The minor population of M3-receptors mediate contraction of human detrusor muscle in vitro

1 The objective was to determine the role of muscarinic receptor subtypes in mediating contraction of the human detrusor smooth muscle in vitro. 2 Contractile responses of human detrusor muscle strips to carbachol were obtained in the absence and presence of a range of muscarinic antagonists (pirenzepine, methoctramine, 4-diphenylacetoxy-N-methyl piperidine methiodide (4-DAMP), tropicamide, oxybutynin and tolterodine). Affinity estimates (pKB values) were calculated for the antagonists and correlated with values at the cloned muscarinic receptor subtypes quoted in the literature. 3 Pirenzepine, methoctramine and tropicamide drugs that have high affinities at M1, M2 and M4-receptors, respectively, all had low affinities on the human detrusor (pKB values of 6.8, 6.9 and 6.5, respectively), whilst the M3-selective antagonist 4-DAMP had a high affinity (9.5). Schild plots for all four antagonists had slopes of unity indicating an action at a single receptor. Oxybutynin and tolterodine also acted as competitive antagonists with affinity estimates of 7.6 and 8.1, respectively. 4 When the antagonist affinities obtained on the bladder were plotted against the values published for these antagonists at the cloned muscarinic receptor subtypes, the best correlations were obtained for the m3- and m5-muscarinic receptor subtypes. 5 These data suggest that direct contractile responses of the human detrusor muscle to muscarinic receptor stimulation in vitro are mediated solely via the M3-muscarinic receptor subtype with no contribution from the major M2-receptor population.     

Adrenaline hypothesis: effect of formoterol on noradrenaline release

1 Originally, the so‐called `adrenaline hypothesis' related the release of noradrenaline (NA) to stimulation of presynaptic β₂‐receptors in nerve endings; now it confers a possible role to adrenaline taken up then released by nerves endings. It represents a potentially useful therapeutic pathway. The present study aims to investigate the effects of formoterol, a highly selective β₂‐adrenoceptor agonist. 2 It was carried out in freely moving rats, the isotope dilution technique being used to measure the NA spill over rate (NA‐SOR) and metabolic clearance rate (MCR). 3 A series of three results are reported. (a) When compared with adrenaline on equimolar basis, formoterol (2.3 μg kg^?1 min^?1) increased NA‐SOR while mean arterial blood pressure was decreased and heart rate increased. Thus, it was difficult to separate a direct presynaptic effect from indirect baroreflex‐dependent activation of the sympathetic system. (b) When formoterol was infused at 1 ng kg^?1 min^?1, a dose empirically defined to induce no haemodynamic effect, NA‐SOR was significantly increased, while NA‐MCR remained unchanged. (c) The NA‐SOR response to formoterol was not amplified by the presynaptic α₂‐adrenoceptor blocker, yohimbine, in contrast to the NA‐SOR response to adrenaline. 4 In conclusion, formoterol, a β₂‐adrenoceptor agonist, is shown to increase the release and plasma concentration of NA while its clearance was not changed.     

Age-dependent contribution of Rho kinase in carbachol-induced contraction of human detrusor smooth muscle in vitro

Aim： Activation of muscarinic receptors on the detrusor smooth muscle is followed by contraction, which involves both myosin light chain kinase （MLCK） and Rho kinase （ROCK）. The aim of this study was to determine the relative contributions of MLCK and ROCK to carbachol-induced contraction of human detrusor smooth muscle in vitro.
Methods： Detrusor smooth muscle strips were prepared from the macroscopically unaffected bladder wall of patients underwent cystectomy. The strips were fixed in an organ bath, and carbachol or KCl-induced isometric contractions were measured by force transducers.

