Binding of beta-carboline-3-carboxylic acid ethyl ester to mouse brain benzodiazepine receptors in vivo

beta-Carboline-3-carboxylic acid ethyl ester (beta-CEE) displaced [3H] flunitrazepam from the mouse brain benzodiazepine receptor in vivo with an i.v. ED50 of 2.1 mg/kg, an i.p. ED50 and 53.4 mg/kg, and an i.g. ED50 of 450 mg/kg. At 2.1 mg/kg i.v. beta-CEE displaced 37 +/- 9% of the label from hippocampal membranes and 76 +/- 7% from cerebellar membranes. The results suggest that the in vitro binding specificity of beta-CEE is also expressed in vivo and that the drug may act in vivo by binding to the benzodiazepine receptor.     

Effects of beta-carboline-3-carboxylic acid ethyl ester on suppressed and non-suppressed responding in the rhesus monkey

The effects of the putative 'anxiogenic' compound beta-carboline-3-carboxylic acid ethyl ester (beta-CCE) were studied on non-suppressed and suppressed responding of rhesus monkeys. Responding was maintained under a fixed-interval 2 min schedule of food presentation (non-suppressed responding) and a fixed-interval 2 min schedule of food presentation where each 10th response produced a 3-5 mA footshock (suppressed responding). Rates of suppressed responding were 60-90% lower than those for non-suppressed responding. beta-CCE reliably decreased non-suppressed responding over the range of 0.03-0.3 mg/kg (ED50 approximately 0.04 mg/kg) while consistent decreases in suppressed responding were not obtained in all animals until doses of 1-3 mg/kg (ED50 approximately 2 mg/kg) were given. Doses of beta-CCE greater than 0.01 mg/kg slightly increased blood pressure, moderately increased heart rate and greatly increased plasma ACTH levels for both types of response. In contrast, diazepam increased suppressed responding without affecting non-suppressed responding at low doses (0.3-1 mg/kg), while higher doses were required to decrease suppressed responding. Diazepam had little effect on blood pressure or mean heart rate. Ro 15-1788 (1 mg/kg) blocked the rate-decreasing effects of beta-CCE on non-suppressed responding, suggesting the decrease is mediated via a benzodiazepine recognition site. These results show that under conditions where relatively low doses of diazepam have an anxiolytic effect (i.e. selectively increase rates of suppressed responding), relatively low doses of beta-CCE selectively decrease non-suppressed responding, questioning current notions of how to define an anxiogenic drug effect.     

Non-benzodiazepine, Ag-dependent, brain-specific binding sites for [3H]beta-carboline-3-carboxylic acid ethyl ester

Specific, saturable and reversible binding of [3H]beta + CCE ([3H]beta-carboline-3-carboxylic acid ethyl ester) in buffer containing 20 microM AgNO3 and 10 microM diazepam was detected in rat brain membranes. The binding of [3H]beta CCE to non-benzodiazepine binding sites is Ag+ (Cu)-dependent, stimulated by NaCl and ascorbic acid and inhibited by dithiothreitol. The concentration of non-benzodiazepine [3H]beta CCE binding sites (Bmax) determined in the brain membranes was 1180 +/- 320 pmol/g tissue, and Kd = 77 +/- 19 nM. [3H]beta CCE bound to benzodiazepine receptors in the same membranes with Bmax = 81 +/- 9 pmol/g tissue and Kd = 3.2 +/- 0.4 nM.     

Differential pharmacological effects of beta-carboline-3-carboxylic acid esters

The effects of the methyl, ethyl and propyl esters of beta-carboline-3-carboxylic acid have been studied in vitro and in vivo. All three esters were found to be potent inhibitors of 3H-flunitrazepam binding in the rat cerebellum and cerebral cortex in vitro. In vivo, the methyl and ethyl esters were potent proconvulsant agents, whereas the propyl ester was not. Furthermore, the methyl ester produced convulsions which were blocked by the ethyl and propyl esters as well as by diazepam. These in vivo differences may be due to the beta-carboline esters having different proportions of agonistic and antagonistic actions at their recognition sites.     

