北京鸭红细胞膜β肾上腺素受体放射配基结合测定法

本文以[~3H]dihydroalprenolol([~3H]DHA)为放射配基。对北京鸭红细胞(RBC)膜β受体进行了系统研究。结果表明,[~3H]DHA与北京鸭RBC结合具有饱和性,肾上腺素能激动剂与[~3H]DHA竞争结合的强弱顺序与它们的生物活性一致,提示结合有结构特异性和立体特异性,[~3H]DHA与鸭RBC结合迅速、可逆。可见北京鸭RBC膜存在β受体,用于放射配基结合测定有取材方便,受体较丰富,膜蛋白得率高,[~3H]DHA非特异结合低和易于保存等优点,可以代替火鸡RBC。     

新抗胆碱药[3H]三环哌酯对人脑M受体的作用

用放射配体受体结合试验法,研究了新化合物三环哌酯与人大脑皮质M受体的结合特性,并与QNB作了比较。饱和实验结果显示,[³H]三环哌酯的结合参数与[³H]QNB相近,两种配体的作用均符合单位点模型。竞争性抑制实验结果表明二者作用强度相当。[³H]三环哌酯的结合和解离速率常数均较[³H]QNB大,且其与皮质M受体的解离受季铵酚的变构调节,结果提示,两种配体与M受体有一些不同的结合特性,在M受体研究中,[³H]三环哌酯可以作为[³H]QNB的补充工具。     

氚标记强效镇痛剂N-[1-(β-羟基-β-苯乙基)-3-甲基-4-哌啶基]-N-丙酰苯胺的制备以及与阿片受体的结合试验

本文报道以N-[1-(对-溴苯甲酰甲基)-3-甲基-4-哌啶基]-N-丙酰苯胺(Ⅲ)为前体,以PdO/BaSO₄作催化剂,用氚气进行卤—氚置换,氚化还原羰基的反应。反应产物经硅胶纸层析纯化后,用甲基橙比色法定量测定,得到N-{1-[β-羟基-β-氚-β-(对-氚苯基)乙基]-3-甲基-4-哌啶基}-N-丙酰苯胺(Ⅳ,[³H]F-7302),其比放射性为59 Ci/mM,放化纯度为98%。[³H]F-7302与小鼠脑内阿片受体的特异性结合在浓度为4.5×10^-9M时达到饱和,解离常数Kd=1.25×10^-9M,最大结合量B_max=93.08×10¹²M/g蛋白,其特异性结合与非特异性结合比值达10～15。     

可乐定镇痛与中枢Ca2+的关系

用大鼠甩尾法和放射配基结合实验,探讨了可乐定镇痛与中枢Ca²⁺的关系。CaCl₂(1μmol/rat,icv)和EGTA(0.2μmol/rat,icv)分别拮抗和增强可乐定(1mg/kg,sc)的镇痛。戊脉安(0.1μmol/rat,icy)对可乐定(1 mg/kg,sc)镇痛无明显影响,但可部分翻转CaCl₂对可乐定镇痛的拮抗。CaCl₂(1×10^-3mol)对[³H]-可乐定结合无明显抑制。结果表明可乐定镇痛与脑室周围组织中Ca²⁺浓度变化密切相关,Ca²⁺至少部分需经对戊脉安敏感的钙通道进入细胞内方可拮抗可乐定镇痛。推沦:可乐定镇痛与神经元内Ca²⁺有关。     

新的高强度高选择性阿片μ受体激动剂[11C]-羟甲芬太尼的合成

羟甲芬太尼(I)是一个新的高强度高选择性阿片μ受体激动剂。本文用cis-A-N-[1-(2-羟基-2-苯乙基)-3-甲基-4-哌啶基]-苯胺(II)或cis-N-[1-(苯甲酰甲基)-3-甲基-4-哌啶基]-苯胺(III)作为前体合成了[¹¹C]-羟甲芬太尼,以便用正电子发射断层扫描(PET)来观察μ受体。通过水解cis-A-羟甲芬太尼(I)和cis-N-[1-(苯甲酰甲基)-3-甲基-4-哌啶]-N-苯基丙酰胺(cis-IV)的4-N-丙酰基分别获得II和III。溴乙烷的格氏试剂与回旋加速器产生的[¹¹C]-二氧化碳反应后继而直接加入邻苯二甲酸二酰氯和2,6-二叔丁基吡啶生成同位素标记中间体[¹¹C]-丙酰氯。[¹¹C]-丙酰氯与OH-前体(II)反应后再经HPLC分离纯化直接得[¹¹C]-羟甲芬太尼;[¹¹C]-丙酰氯与酮-前体(III)反应后,再用硼氢化钠甲醇溶液处理,然后进行HPLC分离纯化得[¹¹C]-羟甲芬太尼。两种方法均可获得ll.1～14.8GBq/μmol的特异性放射化学纯[¹¹C]-羟甲芬太尼。总共耗时为40～50min(EOB)。     

