M1 Muscarinic Receptor Antagonist Research Articles

Cholinergic modulation of non-adrenergic, non-cholinergic relaxation in isolated, small coronary arteries from lambs.

The presence of cholinergic innervation of small coronary arteries in the lamb was investigated by measuring choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) activities and by performing in vitro experiments in a microvascular myograph to establish whether or not there is a cholinergic component in the response to electrical field stimulation (EFS). ChAT-specific activity was present in proximal coronary segments, but was significantly higher in small coronary arteries. AChE-positive ganglia and fibres were distributed within the adventitia and outer third of the media in proximal coronary segments, and dense perivascular nerve plexuses were observed in small coronary arteries. Acetylcholine induced contractions in all preparations examined and relaxations in 20% of the segments contracted with the thromboxane analogue U46619. EFS did not induce neurogenic contractions in lamb small coronary arteries. In the presence of the alpha-adrenoceptor antagonist, phentolamine, EFS caused frequency-dependent reproducible relaxations that were enhanced by the blocker of cholinergic transmission, botulinum neurotoxin. An inhibitor of AChE, physostigmine, had no significant effect on the relaxations caused by EFS, while both the muscarinic receptor antagonist, atropine, and the muscarinic M2-receptor antagonist, AFDX 116, enhanced these responses. Blockade of sympathetic neurotransmission with guanethidine or incubation with the P2-receptor antagonist, suramin, abolished the relaxations induced by EFS, whereas propranolol was without effect. Low-frequency EFS caused less relaxation in preparations activated by acetylcholine than in those contracted with U46619, while sensitivity and maximal relaxation induced by adenosine 5'-triphosphate (ATP) were not different in U46619- and acetylcholine-contracted arteries. The presence of the enzymes necessary for both biosynthesis and degradation of acetylcholine and the finding that blockers of cholinergic neurotransmission enhance EFS-induced relaxations suggest that small coronary arteries are cholinergically innervated.

Inhibition of GTPase activity of Gi proteins and decreased agonist affinity at M2 muscarinic acetylcholine receptors by spermine and methoctramine.

1. The effects of spermine and methoctramine, a selective M2 muscarinic receptor antagonist, were studied on the high-affinity GTPase activity of G proteins, and on ligand binding to M2 muscarinic receptors in pig heart sarcolemma. 2. The spontaneous GTP hydrolysis by pig heart sarcolemma and its stimulation by mastoparan or carbachol were prevented by pertussis toxin and inhibited by methoctramine (IC50s: 21, 13 and 0.005 microM, respectively), and spermine (IC50s: 967, 278 and 11 microM). Spermine and methoctramine also inhibited spontaneous GTP hydrolysis by rat peritoneal mast cell membranes which do not respond to carbachol. 3. The neutral muscarinic antagonists, AF-DX 116 and atropine, did not modify the inhibitory effect of high concentrations of methoctramine, indicating that this effect was not related to the antagonist binding site of muscarinic receptors. We suggest that methoctramine behaves as a receptor antagonist at nanomolar concentrations and interacts with G proteins at micromolar concentrations. 4. Spermine did not modify the binding of the tritiated muscarinic antagonist [3H]-NMS, but decreased the binding of the agonist [3H]-Oxo-M. Spermine elicited a rightward shift of the carbachol/[3H]-NMS binding isotherm with a decrease in the proportion of sites with high-affinity for carbachol, suggesting that polyamines uncouple Gi proteins from receptors. 5. The inhibition of GTPase activity by polyamines, preventing the re-association of alpha and betagamma subunits of Gi proteins, might sustain the regulatory effect of Gi subunits on downstream effectors. The level of intracellular polyamines might be important for the control of the transduction of extracellular signals through Gi protein-coupled receptors.

Nitric oxide-dependent relaxation induced by M1 muscarinic receptor activation in the rat small intestine.