Results： Addition of carbachol （0.4-4 μmol/L） into the bath induced concentration-dependent contractions of detrusor specimens, which was completely abolished by atropine （1 μmol/L）. Pre-incubation of detrusor specimens with either the MLCK inhibitor ML-9 or the ROCK inhibitors HA1100 and Y-27632 （each at 10 μmol/L） significantly blocked carbachol-induced contractions as compared to the time-control experiments. Moreover, MLCK and ROCK inhibition were equally effective in reducing carbachol-induced contractions. The residual carbachol-induced contractions in the presence of both MLCK and ROCK inhibitors were significantly smaller than the contractions obtained when only one enzyme （either MLCK or ROCK） was inhibited, suggesting an additive effect of the two kinases. Interestingly, ROCK-mediated carbachol-induced contractions were positively correlated to the age of patients （r=0.52, P〈0.05）.

Conclusion： Both MLCK and ROCK contribute to carbachol-induced contractions of human detrusor smooth muscle. ROCK inhibitors may be a new pharmacological approach to modulate human bladder hyperactivity.     

M3 muscarinic receptors but not M2 mediate contraction of the porcine detrusor muscle in vitro

1. The objective of the study was to determine the role of muscarinic receptor subtypes in mediating contraction of the porcine detrusor smooth muscle in vitro. 2. Strips of pig detrusor muscle were set up in physiological salt solution and the tensions developed by the tissues were recorded. Responses to carbachol were obtained in the absence and presence of a range of muscarinic antagonists (4-DAMP, methoctramine, darifenacin, oxybutynin, tolterodine and pirenzepine). Antagonist affinity values (pKB values) were calculated and compared with those quoted in the literature for these antagonists at each of the muscarinic receptor subtypes. 3. The M3-selective antagonists, 4-DAMP and darifenacin had high affinities (pKB values of 9.4 and 8.6, respectively). Oxybutynin, tolterodine and pirenzepine had affinities of 8.2, 8.1 and 6.8, respectively, whilst the M2-selective agent methoctramine had a relatively low affinity (pKB = 6.1). The rank order of affinities was, therefore, 4-DAMP > darifenacin > oxybutynin > tolterodine > pirenzepine > methoctramine for the pig detrusor. Correlation of the antagonist affinities obtained on the bladder with those published for these antagonists at the five muscarinic receptor subtypes identified the M3(m3)-receptor as the muscarinic subtype mediating detrusor contractile responses in vitro. 4. These data suggest that a small population of M3-muscarinic receptors must mediate direct contractile responses of the pig detrusor muscle to muscarinic receptor stimulation in vitro.     

Prejunctional M1 and postjunctional M3 muscarinic receptors in the circular muscle of the guinea-pig ileum

The effects of subtype-selective muscarinic receptor antagonists on electrically evoked release of acetylcholine and muscle contraction were compared in circular muscle preparations of the guinea-pig ileum. Incubation of the preparation with [³H]choline resulted in the formation of [³H]acetylcholine. Electrical stimulation caused the release of [³H]acetylcholine which was abolished by tetrodotoxin and omission of calcium from the medium. 5-Hydroxytryptamine (10 M) and the nicotinic agonist 1,1-dimethyl-4-phenylpiperazinium (300 M) did not change acetylcholine release. The muscarinic antagonists pirenzepine (M1 selective), AF-DX 116 (M2 selective) and hexahydrosiladifenidol (M3 selective) caused concentration-dependent increases in the evoked release of acetylcholine, and inhibitions of the circular muscle contraction. The postjunctional affinity constants (pA₂ values) obtained for hexahydrosiladifenidol (8.06), pirenzepine (6.95) and AF-DX 116 (6.60) identified the muscular receptor as an M3 subtype. Pirenzepine was more potent in facilitating the evoked release than hexahydrosiladifenidol and AF-DX 116. These findings suggest that the release of acetylcholine in the circular muscle is inhibited by M1 muscarinic autoreceptors whereas muscle contraction is mediated by M3 receptors.     