13C核磁共振技术测定鱼肝油中ω-3脂肪酸α(1,3)-酰基和β(2)-酰基的位次分布

目的 应用¹³C核磁共振（¹³C-NMR）光谱测定鱼肝油中ω-3脂肪酸[十八碳四烯酸（C18：4 n-3,moroctic acid,MA）、二十碳五烯酸（C20：5 n-3,eicosapentaenoic acid,EPA）、二十二碳六烯酸（C22：6 n-3,docosahexaenoic acid,DHA）]形成的甘油三脂中α（1,3）-酰基、β（2）-酰基的位次分布。方法 用CDCl₃溶解样品,利用高分辨核磁共振光谱直接测定。结果 鱼肝油中MA、EPA和DHA α（1,3）-酰基、β（2）-酰基在δ 171.5~173.5 ppm的化学位移与文献报道基本一致。2批次鱼肝油未发现上述ω-3脂肪酸特征峰,4批次鱼肝油有特征峰检出,但是β（2）-酰基的位次分布不同。按照β（2）-酰基的比例,4批鱼肝油来源于家养鱼类提取的可能性较大。结论 应用该¹³C核磁共振技术可以鉴别鱼肝油的优劣,方法简单、快捷。     

蝙蝠葛碱和粉防己碱对[3H]WEB 2086与体外牛脑前动脉平滑肌细胞结合的影响(英文)

用标记的血小板活化因子拮抗剂[³H]WEB 2086,在培养的牛脑前动脉平滑肌细胞上鉴定了血小板活化因子受体。结果表明在25℃时该细胞上存在两种与配基具有不同亲和力的受体结合位点,其中K_d-1=22.8±5.0 nmol·L^-1,K_d-2=186+20.5 nmol·L^-1;B_max-1=2.1±0.3 pmol/10⁴细胞,B_max-2=12.1±1-5 pmol/10⁶细胞。蝙蝠葛碱和粉防己碱均能抑制[³H]WEB2086与上述细胞的结合。     

四氢巴马汀等异喹啉生物碱对突触体及囊泡摄取[3H]多巴胺的影响

本文报道四氢巴马汀(THP)等异喹啉生物碱对[³H]DA摄取的作用。突触体对[³H]DA的亲和力常数Km为0.23μmol。最大摄取速率Vmax为1.2 nmol/g(5 min)。d-THP,1-SPD和1-THP对突触体摄取[³H]DA均有抑制作用,其IC₅₀分别为0.44,2.24和18.5μmol。d-THP的作用比1-THP约强20倍。nomifensine,苯丙胺和氟哌啶醇亦能有效地抑制[³H]DA摄取(IC₍₅₀₎分别为0.05,0.27和3.18μmol)。动力学研究表明,d-THP和nomifensine为DA摄取的竞争性抑制剂。用低渗溶液处理溶胀的方法研究药物对递质贮存的作用发现,与利血平相似,d-THP显著降低贮存囊泡[³H]DA含量并使囊泡与突触体[³H]DA含量之比明显减小。实验结果表明,THP等能抑制突触体摄取,并直接作用于贮存囊泡抑制[³H]DA贮存,因此具有利血平样排空作用。     

7α-和7β-甲基-10β,17β-二乙酰氧基-△4-雌甾烯-3酮的抗早孕作用

7α-和7β-甲基-10β,17β-二乙酰氧基-△⁴-雌甾烯-3酮(简称7α-和7β-甲-乙氧雌酮)对小鼠抗早孕ED₅₀分别为1.6和5.5 mg/kg。7α-甲-乙氧雌酮在大鼠也有抗早孕作用并使血浆孕酮浓度降低,应用10 μg/ml浓度能抑制离体妊娠大鼠卵巢孕酮合成。7α-和7β-甲-乙氧雌酮与兔子宫胞浆雌二醇受体的相对结合亲和力(RBA)分别为10.8和1.5,与孕酮受体的RBA均<1.7α-和7β-甲-乙氧雌酮都有较弱的雌激素和抗雌激素活性。     

