The effect of neostigmine on metoclopramide-induced aldosterone secretion after the administration of muscarinic antagonists in man

Summary In this study the effects of neostigmine on metoclopramide-induced aldosterone secretion were examined in the presence of a relatively selective M₁-antagonist, pirenzepine and of a non-selective muscarinic antagonist, atropine.Six normal male volunteers received metoclopramide, 10 mg i.v. on three different occasions, each study day being preceded by a day in which the intake of sodium and potassium was limited. The dosing was either metoclopramide alone or combined with either neostigmine and pirenzepine or with neostigmine and atropine.Serum aldosterone increased significantly with all three regimens. The highest levels were attained with the metoclopramide/neostigmine/prienzepine regimen and those were significantly higher than those after metoclopramide alone and also, from 45 min onwards, from those after the metoclopramide/neostigmine/atropine regimen.The results of this investigation suggest that the metoclopramide-induced aldosterone secretion in humans is augmented by an accumulation of acetylcholine at the nerve-zona glomerulosa junctions and that the receptors mediating aldosterone secretion are of the M₂-subclass of muscarinic receptors.     

Dose-response curves of pirenzepine in man in relation to M1- and M2-cholinoceptor occupancy

Summary The aim of the present study was to investigate the M-cholinoceptor subtype selectivity of pirenzepine in man. In parallel with effects on the heart rate and salivary flow, M-cholinoceptor subtype occupancy by antagonist present in plasma samples was detected in radioreceptor assays. Bovine cerebral cortex membranes labelled with ³H-pirenzepine (M₁) and rat salivary gland membranes labelled with ³H-N-methylscopolamine (M₂) were used in these in vitro assays. A half-maximal occupancy of M₁-cholinoceptors in the in vitro assay of plasma samples was detected after 0.25 mg of pirenzepine i.v. The respective half-maximal M₂-cholinoceptor occupancy was observed after 10 mg. Doses < 3 mg decreased the heart rate by maximally 10.7 beats/min with an ED₅₀ of about 0.1 mg. An increase in heart rate (relative to control values) was observed at doses > 10 mg. This bivalent dose-response relationship was also observed after -blockade. Salivary flow tended to increase at doses < 1 mg and was half-maximally inhibited after 10 mg. Combining the in vitro and in vivo results, the typical antimuscarinic effects (tachycardia and inhibition of salivary flow) can be attributed to the blockade of M₂-cholinoceptors, whereas the reduction of heart rate coincides with the blockade of the M₁-subtype. With respect to the typical antimuscarinic effects, pirenzepine was 70-fold less potent than atropine; in contrast, with respect to the reduction of heart rate, pirenzepine was equipotent with atropine. It is concluded that pirenzepine does not discriminate between cardiac and salivary gland M₂-cholinoceptors but shows pronounced selectivity for the M₁-cholinoceptors through which it mediates the decrease in heart rate. The latter effect may be explained by inhibitory M₁-auto-receptors. Send offprint requests to H. F. Pitschner at the above addressParts of the results were reported at the spring meeting 1987 of the Deutsche Gesellschaft für Pharmakologie und Toxikologie (Wellstein et al. 1987)     

Subclassification of muscarinic receptors in the heart,urinary bladder and sympathetic ganglia in the pithed rat