Postjunctional modulation by muscarinic M2 receptors of responses to electrical field stimulation of rat detrusor muscle preparations

1. The aim of the present study was to examine the modulator influence of muscarinic M(2) receptors on responses of rat urinary bladder detrusor muscle evoked by endogenous stimuli, i.e. by stimulation of the bladder innervation. 2. Responses were evoked by electrical field stimulation (EFS; 2-20 Hz, 0.8 ms, 60 V) of isolated strip preparations mounted in organ baths. The tension of the muscle strips was recorded digitally. EFS was performed by applying stimulation with either a short duration (5 s) or a longer duration (to reach peak response; approximately 20 s). 3. Effects of muscarinic receptor antagonists (muscarinic M(1)/M(3) receptor selective: 4-diphenylacetoxy-N-methylpiperidine methobromide (4-DAMP); muscarinic M(2) receptor selective: methoctramine), a beta-adrenergic antagonist (propranolol) and an adenosine receptor antagonist (8-p-sulfophenyltheophylline) were assessed on contractile activity and on poststimulatory relaxations. 4. Low concentrations of methoctramine (10(-8) m) reduced or tended to reduce the EFS-induced contraction, e.g. at 2 Hz by 12% while methoctramine at 10(-7) m had no significant effect. In addition, in the presence of 4-DAMP (10(-9) m), which tended to inhibit contractions at all frequencies (2-20 Hz; -17 to -25%), methoctramine at 10(-8) and 10(-7) m induced a further reduction of the contractile responses (-5 to -10%; 2-20 Hz). 5. The beta-adrenergic receptor antagonist propranolol (10(-6) m) and the adenosine receptor antagonist 8-p-sulfophenyltheophylline (10(-6) m) both increased contractile responses by 9-21% (2-10 Hz, long duration; P < 0.05-0.001) as a consequence of antagonizing relaxatory stimuli. Neither antagonist affected the contractile responses to EFS with the short duration stimulation. Poststimulatory relaxations were reduced by 30-60% (P < 0.05) by propranolol and by 40-60% (P < 0.001) by 8-p-sulfophenyltheophylline, but for 8-p-sulfophenyltheophylline only after stimulation with the short duration. 6. In the presence of methoctramine (10(-7) m), the 8-p-sulfophenyltheophylline-induced increases of the contractile response to long duration EFS were significantly enhanced at 10 Hz (+12 +/- 4%; P < 0.05), whereas no such enhancement of the propranolol inhibitory effect occurred in the presence of methoctramine. However, poststimulatory beta-adrenoceptor-evoked relaxations after short duration EFS were increased by about 35% in the presence of methoctramine, but not those after long duration. 7. Thus, muscarinic M(2) receptor activation inhibits adenosine receptor- and beta-adrenoceptor-evoked relaxations of the rat detrusor muscle. The inhibition occurs via a transient postjunctional mechanism that mainly affects responses with a short latency.     

Bladder instability: a re‐appraisal of classical experimental approaches and development of new therapeutic strategies

1 Despite the growing social interest in human urinary tract disorders, the aetiology of detrusor instability remains poorly understood. Myogenic and neural impairment of detrusor activity caused by CNS or autonomic injuries can results in dysfunctions of normal voiding of the bladder such as urinary incontinence. 2 The contractility of human detrusor smooth muscle is critically dependent on acetylcholine‐induced muscarinic receptor activation. Biochemical and functional in vivo and in vitro studies suggest the presence of an heterogeneous population of muscarinic receptor subtypes (M₁–M₄) localized at muscular and neutral sites. There is increasing evidence on the prejunctional auto‐ and hetero‐regulation of acetylcholine release from parasympathetic nerve endings in modulating detrusor muscle contraction during micturition. 3 Activation of P2X purinoreceptors closely associated with the parasympathetic varicosities seems to be implicated to varying extent in the contractility in normal or instable human detrusor. Interestingly, P2X₁ subtype expression on smooth muscle increases considerably in the symptomatically obstructed bladder. A striking absence of P2X₃ and P2X₅ subtypes was observed in the cholinergic innervation of detrusor from patients with urgent incontinence. Thus, it is likely that alteration of the neural acetylcholine control can play a critical role in pathological states. 4 If the failures in storage and voiding can be recognized urodynamically, considerable difficulties remain in investigating the underlying functional changes especially because the study of the pathophysiology requires techniques that can be justified in animals but not in humans. 5 Recently, to solve this problem an alternative technique using human smooth muscle cells in culture has been developed. Human cell lines may be relevant in investigating the molecular pathways in physiological and pathological conditions. 6 The potential development of novel molecular therapeutic strategies such as gene therapy and tissue engineering is also discussed.     

