High Affinity For Benzodiazepine Receptors Research Articles

Behavioral and subjective effects of DN-2327 (pazinaclone) and aiprazolam in normal volunteers

DN-2327 (pazinaclone) is a new non-benzodiazepine compound which has high affinity for benzodiazepine receptors. The acute behavioral effects and abuse liability of DN-2327 (2, 4 and 8mg) were compared to those of the benzodiazepine anxiolytic alprazolam (0.25, 0.5 and 1mg) in ten normal adult male volunteers using a double-blind, placebo-controlled, outpatient design. For both drugs, the neck effect occurred approximately 1.5h after drug administration. Both drugs also produced comparable dose-related effects on several measures related to sedation, as well as on subject- and observer-rated strength of drug effect. Both alprazolam and DN-2327 produced dose-related impairments on various performance measures; on some tasks, the impairment was greater for DN-2327. In contrast, there were no differences between DN-2327 and alprazolam on observer-rated measures. Although no measures of drug-taking were made in this study, to the extent that self-reported effects predict reinforcing effects, the data suggest little liability for abuse of the two compounds in this subject population. Ratings of 'drug liking' and 'willing to take the drug again' were not increased following alprazolam. Although DN-2327 did not increase ratings of 'willing to take the drug again', DN-2327 did produce small but significant increases on ratings of 'drug liking'. Overall, these results suggest that the non-benzodiazepine DN-2327 has a pharmacological profile that is similar to that of benzodiazepines.

Human studies on abecarnil a new beta‐carboline anxiolytic: safety, tolerability and preliminary pharmacological profile.

1. Abecarnil (isopropyl-6-benzyloxy-4-methoxymethyl-beta-carboline-3-carboxylate), a beta-carboline with high affinity for benzodiazepine receptors, was tested in healthy male subjects; single doses of abecarnil were given in five dosage levels (1 mg, 5 mg, 10 mg, 20 mg, 40 mg) and in a multiple dose study in four dosage levels (15 mg, 30 mg, 60 mg, 90 mg day-1) for 7 days. On two days following multiple dose treatment, placebo was given in single-blind conditions (follow-up). In each dosage level, in both studies drug was given to 10 subjects (7: verum, 3: placebo). 2. Safety and tolerability were evaluated by changes in vital signs, incidence and severity of adverse reactions and biochemical and haematological screening. Drug effects were estimated utilizing a bipolar visual analogue scale (poles: 'sleepy'-'alert') and a psychomotor task, the digit symbol substitution task. The pharmacokinetics of single and multiple doses were also determined in the multiple dose study. 3. Abecarnil was generally well tolerated. In the single dose study the most frequently reported side effects associated with abecarnil at high doses (20 and 40 mg) were dizziness, unsteady gait, and lack of concentration. A decrement in performance on the digit symbol substitution task was also observed in the two high dosage groups 20 mg and 40 mg. Evaluation of visual analogue scale ratings did not reveal a sedative effect even at higher doses. 4. In the multiple dose study the most frequently reported side effects during the treatment period were dizziness, unsteady gait, and lack of concentration.(ABSTRACT TRUNCATED AT 250 WORDS)

High affinity ligands for ‘diazepam-insensitive’ benzodiazepine receptors

Structurally diverse compounds have been shown to possess high affinities for benzodiazepine receptors in their ‘diazepam-sensitive’ (DS) conformations. In contrast, only the imidazobenzodiazepinone Ro 15-4513 has been shown to exhibit a high affinity for the ‘diazepam-insensitive’ (DI) conformation of benzodiazepine receptors. We examined a series of 1,4-diazepines containing one or more annelated ring systems for their affinities at DI and DS benzodiazepine receptors, several 1,4-diazepinone carboxylates including Ro 19-4603, Ro 16-6028 and Ro 15-3505 were found to possess high affinities (K i ∼ 2.6–20 nM) for DI. Nonetheless, among the ligands examined, Ro 15-4513 was the only substance with a DI/DS potency ratio ∼ I: other substances had ratios ranging from 13 to > 1000. Ligands with high to moderate affinities at Di were previously classified as partial agonists, antagonists, or partial inverse agonists at DS benzodiazepine receptors, but behaved as ‘GABA neutral’ (antagonist) substances at DI. The identification of several additional high affinity ligands at DI benzodiazepine receptors may be helpful in elucidating the pharmacological and physiological importance of these sites.