1-甲基-3-正丙基吡唑-5-甲酸乙酯的合成

目的：合成标题化合物,并进行工艺改进。方法：以2－戊酮和草酸二乙酯为原料,经三步反应合成了产物。结果：反应的总收率为47．7％。合成的各步产品经元素分析、红外和核磁共振光谱确证。结论：改进的工艺具有反应温度低、时间短等优点,并提高了收率。     

Biological properties of 2,4-dioxo-3H-quinoline-3-carboxylic acid and its ethyl ester

Translated from Khimiko-farmatsevticheskii Zhurnal, Vol. 26, No. 2, pp. 33–35, February, 1992.     

Behavioural and convulsant actions of two methyl esters of beta-carboline-3-carboxylic acid in photosensitive baboons and in DBA/2 mice

The proconvulsant and convulsant actions of methyl 6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM) and of methyl-beta-carboline-3-carboxylate (beta-CCM) have been evaluated in two animal models of reflex epilepsy, the photosensitive baboon, Papio papio, and the audiogenic seizure prone DBA/2 mouse. In baboons, myoclonic responses to photic stimulation are markedly enhanced 1 min after DMCM, 0.25 mg/kg i.v. In the absence of photic stimulation DMCM, 0.5 mg/kg i.v. induces a single brief tonic clonic seizure within 10-90 s. beta-CCM, 0.025-0.05 mg/kg i.v. similarly enhances myoclonic responses to photic stimulation. Generalised seizures occur without photic stimulation 0.5-3 min after beta-CCM, 0.1-0.2 mg/kg. Pretreatment with the excitatory amino acid antagonist, 2-amino-7-phosphonoheptanoic acid (2-APH), 110 mg/kg i.v., prevents the generalised seizures induced by DMCM, 0.5 mg/kg, but not those induced by beta-CCM, 0.1-0.2 mg/kg. In DBA/2 mice beta-CCM and DMCM are indistinguishable in potency as convulsants (ED50 values for clonic seizures: 4.4 and 4.6 mg/kg i.p. respectively) and as proconvulsants (ED50 values for facilitation of clonic seizure responses to an 83 dB sound stimulus: 0.25 and 0.23 mg/kg). Pretreatment with 2-APH gives equipotent protection against audiogenic seizures induced by beta-CCM, 1 mg/kg or DMCM, 1 mg/kg. The differences in relative potency of beta-CCM and DMCM in the two species are probably accountable for in terms of differing metabolism. A differential action of the two beta-carbolines on receptor subtypes, with enhancement of excitatory amino acid release playing a more important role in epileptogenesis after DMCM, is proposed.     

Ethanol effects in pigeons trained to discriminate MK-801, PCP or CGS-19755

The non-competitive N-methyl-D-aspartate (NMDA) antagonists MK-801, PCP and ketamine have recently been found to produce full drug-appropriate responding in pigeons trained to ethanol (1.5g/kg) in a two-key operant drug discrimination procedure. In the present study, ethanol (0.56-3.2g/kg i.g.) was administered to pigeons trained to discriminate MK-801 (0.18mg/kg, n = 5), PCP (1.0mg/kg, n = 4) or the competitive NMDA antagonist CGS-19755 (1.8mg/kg, n = 4) from vehicle. Up to doses that caused large reductions in response rates, ethanol produced only vehicle-appropriate responding in the pigeons trained to PCP and only low levels of drug-appropriate responding in pigeons trained to MK-801 and CGS-19755. The present results suggest there could be asymmetric generalization between the discriminative stimulus effects of i.g. ethanol and NMDA antagonists.     