Summary In pithed normotensive rats muscarinic receptors were characterized heart, urinary bladder and sympathetic ganglia; the selectivity of some classical muscarinic agents for these subtypes was investigated. The potencies in decreasing heart rate, increasing bladder pressure and increasing diastolic blood pressure were measured for the following, intraarterially administered cholinergic agonists: McN-A-343 ([4-m-chlorophenylcarbamoyloxy]-2-butynyltrimethylammonium), pilocarpine, carbachol, oxotremorine, arecoline, acetyl--methylcholine and acetylcholine. The selective M₁-antagonist pirenzepine, the mixed M₁/M₂-antagonist dexetimide and the cardioselective M₂-antagonist gallamine were used as tools for identification of the receptors. All data were obtained after intravenous pretreatment with a high dose of atenolol to eliminate tachycardia induced by stimulating sympathetic ganglionic muscarinic receptors.Dexctimide strongly antagonized the bradycardia as well as the increase in bladder pressure induced by pilocarpine, carbachol, oxotremorine, arecoline, acetyl--methylcholine and acetylcholine, whereas pirenzepine was much less effective. Gallamine antagonized the bradycardia, whereas no influence was found on the bladder contraction. Pilocarpine acted as a partial agonist in reducing heart rate as well as in increasing bladder pressure, whereas McN-A-343 was almost ineffective in doses up to 1 mg/kg.The hypertensive response to pilocarpine and carbachol was less pronounced than that produced by McN-A-343. Pirenzepine and dexetimide significantly antagonized the hypertensive response to McN-A-343 and pilocarpine, whereas gallamine was much less effective. The hypertensive response induced by carbachol was totally blocked by hexamethonium. The other agonists used in this study did not produce a significant increase in diastolic blood pressure in doses that produced a maximal effect on heart rate and urinary bladder pressure.Simultaneously, intraarterially infused acetylcholine dose-dependently and reversibly decreased the pressor response to intravenously administered McN-A-343.These data suggest that muscarinic receptors in rat sympathetic ganglia belong to the M₁-subtype, whereas the muscarinic receptors in rat heart and urinary bladder represent heterogenous populations of M₂-receptors. The agonists used in this study, though, could not discriminate between these heterogenous M₂-receptors.Like McN-A-343, pilocarpine appears to be a rather selective M₁-agonist. In this study the M₁/M₂ selectivity of muscarinic agents with pronounced M₂-agonist activity could not be evaluated since M₂-receptor stimulation interferes with the hypertensive response to M₁-receptor stimulation.     

The effect of selective and non-selective cholinergic blockade on bombesin- and peptone-stimulated gastrin release

Summary We have studied the effects of the selective muscarinic M₁-receptor antagonist pirenzepine and the non-selective muscarinic antagonist atropine on bombesin- and peptone-stimulated gastrin release in healthy subjects.Pirenzepine (i. v. bolus 0.6 mg/kg) and atropine (i. v. bolus 15 µg/kg, followed by an infusion of 5 µg·kg^–1·h^–1) were given in doses equipotent in terms of reduction of gastric acid secretion. Neither affected bombesin- or peptone-stimulated gastrin release.These findings do not support the involvement of M₁-receptors in the cholinergic regulation of gastrin release and suggest that the reduction in acid secretion caused by pirenzepine is not mediated by inhibition of gastrin release.     

Comparison of the effects of atropine in vivo and ex vivo (radioreceptor assay) after oral and intramuscular administration to man

The effects of an oral dose of atropine (0.03 mg/kg body weight) and an IM (0.02 mg/kg) dose on the heart rate and salivary flow in seven healthy adult volunteers were compared to see whether the oral dose was sufficient to inhibit vagal reflexes of the heart. Atropine concentrations in plasma were determined by an M₂-selective radioreceptor assay, and the in vitro occupancy of porcine cardiac M₂-cholinoceptors was measured in parallel.In ligand-binding studies, atropine has been shown to have a comparable affinity for human and porcine cardiac M₂-cholinoceptors (K_i 4.0 and 5.9, respectively). Slight changes in heart rate after oral administration were not significant. After IM administration, however, the heart rate increased significantly, by a maximum of 22 beats·min^–1 after 45 min. The slight increase in heart rate after the oral dose corresponded to a receptor occupancy in vitro near the lower limit of detection, whereas the significant increase in heart rate after the IM dose corresponded to a receptor occupancy of up to 47%. The maximum reduction in salivary flow was similar after the oral and IM doses (84.3 and 87.5%, respectively).The almost complete inhibition of salivary flow could be explained by the lower vagal tone in the salivary glands compared with to the heart. The difference in the effect on heart rate was probably due to lower absorption of the oral dose. Thus, an oral dose greater than 0.03 mg atropine/kilogram body weight is required to compensate for low gastrointestinal absorption and to overcome the high vagal tone of the heart.The results form part of the thesis of T. Waldhäuser     

Cardiovascular effects of the adenosine A<Subscript>1</Subscript> receptor agonist<Emphasis Type="Italic"> N</Emphasis><Superscript>6</Superscript>-cyclopentyladenosine (CPA) decisive for its therapeutic efficacy in sarin poisoning