远程二维核磁共振和NOE差谱法研究新生物碱马尾杉碱N的结构

自华南马尾杉[Phlegariurus fordii(Baker)Ching]中分得新生物碱—马尾杉碱N(Ⅰ)。它含有七个季碳。本工作采用远程¹³C—¹HCOSY和远程增强型¹H—¹HCOSY新技术与NOE差谱相配合的方法成功地连接被季碳和杂原子分隔开的自旋系统,推定了其结构。     

Influence of eicosanoids on serotonin release in the rat brain: inhibition by prostaglandins E1 and E2

Summary Cardiac alpha- and beta-adrenoceptor sensitivities were examined after chronic pretreatment of rats with reserpine. Increases in sensitivity would indicate that the receptor is under the influence of the sympathetic innervation, removal by catecholamine depletion with reserpine of the tonic effect of neurotransmitter release would permit receptor upregulation. The positive inotropic responses of paced left atria and papillary muscles and the positive chronotropic responses of spontaneously beating right atria were recorded. A concentration-response curve to isoprenaline (beta-adrenoceptor-mediated) was followed, in the presence of beta-blockade, by one to methoxamine (alpha-adrenoceptor-mediated). Methoxamine exerted positive inotropy of left atria and papillary muscles, the maxima being 43.2 ± 2.7 and 26.8 ± 4.4% of the isoprenaline maxima. A small positive chronotropy (16.5 ± 5.6% maximum) of right atria occurred. After pretreatment with reserpine (1.0 mg kg^–1 i.p. daily) for 7 days, the three preparations displayed supersensitivity to isoprenaline, revealed as a significant displacement (P < 0.05) of the concentration-response curves to the left of those for control rats. Reserpine pretreatment, however, had no effect on the sensitivity to methoxamine. The increase in beta-adrenoceptor sensitivity to isoprenaline after reserpine pretreatment was accompanied by a significant 41.3% increase (P < 0.05) in the number of [³H]-dihydroalprenolol ([³H]-DHA) binding sites (B _max) in ventricular membranes, although the dissociation constant (_{K D}) was unaffected. There were more alpha-adrenoceptor [³H]-prazosin binding sites in ventricular than atrial membranes. However, there was no difference in K _D or B _max between reserpine-pretreated and control tissues. It is proposed that the increase in beta-adrenoceptor sensitivity and binding sites arises from the depletion-induced loss of neuronal noradrenaline release onto the beta-adrenoceptors which are therefore directly under the influence of the neurotransmitter. The failure of cardiac alpha-adrenoceptors to exhibit supersensitivity and increased numbers suggests that they are not directly affected by the sympathetic innervation. Send offprint requests to K. J. Broadley at the above address     

Comparison of three radioligands for the labelling of human β-adrenoceptor subtypes

We have compared the ability of three radioligands, [¹²⁵I]-cyanopindolol, [³H]-CGP 12,177 and [³H]-dihydroalprenolol, to label the three human β-adrenoceptor subtypes. Saturation and competition binding experiments were performed using membrane preparations from Chinese hamster ovary cells stably transfected with the three subtypes. While [³H]-CGP 12,177 had very similar affinity for β₁- and β₂-adrenoceptors (about 40 pM), [¹²⁵I]-cyanopindolol and [³H]-dihydroalprenolol had 4- to 6-fold higher affinity for β₂- as compared to β₁-adrenoceptors (10 vs 45 and 187 vs 1,021 pM, respectively). The affinity of [¹²⁵I]-cyanopindolol at β₃-adrenoceptors was considerably lower (440 pM) than at the other two subtypes. The β₃-adrenoceptor affinity of [³H]-CGP 12,177 and [³H]-dihydroalprenolol was so low that it could not be estimated within the tested range of radioligand concentrations (up to 4,000 pM and 30,000 pM for [³H]-CGP 12,177 and [³H]-dihydroalprenolol, respectively). We conclude that all three radioligands are ill-suited to label β₃-adrenoceptors, particularly in preparations co-expressing multiple subtypes. In the absence of alternatives, [¹²⁵I]-cyanopindolol appears the least unsuitable to label β₃-adrenoceptors. There is a need for high-affinity radioligands which are either selective for β₃-adrenoceptors or reasonably non-selective among all three β-adrenoceptor subtypes.     

