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。     

可变误差多面体法用于多种维生素的同时测定

本文基于对多元校正分析模型的简要讨论,探索了应用可变误差多面体法同时测定维生索B₁,B₂,B₆和烟酰胺的可行性。其结果准确度和精密度均较满意。维生素B₁,B₂,B₆及烟酰胺的回收率分别是99.8±0.9%(CV),100.1±0.8%(CV),100.2±2.1%,100.1±0.7%(CV)。结果表明,通过公式K_S=A_SC_S^T(C_SC_S^T)^-1计算校正系数矩阵K_S,并结合可变误差多面体法这一直接求解方法,能有效地提高分析结果的准确度,克服组分间的交互作用及病态,是多元校正分析的较佳策略之一。     

Activation of recombinant h 5-HT1B and h 5-HT1D receptors stably expressed in C6 glioma cells produces increases in Ca2+-dependent K+ current

The putative coupling between stably expressed recombinant h 5-HT_1B or h 5-HT_1D receptors and K⁺ channels which regulate excitability was investigated in C6 glioma cells. Outward K⁺ currents (I _K) were examined in non-transfected C6 glioma cells and in cells expressing cloned h 5-HT_1B or h 5-HT_1D receptors using the patch-clamp technique in the whole-cell configuration. I _K was elicited by a depolarizing step from a holding potential of –60 mV. In C6 glioma cells expressing either recombinant h 5-HT_1B or h 5-HT_1D receptors, sumatriptan similarly increased I _K in a concentration-dependent manner (maximum increase 19.4±7.2%, n=8, P<0.05 and 25.1±3.9%, n=6, P<0.001, respectively) with EC₅₀ values (geometric mean with 95% confidence intervals in parentheses) of 56.3 nM (7.9–140 nM) and 68.7 nM (16–120 nM), respectively. Sumatriptan failed to elicit increases in I _K in non-transfected cells, confirming a specific involvement of the respective membrane h 5-HT_1B and h 5-HT_1D receptors in transfected C6 cells. In the presence of the mixed 5-HT_1B/D receptor antagonist GR 127935 (0.1 μM), sumatriptan (1 μM) failed to significantly increase I _K in C6 cells expressing h 5-HT_1B receptors (–7.5±3.5%, P=NS, n=6), although a higher concentration of GR 127935 (1 μM) was required to significantly inhibit sumatriptan-evoked increases in I _K in C6 cells expressing h 5-HT_1D receptors (–1.8±3.5%, P=NS, n=6), confirming that sumatriptan-evoked responses were indeed mediated by h 5-HT_1B and h 5-HT_1D receptors, respectively. In C6 cells expressing either cloned h 5-HT_1B or h 5-HT_1D receptors, sumatriptan-induced increases in I _K were prevented by the calcium chelator EGTA (5 mM) when included in the patch pipette (maximum increase 0.57±0.6%, n=3, P=NS and –2.8±1.6%, n=5, P=NS, respectively). In C6 cells expressing cloned h 5-HT_1B receptors, sumatriptan (1 μM) similarly failed to significantly increase I _K in the presence of dibutyryl cAMP (10 μM) or when a nominally Ca²⁺-free medium was included in the patch pipette (–19.4±5.1%, n=5 and –5.2±4.3%, n=5, respectively, P=NS in each case). In addition, the Ca²⁺-dependent K⁺ channel blockers iberiotoxin (0.1 μM) and tetraethylammonium (TEA, 1 mM) abolished sumatriptan-induced increases in I _K (–0.5±1.0%, n=4 and –3.9±3.1%, n=4, respectively, P=NS in each case) in C6 cells expressing h 5-HT_1B receptors, confirming the involvement of Ca²⁺-dependent K⁺ channels. In C6 cells expressing cloned h 5-HT_1B receptors, sumatriptan (1 μM) similarly failed to significantly increase I _k after 30-min incubation with thapsigargin (1 μM) or when heparin (2 mg/ml) was included in the patch pipette (1.1±0.4%, n=5 and 1.2±2.4%, n=5, respectively, P=NS). In conclusion, evidence is provided that both recombinant h 5-HT_1B and h 5-HT_1D receptors stably transfected in C6 glioma cells are positively coupled to Ca²⁺-dependent K⁺ channels, and the outward hyperpolarizing current mediated by these channels is dependent upon IP₃ receptor-mediated intracellular Ca²⁺ release. Received: 15 April 1998 / Accepted: 9 September 1998     

