigen challenged guinea‐pigs compared to controls. Pretreatment of sensitized animals with CP99994 before challenge prevented the increase in histamine‐induced reactivity.

Bronchoalveolar lavage and histological studies showed that after antigen challenge significant numbers of eosinophils accumulated in the airways and around airway nerves. This eosinophilia was not altered by pretreatment with the NK1 receptor antagonist CP99994.

These data indicate that pretreatment of antigen‐sensitized guinea‐pigs with NK1, but not with NK2 receptor antagonists before antigen challenge prevented the development of hyperreactivity by protecting neuronal M2 receptor function. NK1 receptor antagonists do not inhibit eosinophil accumulation around airway nerves.
Dual effects of dichloroacetate on cardiac ischaemic preconditioning in the rat isolated perfused heart
Authors: Randall Michael D , Keon Claudia A , Greenhaff Paul L , Constantin‐Teodosiu Dumitru
Published: January 2005

Ischaemic cardiac preconditioning represents an important cardioprotective mechanism which limits myocardial ischaemic damage. The aim of this investigation was to assess the impact of dichloroacetate (DCA), a pyruvate dehydrogenase complex activator, on preconditioning.

Rat isolated hearts were perfused by use of the Langendorff technique, and were subjected to either preconditioning (3×4 or 3×6 min ischaemia) or continuous perfusion, followed by 30 min global ischaemia and 60 min reperfusion. DCA (3 mM) was either given throughout the protocol (pretreatment), during reperfusion only (post‐treatment), or not at all. Throughout reperfusion mechanical performance was assessed as the rate‐pressure product (RPP: left ventricular developed pressure×heart rate).

In non‐preconditioned control hearts, mechanical performance was substantially (P<0.001) depressed on reperfusion (the RPP after 60 min of reperfusion (RPPt=60) was 4,246±974 mmHg beats min−1 compared to baseline value of 21,297±1,728 mmHg beats min−1). Preconditioning with either 3×4 min or 3×6 min cycles caused significant protection, as shown by enhanced recovery (RPPt=60=7,818±1,138, P<0.05, and 11,123±587 mmHg beats min−1, P<0.001, respectively).

Addition of DCA (3 mM) to hearts under baseline conditions significantly (P<0.001) enhanced systolic function with an increased left ventricular developed pressure of 108±5 mmHg compared to 88.3±3.0 mmHg in the controls.

Pretreatment with 3 mM DCA had no effect on recovery of mechanical performance in the non‐preconditioned hearts (RPPt=60=3,640±1,235 mmHg beats min−1) while the beneficial effects of preconditioning were reduced in the preconditioned hearts (3×4 min: RPPt=60=2,919±1,060 mmHg beats min−1; 3×6 min: RPPt=60=8,032±1,367 mmHg beats min−1). Therefore, DCA had increased the threshold for preconditioning.

By contrast, post‐treatment of hearts with 3 mM DCA substantially improved recovery on reperfusion in all groups (RPPt=60=5,827±1,328 (non‐preconditioned), 14,022±3,743 (3×4 min; P<0.01) and 23,219±1,374 (3×6 min; P<0.001) mmHg beats min−1).

The results of the present investigation clearly show that pretreatment with DCA enhances baseline cardiac mechanical performance but increases the threshold for cardiac preconditioning. However, post‐treatment with DCA substantially augments the beneficial effects of preconditioning.
Inhibition of nitric oxide generation unmasks vascular dysfunction in insulin‐resistant, obese JCR:LA‐cp rats
Authors: McKendrick Joyce D , Salas Eduardo , Dubé Gregory P , Murat Jesus , Russell James C , Radomski Marek W
Published: January 2005

The effects of nitric oxide (NO) on vascular reactivity and platelet function in the obese (cp/cp) and lean (+/?) JCR:LA‐cp rats were investigated.