Reinforcement schedule effects in rats trained to discriminate 3,4-methylenedioxymethamphetamine (MDMA) or cocaine

Rationale Relatively few studies have compared the discriminative stimulus effects of 3,4-methylenedioxymethamphetamine (MDMA) and cocaine, and findings from different laboratories are somewhat inconsistent. One possible reason for discrepant results may be the use of different reinforcement schedules during discrimination training.Objective The present study compared fixed ratio (FR) 20 and variable interval (VI) 15-s reinforcement schedules to determine their influence on discrimination acquisition, response rates, frequency of reinforcements, and stimulus generalization in rats trained to discriminate cocaine or MDMA.Materials and methods Thirty-two male Sprague–Dawley rats were trained to discriminate cocaine (10 mg/kg; n=16) or MDMA (1.5 mg/kg; n=16) from saline under either a FR 20 or a VI 15-s schedule of food reinforcement. Stimulus generalization tests were conducted with a range of doses of cocaine, MDMA, d-amphetamine, and lysergic acid diethylamide in all four training groups.Results The FR 20 schedule facilitated more rapid discrimination acquisition compared to the VI 15-s schedule and established differential response rates and frequency of reinforcement under drug and vehicle conditions. However, reinforcement schedule had little influence on stimulus generalization between MDMA and cocaine. Cocaine produced partial substitution for MDMA in both training groups (FR 20, 51%; VI 15-s, 58%). Likewise, MDMA produced only partial substitution for cocaine in both training groups (FR 20, 40%; VI 15-s, 72%).Conclusions The present findings suggest that the number of sessions required to establish discriminative stimulus control varies with different reinforcement schedules. Nevertheless, training schedules alone do not appear to have significant effects on stimulus generalization between MDMA and cocaine.     

Chronic administration of beta-carboline-3-carboxylic acid methylamide by continuous intraventricular infusion increases GABAergic function.

The repeated, intraperitoneal administration of the benzodiazepine receptor inverse agonist, FG 7142 (beta-carboline-3-carboxylic acid methylamide), leads to pharmacological kindling and an associated decrease in GABA-stimulated influx of 36Cl- into cortical membrane preparations. The chronic administration of benzodiazepine agonists results in the development of tolerance and also results in a decrease in GABA-stimulated uptake of 36Cl-. The present study was designed to evaluate further the paradoxical reports that both chronic treatment with benzodiazepine receptor agonists and inverse agonists results in a decreased ability of GABA to stimulate uptake of 36Cl- into cortical membrane preparations. The effects of continuous administration of FG 7142 on GABA-stimulated uptake of 36Cl-, the threshold for bicuculline-induced seizures and the proconvulsant actions of acute administration FG 7142 were evaluated. The continuous administration of FG 7142 resulted in an increased capacity of GABA to stimulate the uptake of 36Cl- into cortical membrane preparations and a significant increase in the seizure threshold for bicuculline following the acute administration of FG 7142. These data, therefore, indicate that changes in GABAergic function following chronic administration of GF 7142 are dependent on the regimen of administration of drug. The results also suggest that the GABA receptor homeostatically responds to continuous occupation by inverse agonists by an upregulation of its functional response to GABA.     

6-溴-5-羟基吲哚-3-羧酸酯类衍生物的合成及其抗病毒活性

目的 设计合成6-溴-5，羟基吲哚-3-羧酸酯类衍生物。并对其体外抗呼吸道病毒活性进行初步评价。方法 以对苯醌、3-乙氨基-2-丁烯酸乙酯为起始原料，经多步反应合成目标化合物；并在MDCK和HeLa细胞内。采用细胞病变法，检测目标化合物对甲3型流感病毒和呼吸道合胞病毒的抑制作用。结果 与结论共制得12个新化合物，经^1H—NMR、MS确证其结构。体外试验表明。目标化合物具有不同的抗病毒活性，其中Ⅵj的体外抗病毒作用与阳性对照药金刚烷胺相当。     

Gamma-hydroxybutyric acid in male and female cynomolgus monkeys trained to discriminate 1.0 or 2.0 g/kg ethanol