Mortality and occurrence of cholinergic symptoms upon sarin intoxication (144 µg/kg s.c., ~2×LD₅₀) in rats is completely prevented by treatment with the adenosine A₁ receptor agonist N⁶-cyclopentyladenosine (CPA, 2 mg/kg i.m.). Previously, we have shown that CPA treatment altered the distribution of sarin into the brain, presumably through its cardiovascular side effects. Therefore, the objective of the present study was to evaluate the contribution of the cardiodepressant effects of CPA to its therapeutic efficacy against sarin intoxication. Intramuscular treatment of rats with 0.5 and 2.0 mg/kg CPA 1 min after sarin poisoning attenuated most cholinergic symptoms and prevented mortality, which seemed to be directly associated with an immediate strong and long-lasting bradycardia and hypotension caused by CPA. Treatment with lower doses of CPA (0.1 and 0.05 mg/kg i.m.) caused similar levels of bradycardia and hypotension, albeit a few minutes later than at the higher doses of CPA. Upon sarin intoxication, this was correlated with increased incidence of cholinergic symptoms and decreased survival rates. Pretreatment with the peripheral adenosine A₁ receptor antagonist 8-p-sulphophenyltheophylline (8-PST, 20 mg/kg i.p.) counteracted the cardiodepressant effects of 0.05 mg/kg CPA almost completely, thereby nearly abolishing its therapeutic efficacy against sarin poisoning. In conclusion, the present results strongly indicate that bradycardia and hypotension induced by the peripheral adenosine A₁ receptor play a prominent role in the therapeutic efficacy of CPA in cases of sarin poisoning.     

Pharmacological analysis of the interaction of antimuscarinic drugs at M<Subscript>2</Subscript> and M<Subscript>3</Subscript> muscarinic receptors in vivo using the pithed rat assay

Muscarinic receptor antagonists form the mainstay of the therapeutic options for airway, bladder, and gastrointestinal smooth muscle disorders. Both M₂ and M₃ muscarinic receptors are involved in mediating smooth muscle contractility, although the relative functional contribution of each subtype, especially in the disease state, is unclear. Because the potency and selectivity of compounds for a given receptor in an in vivo setting can be dissimilar to that observed in an in vitro system, we developed an in vivo assay to simultaneously determine the absolute potency and selectivity of muscarinic receptor antagonists at M₂ and M₃ receptors using the pithed rat. Methacholine (MCh)-induced bradycardia and depressor responses were used as surrogate functional endpoints for M₂ and M₃ receptor activation, respectively. The influence of the muscarinic antagonists, tolterodine, oxybutynin, darifenacin, Ro 320-6206, solifenacin, or tiotropium on the MCh-induced responses were studied. The estimated DR₁₀ values (dose producing a tenfold shift in the MCh curve) of tolterodine, oxybutynin, darifenacin, Ro 320-6206, solifenacin, and tiotropium for the M₂ muscarinic receptor-mediated bradycardia were 0.22, 1.18, ∼2.6, 0.025, 0.40, and 0.0026 mg/kg, respectively, and 0.14, 0.18, 0.11, 3.0, 0.18, and 0.0017 mg/kg, respectively, for the M₃ muscarinic receptor-mediated depressor response. In a separate set of experiments, a single intravenous dose of tiotropium was administered before a MCh curve at 1, 3, 6, or 9 h to determine if tiotropium exhibited time-dependent selectivity for the M₃ receptor as has been reported from in vitro studies. The results indicate a slight preference of tiotropium for the M₃ receptor at later time points. The pithed rat assay may serve useful for elucidating the functional contribution of M₂ and M₃ receptors to the in vivo pharmacological effects of antagonists in disease animal models.     

Muscarinic M3 receptors mediate contractions in rabbit endothelium-denuded aorta in vitro

1 Muscarinic receptors mediating contraction of rabbit endothelium-denuded aorta have been characterized functionally, in vitro, using a range of antagonists (atropine, pirenzepine, methoctramine, himbacine, 4-diphenyl-acetoxy-N-methyl piperidine methiodide (4-DAMP) and para-fluoro-hexahydro-siladifenidol (p-F-HHSiD). 2 The non-selective muscarinic agonist, (+)cis-dioxolane, induced concentration-dependent contractions of endothelium-denuded aortic rings. The potency (EC₅₀) of (+)cis-dioxolane was 1.0 ± 0.4 μM and the maximal increase in isometric tension was 944 ± 98 mg (mean ± SEM, n= 25). The concentration–effect curves to (+)cis-dioxolane were shifted to the right in the presence of antagonists, in a concentration-dependent manner. The following affinities (– log K_B) were calculated; atropine, 9.4; pirenzepine, 6.6; methoctramine, 5.9; himbacine, 7.1; 4-DAMP, 9.2; and p-F-HHSiD, 7.7. 3 It is concluded that muscarinic M₃ receptors mediate contractions of endothelium-denuded aorta. The low potency of (+)cis-dioxolane, when compared to its potency in other M₃ receptor assays, suggests that the efficiency of receptor coupling, associated with contraction of this tissue, is poor.     