Thienylpyrazoloquinolines with high affinity to benzodiazepine receptors: continuous shift from inverse agonist to agonist properties depending on the size of the alkyl substituent

2-(5-Alkylthien-3-yl)-(1),2-(4-alkylthien-2-yl)-(2), and 2-(5-alkylthien-2-yl)-2,5-dihydro-3H-pyrazolo[4,3-c]quinolines (3) were prepared in four steps starting from ethyl 4-chloroquinoline-3-carboxylate (4) and hydrazinothiophene-carboxylates 5, 8, and 9. All the assayed compounds possessed high affinities for benzodiazepine receptors (Ki = 0.3-2.6 nM). The activities of agonists and inverse agonists were assessed on the basis of inhibition or facilitation of pentylenetetrazole-induced convulsions, respectively. Introduction of alkyl groups of different sizes into the unsubstituted inverse agonistic compounds results in a corresponding shift in the activity from an inverse agonist to an antagonist to an agonist. The susceptibility of such a shift increases in the order of 1 less than 2 less than 3. This tendency may be explained by slight differences in the geometry of the alkyl substituents among the three series.

Synthesis of novel 3-substituted .beta.-carbolines as benzodiazepine receptor ligands: probing the benzodiazepine receptor pharmacophore

The 3-substituted beta-carbolines 2-4 and 5-7 were prepared from 3-amino-beta-carboline (8) in one step via diazotization, followed by reaction with the appropriate nucleophile in order to determine their binding affinity for benzodiazepine receptors (BzR). All three of the 3-alkoxy-beta-carbolines 2 (IC50 = 124 nM), 3 (IC50 = 24 nM), and 4 (IC50 = 11 nM) have high affinities for BzR. The beta-carbolines substituted with electron-withdrawing groups including 5 (Cl; IC50 = 45 nM), 6 (NO2; IC50 = 125 nM), and 7 (N = C = S; IC50 = 8 nM) also had high affinities for BzR. The affinities of 5-8 clearly indicate that a carbonyl moiety at position 3 of a beta-carboline is not required for high-affinity binding to BzR. These findings have led to the development of a model for the binding of ligands to an inverse agonist domain at BzR. This model is supported by the recent synthesis of 3-ethoxy-beta-carboline (3), a potent, long-lived partial inverse agonist, and 7, an irreversible BzR ligand.

Behavioral interactions between ethanol and imidazodiazepines with high affinities for benzodiazepine receptors

The intrinsic effects of two imidazodiazepines RO 15–3505 and RO 17–1812 on the behavior of mice in a holeboard test were investigated. The interactions of these two drugs with ethanol were also studied. RO 15–3505 (0.75–6.0 mg/kg) failed to significantly alter either exploratory head-dipping or locomotor activity when administered alone but doses of 0.75 and 1.5 mg/kg reversed the reduction in the number of head-dips caused by ethanol (2 g/kg) and partially reveresed ethanol's locomotor stimulant action. In contrast, RO 17–1812 (0.75–6.0 mg/kg) increased locomotor activity when administered alone, and enhanced the reduction in exploration caused by ethanol. Neither RO 15–3505 nor RO 17–1812 altered blood alcohol concentrations suggesting a pharmacodynamic basis for these interactions. The results suggest that in the holeboard test the interactions of imidazodiazepines with ethanol are related to the nature of their interaction with benzodiazepine receptors, inverse agonists antagonising and agonists enhancing ethanol's effects on exploration.