Phenylephrine (PE; 0.1 nM–10 μM) induced contraction of isolated aortic rings in both genotypes (cp/cp and +/?) of JCR:LA‐cp rats. The sensitivity to contraction with PE was enhanced in cp/cp compared with +/? rings. Rings from both genotypes showed an increased contraction upon removal of the endothelium.

Acetylcholine (ACh; 0.1 nM–10 μM)‐induced endothelium‐dependent relaxation of rings was not significantly different in the two genotypes. Both were inhibited to a similar extent by NG‐nitro‐L‐arginine methyl ester (L‐NAME; 0.01–1 mM) when administered in vitro.

The nitric oxide synthase (NOS) inhibitor (L‐NAME; 0.3, 1 or 3 mg ml−1, p.o.) when administered in vivo increased blood pressure in cp/cp rats but not in +/? rats.

L‐NAME resulted in greater inhibition of ACh‐induced relaxation in cp/cp rings compared with +/? rings.

L‐NAME treatment in vivo caused a decrease in cyclic GMP and NOS activity in rings from cp/cp but not +/? rats.

The NO donor, S‐nitroso‐N‐acetyl‐DL‐penicillamine (SNAP; 0.1 nM–10 μM)‐induced relaxation of rings from +/? rats, an effect enhanced by the treatment with L‐NAME in vivo.

Oral administration of L‐NAME did not enhance the vasorelaxant effect of SNAP on rings of aorta from cp/cp animals.

Platelet aggregation and NOS activity were similar in both genotypes and were not modified by oral administration of L‐NAME.

These results show that unimpaired generation of NO is crucial for maintenance of vascular tone particularly under conditions of vascular insult exemplified by insulin resistance, obesity and dyslipidemia detected in cp/cp rats.
Characterization of the phencyclidine‐induced increase in prefrontal cortical dopamine metabolism in the rat
Authors: Umino Asami , Takahashi Kiyohisa , Nishikawa Toru
Published: January 2005

We have investigated the effects of a schizophrenomimetic drug phencyclidine (PCP) and N‐methyl‐D‐aspartate (NMDA)‐related agents alone or in combination on dopamine metabolism in the medial prefrontal cortex and striatum of the rats by measuring the tissue concentrations of dopamine and its metabolite, 3,4‐dihyroxyphenylacetic acid (DOPAC), and the rate of dopamine disappearance (dopamine utilization) after its synthesis inhibition.

Systemic injection of PCP and selective, non‐competitive, NMDA antagonists caused an increase of both tissue concentrations of DOPAC and dopamine utilization in the prefrontal cortex but not in the striatum. The PCP‐induced augmentation of cortical dopamine metabolism was not influenced by selective lesion of ascending noradrenergic neurones.

Intra‐prefrontal cortical infusion of PCP or selective competitive or non‐competitive antagonists of the NMDA receptor mimicked the ability of systemic PCP injection to enhance DOPAC levels and dopamine utilization in the prefrontal cortex. However, an NMDA antagonist injected into the cell body area of the mesocortical dopaminergic neurones failed to affect dopamine metabolism in the prefrontal cortex.

The increasing effects of PCP and selective NMDA antagonists on cortical dopamine utilization were not additive, although a dopamine receptor antagonist, haloperidol, still accelerated the disappearance of dopamine, even in the presence of PCP.

Intra‐cortical or intra‐ventricular infusion of NMDA or D‐alanine but not L‐alanine, attenuated the ability of systemic PCP administration to facilitate prefrontal dopamine utilization.

These data suggest that PCP might activate prefrontal cortical dopaminergic neurones, at least in part, by blocking the NMDA receptor in the prefrontal cortex which participates in a tonic inhibitory control of the mesoprefrontal dopaminergic projections.
Effects of pravastatin on cholesterol metabolism of cholesterol‐fed heterozygous WHHL rabbits
Authors: Harsch Michael , Gebhardt Angelika , Reymann Andreas , Lang Gerhard , Schliack Michael , Löser Roland , Hinrich Braesen Jan , Niendorf Axel
Published: January 2005

We administered the 3‐hydroxy‐3‐methylglutaryl coenzyme A reductase inhibitor pravastatin at a daily dose of 1 mg kg−1 body weight to cholesterol‐fed (0.03%) heterozygous Watanabe heritable hyperlipidaemic rabbits, an animal model for heterozygous familial hypercholesterolaemia.