Characterization of the muscarinic receptor mediating contraction of the dog prostate

1 We have studied the effects of muscarinic cholinoceptor agonists and subtype-preferring antagonists on the isometric contraction of smooth muscle strips from dog prostate. 2 Acetylcholine and carbachol induced contraction of prostate strips from the peripheral zone, (‘the capsule’). Bethanechol contracted the tissue but not at lower doses. McN-A-343 and oxotremorine-M showed the same effects. 3 Blocking α- and β-adrenoceptors with phentolamine and propranolol, respectively, did not modify carbachol-induced contractions. 4 The nicotinic receptor blocker, hexamethonium (10^??6–10^??4 m ) did not affect the contractile response evoked by a single dose of carbachol (10^??5 m ), whilst the muscarinic receptor antagonist, atropine (10^??11–10^??9 m ), inhibited it in a competitive manner. 5 The muscarinic M₁ (pirenzepine), M₂[AF-DX 116, himbacine (M₂/M₄) and methoctramine], M₃ (HHSID and f-F-HHSID), and putative M₄ (tropicamide) antagonists reduced significantly the carbachol-induced contractions. The pIC₅₀ values were: atropine (10.01) > himbacine (8.3) > methoctramine (7.85) > AF-DX 116 (7.60) > HHSID (7.21) > p-F-HHSID (7.10) > pirenzepine (7.30) > tropicamide (7.00). 6 The antagonist profile indicates that an predominant M₂ receptor subtype could mediate the muscarinic contraction in the canine prostate.     

A study on the cause of death produced by Angusticeps-type toxin F7 isolated from eastern green mamba venom

The cause of death due to toxin F₇, an angusticeps-type toxin, isolated from the venom of Dendroaspis angusticeps was studied in anesthetized mice. The carotid arterial blood pessure, the ECG and the respiratory movements were recorded. Within a few minutes after i.v. injection of F₇ (1 mg/kg), both the rate and amplitude of the respiratory movements decreased and respiratory arrest took place within 15 min in most cases. Before respiratory arrest, marked bradycardia with various types of arrhythmia and oscillation of blood pressure were observed. Artificial ventilation could abolish these cardiovascular changes and maintain the blood pressure for a long period. Toxin F₇ caused a transient and slight increase of arterial blood pressure which could be prevented by hexamethonium. Intracisternal application of F₇ (1 mg/kg) caused a long-lasting hypertension and bradycardia and the respiratory arrest time was significantly longer than after i.v. injection. A large dose (50 mg/kg i.p.) of atropine, but not smaller doses (5–10 mg/kg), protected mice against respiratory failure induced by F₇. In rats, the phrenic nerve discharge was prolonged during respiratory depression. Since F₇ has a potent anticholinesterase activity, it is concluded that the respiratory failure induced by F₇ is peripheral in origin, chiefly, if not entirely, due to its anticholinesterase activity.     

Central muscarinic receptor subtypes involved in pilocarpine-induced salivation, hypertension and water intake

Background and purpose:

Recent evidence has suggested that pilocarpine (ACh receptor agonist) injected peripherally may act centrally producing salivation and hypertension. In this study, we investigated the effects of specific M₁ (pirenzepine), M₂/M₄ (methoctramine), M₁/M₃ (4-DAMP) and M₄ (tropicamide) muscarinic receptor subtype antagonists injected into the lateral cerebral ventricle (LV) on salivation, water intake and pressor responses to peripheral pilocarpine.

Experimental approach:

Male Holtzman rats with stainless steel cannulae implanted in the LV were used. Salivation was measured in rats anaesthetized with ketamine (100 mg per kg body weight) and arterial pressure was recorded in unanaesthetized rats.