Effects of benzodiazepines on responses of guinea-pig ileum and gall-bladder and rat pancreatic acini to cholecystokinin

Bradwejn and De Montigny have recently shown that benzodiazepines selectively inhibit excitation of hippocampal neurones by cholecystokinin (CCK). We show here that lorazepam and chlordiazepoxide selectively inhibit the nerve-mediated response of ileal longitudinal muscle to CCK, but have no effect on the direct stimulation of gall-bladder muscle or pancreatic acini by this peptide. Lorazepam (1 and 10 μM) and chlordiazepoxide (0.1 and 1 μM) inhibited responses of guinea-pig ileum, but not gall-bladder to CCK. Responses of both tissues to acetylcholine (ACh) were unaffected and lorazepam (10 μM) did not inhibit ileal responses to neurotensin, 5-hydroxytryptamine and substance P which act entirely or in part by stimulating myenteric nerves. Chlordiazepoxide (1 and 10 μM) did not inhibit CCK-stimulated amylase release from dispersed rat pancreatic acini. Higher concentration of the same drugs and diazepam (1 and 10 μM) which has high affinity for benzodiazepine receptors on gastrointestinal muscle, inhibited responses of ileum and gall-bladder to both CCK and acetylcholine.

Detection and determination of active metabolites of 1-(2- o-chlorobenzoyl-4-chlorophenyl)-5-glycyl-aminomethyl-3-dimethylcarbamoyl-1 H-1,2,4-triazole hydrochloride dihydrate, (450191-S), in rat tissues, using a radioreceptor assay for benzodiazepines

1-(2- o-Chlorobenzoyl-4-chlorophenyl)-5-glycyl-aminomethyl-3-dimethylcarbamoyl-1 H-1,2, 4-triazole hydrochloride dihydrate, (450191-S), exhibits pronounced central nervous system (CNS) activities similar to those of benzodiazepines, but it has only low affinity for benzodiazepine receptors. However, when 450191-S was administered to rats at a dose of 10 mg/kg, brain extracts markedly inhibited [ 3H]diazepam binding to the receptors. Thin-layer chromatography (TLC), high performance liquid chromatography (HPLC), and radioreceptor assay (RRA) were used to isolate three metabolites that could inhibit [ 3H]diazepain binding prominently. These were identified by gas chromatographymass spectrometry (GC/MS) as compounds having the triazolo-benzodiazepine skeleton. They showed high affinities for benzodiazepine receptors ( K i = 0.9 to 2.1 nM) and exerted potent pharmacological effects similar to those of 450191-S. In addition, their levels in the brain were sufficient to explain the pharmacological activity of 450191-S, which could not be detected in tissue extracts 15 min after administration. These results indicate that the pharmacological activity of 450191-S is largely due to the action of active metabolites, although some points remain to be elucidated to fully account for the large attenuation of the side effect (ataxia) compared with the major effects (anti-convulsant and hypnotic). We also determined the brain levels of metabolites following the administration of 450191-S and evaluated the extent to which each active metabolite contributes to the pharmacological activities of this drug.

A novel chemically induced animal model of human anxiety.

The ethyl ester of beta-carboline-3-carboxylic acid (beta-CCE) has a high affinity for benzodiazepine receptors and can antagonize some of the pharmacologic actions of benzodiazepines in rodents. Administration of beta-CCE (2.5 mg/kg) to chair-adapted, male rhesus monkeys (7-9 kg) elicited a behavioral syndrome characterized by extreme agitation, head and body turning, distress vocalization and other behaviors which might be termed 'anxious'. Concomitant increases in plasma cortisol, epinephrine, norepinephrine, heart rate, and mean arterial blood pressure were observed. Pretreatment of animals with the benzodiazepine receptor antagonist Ro 15-1788 (5 mg/kg) antagonized the behavioral, endocrine, and somatic changes produced by beta-CCE, but did not elicit any significant changes in these parameters when administered alone. Thus, administration of beta-CCE to primates may be a reliable and reproducible model of human anxiety and, as much, may prove valuable for studying the postulated role of 'stress' or 'anxiety' in a variety of human disorders. In toto, these results strongly suggest that benzodiazepine receptors not only mediate the pharmacologic actions of benzodiazepines, but may also subserve both the affective and physiologic expression of anxiety.

