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1.
[^3H]Cortisol及[^3H]Dexamethasone在大鼠肝细胞膜上的特异结合位点 总被引:1,自引:0,他引:1
本研究应用[3H]cortisol和[3H]dexamethasone(DEX)两种配基,观察到大鼠肝细胞膜上存在一类糖皮质激素(GC)特异结合位点。这些位点与GC的结合具有饱和性、高亲和力及低容量。其平衡解离常数(Kd)分别为12.84±6.58nmol/L和40.27±23.44nmol/L;最大结合容量(Bmax)分别为2.57±1.84pmol/mg蛋白质与0.64±0.18pmol/mg蛋白质(cortisol,n=4;DEX,n=3;±SE)。动力学实验数据所得的Kd值与Scatchard分析所得的Kd值结果基本一致。[3H]cortisol和[3H]DEX饱和结合实验数据用Scatchard作图分析,均显示为直线。Hill系数则分别为0.9880和0.9990.竞争抑制实验结果表明,cortisol对[3H]cortisol的结合位点有高度特异性竞争,比其它几种类固醇(强的松、黄体酮、RU486、DEX)的竞争力至少强40倍以上.用放射自显影技术进行研究,也提供了[3H]cortisol特异结合银粒位于大鼠肝细胞膜上的依据。 相似文献
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L. A. Nesterova B. N. Manukhin 《Biochemistry (Moscow) Supplemental Series A: Membrane and Cell Biology》2007,1(4):318-324
The binding of nonselective α1- and β-adrenoreceptor antagonists [3H]prazosin and [3H]dihydroalprenolol ([3H]DHA) to rat cerebral cortex synaptosomal membranes has been studied. It is found that ligand-receptor interactions of α1-adrenoreceptors fit into a single receptor pool model, which assumes the binding of two ligand molecules to one receptor molecule. The parameters of [3H]prazosin binding to α1-adrenoreceptors are as follows: K d = 2.58 ± 0.20 nM; B m = 2.95 ± 1.12 fmol/mg protein; Hill coefficient, n = 2. For β-adrenoreceptors, ligand-receptor interactions fit into a model assuming the presence of two receptor pools in the same effector system and binding of two ligand molecules to one receptor molecule. The corresponding parameters of the [3H]DHA binding to β-adrenoreceptors are as follows: K d1 = 0.74 ± 0.09 nM; K d2 = 7.63 ± 0.70 nM; B m1 = 25 ± 2 fmol/mg, B m2 = 48 ± 2 fmol/mg, n 1 = 2; n 2 = 2. We suggest that in rat cerebral cortex membranes α-and β-adrenoreceptors exist as dimers. 相似文献
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答:这里的[H]并不是指氢离子(H+),而是指辅酶中的氢(还原性氢),[H]的产生与利用即NADH、FADH。和NADPH的产生与利用。光合作用与呼吸作用中产生和利用的[H]不同,光合作用产生及利用NADPH.而呼吸作用产生与利用的[H]主要是NADH与FADH2。 相似文献
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E. Sylvester Vizi George T. Somogyi Laszlo G. Harsing Jr. Ildiko Zimanyi 《Neurochemical research》1986,11(1):71-84
Mouse isolated vas deferens preincubated with [3-H]noradrenaline was superfused and the effect of 1-adrenoceptor agonists was studied on the release of total radioactivity ([3H]noradrenaline +3H-metabolites) and [3H]noradrenaline. Reverse phase high pressure liquid chromatography (HPLC) combined with scintillation spectrometry was used to separate [3H]noradrenaline from its metabolites. Among the 1-adrenoceptor agonists (1-phenylephrine, ST-587(2-(2-chloro-5-trifluoromethyl phenylimino)-imidazole), (–)-amidephrine, methoxamine, cirazoline and l-noradrenaline) studied l-phenylephrine, ST-587 and l-noradrenaline were capable of releasing3H-noradrenaline. The effect of noradrenaline was stereospecific. As determined by HPLC combined with scintillation spectrometry the release of total radioactivity in response to l-noradrenaline is mainly due to [3H]noradrenaline. It is suggested that l-noradrenaline, l-phenylephrine, and ST-587 in addition to their direct effect on different receptors they also have indirect action through the release of noradrenaline which might be partly involved in the pharmacological responses. The mechanisms whereby l-noradrenaline and l-phenylephrine release noradrenaline would appear to involve a saturable Ca-independent and a cocaine and temperature sensitive process. On the basis of our findings among the 1-adrenoceptor agonist studied (–)-amidephrine, methoxamine and cirazoline is a better choice than l-phenylephrine or ST-587 for selective stimulation of postjunctional 1-adrenoceptor, they do not release noradrenaline. 相似文献