Properties of agonist binding at theβ-adrenoceptor of the rat reticulocyte

Summary To study the fundamental differences between agonist and antagonist interaction with the -adrenoceptor of the rat reticulocyte the radiolabeled agonist³H hydroxybenzylisoprenaline (³H HBI) and the radiolabeled antagonist³H dihydroalprenolol (³H DHA) were used.Equilibrium binding experiments with³H HBI revealed all characteristics expected to a -adrenoceptor site, i. e. high affinity binding (K _{D high}=7.4±0.9×10^–9 M), saturability (B _{max high}=230±24 fmoles/mg protein), and stereoselectivity. The rank order of potency for competing agonists was isoprenaline > adrenaline > noradrenaline > dopamine.³H HBI high affinity binding sites amounted to about 25% of -adrenoceptor sites detectable with³H DHA.In competition experiments with³H HBI and (-)isoprenaline[(-)Ipn]aK _{D high}-value for (-)Ipn of 3.1±0.6×10^–8M was obtained corresponding to theK _{D high}-value of (-)Ipn obtained from competition experiments using³H DHA. For (-)propranololK _D-values of 0.9±0.5×10^–8 M and 1.0 ±1.0×10^–8 M were measured using³H HBI and³H DHA respectively.Agonist affinity derived from competition experiments with (-)Ipn versus³H DHA was not affected by temperature changes.Guanylyl-imidodiphosphate [Gpp(NH)p] decreased concentration dependently the number of high affinity binding sites of³H HBI not affecting the respectiveK _D-value. Similar effects were observed after omission of Mg²⁺ from the binding assay or inclusion of Na⁺ in the Mg²⁺-free incubation mixture.The association reaction of³H HBI at the -adrenoceptor revealed two different velocities. The slower phase of the association reaction which represents high affinity binding (80% of equilibrium binding) is not observed in the presence of Gpp(NH)p.A biphasic dissociation of³H HBI binding was induced by 10^–4 M (±)propranolol: 25% dissociated with at _1/2 of 1.3 min whereas the high affinity binding was reversed with at _1/2 of 150 min. This slowly reversible binding of³H HBI however was rapidly reversed by Gpp(NH)p (t _1/2<1 min).It is concluded that the agonist ligand³H HBI permits a direct qualitative and quantitative characterization of the agonist induced high affinity state of the -adrenoceptor. In particular, the kinetic studies strongly support a two step binding model for the agonist--adrenoceptor interaction.This work was supported by a grant from the Deutsche ForschungsgemeinschaftParts of this work were presented at the Spring Meetings of the German Pharmacological Society (Wiemer et al. 1978, 1981 b)Herrn Professor Dr. med. Hans Herken, Pharmakologisches Institut der Freien Universität Berlin, zum 70. Geburtstag gewidmet.     

Platelet 5-HT uptake sites in depression: three concurrent measures using [3H] imipramine and [3H] paroxetine

Platelet 5-HT uptake sites were measured in 40 depressed patients and 40 controls using [³H] imipramine binding, defined with desmethylimipramine (DMI) and Na⁺ dependence, and [³H] paroxetine binding. In control subjects the B_max of DMI defined [³H] imipramine binding was significantly higher than both Na⁺ dependent [³H] imipramine (by 30%) and [³H] paroxetine binding (by 22%). The B_max of Na⁺ dependent [³H] imipramine and [³H] paroxetine binding did not differ significantly. The K_d of Na⁺ dependent [³H] imipramine binding was significantly lower than the K_d of DMI defined [³H] imipramine binding. The binding of DMI defined and Na⁺ dependent [³H] imipramine and [³H] paroxetine did not differ significantly between depressed patients and controls in the total group, in those depressed patients who had never taken antidepressants or in those depressed patients who had been recently with-drawn from antidepressants. This study provides no support for the view that the number of platelet 5-HT uptake sites are reduced in depression.     