Beta-Carboline-induced anxiety states.

Several C-3-substituted beta-carbolines (e.g., the ethyl ester of beta-carboline-3-carboxylic acid) have high affinities for benzodiazepine receptors and can antagonize the principal pharmacologic actions of the benzodiazepines. The observation that some of these compounds also possess 'intrinsic' actions, best described as pharmacologically opposite to the benzodiazepines, has provided a chemically induced model of extreme stress or anxiety. The actions of such compounds in currently used animal models of anxiety are reviewed, as well as the effects of these compounds in primates, including man.

Benzodiazepine receptor-mediated experimental "anxiety" in primates.

The ethyl ester of beta-carboline-3-carboxylic acid has a high affinity for benzodiazepine receptors in the brain. In the rhesus monkey this substance produces an acute behavioral syndrome characterized by dramatic elevations in heart rate, blood pressure, plasma cortisol, and catecholamines. The effects are blocked by benzodiazepines and the specific benzodiazepine receptor antagonist Ro 15-1788. The benzodiazepine receptor may consist of several subsites or functional domains that independently recognize agonist, antagonists, or "active" antagonists such as beta-carboline-3-carboxylic acid ethyl ester. These results suggest that the benzodiazepine receptor is involved in both the affective and physiological manifestations of anxiety, and that the administration of beta-carboxylic acid ethyl ester to monkeys may provide a reliable and reproducible animal model of human anxiety.

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

The effects of the methyl, ethyl and propyl esters of β-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 β-carboline esters having different proportions of agonistic and antagonistic actions at their recognition sites. There is good evidence that the pharmacological actions of benzodiazepines are mediated via specific binding sites found in mammalian brain (Braestrup & Squires, 1978; Mohler & Okada, 1977; Squires & Braestrup, 1977). In an attempt to isolate an endogenous ligand for these receptors, Braestrup, Nielsen & Olsen (1980) discovered ethyl-β-carboline-3-carboxylate (β-CCE) in human urine. This compound, although now known to be an extraction artifact, shows a very high affinity for benzodiazepine receptors (Braestrup, Nielsen and Olsen, 1980: Nielsen & Braestrup, 1980). More recently, β-CCE has been shown to have pharmacological actions opposite to those of the benzodiazepines and can reverse some of their actions (Cowen, Green, Nutt & Martin, 1981; Oakley & Jones 1980; Tenen & Hirsch 1980). In the present study, the pharmacological effects of two closely related β-carbolines, methyl-β-carboline-3-carboxylate (β-CCM) and propyl-β-carboline -3-carboxylate (β-CCP), were investigated.

Detection and determination of pharmacologically active benzodiazepines in rat brain after the administration of 2-o-chlorobenzoyl-4-chloro-N-methyl-N'-glycylglycinanilide, using a combination of high-pressure liquid chromatography and radioreceptorassay.

In the present work we attempted to detect and quantitate central nervous system (CNS)-active benzodiazepines in rat brain after administration of a peptido-aminobenzophenone. The results can be summarized as follows. 1. Radioreceptorassay (RRA) using 3H-diazepam as the labeled ligand and ratbrain crude synaptosomes was a simple, reliable and sensitive assay method for benzodiazepines. 2. A benzene extract of a rat brain homogenate prepared 15 min after i.p. administration of 2-o-chlorobenzoyl-4-chloro-N-methyl-N'-glycylglycinanilide (1) was subjected to high pressure liquid chromatography (HPLC). By RRA analysis of the eluted fractions, four benzodiazepines having high affinity for benzodiazepine receptors were detected. 3. Changes in the levels of each benzodiazepine in rat brains after i.p. administration of 1 were determined by a combination of HPLC and RRA. The levels can account for the CNS activities of 1. 1 itself could not be detected throughout the experiments. These results strongly indicate that 1 exerts its CNS activities through conversion into CNS-active benzodiazepines in vivo.