Key results:

Salivation induced by i.p. pilocarpine (4 μmol per kg body weight) was reduced only by 4-DAMP (25–250 nmol) injected into the LV, not by pirenzepine, methoctramine or tropicamide at the dose of 500 nmol. Pirenzepine (0.1 and 1 nmol) and 4-DAMP (5 and 10 nmol) injected into the LV reduced i.p. pilocarpine-induced water intake, whereas metoctramine (50 nmol) produced nonspecific effects on ingestive behaviours. Injection of pirenzepine (100 nmol) or 4-DAMP (25 and 50 nmol) into the LV reduced i.v. pilocarpine-induced pressor responses. Tropicamide (500 nmol) injected into the LV had no effect on pilocarpine-induced salivation, pressor responses or water intake.

Conclusions and implications:

The results suggest that central M₃ receptors are involved in peripheral pilocarpine-induced salivation and M₁ receptors in water intake and pressor responses. The involvement of M₃ receptors in water intake and pressor responses is not clear because 4-DAMP blocks both M₁ and M₃ receptors.     

Conditional involvement of muscarinic M1 receptors in vagally mediated contraction of guinea-pig bronchi

The involvement of ganglionic muscarinic M₁ receptors in vagally induced bronchoconstriction in guinea-pig airways is controversial. Therefore, we studied the effects of the M₁-selective muscarinic receptor antagonist pirenzepine on vagus nerve (VNS, preganglionic) and electrical field stimulation (EFS, postganglionic)-induced contractions of the guinea-pig main bronchus under various experimental conditions.Using identical stimulation parameters for VNS and EFS (8V, 30 Hz, 0.5 ms, 5s every min), the amplitude of the VNS-induced twitch contractions was 30.4% of the EFS-induced responses, and pirenzepine showed 2.3-fold selectivity (pIC₅₀-values 6.45 and 6.09, respectively) to inhibit vagally induced contractions. With the stimulation frequency for EFS lowered to match contraction levels obtained using VNS, pirenzepine was equipotent to inhibit both types of response at M₃ receptor-selective concentrations, suggesting that M₁ receptors are not involved. By contrast, when the stimulation episode was prolonged until plateau contraction (10–20 s), in the presence of the nicotinic antagonist hexamethomum (5 M), the M₂ receptor antagonist AQ-RA 741 (0.1 M) and the -adrenoceptor antagonist timolol (1 M), and again using matched VNS- and EFS-induced contraction levels, pirenzepine inhibited nerve stimulation-evoked responses in a biphasic manner, yielding (pIC₅₀-values of 8.12 indicative of M₁ receptor blockade) and 6.43 (indicative of M₃ receptor blockade) for the first and second phase, respectively, while postganglionic stimulation showed a purely monophasic inhibition (pIC₅₀ = 6.32).These results show that facilitatory muscarinic M₁ receptors are involved in vagally mediated contraction of guinea-pig bronchi, under conditions of elevated neurotransmission and partial nicotinic receptor blockade.     

Blockade of spatial learning by the M1 muscarinic antagonist pirenzepine

Two experiments were conducted to determine the effects of the M₁ muscarinic receptor antagonist pirenzepine on place navigation in a water maze. In the first experiment rats were required to learn the location of a hidden platform following intracerebroventricular injections of equimolar doses of pirenzepine or scopolamine methylbromide. Both drugs dose-dependently impaired spatial learning according to both escape latency data and transfer test analysis. Pirenzepine was approximately 3 times less potent than scopolamine, a potency ratio which suggests M₁ receptor mediation of the impairment. In the second experiment pirenzepine (192.3 g/rat ICV) was injected prior to training on a simultaneous place dicrimination task in the water maze. Impairments of choice accuracy were found with a dose of 20 g/rat in the absence of any marked increases in either errors of omission or choice latency. These data suggest that M₁ receptor blockade impairs processes which are involved in spatial learning.     