The aim of the present study was to investigate whether muscarinic M1 receptor activation induces intestinal relaxation via nerve-dependent nitric oxide formation. Mechanical activity in longitudinal segments of rat jejunum was recorded isotonically in organ baths. The muscarinic M1 receptor agonist 4-[[[(3-Chlorophenyl)amino]carbonyl]oxy]-N,N,N,-trimethyl-2-butyn- 1-ammonium chloride (McN-A-343, 10(-7)-10(-4) M) induced a concentration-dependent relaxation of rat jejunum. Relaxations induced by McN-A-343 (10(-5) M) were inhibited by the M1 receptor antagonist telenzepine (10(-8) M), and enhanced by the M3 receptor antagonist para-fluorohexahydrosiladifenidol (p-F-HHSiD; 3x10(-7) M). The inhibitory responses induced by McN-A-343 were abolished by the nitric oxide synthase inhibitors Nomega-nitro-L-arginine (L-NOARG; 10(-4) M) and Nomega-monomethyl-L-arginine (L-NMMA; 3x10(-5) M), the guanylyl cyclase inhibitor 1H-[1,2,4]oxadiazolo[4,3,-a]quinoxalin-1-one (ODQ; 10(-5) M), and by tetrodotoxin (TTX; 3x10(-7) M). Guanethidine or hexamethonium did not affect inhibitory responses induced by McN-A-343. In conclusion, McN-A-343 induces nerve-dependent, nitrergic relaxations in rat jejunum, via activation of muscarinic M1 receptors. Hence, selective muscarinic M1 receptor agonists or antagonists might offer possibilities for pharmacological manipulation of the NO system.

Inhibition of nitrergic relaxations by the M3-selective antagonist 4-DAMP

The inhibitory effect of the selective M3 musarinic acetylcholine receptor antagonist, 4-diphenylacetoxy-N-methylpiperidine methobromide (4-DAMP, 0.1–10 μM) on nicotine (100 μM)-induced nitrergic relaxation was investigated in comparison to d-tubocurarine (0.1–10 μM) and hexamethonium (0.1–10 μM) by using phenylephrine (1 μM)-precontracted rat anococcygeus muscles in vitro. Nicotine produced a 60.1± 2.4% (n = 40) inhibition of phenylephrine precontractions. But this relaxant response was at a significantly lower magnitude of 20.2± 4.6% (n = 18, P hexamethonium > d-tubocurarine with IC50 values being 0.47± 0.04 μM, 0.75± 0.06 μM, and 1.02± 0.05 μM, respectively. Therefore, it was concluded that the selective M3 muscarinic acetylcholine receptor antagonist 4-DAMP also possesses potent antagonistic action on nicotinic receptors of peripheral nitrergic neurons that innervate the rat anococcygeus muscle. Drug Dev. Res. 46:148–154, 1999. © 1999 Wiley-Liss, Inc.

The antiallodynic effects of intrathecal cholinesterase inhibitors in a rat model of neuropathic pain.

This study determined the effect of intrathecally administered cholinesterase inhibitors, edrophonium and neostigmine, on nerve injury-induced, touch-evoked allodynia and identified the pharmacologic characteristics of this action. Rats were prepared with tight ligation of the left L5 and L6 spinal nerves and with lumbar intrathecal catheters fitted for long-term monitoring. Edrophonium (3, 10, 30, or 100 microg) or neostigmine (0.3, 1, 3, or 10 microg) was administered intrathecally. Tactile allodynia and motor weakness were assessed. To evaluate the pharmacologic characteristics of the activity, a muscarinic receptor antagonist or a nicotinic receptor antagonist was administered intrathecally before edrophonium or neostigmine was injected. To compare the action of subtype antagonists, the M1 muscarinic receptor antagonist pirenzepine, the M2 antagonist methoctramine, the M3 antagonist 4-DAMP (diphenylacetoxy-N-methypiperidine), and the M4 antagonist tropicamide were administered intrathecally before cholinesterase inhibitors were injected. Intrathecal edrophonium or neostigmine produced a dose-dependent antagonism of the touch-evoked allodynia. Neostigmine resulted in a moderate effect on motor weakness at doses of 3 and 10 microg. Pretreatment with intrathecal atropine but not mecamylamine yielded a complete antagonism of the effects of the cholinesterase inhibitors. In addition, antiallodynia produced by edrophonium (100 microg) was reversed by pretreatment with methoctramine, 4-DAMP, tropicamide, and pirenzepine. In the neostigmine (10 microg) group, only the M1 antagonist pirenzepine had a moderate effect on reversal of increased allodynic threshold. These experiments suggest that intrathecal edrophonium or neostigmine produces an antagonism on touch-evoked allodynia at the spinal level in a rat model of neuropathic pain and that the antiallodynic action of cholinesterase inhibitors is probably mediated by a spinal muscarinic system, especially at the M1 receptor subtype.