1-(2,6-二甲氧基)-2-(3,4-二甲基苯乙氨基)丙烷盐酸盐(DDPH)拮抗α1肾上腺素受体的特性

目的　研究DDPH对α₁-肾上腺素受体(α₁-AR)及其亚型的拮抗作用。方法　放射配体结合实验和离体血管收缩功能实验。结果　DDPH对¹²⁵I-BE2254与大鼠脑皮质和脾脏α₁-AR结合呈竞争性拮抗作用。pK_I值在两者间无显著性差别, Hill系数均接近于1.0。在分别稳定表达α_1A,α_1B或α_1D-AR的克隆HEK293细胞中,其拮抗的pK_I值α_1A和α_1D比α_1B-AR高约2倍,Hill系数均接近于1.0。并拮抗去甲肾上腺素(NE)介导大鼠主动脉,肾动脉和脾脏收缩的pA₂值,在三者间无显著差别,斜率接近1.0。结论　DDPH对α₁-AR有竞争性拮抗作用,但其作用对α₁-AR亚型无选择性。     

安定在家兔体内的肠胃循环药代动力学分析

6只家兔iv安定5 mg·kg^-1后,浓度—时间数据呈现双峰形。本文提出肠胃循环动力学模型,用于分析实测数据,得到了一般动力学参数:T_1/2(α)=0.21±=0.15 h,T_1/2(β)=2.2+0.6 h,K_e=1.5±0.6 h^-1,K₁₂=2.0±1.0 h^-1,K₂₁=1.0±0.4 h^-1,V₁=3.1±1.6 L·kg^-1,AUC=1.7±0.5μg·h^-1·ml^-1。此外,还求得有关肠胃循环的参数,即:重吸收滞后时间T′=0.25±0.24h,重吸收速率常数K_A=3.5±1.4 h^-1,重吸收率R_A=24±7%。     

四氢原小檗碱同类物对α1肾上腺素受体的拮抗作用

用放射配体结合实验与离体血管收缩功能实验相结合的方法,研究了4种四氢原小檗碱(tetrahydroproberberine,THPB)同类物对α₁-肾上腺素受体(α₁-AR)的作用。结果显示左旋四氢巴马汀(l-THP),左旋千金藤立陡碱(l-SPD),四氢原小檗碱-18(THPB-18)和四氢小檗碱(THB)¹²⁵IBE2254¹²⁵I-2-β(4-hydroxyphenyl)-ethyaminomethyl-tetralone,¹²⁵IBE)与大鼠脑皮质α₁-AR的结合呈竞争性拮抗作用,pK_I值分别为5.54±0.36,5.56±0.47,5.75±0.56和6.01±0.60,Hill系数接近于1.0。并拮抗苯肾上腺素(phenylephrine,PE)介导大鼠主动脉的收缩,pA₂值分别为5.48±0.58,5.66±0.54,5.64±0.34和5.45±0.76,斜率与1.0无显著性差别。结果提示,4种四氢原小檗碱同类物对α₁-AR均有非亚型选择性拮抗效应,且亲和性相同。     

粉防己碱,小檗胺等四氢异喹啉类生物碱对大鼠脑内M-胆碱受体的作用

用放射受体结合方法,研究了36种四氢异喹啉类生物碱及其半合成衍生物对大鼠脑内M-胆碱受体的结合特性。实验发现,粉防己碱对M-胆碱受体的亲和力最高,其K_i值为7.3×10^-8mol/L。小檗胺、四氢黄连碱和半合成原小檗碱衍生物B₁亦具有较高亲和力,K_i值分别为1.9×10^-7,6.8×10^-7和8.1×10^-7mol/L。四氢小檗碱半合成原小檗碱衍生物B₂,B₃,B₄,B₅,B₆和半合成小檗胺衍生物E₁及半合成蝙蝠葛碱衍生物D₁对M-胆碱受体的亲和力为中等强度,K_i值在1～2×10^-6mol/L之间。实验结果提示,某些四氢异喹啉类生物碱的药理作用可能与M-胆碱受体有关。     