After 12 months of cholesterol treatment, immunohistochemistry with the monoclonal antibody 9D9 was used to detect hepatic low density lipoprotein (LDL) receptors, which were quantified by densitometry. In addition we determined LDL receptor mRNA by competitive reverse transcriptase polymerase chain reaction. The cholesterol precursor lathosterol and the plant sterol campesterol were analysed by gas‐liquid chromatography.

The drug reduced total plasma cholesterol levels by 51% (P=0.04), when compared to the control group. Unexpectedly, hepatic LDL receptor density and mRNA showed no significant differences between the groups. Total plasma levels of lathosterol and campesterol also revealed no significant differences between the groups, if expressed relative to plasma cholesterol.

The findings suggest that mechanisms other than induced hepatic LDL receptors are responsible for the cholesterol‐lowering effect of pravastatin in this animal model. We propose a reduced cholesterol absorption efficiency compatible with similar campesterol levels between both groups observed in our study.
Modulation of extracellular GABA levels in the retina by activation of glial P2X‐purinoceptors
Authors: Neal M J , Cunningham J R , Dent Z
Published: January 2005

In the rat retina, γ‐aminobutyric acid (GABA) released as a transmitter is inactivated by uptake mainly into glial cells (Müller cells). Activation of P2‐purinoceptors in Müller cells increases [Ca2+]i and the present study was undertaken to see whether this action affected the glial release of [3H]‐GABA from the superfused rat isolated retina.

Adenosine 5′‐triphosphate (ATP) and the P2X‐purinoceptor agonists, α,β‐methylene‐ATP (α,β‐meATP) and β,γ‐methyleneATP (β,γ‐meATP) significantly increased the KCl‐evoked release of [3H]‐GABA from the retina.

Adenosine and the P2Y‐purinoceptor agonist, 2‐chloroATP, had no effect on the KCl‐evoked release of [3H]‐GABA from the retina. However, 2‐methylthioATP (2‐Me‐S‐ATP) significantly enhanced the evoked release of [3H]‐GABA.

The effect of ATP on the glial release of [3H]‐GABA was abolished by the P2‐antagonist, pyridoxalphosphate‐6‐azophenyl‐2′,4′‐disulphonic acid (PPADS).

When the superfused retina was exposed to the GABA uptake inhibitor, SKF89976A, the enhancing effect of α,β‐meATP on the KCl‐evoked release of GABA was abolished.

The KCl‐evoked release of [3H]‐GABA from the frog retina and rat cerebrocortical slices, which take up GABA mainly into neurones, was not affected by ATP or α,β‐meATP.

We concluded that the glial Müller cells in the rat retina possess P2‐receptors, activation of which increases the ‘release’ of preloaded [3H]‐GABA apparently by reducing uptake. On balance, the results suggest the involvement of P2X‐purinoceptors, although we cannot exclude the possibility that P2Y‐purinoceptors may be involved. Our results suggest that ATP, as well as being a conventional transmitter in the retina, may be involved in neuronal‐glial signalling and modulate the extracellular concentration of GABA.
Differential effects of somatostatin and angiopeptin on cell proliferation
Authors: Alderton Forbes , Lauder Heather , Feniuk Wasyl , Fan Tai‐Ping D , Humphrey Patrick P A
Published: January 2005

Somatostatin (SRIF) exerts antiproliferative effects, and angiopeptin (an sst2/sst5 receptor‐selective analogue) has recently been evaluated in clinical trials for the prophylaxis of restenosis following coronary angioplasty. Using an in vitro model of cell growth we have examined the effects of SRIF and angiopeptin on cell proliferation in CHO‐K1 cells stably transfected with the human or rat recombinant sst2 or sst5 receptor and compared these with their effects on rat aortic vascular smooth muscle cells (VSMC) expressing endogenous somatostatin receptors.