This Month in ANESTHESIOLOGY

This Month in ANESTHESIOLOGY

Pharmacological characterization of J-104129, a novel muscarinic M3 receptor antagonist with high selectivity over M2 receptors

Pharmacological characterization of J-104129, a novel muscarinic M3 receptor antagonist with high selectivity over M2 receptors

Synthesis and biological evaluation of 1,2,3,4-tetrahydroisoquinoline derivatives as potent and selective M2 muscarinic receptor antagonists.

A series of 1,2,3,4-tetrahydroisoquinoline derivatives containing the 5,11-dihydro-6H-pyrido[2,3-b][1,4]benzodiazepin-6-one skeleton were prepared and evaluated for their in vitro binding affinities to muscarinic receptors and for antagonism of bradycardia in vivo. Among them, compound 3f had the highest affinity for M2 muscarinic receptors in the heart (pKi = 9.1) with low affinity for M3 muscarinic receptors in the submandibular gland. A structure-activity relationship (SAR) study suggested that the benzene ring fused piperidine and the alkyl linker chain length are crucially important for increased M2 affinity.

Contractile and phosphatidylinositol responses of rat trachea to anticholinesterase drugs.

Some anticholinesterases (anti-ChE) such as neostigmine and pyridostigmine but not edrophonium, stimulate phosphatidylinositol (PI) response. Although a direct relationship was suggested between the increase in PI response and airway smooth muscle contraction, there are no data regarding the effects of anti-ChE drugs on airway smooth muscle. Thus, we examined the contractile properties and PI responses produced by anti-ChE drugs. Contractile response. Rat tracheal ring was suspended between two stainless hooks in Krebs-Henseleit (K-H) solution. (1) Carbachol (CCh), anti-ChE drugs (neostigmine, pyridostigmine, edrophonium) or DMPP (a selective ganglionic nicotinic agonist) were added to induce active contraction. (2) The effects of 4-diphenylacetoxy-N-methyl-piperidine methobromide (4-DAMP), an M3 muscarinic receptor antagonist, on neostigmine- or pyridostigmine-induced contraction of rat tracheal ring were examined. (3) Tetrodotoxin (TTX) was tested on the anti-ChE drugs-induced responses. PI response. The tracheal slices were incubated in K-H solution containing LiCl and 3[H]myo-inositol in the presence of neostigmine or pyridostigmine with or without 4-DAMP, an M3 muscarinic receptor antagonist. 3[H]inositol monophosphate (IP1) formed was counted with a liquid scintillation counter. Carbachol (0.1 microM), neostigmine (1 microM), pyridostigmine (10 microM) but not edrophonium or DMPP, caused tracheal ring contraction. 4-DAMP, but not tetrodotoxin, inhibited neostigmine and pyridostigmine-induced contraction. Neostigmine- or pyridostigmine-induced IP1, accumulation was inhibited by 4-DAMP. The data suggest that anti-ChE drugs activate the M3 receptors at the tracheal effector site.

Presynaptic muscarinic M1 and M2 receptor modulation of auriculotemporal nerve transmission in the rat

Parotid secretory and vascular responses to electrical stimulation of the parasympathetic innervation were measured in anaesthetized rats. Stimulation was performed at 1, 10 and 40 Hz. Atropine (1.5 μmol/kg I.V.) almost abolished the secretion to stimulation of peptide depleted nerves at 40 Hz, thus confirming the existence of a pure cholinergic response. Atropine also reduced secretion by 74% during stimulation of non-depleted nerves at the same frequency. Selective blockade by the muscarinic M1 receptor antagonist pirenzepine and by the muscarinic M2 receptor antagonist methoctramine was found to occur at doses (50 nmol/kg I.V. and of 300 nmol/kg I.V., respectively) that did not inhibit the responses to exogenous acetylcholine. In the presence of methoctramine, the nerve-evoked fluid responses were increased by 200% at 1 Hz independently of the total number of impulses (10–300), suggesting that M2 receptor activation normally has an inhibitory effect on transmitter release. The magnitude of the increase was inversely related to frequency of stimulation, and changes in the secretory responses occurred at 40 Hz only when non-depleted nerves were stimulated over the longest period employed. The fluid response then increased by 35% and protein concentration by 200%. The vasodilator responses increased at 1 and 10 Hz, but not at 40 Hz. Pirenzepine reduced the secretory and vascular responses at 10 and 40 Hz but only during stimulation over short periods of time. This suggests that M1 receptor activation normally has a facilitatory effect on neurotransmitter release. During stimulation of non-depleted nerves at 10 Hz for 10 impulses, the fluid response was reduced by 29% and the protein concentration by 26%. When the peptide depleted nerves were stimulated at 10 Hz, pirenzepine also reduced the fluid response (by 43%), but not the protein concentration. It is concluded that the release of transmitter from postganglionic nerve fibres in the rat auriculotemporal nerve is modulated by presynaptic muscarinic receptors. Muscarinic M1 receptors normally facilitate cholinergic and peptidergic transmission during short, intense stimulation. On the other hand, muscarinic M2 receptors normally inhibit cholinergic transmission at low frequencies; at higher frequencies, peptidergic transmission is also inhibited, but only after some delay.