Gamma-hydroxybutyric acid has been proposed as a pharmacotherapy for alcoholism in part based on similar discriminative stimulus effects as ethanol. To date, drug discrimination studies with gamma-hydroxybutyric acid and ethanol have exclusively used rodents or pigeons as subjects. To evaluate possible differences between species, sex, and route of administration, this study investigated the substitution of gamma-hydroxybutyric acid (intragastrically or intramuscularly) for ethanol 30 or 60 min after administration in male (n=6) and female (n=7) cynomolgus monkeys trained to discriminate 1.0 and 2.0 g/kg ethanol. At least one dose of gamma-hydroxybutyric acid completely or partially substituted for ethanol in three of the 13 monkeys tested, with each case occurring in female monkeys. Ethanol-appropriate responding did not increase with gamma-hydroxybutyric acid dose. Monkeys were more sensitive to the response rate decreasing effects of gamma-hydroxybutyric acid administered intramuscularly compared with intragastrically. The lack of gamma-hydroxybutyric acid substitution for ethanol suggests that these drugs have different receptor bases for discrimination. Furthermore, the data do not strongly support shared discriminative stimulus effects as the rationale for gamma-hydroxybutyric acid pharmacotherapy for alcoholism.     

Beta-carbolines as benzodiazepine receptor ligands II: Synthesis and benzodiazepine receptor affinity of beta-carboline-3-carboxylic acid amides

Numerous beta-carboline-3-carboxamides were synthesized by amidation of beta-carboline-3-carboxylic acid, with various amino acids and amino acid esters serving as amine components, and tested in respect to their affinity for the benzodiazepine receptor in mouse brain membranes. The title compounds have affinities in the low micromolar range. The results are discussed with respect to their relevance for a possible beta-carboline structure containing the endogenous ligand of the benzodiazepine receptor.     

Competitive and noncompetitive NMDA antagonist effects in rats trained to discriminate lever-press counts

The glutamate activated N-methyl-D-aspartate (NMDA) receptor may play a role in short-term memory processing. Among the evidence for this is that NMDA antagonists can impair accuracy in fixed consecutive number (FCN) tasks. This study was designed to further characterize this effect by examining NMDA antagonists differing in their cellular mechanisms of action. Rats were trained to respond under an FCN operant schedule, which required eight presses on one lever (counting lever) before one press at an alternate lever (reinforcement lever) would produce food reinforcement. The effects of three noncompetitive [MK-801 (0.01-0.56 mg/kg); phencyclidine (0.3-3.0 mg/kg); memantine (1-10 mg/kg)] and two competitive [SDZ EAA 494 (0.3-3.0 mg/kg) and NPC 17742 (2.0-16 mg/kg)] NMDA antagonists were analyzed. MK-801 and phencyclidine decreased accuracy at doses not reducing response rates. Memantine, and both of the competitive antagonists, also reduced accuracy, but did so only at doses that markedly reduced response rates. These results suggest that both the affinity and the site bound on the NMDA glutamate receptor by antagonists can determine their effects on FCN performance. Subsequent studies investigated whether SCH 23390, a dopamine D1 receptor antagonist, and NMDA could modulate the effects by phencyclidine and SDZ EAA 494, respectively, on FCN performance.     

beta-Carbolines as benzodiazepine receptor ligands. 1. Synthesis and benzodiazepine receptor interaction of esters of beta-carboline-3-carboxylic acid

Several esters of beta-carboline-3-carboxylic acid were synthesized and tested in respect to their affinity for the benzodiazepine receptor in bovine cortex membranes. Out of these derivatives, the methyl, ethyl, and n-propyl ester were clearly the most potent, while the n-butyl, benzyl, and 3-pyridylmethyl ester were considerably less active. Moreover, several beta-carboline-3-carboxylates with ethanol derivatives as ester alcohol components were all less active than the ethyl or n-propyl ester themselves. It is concluded that the affinity of beta-carboline-3-carboxylates to the benzodiazepine receptor is profoundly dependent on molecular size, as well as hydrophobic and electronic parameters of the ester alcohol component.     