Muscarinic receptors of the urinary bladder: detrusor,urothelial and prejunctional

1. The parasympathetic nervous system is responsible for maintaining normal bladder function, contracting the bladder smooth muscle (detrusor) and relaxing the bladder outlet during micturition. 2. Contraction of the bladder involves direct contraction via M3 receptors and an indirect 're-contraction' via M2-receptors whereby a reduction in adenylate cyclase activity reverses the relaxation induced by beta-adrenoceptor stimulation. 3. Muscarinic receptors are also located on the epithelial lining of the bladder (urothelium) where they induce the release of a diffusible factor responsible for inhibiting contraction of the underlying detrusor smooth muscle. The factor remains unidentified but is not nitric oxide, a cyclooxygenase product or adenosine triphosphate. 4. Finally, muscarinic receptors are also located prejunctionally in the bladder on cholinergic and adrenergic nerve terminals, where M1-receptors facilitate transmitter release and M2 or M4-receptors inhibit transmitter release. 5. In pathological states, changes may occur in these receptor systems resulting in bladder dysfunction. Muscarinic receptor antagonists are the main therapeutic agents available for treatment of the overactive bladder, but whether their therapeutic effect involves actions at all three locations (detrusor, prejunctional, urothelial) has yet to be established.     

Tramadol inhibits the contractility of isolated caprine detrusor muscle

1 The atypical opioid analgesic tramadol has been shown to provide beneficial clinical and urodynamic effects in patients with detrusor overactivity. The effect of tramadol on isolated detrusor muscle has not been studied. This study investigated the ability of tramadol to inhibit acetylcholine (ACh)‐induced contractility of the isolated caprine (goat) detrusor muscle. The effect of three concentrations (30, 100 and 300 μm ) of tramadol on 10 caprine detrusor strips contracted by the addition of 100, 200 or 400 μm ACh was studied. The sensitivity of tramadol‐induced inhibition of ACh responses to treatment with the β‐adrenoceptor antagonist propranolol (1 μm ) and the opioid receptor antagonist naloxone (100 μμ ) was also studied. 2 Tramadol caused a concentration‐dependent inhibition of ACh‐induced detrusor contraction that was reversed by raising the concentration of ACh. Propranolol, but not naloxone, reversed the tramadol‐induced inhibition of contractions to ACh in the detrusor. 3 These results suggest that tramadol inhibits ACh‐induced contractility of the isolated detrusor. They also suggest that tramadol does so by an indirect anticholinergic mechanism involving the stimulation of β‐adrenoceptors. Tramadol may be useful in managing clinical conditions requiring relaxation of the detrusor muscle. Although the concentrations of tramadol needed to relax the detrusor were relatively high, these could be clinically attained via intravesical administration.     

Muscarinic M3 receptors mediate contraction of human central and peripheral airway smooth muscle

The muscarinic receptor subtype involved in human airway smooth muscle contraction was characterised for the first time, using subtype-selective muscarinic antagonists. It was demonstrated that methacholine-induced contraction of central (trachea) and peripheral (small bronchi) airway smooth muscle preparations was antagonised by pirenzepine, AF-DX 116, 4-DAMP methobromide, hexahydrosiladifenidol, and methoctramine with pA₂-values characteristic of M₃ (smooth muscle/glandular) muscarinic receptors. Since these pA₂-values demonstrate significant correlations with those found in bovine and guinea-pig tracheal smooth muscle contraction, it is concluded that these animal tissues provide a good model for the study of M₃ subtype-selective muscarinic antagonists to be used as bronchodilators.     