DIFFERENCES IN THE EFFECTS OF VASOPRESSIN AND OXYTOCIN ON FELINE GASTRIC CORPUS MOTILITY: SELECTIVE ACTION OF VASOPRESSIN ON LONGITUDINAL MUSCLE

The effect of vasopressin and oxytocin on the contractile activity of preparations isolated from the feline gastric corpus wall was investigated. Vasopressin (1.5×10−9–2.1×10−7m), but not oxytocin, evoked concentration-dependent tonic contractions only of longitudinal muscle strips. At the same time, vasopressin (1.5×10−9–2.1×10−7m) potentiated the magnitude of amplitudes, but not the frequency, of spontaneous contractions. Both the vasopressin V1 receptor antagonistd(CH2)5-(Me)2-Tyr-AVP and the predominantly vasopressin V2 receptor antagonistd(CH2)5,d-Ile2, Ile4-AVP, the non-selective muscarinic receptor antagonist, atropine, the predominantly selective muscarinic M1 receptor antagonist, pirenzepine, the predominantly selective muscarinic M2 antagonist, methoctramine, the predominantly selective muscarinic M3 receptor antagonist, para-fluoro-hexahydro-siladifenidol, and the calcium channel blocker, nifedipine, but not the ganglion blocking agent, mecamylamine, depressed or blocked the tonic contractions induced by vasopressin. Among the antagonists, only atropine and nifedipine inhibited the spontaneous contractions. On the other hand, the anticholinesterase, physostigmine, potentiated both the vasopressin-induced tonic and spontaneous contractions. With regard to the receptors, the vasopressin-induced tonic contractions are mediated at least in part through vasopressin V1and V2receptors, non-selective muscarinic and selective muscarinic M1, M2and M3receptors. The increase in amplitudes of spontaneous contractions is mediated only via-nonselective muscarinic receptors. Vasopressin receptors appear to be located mostly pre-synaptically, although the direct effect of vasopressin on post-synaptic receptors cannot be excluded. The pA2values suggests rather V1athan V1bvasopressin receptor subtype involvement in tonic contractions vasopressin had produced. The tonic as well as spontaneous contractions are calcium-dependent. In addition, these results point to the existence of non-selective muscarinic receptors, which participate in the regulation of both tonic and spontaneous contractions, while muscarinic M1, M2and M3receptors subserve only the tonic contractions.

Cholinergic regulation of the rat adrenal zona glomerulosa.

Using histochemical and immunocytochemical methods, cholinergic nerve fibres were demonstrated in the rat adrenal cortex, primarily in the capsule and zona glomerulosa, and in the medulla. Some terminated among the glomerulosa cells or around blood vessels. Occasional fibres were also seen in the fasciculata, ending in islets of chromaffin tissue without ramifications on cortical cells. To clarify the role of cholinergic innervation, a microvolume perifusion system was used to study steroid production by the rat adrenal capsule-glomerulosa. Acetylcholine (ACh) itself had no reproducible effects; however, since variable amounts of endogenous ACh were present, the actions of antagonists were also studied. The M1 muscarinic receptor antagonist pirenzepine (10 and 100 microM) stimulated aldosterone secretion. This stimulation was abolished by co-incubation with carbachol, the M1 agonist McN A-343 and by atropine. We found that the action of pirenzepine was blocked by nifedipine (Ca2+ channel blocker), suggesting that pirenzepine (through release of endogenous ACh) provides an acute stimulus by enhancing Ca2+ inflow. Hemicholine, a choline uptake blocker, reduced the stimulatory effect of pirenzepine on steroid secretion, confirming that stimulation was of neural origin. Neither the non-selective muscarinic receptor antagonist atropine, the selective M1-M3 muscarinic receptor antagonist 4-DAMP, nor the selective M2 muscarinic receptor antagonist methoctramine influenced aldosterone output. Receptor-binding studies revealed the existence of M3 receptors in capsule-glomerulosa homogenates. We conclude that pirenzepine acts on presynaptic M1 autoreceptors to increase spontaneous ACh release from varicose axon terminals that lie in close proximity to the glomerulosa cells. In turn ACh may thus stimulate steroidogenesis acutely through M3 receptors. These results support the concept of a direct cholinergic influence on zona glomerulosa function in the rat.