Interaction of avermectins with [3H]beta-carboline-3-carboxylate ethyl ester and [3H]diazepam binding sites in rat brain cortical membranes

The binding of [3H] beta-carboline-3-carboxylate ethyl ester ([3H] beta-CCE), a ligand for the benzodiazepine receptor in the mammalian CNS, to rat cortical membranes, is enhanced by avermectin B1a and its therapeutic formulation, Ivermectin. In contrast to the effects of the avermectins on [3H]diazepam binding, which involve changes in both receptor affinity and number, increases in beta-CCE binding, which are much less than those observed for the benzodiazepine ligand, involve only alterations in receptor number. This Bmax increase is bicuculline insensitive whereas Ivermectin effects on benzodiazepine binding are partially antagonized by GABA antagonist. The data suggest a differential interaction by the avermectins on benzodiazepine and beta-CCE binding sites in rat cortical membranes and indicate that these macrolide anthelmintics may be a useful tool for characterizing benzodiazepine/anxiolytic receptor subtypes.     

7-苯乙酰氨基-3-吡啶甲基头孢-4-羧酸对甲氧苄酯合成及反应动力学研究

以GCLE和吡啶为原料合成头孢他啶中间体7-苯乙酰氨基-3-吡啶甲基头孢-4-羧酸对甲氧苄酯,通过高效液相色谱仪对反应过程进行监测.研究了该反应的动力学,确定反应为SN2历程.通过测定25和35℃在二氯甲烷中反应的速率常数,求得表观活化能为76.882kJ/mol.     

HPLC法测定盐酸阿比朵尔中间体6-溴-5-羟基-1-甲基-2-苯硫甲基吲哚-3-羧酸乙酯

目的自主合成得到盐酸阿比朵尔中间体6-溴-5-羟基-1-甲基-2-苯硫甲基吲哚-3-羧酸乙酯(Sr-5),建立Sr-5的高效液相色谱测定方法,为工业生产的质量控制提供依据。方法采用Kromasil sum C18柱(150mm×4.6mm,5μm);流动相:0.2g·L-1磷酸二氢钾水溶液(pH 3.0)-乙腈(65∶35);检测波长:310nm;流速:1.0mL·min-1。结果 Sr-5中间体在1.6～8.0μg范围内线性关系良好,回收率为98.1%。结论 HPLC测定Sr-5中间体的方法准确,简单可行,重复性好,适用于Sr-5的质量控制。     

Differential generalization to pentobarbital in rats trained to discriminate lorazepam,chlordiazepoxide, diazepam,or triazolam

In drug discrimination studies benzodiazepinetrained animals have typically responded on the drug lever when tested with barbiturates. In a recent study, greater specificity appeared to be shown when lorazepam was used as a training drug. The generality and limits of this finding were explored in the present set of experiments. The asymmetrical cross-generalization found in lorazepam-and pentobarbital-trained baboons was replicated in rats and was shown not to be a function of either lorazepam (0.1., 0.32, or 1.0 mg/kg) or pentobarbital (10 or 25 mg/kg) training dose (i.e., pentobarbital-trained rats responded on the drug lever in tests with lorazepam, but lorazepam-trained rats did not show comparable pentobarbital generalization). In the next experiment, groups of rats were trained to discriminate chlordiazepoxide (10 mg/kg), triazolam (0.1 mg/kg), or diazepam (1.0 mg/kg). Generalization to both lorazepam and pentobarbital was shown by these rats. Finally after daily pentobarbital administration, lorazepam-trained rats made a sufficient number of responses after high pentobarbital doses to permit extension of the range of pentobarbital doses tested. Pentobarbital generalization increased, but still did not occur in all rats and was unreliable in successive tests in the same rats. These results suggest less homogeneity in the discriminative stimulus effects of depressant drugs than generally has been recognized.Portions of these data were presented at the meetings of the Federation of American Societies of Experimental Biology, Anaheim, 1985 (Ator and Griffiths 1985) and Behavioral Pharmacology Society, Rockville MD (1987)