四氢异喹啉类生物碱对大鼠脑内α肾上腺素受体的作用

应用放射受体结合法研究了近30种四氢异喹啉类(TIQs)生物碱对大鼠脑内α肾上腺素受体的作用。其中l-CBN,l-THC和l-STP对α₁受体亲和力最高,K_i值为～2.0×10^-7mol/L。其次是DHS,XLP和l-DCT,K_i值分别为4.7×10^-7,6.5×10^-7和7.6×10^-7mol/L。DHS对α₂受体亲和力最高(K_i=1.25×10^-6mol/L),l-REM次之。对α受体亚型亲和力选择比K_i(alpha-2)/Ki(alpha-1)最高的是l-STP(357) 和XLP(154),它们对α₂受体几无亲和力(K_i>10^-4mnl/L)。提示l-STP和XLP对α₁受体有较高的选择性。l-SPD和l-THP对α₁和α₂受体亲和力相近,均为中等强度。THJ,DRC及l-TTD等6种TIQs对α₁和α₂受体均无亲和力(K_i>10^-4mnl/L)。     

7-甲氧基-4′-羟基-3′-二乙胺甲基异黄酮(MHDF)对大鼠血流动力学和主动脉的作用

本实验观察了MHDF对整体大鼠血流动力学和离体大鼠胸主动脉的作用。结果表明iv MHDF(3～12.8 mg/kg)能降低大鼠左心室±dp/dt_max,V_max,V_pm和LVSP,延长T-dp/dt_max,减慢心率。MHDF还能舒张大鼠胸主动脉,ED₅₀为6.5×10^-6mol/L;非竞争拮抗NA和CaCl₂致主脉收缩,pD₂′为3.11±0.21和3.73±0.07;抑制高K⁺致主动脉收缩,IC₅₀为1.76×10^-5mol/L。提示MHDF对血管的作用与α受体阻断剂不同,而可能与钙拮抗有关。     

卡尔曼滤波分光光度法测定复方维生素B片四组分含量

复方维生素B片中的主要成分维生素B₁,B₂,B₆及烟酰胺各具有一定的不稳定性,且配方比例差较大,因而给多组分同时测定带来一定的困难。本文研究了应用卡尔曼滤波分光光度法同时测定复方维生素B片中的主要成分的可行性,并在处理由于分子间的相互作用引起的吸收度偏离Beer-Lambert定律的现象作了探索,得到较满意的结果。维生素B₁,B₂,B₆及烟酰胺的回收率分别是:100.2±0.90%(CV),100.3±1.7%(CV),101.1±3.3%(CV),99.90±0.65%(CV)。较已有的报道结果均好,表明方法可行。     

碘化二甲基木防己碱的药物—受体相互作用研究

碘化二甲基木防己碱(Dimethyltrilobine iodide简称DTI),具有降压作用,抗心律失常作用和骨骼肌松弛作用。初步证明,DTI属非去极化型肌松剂,拮抗性质推测为竞争性拮抗。究竟是否确属竞争性拮抗,强度如何,尚待研究。为此,我们用蟾蜍腹直肌标本作了进一步研究,并与箭毒作了比较。     

Antagonism of the trypanocide,berenil, and the nematocide,morantel, by tubocurarine

Summary The tubocurarine antagonism, (pA₂ values) of acetylcholine, suxamethonium, morantel and berenil were calculated as 6.5, 6.3, 6.4 and 6.2 respectively with the toad rectus abdominis muscle. It is suggested that tubocurarine competitively inhibits the morantel and berenil induced contracture of the toad rectus abdominis muscle.     