Modification of cardiovascular responses to adenosine A1 receptor stimulation in the posterior hypothalamus of anaesthetized rats by cAMP and by GABAB receptor blockade

1 Injection of N⁶‐cyclohexyladenosine (CHA; 1, 5 and 10 nmol), an adenosine A₁ receptor agonist, into the posterior hypothalamus of rats produced a dose‐dependent decrease in blood pressure (BP) and heart rate (HR). 2 Pretreatment with 8‐cyclopentyl‐1,3‐dimethylxanthine (CPDX; 50 nmol), an adenosine A₁ receptor antagonist, blocked the depressor and bradycardic effects of CHA (10 nmol). 3 Pretreatment with 8‐bromo‐cyclic adenosine monophosphate (AMP) (10 nmol), a cAMP analogue, attenuated the depressor and bradycardic effect of CHA (10 nmol); 8‐bromo‐cyclic guanosine monophosphate (GMP) (10 nmol), a cGMP analogue, did not modify those effects of CHA. 4 In addition, pretreatment with 5‐aminovaleric acid (25 nmol), a γ‐aminobutyric acid (GABA)_B receptor antagonist, attenuated the depressor and bradycardic effects of CHA (10 nmol). 5 These results suggest that adenosine A₁ receptors in the posterior hypothalamus have an inhibitory role in the central cardiovascular regulation and that these vasodepressive and bradycardic actions are modified by raised cAMP and by GABA_B receptor inhibition.     

Effects of phorbol ester on carbachol-induced contraction in bovine ciliary muscle: possible involvement of protein kinase C

The aim of the research was to characterize muscarinic receptors of bovine ciliary muscle and to investigate the desensitization process. The role of protein kinase C was analyzed. The results show that muscarinic receptors of bovine ciliary muscle have the pharmacological characteristics of the M₃ subtype. Acute exposure to phorbol esters (1 μM phorbol 12,13-dibutyrate, PDB, or 0.1 μM phorbol 12-myristate 13-acetate, PMA, for 15 and 5 min, respectively) resulted in antagonism of muscarinic receptor-mediated contraction. Long-term pretreatment (18 h) with PMA to down-regulate protein kinase C resulted in potentiation of carbachol-induced contraction, reduction of agonist-induced desensitization and loss of phorbol ester-induced desensitization. Staurosporine (3 μM) and H₇ [1-(5-isoquinolinesulfonyl)-2-methyl-piperazine] (1 μM), protein kinase C inhibitors, produced a significant potentiation of the contractile effect of carbachol, reduced the desensitization produced by repeated addition of carbachol and suppressed that induced by phorbol esters. In vitro incubation with carbachol, PDB or PMA did not cause any modification of the binding of labeled [³H]quinuclidinyl benzilate. In vitro incubation with PDB and PMA produced, as expected, a significant translocation of protein kinase C from the cytosol to the membrane. The incubation of the ciliary muscle with carbachol, using the protocol of exposure that induced maximal desensitization of contractile responses, produced a significant redistribution of the enzyme from the cytosol to the membrane. These findings suggest that agonist-induced modulation of functional cholinergic sensitivity in ciliary muscle is correlated, at least partially, to the translocation of protein kinase C from the cytosol to the membrane. The desensitization by phorbol esters is completely due to protein kinase C activation; during the desensitization process, direct modification of the density and affinity of muscarinic receptors is not involved.     

Characterization of postjunctional muscarinic receptors mediating contraction in rat anococcygeus muscle