6β-乙酰氧基去甲托烷呼吸抑制效应

6β-乙酰氧基去甲托烷(6β-acetoxynortropane,6β-AN)是一种新型M2受体激动剂,兔10ug·kg^-1iv,犬2,5,20ug·kg^-1iv均导致呼吸频率、潮气量、每分通气量明显减少(P<0.05或0.01),呈剂量依赖关系。pO₂降低,pCO₂升高。较小剂量给兔0.5,1.0,2.0,4.0ug·kg^-1iva和犬0.25,0.5,1.0ug·kg^-1iva亦产生与静脉给药相似的呼吸抑制效应。AF-DX116能拮抗6β-AN的呼吸抑制作用,PZ则与6β-AN产生协同作用。表明6β-AN有呼吸抑制作用,并可能与其激动呼吸中枢M2受体有关。     

The relative potencies of cholinomimetics and muscarinic antagonists on the rat iris in vivo: effects of pH on potency of pirenzepine and telenzepine

Summary The effects of cholinomimetics and muscarinic antagonists were compared following topical administration to the eyes of anaesthetized rats. For tests with cholinomimetics, clonidine (0.3 mg/kg) was used to induce mydriasis via central inhibition of parasympathetic tone. Full, dose-dependent miosis was induced by acetylcholinesterase inhibitors [physostigmine > neostigmine > tetrahydroaminoacridine (THA)] and by membrane channel blockers (4-aminopyridine > 3,4-diaminopyridine). Oxotremorine was the most potent direct agonist tested [oxotremorine > arecaidine propargylester (APE) > arecolne > carbachol > ethoxyethyltrimethyl-ammonium iodide (EOE) > RS 86]. Some putative M1 selective agonists were weakly active or behaved as partial agonists (pilocarpine > AH6405 > Mc-A-343 > isoarecoline). Of the antagonists, compared in non-clonidine treated rats, scopolamine hydrochloride was the most potent. Of the receptor selective antagonists the M₂ (ileal) selective compounds hexahydrosiladifenidol and 4-DAMP were more potent than either M₁ selective (pirenzepine, telenzepine) or M₂ (atrial) selective (AF DX 116) drugs. These data tentatively suggest the involvement of an M₂ (ileal) type muscarinic receptor. Potency was lower for quaternary structures, probably due to impaired corneal penetration. The potency of pirenzepine and telenzepine was increased 60-fold at low pH following topical administration. Acid induced corneal damage does not appear to account for this potency shift as the effects of scopolamine and several agonists (oxotremorine, pilocarpine and McN-A-343) were not substantially altered by acid media. For pirenzepine the potency shift appears to be related to protonation of the second amino group (N1) in the piperazine tail (pK _a = 2.05). Intraocular injections suggest that diprotonation facilitates penetration through the cornea. This anomalous behaviour of pirenzepine may contribute to its potency in gastric acid inhibition where the acid environment of the stomach would favour the diprotonated state and therefore penetration through the epithelium. Send offprint requests to J. J. Hagan at the above address     

A presynaptic excitatory M1 muscarine receptor at postganglionic cardiac noradrenergic nerve fibres that is activated by endogenous acetylcholine

Summary Rabbit atria were isolated with the extrinsic right vagus and sympathetic nerves intact and perfused with Tyrode solution. Noradrenaline overflow evoked by sympathetic nerve stimulation (SNS) at 3 Hz for 3 min was determined before, during, and after vagus nerve stimulation (VNS), also at 3 Hz and for 3 min. The VNS pulses preceded the SNS pulses by 3, 100 and 233 ms. Acetylcholine overflow was determined after labelling of the transmitter stores with [¹⁴C]choline.Pirenzepine 80 nmol/l failed to alter the muscarinic inhibition of noradrenaline overflow when the vago-sympathetic impulse intervals were 3 and 233 ms. At an interval of 100 ms VNS did not significantly inhibit noradrenaline overflow in the absence of pirenzepine but produced an inhibition in the presence of the drug. When the pirenzepine concentration was varied (0.4–300 nmol/l) the largest inhibition of noradrenaline overflow was observed at 5.7 nmol/l whereas 300 nmol/l fully antagonized the inhibition. Acetylcholine overflow evoked by VNS was not altered by pirenzepine 0.4–300 nmol/l.AF-DX 116 (11-[{2[oi(diethylamino)methyl]-1-piperidinyl}-acetyl]-5,11-dihydro-6H-pyrido-[2,3-b]-[1,4]benzodiazepine-6-one), an M₂ receptor selective antagonist, concentration-dependently (100–800 nmol/l) inhibited the decrease of tension development elicited by VNS. At the 100 ms vago-sympathetic impulse interval noradrenaline overflow was enhanced in the presence of AF-DX 116 400 and 800 nmol/l. However, already 100 nmol/l of the drug caused a maximum (fourfold) increase of acetylcholine overflow.It is concluded that acetylcholine released onto noradrenergic nerve fibres causes a small facilitation of noradrenaline overflow at a vago-sympathetic impulse interval of 100 ms. This response is mediated by an M₁ receptor and is superimposed on the well-known M₂ receptor mediated inhibition of noradrenaline release which is obtained at vago-sympathetic impulse intervals ranging between 3 and 233 ms. The M₂ autoreceptor regulating acetylcholine release is activated by lower synaptic concentrations of the transmitter than the M₂ heteroreceptor regulating noradrenaline release.Abbreviations SNS sympathetic nerve stimulation - VNS vagus nerve stimulation Send offprint requests to: E. Muscholl at the above address     