Cardiac muscarinic receptors decrease with age. In vitro and in vivo studies.

The M1 muscarinic receptor antagonist pirenzepine in low doses decreases resting heart rate; this effect declines with age (Poller, U., G. Nedelka, J. Radke, K. Pönicke, and O.-E. Brodde. 1997. J. Am. Coll. Cardiol. 29:187-193). To study possible mechanisms underlying this effect, we assessed (a) in six young (26 yr old) and six older volunteers (61 yr old), pirenzepine effects (0.32 and 0.64 mg intravenous [i.v.] bolus) on isoprenaline-induced heart rate increases; (b) in five heart transplant recipients, pirenzepine effects (0.05-10 mg i.v. bolus) on resting heart rate in the recipient's native and transplanted sinus nodes; and (c) in right atria from 39 patients of different ages (5 d-76 yr) undergoing open heart surgery, M2 muscarinic receptor density (by [3H]N-methyl-scopolamine binding) and adenylyl cyclase activity. (a) Pirenzepine at both doses decreased heart rate in young volunteers significantly more than in older volunteers; (b) pirenzepine (< 1 mg) decreased resting heart rate in the recipient's native but not transplanted sinus node; and (c) M2 receptor density and carbachol-induced inhibition of forskolin-stimulated adenylyl cyclase activity decreased significantly with the age of the patients. We conclude that pirenzepine decreases heart rate via inhibition of presynaptic M1 autoreceptors, thereby releasing endogenous acetylcholine, and that the heart rate-decreasing effect of acetylcholine declines with age because right atrial M2 receptor density and function decrease.

Synthesis and Biological Evaluation of Phenylacetyl Derivatives Having Low Central Nervous System Permeability as Potent and Selective M2 Muscarinic Receptor Antagonists.

A series of phenylacetyl derivatives containing the 5,10-dihydro-11H-dibenzo[b,e,][1,4]diazepin-11-one or 5,11-dihydro-6H-pyrido[2,3-b][1,4]benzodiazepin-6-one skeleton was prepared and evaluated for their binding affinities to muscarinic receptors in vitro and for antagonism of bradycardia, salivation and tremor in vivo. Among them, compounds 56 and 66 had high affinity for M2 muscarinic receptors in the heart (pKi = 8.7 and 8.9, respectively) with low affinity for M3 muscarinic receptors in the submandibular gland. A structure-activity relationship (SAR) study suggested that the high M2 selectivity over the M3 muscarinic receptors of 56 may be attributed to the direction of the carboxamide carbonyl group. In in vivo studies, 56 and 66 antagonized oxotremorine-induced bradycardia in rats on both intravenous and oral administration, and their heart rate increasing effect in dogs with nocturnal bradycardia was about 3-fold greater than that of AF-DX 116. Furthermore, they had almost no influence on oxotremorine-induced tremor in mice, presenting no evidence of central transfer.

Endogenous acetylcholine in the dorsal hippocampus reduces anxiety through actions on nicotinic and muscarinic1 receptors.

Dorsal hippocampal cholinergic modulation of behavior in different tests of anxiety was investigated by direct injection of the muscarinic M1 and M2 receptor antagonists, pirenzepine and gallamine, and the nicotinic receptor antagonist mecamylamine. In the social interaction test, the anxiogenic effect of pirenzepine (30-100 ng) provided evidence for a tonic cholinergic anxiolytic action mediated by postsynaptic M1 receptors. The anxiogenic action of mecamylamine (30 and 100 ng) was most likely mediated by its action of presynaptic nicotinic receptors to reduce acetylcholine release. Gallamine (10-1,000 ng) was without effect, suggesting that M2 receptors in this brain region do not play a significant role in this behavioral test. On Trial 1 in the elevated plus-maze, the receptor antagonists were without any effect, but in those with a previous 5-min experience of the plus-maze pirenzepine and mecamylamine had anxiogenic effects in the dose range of 30-300 ng; gallamine (100 and 300 ng) was without significant effect.