Fourth international symposium on quantitative mass spectrometry in life sciences

The compound 4(5)-[2-(4-azido-2-nitroanilino)ethyl]imidazole (AAH), a photoaffinity label analog of histamine, has been observed previously to produce a photolysis-dependent, specific and irreversible antagonism of histamine-induced contractile responses of the guinea-pig vas deferens. The pharmacological effects of AAH in isolated guinea-pig aorta (GPA) and dog trachealis (DT) were evaluated in the present study. Photolysis of 3 × 10^?5 M AAH for 5 min with visible light in organ baths containing the GPA resulted, after washout of AAH, in a nonequilibrium competitive antagonism of contractile responses to histamine; responses to norepinephrine and KCl were not affected. In contrast, photolyzed AAH (3 × 10^?5 M, one or two photoalysis treatments; 10^?4 M AAH, two treatments) did not antagonize contractile responses of the DT to histamine, nor were responses to acetylcholine or KCl affected. To test the possibility that AAH lacks affinity for histamine receptors in the DT, molar K_B values were obtained for nonphotolyzed AAH used as a conventional equilibrium competitive antagonist; K_B values for GPA and DT were not different (?log K_B were 5.48±0.18 and 5.04±0.23 for GPA and DT, respectively; P>0.05). pA₂ values for diphenhydramine, an H₁-histamine receptor antagonist, indicated a moderately greater potency in GPA compared to DT (pA₂ were 7.58±0.10 and 6.89±0.13 for GPA and DT, respectively; P<0.05); however the diphenhydramine: AAH potency ratio did not explain the resistance of the DT to photolyzed AAH. Cimetidine (10^?6?10^?4 M), an H₂-receptor antagonist, potentiated contractile responses of the GPA to histamine but was without an effect in the DT. No evidence for relaxation by histamine in preparations with induced tone was obtained; the resistance of the DT to photolyzed AAH therefore does not involve an inhibition of H₂-inhibitory receptors. Photolyzed AAH thus demonstrates tissue-selective antihistamine activity, perhaps as a result of differences in the characteristics of H₁-receptors in the GPA and DT.     

M3-Type Muscarinic Receptors Predominantly Mediate Neurogenic Quick Contraction of Bovine Ciliary Muscle

• 1.The present experiments were designed to investigate which subtypes of muscarinic receptors are involved in the neurogenic quick contraction of bovine ciliary muscle in connection to quick eye focal accommodation.
• 2.Transmural electrical stimulation (TES) produced a transient contraction, which was abolished in the presence of 3×10⁻⁷ M tetrodotoxin and 10⁻⁶ M atropine, but greatly augmented by 3×10⁻⁷ M physostigmine.
• 3.The exogenously applied acetylcholine (ACh: 10⁻⁹ to 3×10⁻⁶ M) produced a concentration-dependent contraction, which was competitively antagonized by 10⁻⁶ M atropine and augmented by 3×10⁻⁷ M physostigmine, but unaffected by 3×10⁻⁷ M tetrodotoxin.
• 4.The magnitude and time to peak of the maximal contraction produced by TES were significantly greater (1267.5±86.0 mg, P<0.005) and shorter (9.0±0.2 sec, P<0.005) than corresponding values (97.0±9.9 mg and 20.3±2.1 sec, respectively) of the phasic contraction caused by exogenously applied 10⁻⁵ M ACh, at which concentration the agonist caused the maximal contraction. The velocity (140.6±7.8 mg/sec) of the transient contraction caused by TES was approximately 28-fold greater than that of the phasic contraction caused by ACh (5.1±0.9 mg/sec).
• 5.The contractions produced by TES were greatly attenuated by 4-diphenylacetoxy-N-methylpiperidine (4-DAMP) as an M₃ antagonist and slightly by pirenzepine as an M₁ antagonist (20.2±7.9% inhibition at the highest concentration), but not by methoctramine (MET) as an M₂ antagonist. The IC₅₀ value (−log M) for 4-DAMP was determined to be 7.17±0.14.
• 6.Scatchard plot analysis of [³H]-quinuclidinylbenzilate (QNB) binding revealed that the binding sites constituted a single population with a K_d of 31.2±0.8 pM and a B_max of 895.5±93.2 fmol/mg protein. The activity in inhibiting [³H]-QNB binding was most potent with 4-DAMP (-log K_i=7.98±0.02), but less potent with pirenzepine (−log K_i=6.43±0.04) and MET (−log K_i=7.32±0.16). 4-DAMP was approximately 35- and 5-fold more potent than pirenzepine and MET in terms of −log K_i values, respectively, suggesting the predominant localization of M₃ receptor subtypes in the bovine ciliary muscle membrane.
• 7.These results suggest that TES produces a neurogenic quick contraction of the bovine ciliary muscle, which would be mediated mainly by ACh released from the intramural nerve terminals and subsequent excitation of M₃ receptor subtypes localized on the ciliary muscle cells, and that neurogenic quick contraction of the ciliary muscle is possibly involved in part in eye focal accommodation.