In CHO‐K1 cells, expressing either human or rat recombinant sst2 or sst5 receptors, or in rat aortic VSMC, SRIF and angiopeptin (0.1–1000 nM) had no effect on basal re‐growth of cells into a denuded area of a previously confluent monolayer. In contrast, basic fibroblast growth factor (bFGF, 10 ng ml−1) stimulated re‐growth of these cells.

SRIF (0.1–1000 nM) caused a concentration‐dependent inhibition of the bFGF‐stimulated re‐growth in CHO‐K1 cells expressing human sst2 (h sst2) or sst5 (h sst5) receptors (pIC50=8.05±0.03 and 8.56±0.12, respectively). In contrast, angiopeptin (0.1–1000 nM) acted as a partial agonist at the h sst2 receptor (44.6±2.7% inhibition of the bFGF‐stimulated re‐growth at 100 nM; pIC50=8.69±0.25) but was devoid of any agonist activity at the h sst5 receptor.

In CHO‐K1 cells stably expressing rat recombinant sst2 (r sst2) or sst5 (r sst5) receptors, SRIF (0.1–1000 nM) was able to inhibit the bFGF‐stimulated re‐growth (pIC50=7.98±24 and 8.50±0.12, respectively). Angiopeptin (0.1–1000 nM) caused a concentration‐dependent inhibition of bFGF‐stimulated re‐growth at the r sst2 receptor (pIC50=8.08±0.24) but acted as a partial agonist at the r sst5 receptor (maximum response=57.7±3.6% inhibition of bFGF‐stimulated re‐growth at 100 nM; pIC50=8.60±0.16).

Although angiopeptin was inactive as an agonist at the h sst5 receptor, 100 nM angiopeptin potently antagonized the SRIF‐induced inhibition of proliferation in CHO h sst5 (estimated pKB=10.4±0.3). 5‐Hydroxytryptamine (0.1 nM–10 μM) also inhibited bFGF‐stimulated re‐growth (pIC50=8.36±0.11) and angiopeptin had no effect on this response (pKB<7).

SRIF (0.1–1000 nM) caused a concentration‐dependent (pIC50=8.04±0.08) inhibition of bFGF‐stimulated re‐growth in VSMC, whereas angiopeptin displayed weak agonist activity, only inhibiting bFGF‐stimulated re‐growth at concentrations greater than 100 nM. Angiopeptin (100 nM) caused a rightward displacement of the concentration‐effect curve to SRIF with an estimated pKB value of 7.70±0.12.

These findings suggest that the low intrinsic activity of angiopeptin at the h sst2 receptor, combined with its lack of agonist activity at the h sst5 receptor, may explain the poor clinical efficacy of angiopeptin in trials for coronary artery restenosis, which contrasts with encouraging data found in equivalent in vivo animal studies.
Hepatobiliary and intestinal clearance of amphiphilic cationic drugs in mice in which both mdr1a and mdr1b genes have been disrupted
Authors: Smit Johan W , Schinkel Alfred H , Weert Betty , Meijer Dirk K F
Published: January 2005

We have used mice with homozygously disrupted mdr1a and mdr1b genes (mdr1a/1b (−/−) mice) to study the role of the mdr1‐type P‐glycoprotein (P‐gp) in the elimination of cationic amphiphilic compounds from the body. These mice lack drug‐transporting P‐gps, but show no physiological abnormalities under laboratory conditions and have normal bile flow.

3H‐labelled cationic drugs were administered intravenously (i.v.) to mice as a single bolus dose and the disposition of the studied cationic drugs was investigated by focusing on drug secretion into bile, intestinal lumen and urine.