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光合作用与呼吸作用所涉及的[H]指NADH、FADH2与NADPH中的还原性氢。光合作用与呼吸作用中产生和利用的[H]不同,光合作用产生及利用NADPH,而呼吸作用产生与利用的[H]主要是NADH与FADH2。 相似文献
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《Journal of receptor and signal transduction research》2013,33(1-2):191-198
AbstractThe ionic and drug specificities of the [3H]nifedipine binding site in rabbit cardiac homogenates were investigated. Divalent cations inhibited specific [3H]nifedipine binding in the potency order: Ni+2 > Ca+2 ≥ Mg+2. Monovalent cations did not affect binding. The inorganic calcium entry blocker La+3 (IC50 = 1.1 mM) was the most potent cation in inhibiting radioligand binding. Calcium entry blocking drugs of different chemical classes inhibited [3H]-nifedipine binding, with a rank potency order of: nifedipine >> D600 = verapamil > tiapamil > cinnarizine = prenylamine. The same potency order was observed for these drugs in inducing negative inotropic activity of isolated, electrically stimulated rabbit papillary muscle. The stereoselectivity of verapamil and D600 ((?) >> (+) isomers) in depressing papillary muscle contractions was not seen in [3H]nifedipine competition experiments. This presents an obstacle to accepting the equivalence of the [3H]nifedipine binding site with the myocardial Ca+2 channel. It is, however, possible that the myocardial Ca+2 channel may be associated with multiple sites of action for calcium entry blockers. 相似文献
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Background
GABAA receptors (GABAAR) are composed of several subunits that determine sensitivity to drugs, synaptic localisation and function. Recent studies suggest that agonists targeting selective GABAAR subunits may have therapeutic value against the cognitive impairments observed in schizophrenia. In this study, we determined whether GABAAR binding deficits exist in the dorsolateral prefrontal cortex (DLPFC) of people with schizophrenia and tested if changes in GABAAR binding are related to the changes in subunit mRNAs. The GABA orthosteric and the benzodiazepine allosteric binding sites were assessed autoradiographically using [3H]Muscimol and [3H]Flumazenil, respectively, in a large cohort of individuals with schizophrenia (n = 37) and their matched controls (n = 37). We measured, using qPCR, mRNA of β (β1, β2, β3), γ (γ1, γ2, γ2S for short and γ2L for long isoform, γ3) and δ subunits and used our previous measurements of GABAAR α subunit mRNAs in order to relate mRNAs and binding through correlation and regression analysis.Results
Significant increases in both [3H]Muscimol (p = 0.016) and [3H]Flumazenil (p = 0.012) binding were found in the DLPFC of schizophrenia patients. Expression levels of mRNA subunits measured did not show any significant difference in schizophrenia compared to controls. Regression analysis revealed that in schizophrenia, the [3H]Muscimol binding variance was most related to α4 mRNA levels and the [3H]Flumazenil binding variance was most related to γ2S subunit mRNA levels. [3H]Muscimol and [3H]Flumazenil binding were not affected by the lifetime anti-psychotics dose (chlorpromazine equivalent).Conclusions
We report parallel increases in orthosteric and allosteric GABAAR binding sites in the DLPFC in schizophrenia that may be related to a “shift” in subunit composition towards α4 and γ2S respectively, which may compromise normal GABAergic modulation and function. Our results may have implications for the development of treatment strategies that target specific GABAAR receptor subunits. 相似文献10.