甲基黄酮醇胺盐酸盐对β受体的阻断作用

The β-receptor blocking action of methylflavonolamine hydrochloride(MFA) was studied and compared with those of propranolol. The doseresponse curves of isoproterenol were shifted to the right by MFA on isolated rabbit atrium in this experiment. The pA₂ value and the slope of the regression line of MFA calculated from Schild plot were 5.53 and -0.84 respectively. The effects of MFA and propranolol on duck erythrocyte membranes were studied by the radioligand binding method. Both MFA and propranolol inhibited the binding of [³H] dihydroalprenolol to β-receptors. Their apparent equilibrium dissociation constants were 1.12×10^-5 mol/L and 5.50×10^-9 mol/L respectively. The affinity of propranolol to β-receptors of duck erythrocyte membranes was 2039-fold higher than that of MFA. These results demonstrates that MFA is a weak competitive β-receptor blocking agent.     

Dopamine D2 receptors selectively labeled by a benzamide neuroleptic: [3H]-YM-09151-2

Summary In order to label dopamine D₂ receptors selectively we tritiated the potent benzamide neuroleptic, YM-09151-2 (26.7 Ci/mmol). The binding of [³H]-YM-09151-2 to canine striatal membranes was saturable and specific with a K _D of 57 pmol/l and B _max of 36 pmol/g tissue as determined by Scatchard analysis. The K _D, but not the B _max, of [³H]-YM-09151-2 increased 6-fold in the absence of sodium chloride. [³H]-YM-09151-2 labeled 40% more sites than [³H]-spiperone in the same tissue homogenate. [³H]-YM-09151-2 binding was inhibited by dopaminergic drugs in a concentration and stereoselective manner with the appropriate dopamine D₂ receptor profile. Thus, dopamine agonists inhibited [³H]-YM-09151-2 binding to canine striatal membranes with the following rank order of potency: (–)-N-n-propylnorapomorphine > apomorphine > (±)-6,7-dihydroxy-2-aminotetralin > (+)-N-n-propylnorapomorphine > dopamine > (–)-noradrenaline > serotonin > (–)-isoprenaline. Dopaminergic antagonists competed for [³H]-YM-09151-2 binding with the following order of potency: spiperone > (+)-butaclamol > haloperidol > clebopride > (–)-sulpiride > SCH-23390 > (–)-butaclamol. Furthermore, dopamine agonists recognized 2 states of the receptor labeled by [³H]-YM-09151-2, D ₂ ^high and D ₂ ^low . The D ₂ ^high state of the receptor could be converted to D ₂ ^low by guanine nucleotides and sodium ions as is the case for [³H]-spiperone binding to D₂ receptors. [³H]-YM-09151-2 appears to be a more selective ligand for dopamine D₂ receptors than [³H]-spiperone, since YM-09151-2 displays approximately 9-fold lower affinity than spiperone for cortical serotonergic (S₂) receptors. [³H]-YM-09151-2 may become a useful tool for the selective characterization of dopamine D₂ receptors.Abbreviations used (±)ADTN (±)-2-amino-6,7-dihydroxy-1,2,3,4-tetrahydronaphthalene - NPA N-n-propylnorapomorphine - Gpp(NH)p 5-guanylylimidodiphosphate     

Characterization of the [3H]-desipramine binding site of the bovine adrenomedullary plasma membrane

Summary The specific (i.e. nisoxetine-sensitive) binding of [³H]desipramine was studied in membranes prepared from bovine adrenal medullae. (1) [³H]desipramine bound reversibly and with high affinity (K _D = 2.8 nmol/l) to a single class of non-interacting binding sites (Hill coefficient = 0.96); the maximal number of binding sites (B_max) was 2.1 pmol/mg protein. (2) Binding of [³H]desipramine was dependent on [Na⁺] and [Cl^–]. Increasing the concentrations of these ions increased binding. (3) Substrates and inhibitors of the neuronal noradrenaline transport system (uptake,) inhibited binding of [³H]desipramine with a rank order of potency typical for an interaction with the uptake, carrier.The characteristics of [³H]desipramine binding remained essentially unchanged after solubilization of adrenomedullary membranes with the non-ionic detergent digitonin.The results indicate that the plasma membrane of bovine adreno-medulary cells is endowed with the neuronal uptake₁ transporter. Correspondence to: H. Bönisch     

Pharmacological comparison of the sigma recognition site labelled by [3H]haloperidol in human and rat cerebellum