The present study was designed to characterize the postjunctional muscarinic receptors mediating contraction in rat anococcygeus muscle by means of a series of muscarinic agonists and subtype-preferring key muscarinic antagonists. Cumulative addition of muscarinic agonists elicited concentration-dependent contractions with the following rank order of potency (pD₂ values): (+)-muscarine (6.36) ≥ oxotremorine M (6.21) ≥ arecaidine propargyl ester (APE) (6.18) > carbachol (5.68)=(±)-methacholine (5.65) > 4-(4-chlorophenyl-carbamoyloxy)-2-butynyltrimethylammonium chloride (4-Cl-McN-A-343) (4.28) > 4-(3-chlorophenylcarbamoyloxy)-2-butynyltrimethylammonium chloride (McN-A-343) (3.89). (+)-Muscarine, oxotremorine M, carbachol and (±)-methacholine behaved as full agonists, whereas APE, 4-Cl-McN-A-343 and McN-A-343 displayed partial agonism. The contractile responses of the rat anococcygeus muscle to (±)-methacholine were competitively antagonized by pirenzepine (pA₂=6.92), 11-[[4-[4-(diethylamino)butyl]-1-piperidinyl]acetyl] 5,11-dihydro-6H-pyrido(2,3-b)(1,4)-benzodiazepine-6-one (AQ-RA 741; pA₂=6.75), himbacine (pA₂=7.11), (±)-p-fluoro-hexahydro-sila-difenidol (p-F-HHSiD; pA₂=7.68) and the (R)- and (S)-enantiomers of hexahydro-difenidol [(R)-HHD: pA₂=8.52; (S)-HHD: pA₂=6.06]. A comparison of the pA₂ values derived from studies of contraction in rat anococcygeus muscle with literature binding (pK_i values) and functional affinities (pA₂ values) obtained at native M₁-M₄ receptors strongly suggests that the postjunctional muscarinic receptors mediating contraction in rat anococcygeus muscle are of the M₃ subtype. Received: 18 April / Accepted: 18 July 1997     

Neuronal nitric oxide synthase activity in rat urinary bladder detrusor: participation in M3 and M4 muscarinic receptor function

1. The aim of this paper was to determine the different signalling cascades involved in contraction of the rat urinary bladder detrusor muscle mediated via muscarinic acetylcholine receptors (muscarinic AChR). Contractile responses, phosphoinositides (IPs) accumulation, nitric oxide synthase (NOS) activity and cyclic GMP (cGMP) production were measured to determine the reactions associated with the effect of cholinergic agonist carbachol. The specific muscarinic AChR subtype antagonists and different inhibitors of the enzymatic pathways involved in muscarinic receptor-dependent activation of NOS and cGMP were tested. 2. Carbachol stimulation of M(3) and M(4) muscarinic AChR increased contractility, IPs accumulation, NOS activity and cGMP production. All of these effects were selectively blunted by 4-DAMP and tropicamide, M(3) and M(4) antagonists respectively. 3. The inhibitors of phospholipase C (PLC), calcium/calmodulin (CaM), neuronal NOS (nNOS) and soluble guanylate cyclase, but not of protein kinase C and endothelial NOS (eNOS), inhibited the carbachol action on detrusor contractility. These inhibitors also attenuated the muscarinic receptor-dependent increase in cGMP and activation of NOS. 4. In addition, sodium nitroprusside and 8-bromo-cGMP, induced negative relaxant effect. 5. The results obtained suggest that carbachol activation of M(3) and M(4) muscarinic AChRs, exerts a contractile effect on rat detrusor that is accompanied by an increased production of cGMP and nNOS activity. The mechanism appears to occur secondarily to stimulation of IPs turnover via PLC activation. This in turn, triggers cascade reactions involving CaM, leading to activation of nNOS and soluble guanylate cyclase. They, in turn, exert a modulator inhibitory cGMP-mediated mechanism limiting the effect of muscarinic AChR stimulation of the bladder.     

M(3) muscarinic receptors mediate contraction of human urinary bladder

Since muscarinic receptors appear to be the physiologically most important control system for urinary bladder contraction, we have characterized the receptor subtype mediating contraction in response to the muscarinic agonist carbachol in the human bladder. Experiments were based on four antagonists, the non-selective atropine, the M(1)-selective pirenzepine, the M(2)-selective methoctramine and the M(3)-selective darifenacin. All antagonists yielded Schild-plots with a slope close to unity. The order of potency (atropine> or =darifenacin>pirenzepine>methoctramine) as well as the estimated antagonist affinities suggested that contraction of the human bladder occurs predominantly if not exclusively via the M(3) receptor.     