ChemInform Abstract: Synthesis of Novel Succinamide Derivatives Having the 5,11‐Dihydro‐6H‐ pyrido(2,3‐b)(1,4)benzodiazepin‐6‐one Skeleton as Potent and Selective M2 Muscarinic Receptor Antagonists. Part 1.

AbstractChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.

Bradycardia produced by pyridostigmine and physostigmine.

The bradycardia produced by pyridostigmine and physostigmine in an animal model of acute cardiac denervation was examined according to its relation to cholinesterase inhibition and sensitivity to block by cholinergic receptor antagonists. Cats were anaesthetised, vagotomised and propranolol-treated. Heart rate was continuously recorded. Erythrocyte cholinesterase activity of arterial blood was measured using a radiometric technique. Nicotinic and muscarinic M1 receptors were blocked with hexamethonium and pirenzepine, respectively. M2 receptors were blocked with gallamine, pancuronium and AFDX-116. With pyridostigmine and physostigmine the dose-response relationship for the decrease in heart rate (ED50 1.05 +/- 0.25 and 0.198 +/- 0.03 mg.kg-1, respectively) was shifted to the right of that for the inhibition of cholinesterase activity (ED50 0.094 +/- 0.03 and 0.032 +/- 0.01 mg.kg-1, respectively). The decrease in cholinesterase activity reached a plateau at a cumulative dose of 0.56 +/- 0.08 and 0.32 +/- 0.08 mg.kg-1, respectively. In contrast, there did not appear to be a plateau in the bradycardic effect. The bradycardia produced by pyridostigmine and physostigmine was blocked by hexamethonium (ED50 10 +/- 1.3 and 15.3 +/- 2.4 mg.kg-1, respectively), pirenzepine (ED50 68 +/- 16 and 138 +/- 32 micrograms.kg-1, respectively), gallamine (56 +/- 11 and 67 +/- 17 micrograms.kg-1, respectively), pancuronium (32 +/- 10 and 30 +/- 4 micrograms.kg-1, respectively), and AFDX-116 (31 +/- 4 and 28 +/- 4 micrograms.kg-1, respectively). The bradycardia produced by reversible anticholinesterase drugs containing a carbamyl group is not clearly related to the degree of cholinesterase activity, and has a low sensitivity to nicotinic and muscarinic M1 and a high sensitivity to muscarinic M2 receptor antagonists.

Alterations of physostigmine-induced yawning by chronic lithium administration in rats.

The effect of chronic lithium pretreatment on physostigmine-induced yawning was investigated in male rats. Intraperitoneal administration of physostigmine to rats induced yawning in a biphasic manner. However the maximum response was obtained by 0.2 mg/kg of the drug. Intracerebroventricular administrations of a putative M1 and M2 muscarinic receptor antagonists, pirenzepine and methoctramine decreased physostigmine-induced yawning. Intraperitoneal administration of a non-selective muscarinic receptor antagonist, atropine, also decreased the physostigmine-induced yawning significantly. Chronic lithium pretreatment (30 days) reduced yawning induced by physostigmine. The inhibitory effect of pirenzepine, methoctramine and atropine on physostigmine-induced yawning increased in rats pretreated with chronic lithium. These findings indicate that yawning is induced by a central cholinergic mechanism and that chronic pretreatment of lithium may interact with the cholinergic-induced behaviour.