Summary The radioligand binding characteristics of [³H]haloperidol (in the presence of spiperone, 25 nmolL^–1) were investigated in rat and human cerebellar membranes.In both rat and human cerebellar membrane preparations saturation studies with [³H]haloperidol (non-specific binding defined by pentazocine, 10 molL^–1) demonstrated high affinity saturable specific binding to a homogenous population of binding sites (rat, Bmax 6693 ± 1242 fmol mg^–1 protein, pK_D 8.33 ± 0.08; human, Bmax 2550 ± 437 fmol mg^–1 protein, pK_D 8.59 ± 0.11; mean ± SEM, n = 3–6). Competition studies employing a wide range of structurally diverse competing compounds displayed that the [³H]haloperidol binding site was pharmacologically similar in both preparations and comparable to sigma recognition sites previously identified in various tissues originating from different species. In addition, with reference to the potential subtypes of sigma recognition sites, the labelling of these sites by low nanomolar concentrations of [³H]haloperidol provides evidence that they belong to the sigma-1 recognition site subtype.The present findings suggest that the pharmacology of the rat and human cerebellar sigma recognition site are directly comparable and provides further supporting evidence towards the use of [³H]haloperidol radioligand binding studies in the rat to detect sigma receptor ligands with potential therapeutic activity. Send offprint requests to: N.M. Barnes at the above address     

Binding studies with [3H]cis-methyldioxolane in different tissues

Summary Special conditions - tricine buffer containing Ca²⁺ and Mg²⁺, 22°C (TCM) — allow to label a much higher proportion of muscarinic receptors by [³H]cis-methyldioxolane (CD) than hitherto described (Vickroy et al. 1984 a). Taking the maximum number of binding sites, B _max, of [³H]QNB as 100%, B _max of [³H]CD amounts to 83% in the rat heart instead of the reported 17%, 33% in the cerebral cortex instead of 6%, 20% in hippocampus and 55% in pons/medulla. In the salivary glands specific binding was negligible. The affinities of a number of muscarinic agonists and antagonists to [³H]CD and [³H]QNB binding sites in different tissues of the rat are compared. Apparent affinities of agonists are much higher in the [³H]CD system, affinities of antagonists are slightly higher in the [³H]QNB system. In both assay systems receptors of heart and pons/ medulla membranes seem to have similar drug specificity. They differ somewhat from those in the cortex. Receptors in the salivary glands, however, seem to be completely different from those in the other three tissues. In the heart [³H]CD binding can be abolished almost completely by GppNHp. In the cortex about half of the [³H]CD binding is susceptible to GppNHp. The reduction of binding in the cortex is due to a change in B _max and not in the dissociation constant K _D. Competition of unlabelled pirenzepine with [³H]CD: In heart and pons/medulla only low affinity sites for pirenzepine (M₂-receptors) are labelled by [³H]CD. In regions rich in M₁ receptors like hippocampus (80% M₁ receptors) or cortex (65–70% M₁ receptors) the proportion of M₁ receptors labelled by [³H]CD is smaller than expected considering the concentration of M₁ receptors present in these tissues. Thus [³H]CD, under the conditions described in this paper, seems to label preferentially but not exclusively M₂ receptors in their agonist high affinity form. Send offprint requests to A. Closse at the above address     

Interaction of antihistaminics with muscarinic receptor (III)

The muscarinic antagonist 1-[benzilic 4,4′-³H]quinuclidinyl benzilate ([³H] QNB) bound to a single class of muscarinic receptors with high affinity in rabbit ileal membranes. The K _D and B _max values for [³H]QNB calculated from analysis of saturation isotherms were 52.5 pM and 154 fmol/mg, respectively. Chlorpheniramine (CHP), histamine H₁ blocker, increased K _D value for [³H]QNB without affecting the binding site concentrations and Hill coefficient. The K _i value of CHP for inhibition of [³H]QNB binding in ileal membranes was 1.44μM and the pseudo-Hill coefficient for CHP was close to unit. In the functional assay carbachol, muscarinic agonist, increased the contractile force of ileum with ED₅₀ value of 0.11μM. CHP caused the rightward shift of the dose-response curve to carbachol. The pA₂ value of CHP determined from Schild analysis of carbachol-induced contraction was 5.77 and the slope was unity indicating competitive antagonism with carbachol. The dissociation constant (K _i ) of CHP obtained in competitive experiments with [³H]QNB was similar to the K _A value (1.69μM) of CHP as inhibitor of carbachol-induced contraction in rabbit ileum. This result suggests that the binding of H₁ blocker, CHP, vs [³H]QNB to muscarinic receptors in ileal membranes represents an interaction with a receptor of physiological relevance.