In vivo antimuscarinic actions of the third generation antihistaminergic agent, desloratadine

Background

Muscarinic receptor mediated adverse effects, such as sedation and xerostomia, significantly hinder the therapeutic usefulness of first generation antihistamines. Therefore, second and third generation antihistamines which effectively antagonize the H₁ receptor without significant affinity for muscarinic receptors have been developed. However, both in vitro and in vivo experimentation indicates that the third generation antihistamine, desloratadine, antagonizes muscarinic receptors. To fully examine the in vivo antimuscarinic efficacy of desloratadine, two murine and two rat models were utilized. The murine models sought to determine the efficacy of desloratadine to antagonize muscarinic agonist induced salivation, lacrimation, and tremor. Desloratadine's effect on the cardiovascular system was explored in both rodent models.

Results

In the pithed rat, both desloratadine (1.0 mg/kg, i.v.) and the muscarinic M₂ selective antagonist, methoctramine (0.5 mg/kg, i.v.), inhibited negative inotropic (left ventricular dP/dt) effects caused by oxotremorine, a nonselective muscarinic agonist (p < 0.05). Negative chronotropic effects caused by oxotremorine were inhibited by desloratadine, methoctramine, and the muscarinic M₃ selective antagonist, 4-DAMP (1.0 mg/kg, i.v.). A late positive inotropic event observed after the initial decrease was inhibited by all three test compounds with desloratadine and 4-DAMP being the most efficacious. In the conscious animal, inhibition of baroreflex-mediated bradycardia was evaluated. Unlike atropine (0.5 mg/kg, i.v.), desloratadine did not alter this bradycardia. The antimuscarinic action of desloratadine on salivation, lacrimation, and tremor was also explored. In urethane-anesthetized (1.5 g/kg, i.p.) male ICR mice (25–35 g) desloratadine (1.0, 5.0 mg/kg) did not inhibit oxotremorine-induced (0.5 mg/kg, s.c.) salivation, unlike atropine (0.5 mg/kg) and 4-DAMP (1.0 mg/kg). In conscious mice, desloratadine failed to inhibit oxotremorine-induced (0.5 mg/kg, s.c.) salivation, lacrimation, and tremor. However, desloratadine did inhibit oxotremorine-induced tremor in phenylephrine pretreated animals.

Conclusion

The presented data demonstrate that the third generation antihistamine, desloratadine, does not significantly antagonize peripheral muscarinic receptors mediating salivation and lacrimation, therefore, xerostomia and dry eyes should not be observed with therapeutic use of desloratadine. Our data also indicate when administered to a patient with a compromised blood-brain barrier, desloratadine may cause sedation. Patients with compromised cardiovascular systems should be closely monitored when administered desloratadine based on our results that desloratadine has the ability to interfere with normal cardiovascular function mediated by muscarinic receptors.     

In vivo modulation of histamine release by autoreceptors and muscarinic acetylcholine receptors in the rat anterior hypothalamus