Dopaminergic and Cholinergic Interaction in Cataleptic Responses in Mice

The cataleptogenic effects of haloperidol, a dopamine D2 receptor antagonist; SCH23390, a D1 receptor antagonist; physostigmine, a cholinesterase inhibitor; and pilocarpine, a muscarinic M1 receptor agonist, were challenged by pretreatment of mice with SKF38393, a dopamine D1 receptor agonist; apomorphine, a dopamine D1/D2 receptor agonist (mainly D2 receptor); pirenzepine, a muscarinic M1 receptor antagonist; and scopolamine, a muscarinic M1/M2 receptor antagonist. The effect of physostigmine and pilocarpine on haloperidol and SCH23390 cataleptic responses was also examined. Each of the challenging agents blocked one or more of the cataleptogenic agents, but only scopolamine blocked all four. Pirenzepine blocked cataleptic responses induced by SCH23390 and pilocarpine, but not those by haloperidol and physostigmine. The results of this study suggest that the action of physostigmine (endogenous acetylcholine) on M2 receptors might be more potent than that on muscarinic M1 receptors. A further interesting observation was that the haloperidol-induced catalepsy was enhanced by physostigmine pretreatment, but not by pilocarpine pretreatment, whereas the SCH23390-induced catalepsy showed the opposite spectrum of enhancement by the two cholinergic agonists. We conclude that, although the four cataleptogenic agents act via the dopaminergic-cholinergic systems, their pharmacological differences may be due largely to the different receptor subtypes that are involved in the mediation of catalepsy produced by each agent. Thus, dopamine receptors not only influence the cholinergic muscarinic receptors, but muscarinic M1 and M2 receptors also might mediate dopamine D1 and D2 receptor responses, respectively. The results suggest that there are, at the least, relationships between muscarinic M1 receptors and dopaminergic D1 receptors, and between muscarinic M2 receptors and dopaminergic D2 receptors. Dopamine D1 and D2 receptors may interact in a synergistic fashion on dopaminergic systems, but act independently of each other in influencing other system such as cholinergic neurons.

NANC transmitters in the female pig urethra--localization and modulation of release via alpha 2-adrenoceptors and potassium channels.

1. To investigate further the release, localization and identity of a non-nitrergic mediator of smooth muscle relaxation in the female pig urethra, we studied the effects of drugs acting at alpha 2-adrenoceptors or K+ channels, the effects of capsaicin and chemical sympathectomy, and the actions of several transmitter candidates. 2. Electrical field stimulation (EFS; frequencies above 12 Hz) of spontaneously contracted smooth muscle strips from the female pig urethra evoked long-lasting, frequency-dependent relaxations in the presence of prazosin, scopolamine, and NG-nitro-L-arginine. Treatment with the selective alpha 2-adrenoceptor agonist UK-14 304 markedly reduced the relaxations evoked by EFS at all frequencies tested (16-30 Hz). The inhibitory effect of UK-14 304 was completely antagonized by the alpha 2-adrenoceptor antagonist rauwolscine. The muscarinic M1 receptor antagonist, pirenzepine, or exogenously administered carbachol, did not have any effects on the electrically evoked relaxations. 3. Inhibition of high conductance Ca2+ activated K+ channels by iberiotoxin or charybdotoxin significantly enhanced the relaxations evoked by EFS at all frequencies. However, inhibition of voltage-sensitive K+ channels with 4-aminopyridine or dendrotoxin-1, treatment with the ATP-sensitive K+ channel blocker, glibenclamide, or treatment with the high and low conductance Ca2+ activated K+ channel blockers, tetraethylammonium chloride and apamin, had no effect on the relaxations evoked by EFS. 4. Electrically evoked relaxations were not affected by adrenergic denervation with 6-hydroxydopamine (6-OHDA) at any frequency. However, treatment with 6-OHDA abolished prazosin-sensitive electrically induced contractions, and a long-lasting relaxation was revealed. Treatment with capsaicin, believed to damage selectively a subpopulation of primary afferent fibres, did not affect basal tone or relaxations evoked by EFS. 5. Exogenously applied vasoactive intestinal polypeptide (VIP), pituitary adenylate cyclase-activating peptide (PACAP)-27, PACAP-38, adenosine, ATP and 5-hydroxy-tryptamine caused relaxations of the urethral preparations, whereas prostaglandin E2 and calcitonin gene-related peptide had no effects. VIP 10-28, alpha, beta-methylene-ATP, reactive blue-2, suramin or indomethacin did not reduce the electrically-evoked relaxations at any frequency. However, the relaxations were slightly reduced by trypsin or alpha-chymotrypsin. 6. The present results suggest that the release of the unknown mediator in the female pig urethra can be modulated via alpha 2-adrenoceptors and high conductance Ca2+ activated K+ channels. Furthermore, the mediator does not appear to be localized to or released from adrenergic or capsaicin-sensitive sensory nerve-endings. The identity of the transmitter remains to be established.