     

A comparative study of the effects induced by MCN-A-343 and acetylcholine on the isolated toad rectus abdominis.

McN-A-343 (McN), a non nicotinic ganglionic stimulant, induced slow contractile responses of the toad rectus abdominis. A relaxation was also observed when large doses were added in the presence of a contraction caused by acetylcholine (Ach). The relaxation induced by McN could not be overcome by increasing Ach concentration. Bell-shaped log dose-response curves were obtained for McN. d-Tubocurarine caused an unusual change on these curves, suggesting an indirect action of the agonist. This possibility was corroborated by the fact that hemicholinium, procaine, and cold storage of the muscle caused a marked decrease of the organ sensitivity to McN but not to ACh.     

氯化筒箭毒碱抑制蟾蜍椎旁神经节胆碱能传递的时反应量－效关系

应用离体蟾蜍椎旁神经节（ＰＶＧ）细胞内记录技术，测得灌流氯化筒箭毒碱（ｄ－ＴＣ）１５ｍｉｎ抑制ＰＶＧ快突触反应（ｆ－ＥＰＳＰ）的ＥＣＳ５０±Ｌ９５（斜率）为２１±８（３．１９８）μｍｏｌ·Ｌ－１，并发现作用的潜伏期和时程呈浓度依赖性关系（Ｐ＜０．０５）．其双曲线型四参数模型的拟合方程分别为：＝１８９．００５４／（ｌｎ＋Ｏ．２９７２）１．９５３７－２．４８０６和＝１５２．８３８８／（６．３９４５－ｌｎ）２．２１９９－０．３２０９。结果表明ｄ－ＴＣ对离体蟾蜍ＰＶＧ胆碱能传递的抑制作用存在双曲线型时反应量─效关系。     

Kinetic studies on dose-triphosphoinositide responses and dose-contraction responses in rabbit iris.

Correlative studies on the dose-triphosphoinositide (TPI) breakdown and -phosphatidic acid (PA) labeling and dose-tension relationship to acetycholine (ACh). carbachol and acetyl-β-methacholine were investigated in the rabbit iris smooth muscle. Concentrations of these agonists between 1 × 10^?8 M and 1 × 10^?4 M caused a concentration-dependent TPI breakdown and PA labeling, and concentrations between 1 × 10^?7 M and 1 × 10^?3 M caused a concentration-dependent contraction of the iris. The ed₅₀ values for the various cholinergic muscarinic agonists were determined from the dose-response curves. Good correlation between the ed₅₀ values determined by the biochemical and pharmacological methods, being of the order of 1–8 × 10^?6 M. were observed. The apparent dissociation constants of ACh and atropine were estimated by measuring the effects of these agents on TPI metabolism and iris muscle contraction. Atropine, at concentrations between 1 × 10^?10 M and 1 × 10^?9 M, produced a parallel shift to the right of the ACh dose-response curve in both the biochemical and pharmacological methods. Both biochemical and pharmacological responses were inhibited in a competitive manner by atropine. For the TPI, PA and contraction responses, the K_B values (determined with ACh as agonist) were 1.7 × 10^?10, 2 × 10^?9 M and 1.14 × 10^?10 M, respectively, and the corresponding pA₂ values were 10.30, 10.35, and 9.90 respectively. It was concluded that the findings of similar K_B values for the atropine-muscarinic receptor, along with similar pA₂ values from the Schild plots, with the TPI, PA and contraction responses in the rabbit iris could suggest a close relationship between the biochemical and pharmacological responses.     