The modulation of histamine release by histamine and muscarinic acetylcholine receptors was investigated by using the push-pull technique. The anterior hypothalamic area of the conscious, freely moving rat was superfused through the push-pull cannula with CSF or with CSF containing drugs and the release of endogenous histamine was determined in the superfusate.Hypothalamic superfusion with tetrodotoxin (10 mol/1) led to a pronounced and sustained decrease in the histamine release rate. Superfusion with compound 48/80 (100 mg/1) was ineffective. Hypothalamic superfusion with the H₃ agonist (R)--methylhistamine inhibited, while superfusion with the H₃ antagonist thioperamide enhanced the release of histamine. The release of histamine was inhibited on hypothalamic superfusion with the muscarinic receptor agonists carbachol or oxotremorine. Histamine release was enhanced by atropine, and this release-enhancing effect was abolished by oxotremorine. The selective M₁ antagonist pirenzepine (100 mol/I) and 4-diphenylacetoxy-N-methylpiperidine (4-DAMP, 10 ol/1), which blocks M₁ and M₃ receptors, also enhanced the release rate of histamine. On the other hand, 50 and 100 moI/I methoctramine (M₂ receptor antagonist) 10 and 100 moI/l p-fluoro-hexahydro-siladifenidol (p-F-HHSiD, a M₃ receptor antagonist) were ineffective.It is concluded that histamine released in the hypothalamus originates predominantly from neurons. The release of histamine is modulated by H₃ autoreceptors. The histamine release is also modulated by cholinergic neurons which modify histamine release from histaminergic neurons by stimulating M₁ muscarinic acetylcholine heteroreceptors probably located on histaminergic neurons.Supported by the Fonds zur Förderung der wissenschaftlichen Forschung Correspondence to: H. Prast at the above address     

The new muscarinic M1-receptor agonist YM796 evokes yawning and increases oxytocin secretion from the posterior pituitary gland in rats

The present experiments were performed to examine the effects of a new muscarinic M₁-receptor agonist, (−)-YM796 ((−)-S-2,8-dimethyl-3-methylene-1-oxa-8-azaspiro[4.5]decane L-tartrate monohydrate), on yawning and oxytocin secretion from the posterior pituitary gland in rats. YM796, at doses of 2.5–50 mg/kg (SC), elicited yawning. The yawning response was markedly increased by pretreatment with a β-adrenoceptor antagonist, pindolol (20 mg/kg, IP), which per se did not elicit yawning. The yawning induced by YM796 (10 mg/kg, SC) in combination with pindolol (20 mg/kg, IP) was inhibited by scopolamine (0.5 mg/kg, SC), a muscarinic receptor antagonist, and pirenzepine (300 μg/ rat, ICV) and EEDQ (N-ethoxycarbonyl-2-ethoxy-1,2-dihydroquinoline) (5 mg/kg, IP), muscarinic M₁-receptor antagonists, but not by spiperone (0.5 mg/kg, SC), a dopamine D₂-receptor antagonist, 4-DAMP (4-diphenylacetoxy-N-methylpiperidine methiodide) (100 μ/rat, ICV), a muscarinic M₃-receptor antagonist, and [d(CH₂)₅, Tyr(Me)², Orn⁸]-vasotocin (100 ng/rat, ICV), an oxytocin receptor antagonist. YM796 at 2.5–50 mg/kg (SC) did not exert an action on prolactin levels but increased oxytocin secretion from the posterior pituitary gland in rats. This augmentation of oxytocin secretion by YM796 was inhibited by scopolamine (0.5 mg/kg, SC) and pirenzepine (3 mg/kg, SC), but not by mecamylamine (1 mg/kg, IP), a nicotinic receptor antagonist. The present findings obtained with YM796 suggest that the muscarinic M₁-receptor stimulation participates in causing yawning behavior and oxytocin secretion in rats.     

POSSIBLE INVOLVEMENT OF MUSCARINIC M1 AND M3 RECEPTOR SUBTYPES MEDIATING VASODILATION IN ISOLATED, PERFUSED CANINE LINGUAL ARTERIES

1. Using the cannula insertion method, muscarinic receptor subtypes were analysed in isolated, perfused canine lingual arteries preconstricted with phenylephrine. 2. Both acetylcholine and McN-A-343 induced a profound vasodilation in a dose-related manner. Acetylcholine-induced dilations were approximately 1000-times more potent than McN-A-343-induced dilation. 3. Acetylcholine-induced dilations were abolished after removal of the endothelium by intraluminal treatment with 1 mg saponin. 4. Acetylcholine-induced dilations were markedly inhibited by an M₁/M₃ receptor antagonist, 4-DAMP. Moreover, they were slightly, but significantly, inhibited by an M₁ antagonist, pirenzepine, but never influenced by an M₂ antagonist, AF-DX 116. Mc-N-A-343-induced vasodilations were inhibited by both 4-DAMP or pirenzepine. 5. These results suggest that there are abundant functional M₃ and a few M₁ receptors in the canine lingual artery that mediate vasodilation and that this vasodilation is dependent on the presence of an intact endothelium.