Long-term nitric oxide deficiency causes muscarinic supersensitivity and reduces beta(3)-adrenoceptor-mediated relaxation, causing rat detrusor overactivity

BACKGROUND AND PURPOSE: Overactive bladder is a complex and widely prevalent condition, but little is known about its physiopathology. We have carried out morphological, biochemical and functional assays to investigate the effects of long-term nitric oxide (NO) deficiency on muscarinic receptor and beta-adrenoceptor modulation leading to overactivity of rat detrusor muscle. EXPERIMENTAL APPROACH:Male Wistar rats received N(omega)-nitro-L-arginine methyl ester (L-NAME) in drinking water for 7-30 days. Functional responses to muscarinic and beta-adrenoceptor agonists were measured in detrusor smooth muscle (DSM) strips in Krebs-Henseleit solution. Measurements of [(3)H]inositol phosphate, NO synthase (NOS) activity, [(3)H]quinuclidinyl benzilate ([(3)H]QNB) binding and bladder morphology were also performed. KEY RESULTS:Long-term L-NAME treatment significantly increased carbachol-induced DSM contractile responses after 15 and 30 days; relaxing responses to the beta(3)-adrenoceptor agonist BRL 37-344 were significantly reduced at 30 days. Constitutive NOS activity in bladder was reduced by 86% after 7 days and maintained up to 30 days of L-NAME treatment. Carbachol increased sixfold the [(3)H]inositol phosphate in bladder tissue from rats treated with L-NAME. [(3)H]QNB was bound with an apparent K(D) twofold higher in bladder membranes after L-NAME treatment compared with that in control. No morphological alterations in DSM were found. CONCLUSIONS AND IMPLICATIONS:Long-term NO deficiency increased rat DSM contractile responses to a muscarinic agonist, accompanied by significantly enhanced K(D) values for muscarinic receptors and [(3)H]inositol phosphate accumulation in bladder. This supersensitivity for muscarinic agonists along with reductions of beta(3)-adrenoceptor-mediated relaxations indicated that overactive DSM resulted from chronic NO deficiency.     

人结直肠癌平滑肌组织毒蕈碱受体特性的变化

随着受体理论和研究技术的发展 ,人们开始从受体的变化去探寻疾病发生和发展的根源 ,并从受体水平诊治疾病。最近人们发现ＭＡＣｈ Ｒ与肿瘤之间关系密切。一方面 ,ＭＡＣｈ Ｒ过度活化可导致细胞增殖、恶性转化及肿瘤发生[1] ;另一方面 ,在肿瘤状态下ＭＡＣｈ Ｒ的特性也发生变化。本文主要探讨肿瘤状态下ＭＡＣｈ Ｒ特性的变化。1　材料与方法1.1　临床资料　结直肠癌病人 10例 ,男 6例 ,女 4例。中国医科大学附属第一医院肿瘤科提供。1.2　标本采集　结直肠癌患者手术切除的标本 ,分别取癌、癌旁及正常组织各一块。癌旁组织取距癌灶约 2…     

Phenyl‐substituted N6‐phenyladenosines and N6‐phenyl‐5′‐N‐ethylcarboxamidoadenosines with high activity at human adenosine A2B receptors

A series of phenyl‐substituted N⁶‐phenyladenosines and N⁶‐phenyl‐5′‐N‐ethylcarboxamidoadenosines were synthesized and tested at adenosine receptor subtypes. EC₅₀ values were determined for cyclic AMP production in CHO cells expressing human A_2B receptors. Binding affinities were determined for rat A₁ and A_2A receptors and human A₃ receptors. N⁶‐phenyladenosine displayed an EC₅₀ value at A_2B receptors of 6.3 μM. Several N⁶‐phenyladenosine derivatives were more active than N⁶‐phenyladenosine, while two analogs were also more potent than 5′‐N‐ethylcarboxamidoadenosine (NECA, 0.76 μM), i.e., the 4‐iodophenyl ( 10 , 0.37 μM) and the 4‐aminosulfonylphenyl ( 20 , 0.44 μM) derivatives. N⁶‐phenyl‐NECA derivatives were as active as their analogous adenosine derivatives. Drug Dev. Res. 49:85–93, 2000. © 2000 Wiley‐Liss, Inc.