COMPARATIVE STUDY ON α1-ADRENOCEPTOR ANTAGONIST BINDING IN HUMAN PROSTATE AND AORTA

1. Specific binding of [³H]-prazosin in prostatic and aortic membranes of humans was saturable and of high affinity (prostate: apparent dissociation constant, K_d= 0.35 ± 0.03 nmol/L; aorta: K_d= 0.26 ± 0.03 nmol/L). The density of [³H]-prazosin binding sites (B_max) for prostate and aorta was 546 ± 31 and 61.6 ± 1.6 fmol/mg protein, respectively. 2. Prazosin, YM617, naftopidil and urapidil competed with [³H]-prazosin for the binding sites in a dose-dependent manner in the prostate and aorta of humans. The binding affinities of these antagonists in both tissues were compared, based on the inhibition constant, K_i. Both prazosin and urapidil showed similar affinity to [³H]-prazosin binding sites in human tissue, whereas YM617 and naftopidil showed approximately a 12 and two times higher affinity, respectively, to α₁-adrenoceptor sites of prostate than aorta. 3. The chloroethylclonidine treatment reduced partially the B_max values for specific [³H]-prazosin binding in the prostate and aorta of humans with little effect on the K_d values. 4. These data suggest that YM617 is a relatively selective antagonist of human prostatic α₁-adrenoceptors.     

Interaction of Local Anaesthetics with Histamine H1 Receptors in Guinea-pig Ileum

The interaction of amine local anaesthetics and related compounds with histamine H₁ receptors was investigated in guinea-pig ileal longitudinal muscle. Quinacrine, chloroquine, tetracaine and procaine inhibited [³H]mepyramine binding to solubilized membrane from ileal muscle with pK_i values of 5.27 ± 0.11, 5.66 ± 0.01, 4.28 ± 0.08 and 3.97 ± 0.11, respectively. The pK_B values obtained from the initial parallel shift of the dose-response curves for histamine in the presence of these drugs were 549 ± 0.11, 6.14 ± 0.09, 4.86 ± 0.06 and 4.58 ± 0·06, respectively, in reasonable agreement with the pK_i values. The combined dose-ratio test with both local anaesthetics and antagonist (mepyramine) present showed that tetracaine and procaine were competitive and chloroquine was partially competitive, but that quinacrine was not competitive at histamine H₁ receptors. These local anaesthetics inhibited histamine-induced desensitization in guinea-pig ileum. Receptor occupancy (%) by agonist decreased from 95.2 (without inhibitor) to 73.9, 42.8, 35.9 and 33.9 in the presence of quinacrine, chloroquine, tetracaine or procaine, respectively, under the conditions where each inhibitor drug induced half maximum inhibition of desensitization. The results suggested that most of these local anaesthetics interacted competitively at histamine H₁ receptors and inhibited desensitization through their antagonizing actions, whereas quinacrine interacted allo-sterically and inhibited desensitization through a separate action.     

Stereoselective binding of niguldipine enantiomers to α1A-adrenoceptors labeled with [3H]5-methyl-urapidil

[³H]5-Methyl-urapidil, a potent antihypertensive derivative of urapidil, binds to α_1A-adrenoceptors in rat brain cortex membranes with a dissociation constant (K_D) of 0.89 nM and a B_max of 116 fmol/mg protein. The ligand does not bind to purified liver cell membranes (α_1B-adrenoceptors. [³H]5-Methyl-urapidil also labels 5-HT_1A receptors in brain membranes (K_D: 0.84 nM and B_max: 235 fmol/mg protein). (±)-Niguldipine, a novel 1,4-dihydropyridine with Ca²⁺-antagonistic as well as α_1A-adrenoceptor blocking properties, is a competitive inhibitor of [³H]5-methyl-urapidil binding to α_1A-adrenoceptors. In contrast to those for prazosin, the K_i values for niguldipine were highly dependent on the membrane protein concentration, indicating partitioning of niguldipine into hydrophobic compartments unavailable for α-adrenoceptor interaction. The extrapolated, ‘true’ K_i values were as follows: (±)-niguldipine: 0.298 nM, (−)-niguldipine: 3.12 nM, (+)-niguldipine: 0.145 nM.