一、引言自从3',5'-环磷腺苷(cAMP)作为“第二信使”学说提出以来,通过实验不仅证实了它与某些病理过程密切相关,而且还注意到cAMP对肿瘤细胞也有某些影响。这是最近几年受人注意的一个研究课题。它不仅涉及到肿瘤发生,发展的理论,也涉及到肿瘤治疗的一个新的研究方向。近几年来,人们在研究cAMP作用的同时,又对另一种核酸——鸟苷3',5'-环磷酸(cGMP)的作用发生了浓厚兴趣。有人推测,生物的细胞调节作用是受cAMP和cGMP相互作用的影响。这一理论有可能用实验方法加以证实, 相似文献
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Dr. Richard Reynolds Christine Steffen Norbert Herschkowitz 《Neurochemical research》1987,12(10):885-890
Oligodendrocytes were isolated from mixed glial cultures of neonatal mouse forebrain and further grown in serum-free hormone supplemented culture medium. Cell populations were identified by indirect immunofluorescence using a range of specific antibodies, revealing a predominantly immature population of oligodendrocytes, the majority expressing the myelin glycolipids galactocerebroside and sulfatide on their plasma membrane. Astroglial contamination was found to be minimal. Simultaneous autoradiography and immunofluorescence demonstrated the presence of a transport system for the major inhibitory neurotransmitter GABA in the oligodendrocytes. The transport system was found to be energy, sodium and temperature dependent. Kinetic analysis revealed a high affinity system, with aK
m of 6.27 M and aV
max of 0.714 nmol/min/mg protein, which is comparable to that found previously for CNS neurons and astrocytes.Special Issue dedicated to Dr. E. M. Shooter and Dr. S. Varon. 相似文献
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Yoshihiko Shinohara Hiroshi Hasegawa Tomoyoshi Kaneko Yuka Tamura Takao Hashimoto Kimiyoshi Ichida 《Journal of chromatography. B, Analytical technologies in the biomedical and life sciences》2010,878(3-4):417-422
Homocysteine plays a key role in several pathophysiological conditions. To assess the methionine–homocysteine kinetics by stable isotope methodology, we developed a simultaneous quantification method of [2H7]methionine, [2H4]methionine, methionine, [2H4]homocysteine and homocysteine in rat plasma by gas chromatography–mass spectrometry (GC–MS). [13C]Methionine and [13C]homocysteine were used as analytical internal standards to account for losses associated with the extraction, derivatization and chromatography. For labeled and non-labeled homocysteine measurements, disulfide bonds between homocysteine and other thiols or proteins were reduced by dithiothreitol. The reduced homocysteine and methionine species were purified by cation-exchange chromatography and derivatized with isobutyl chlorocarbonate in water–ethanol–pyridine. Quantification was carried out by selected ion monitoring of the molecular-related ions of N(O,S)-isobutyloxycarbonyl ethyl ester derivatives on the chemical ionization mode. The intra- and inter-day precision of the assay was less than 6% for all labeled and non-labeled methionine and homocysteine species. The method is sensitive enough to determine pharmacokinetics of labeled methionine and homocysteine. 相似文献
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《Comparative biochemistry and physiology. C: Comparative pharmacology》1993,104(2):231-238
1. [3H]Batrachotoxinin A-20-α-benzoate ([3H]BTX-b) and [3H]saxitoxin ([3H]STX), radioligands that bind to distinct sites on the voltage-sensitive sodium channel, were bound specifically to saturable sites in rainbow trout (Oncorhynchus mykiss) brain synaptoneurosomes.2. Specific [3H]BTX-B binding was temperature dependent with highest levels of specific [3H]BTX-B binding observed at 7°C. Specific binding was inversely correlated with assay temperature at temperatures above 7°C.3. Saturating concentrations of scorpion (Leiurus quinquestriatus) venom (ScV) stimulated specific [3H]BTX-B binding at 27°C, but not at 7°C. The dihydropyrazole insecticide RH 3421 inhibited specific [3H]BTX-B