Hepatobiliary secretion of the investigated cationic drugs was profoundly reduced in mice devoid of the mdr1‐type P‐gps. In fact, the cumulative biliary output, measured during 1 h, of the small type 1 compounds tri‐butylmethyl ammonium (TBuMA) and azidoprocainamide methoiodide (APM), as well as of the more bulky type 2 cationic drug vecuronium, was reduced by at least 70% in the mdr1a/1b (−/−) mice compared to wild‐type.

The intestinal secretion of TBuMA, APM and vecuronium was also profoundly reduced in mdr1a/1b (−/−) mice compared to wild‐type mice. The absence of the mdr1‐type P‐gp resulted in virtual elimination of intestinal secretion of TBuMA and APM (>90% reduced as compared to wild‐type (P=0.0001 and 0.0022, respectively)). The intestinal secretion of the type 2 cation drug vecuronium was reduced by 58% (P=0.0004) compared to the wild‐type mice.

Increased renal clearances of both the type 1 compounds TBuMA and APM and also of the type 2 cationic compound vecuronium in the mdr1a/1b (−/−) mice were observed. Furthermore, the balance between hepatic, intestinal and renal clearances of small type 1 organic cations clearly shifted towards a predominant role for renal clearance. Increased renal clearance may be explained by (over)expresion of additional mechanisms for renal organic cation secretion, alternatively they may also point to an as yet undefined role of P‐glycoprotein in kidney physiology and renal secretory function.

We conclude that the elimination from the body of a broad spectrum of cationic amphiphilic drugs via liver and intestine, is largely dictated by the activity of mdr1‐type P‐glycoproteins.
Role of reactive oxygen species in the signalling cascade of cyclosporine A‐mediated up‐regulation of eNOS in vascular endothelial cells
Authors: López‐Ongil S , Hernández‐Perera O , Navarro‐Antolín J , Pérez de Lema G , Rodríguez‐Puyol M , Lamas S , Rodríguez‐Puyol D
Published: January 2005

Cyclosporine A (CsA) increases eNOS mRNA expression in bovine cultured aortic endothelial cells (BAEC). As some effects of CsA may be mediated by reactive oxygen species (ROS), present experiments were devoted to test the hypothesis that the CsA‐induced eNOS up‐regulation could be dependent on an increased synthesis of ROS.

CsA induced a dose‐dependent increase of ROS synthesis, with the two fluorescent probes used, DHR123 (CsA 1 μM: 305±7% over control) and H2DCFDA (CsA 1 μM: 178±6% over control).

Two ROS generating systems, xanthine plus xanthine oxidase (XXO) and glucose oxidase (GO), increased the expression of eNOS mRNA in BAEC, an effect which was maximal after 8 h of incubation (XXO: 168±21% of control values. GO: 208±18% of control values). The ROS‐dependent increased eNOS mRNA expression was followed by an increase in eNOS activity.

The effect of CsA on eNOS mRNA expression was abrogated by catalase, and superoxide dismutase (SOD). In contrast, the antioxidant PDTC augmented eNOS mRNA expression, both in basal conditions and in the presence of CsA.

The potential participation of the transcription factor AP‐1 was explored. Electrophoretic mobility shift assays were consistent with an increase in AP‐1 DNA‐binding activity in BAEC treated with CsA or glucose oxidase.

The present results support a role for ROS, particularly superoxide anion and hydrogen peroxide, as mediators of the CsA‐induced eNOS mRNA up‐regulation. Furthermore, they situate ROS as potential regulators of gene expression in endothelial cells, both in physiological and pathophysiological situations.
P2X receptors in cochlear Deiters' cells
Authors: Chen Chu , Bobbin Richard P
Published: January 2005

The ionotropic purinoceptors in isolated Deiters' cells of guinea‐pig cochlea were characterized by use of the whole‐cell variant of the patch‐clamp technique.