binding at 7°C but had no effect on specific binding at 27°C. The sodium channel activators veratridine and aconitine and the local anesthetic dibucaine inhibited specific [3H]BTX-B binding at both 7°C and 27°C.4. Displacement experiments in the presence of ScV at 27°C gave an equilibrium dissociation constant (Kd) for [3H]BTX-B of 710 nM and a maximal binding capacity (Bmax) of 11.3 pmol/mg protein. Kinetic experiments established the rates of association (1.17 × 105min−1 nM−1) and dissociation (0.0514min−1) of the ligand-receptor complex.5. The binding of [3H]STX reached apparent saturation at 7.5 nM. Scatchard analysis of the saturation data indicated a Kd of 3.8nM and a Bmax of 1.9 pmol/mg protein.6. These studies provide evidence for high affinity, saturable binding sites for [3H]BTX-B and [3H]STX in trout brain preparations. Whereas certain neurotoxins modified the specific binding of [3H]BTX-B in trout brain synaptoneurosomes in a predictable fashion, other compounds known to affect specific [3H]BTX-B binding in mammalian brain preparations had no effect on specific [3H]BTX-B binding in the trout. 相似文献
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木质素作为木材的主要组成成分,通常是由3种单体聚合而成,在其生物合成过程中,共有10个酶家族参与负责将苯丙胺酸转化为单体木质素,其中C3H是在对-香豆酰辅酶A(p-coumaroyl CoA)到咖啡酰辅酶A(caffeoyl CoA)的羟基化过程和G/S单体形成中的关键控制酶类,探究PagC3H3基因表达模式,对于进一步了解该基因功能具有重要意义。该研究通过定量PCR对PagC3H3基因的组织特异性表达进行分析;克隆得到了长度为2 035 bp的PagC3H3的启动子序列,预测含有多个顺式作用元件;同时,将获得的PagC3H3的启动子序列构建植物表达载体pBI121-PagC3H3pro::GUS,进行拟南芥瞬时转化,结果显示PagC3H3基因在84K杨的根、中部茎节和基部茎节中的表达量较高;瞬时转化拟南芥,GUS染色表明:在下胚轴和根中GUS活性较强,由此推测PagC3H3基因在木质素合成过程中发挥作用。 相似文献
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本文报道用光镜放射自显影方法观察夜行壁虎网膜感光细胞外段的更新。当注射氚标记的亮氨酸后,这种由其祖先视锥演变来的夜行壁虎视杆(并且至今仍保留着一些形态学上属视锥的特征)其外段呈现的标记图型仍与一般的视杆细胞相似。 相似文献
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Opposing Roles of Dopamine D1 and D2 Receptors in Nigral γ-[3H]Aminobutyric Acid Release? 总被引:2,自引:0,他引:2
Michael Starr 《Journal of neurochemistry》1987,49(4):1042-1049
This study examined the effects of dopamine D1 and D2 receptor agonists and antagonists on the spontaneous and calcium-dependent, K+-induced release of gamma-[3H]aminobutyric acid [( 3H]GABA) accumulated by slices of rat substantia nigra. SKF 38393 (D1 agonist) and dopamine (dual D1/D2 agonist) were without effect on [3H]GABA efflux by themselves (1-40 microM), or in the presence of the phosphodiesterase inhibitor isobutylmethylxanthine (IBMX) (0.5 mM), but potentiated evoked release in the presence of forskolin (0.5 microM), an adenylate cyclase activator. These increases in release were prevented by the D1 antagonist SCH 23390 (0.5 microM), but not by the D2 antagonist metoclopramide (0.5 microM). Higher concentrations of forskolin (10-40 microM) augmented stimulus-evoked [3H]GABA release directly, whereas dibutyryl cyclic AMP (100-200 microM) depressed it. Apomorphine, noradrenaline, and 5-hydroxytryptamine (1-40 microM) had no effect. The D2 stimulants lisuride, RU 24213, LY 171555, and bromocriptine dose-dependently inhibited depolarisation-induced but not basal [3H]GABA outflow. These inhibitory responses were not modified by the additional presence of SKF 38393 (10 microM) or SCH 23390 (1 microM), or by injection of 6-hydroxydopamine into the medial forebrain bundle 42 days earlier, but were attenuated by metoclopramide (0.5 microM). Higher amounts (10 microM) of SCH 23390, metoclopramide, or other D2 antagonists (loxapine, haloperidol) reduced evoked GABA release by themselves, probably by nonspecific mechanisms. These results suggest D1 and D2 receptors may have opposing effects on nigral GABA output and could explain the variable effects of mixed D1/D2 dopaminomimetics in earlier release and electrophysiological experiments. 相似文献