Extracellular application of adenosine 5′‐triphosphate (ATP) induced a dose‐dependent inward current when the cells were voltage‐clamped at −80 mV. The ATP‐induced current showed desensitization and had a reversal potential around −4 mV.

Increasing intracellular free Ca2+ by decreasing the concentration of EGTA in the pipette solution reduced the amplitude of the ATP‐gated current.

The order of agonist potency was: 2‐methylthioATP (2‐meSATP)>ATP>benzoylbenzoyl‐ATP (BzATP)>α,β‐methyleneATP (α,β,meATP>adenosine 5′‐diphosphate (ADP)>uridine 5′‐triphosphate (UTP)>adenosine 5′‐monophosphate (AMP)=adenosine (Ad).

Pretreatment with forskolin (10 μM), 8‐bromoadenosine‐3′,5′‐cyclophosphate (8‐Br‐cyclic AMP, 1 mM), 3‐isobutyl‐1‐methylxanthine (IBMX, 1 mM) or phorbol‐12‐myristate‐13‐acetate (PMA, 1 μM) reversibly reduced the ATP‐induced peak current.

The results are consistent with molecular biological data which indicate that P2X2 purinoceptors are present in Deiters' cells. In addition, the reduction of the ATP‐gated current by activators of protein kinase A and protein kinase C indicates that these P2X2 purinoceptors can be functionally modulated by receptor phosphorylation.
Activation of midbrain presumed dopaminergic neurones by muscarinic cholinergic receptors: an in vivo electrophysiological study in the rat
Authors: Gronier B , Rasmussen K
Published: January 2005

Extracellular single‐unit recording and iontophoresis were used to examine the effects of different cholinoceptor agonists and antagonists on the firing rate and firing pattern of A9 and A10 presumed dopaminergic neurones in the anaesthetized rat.

Administration of low currents (1–5 nA) of the selective muscarinic agonists oxotremorine M (Oxo M) and muscarine and of the non‐selective muscarinic/nicotinic agonist carbamylcholine (CCh) produced a dose‐dependent increase in firing rate in most of the A9 and A10 presumed dopaminergic neurones tested. Oxo M‐induced activation could be completely blocked by iontophoretic application of the muscarinic antagonist butyl‐scopolamine or systemic administration of the muscarinic antagonist scopolamine (300 μg kg−1, i.v.).

Iontophoretic application of the selective nicotinic agonist methylcarbamylcholine (MCCh), but not nicotine, induced a consistent increase in firing rate. Surprisingly, the excitatory effect of MCCh was significantly reduced by the selective muscarinic antagonist scopolamine (300 μg kg−1, i.v.), but not by the selective nicotinic antagonist mecamylamine (2.2 mg kg−1, i.v.). Mecamylamine (3 mg kg−1, i.v.) was also ineffective in reducing the CCh‐induced activation of presumed dopamine neurones, suggesting that both CCh and MCCh increased the activity of dopamine neurones via an interaction with muscarinic receptors.

Iontophoretic application of the endogenous agonist acetylcholine (ACh) had no or little effect on the firing activity of A10 presumed dopaminergic neurones. However, concomitant application of neostigmine, a potent cholinesterase inhibitor, with acetylcholine induced a substantial activation of these neurones. This activation consisted of two components; one, which was prevalent, was scopolamine (300 μg kg−1, i.v.)‐sensitive, and the other was mecamylamine (2 mg kg−1, i.v.)‐sensitive.

In addition to their effect on firing activity, Oxo M, muscarine and concomitant neostigmine/ACh caused a significant increase in burst firing of A10 neurones, but not of A9 neurones.

These data suggest that dopamine cells, both in the A9 and A10 regions, possess functional muscarinic receptors, the activation of which can increase their firing rate and, for A10 neurones, their amount of burst activity. These cholinoceptors would be able to influence the activity of the midbrain dopamine system greatly and may play a role in, and/or be a therapeutic target for, brain disorders in which dopamine is involved (e.g., Parkinson's disease, drug addiction and schizophrenia).