Action Of Benzodiazepines Research Articles

Autoradiography of benzodiazepine receptor binding in the central nervous system of the normal C57BL6J mouse.

[3H]flunitrazepam has been used as a photoaffinity label for the specific, clonazepam-displaceable 1,4-benzodiazepine binding sites in sections of normal C57BL6J mouse brain and spinal cord. Binding was visualized by light microscope autoradiography and quantified by a simple microdensitometric procedure. Specific flunitrazepam binding was seen to be highest in the colliculi, cerebral cortex, hippocampal formation, interpeduncular nucleus, mamillary body, hypothalamus, olfactory tubercle, and in the molecular layer and deep nuclei of the cerebellum. The distribution of specific flunitrazepam binding sites in mouse brain and spinal cord is discussed in terms of the known actions of the benzodiazepines.

Synaptic and non-synaptic actions of benzodiazepines on the crayfish sensory neuron

On the stretch-induced discharge activity of the isolated crayfish sensory neuron flurazepam (⩽ 3 × 10 −4 M) and Ro 15-1788 (⩽ 10 −3 M) produced reversible concentration-dependent excitation, but oxazepam only produced depression (⩽ 5 × 10 −4 M). Similar divergent effects on the membrane properties were observed. Oxazepam increased the threshold to firing without changing resting potential, membrane resistance or the GABA-mediated IPSP. In contrast flurazepam and Ro 15-1788 produced a concentration-dependent decrease in threshold. Flurazepam did not alter membrane resistance or resting potential but facilitated GABA transmission. Ro 15-1788 had the opposite effect on the GABA synapse, and also depolarized the resting potential but did not alter membrane resistance. The change in spike threshold appeared to be an important component in producing discharge excitation or depression. These results not only demonstrate the capability of the sensory neuron to discriminate between structures of benzodiazepines, but also that these agents can produce divergent effects on synaptic and non-synaptic properties of a single neuron.

Synthesis of previously inaccessible quinazolines and 1,4-benzodiazepines as potential anticonvulsants.

A series of 4,6,7,8- tetrasubstituted 3,4- dihydroquinazolines , quinazolines, quinazolin -2-ones, 1,2,3,4- tetrahydroquinazolin -2-ones, and 5,7,8,9- tetrasubstituted 1,4-benzodiazepines have been synthesized by utilizing the Diels -Alder reaction between furan o-amino nitriles and various alkyl or aryl vinyl ketone dienophiles to obtain the anthranilic acid precursors. All of the newly synthesized target compounds were evaluated in mice for anticonvulsant activity. Pro- and anticonvulsant action was quantified by the timed intravenous pentylenetetrazol seizure threshold method. Selected compounds were also evaluated for benzodiazepine receptor binding properties and in vivo antagonist potential. Although the compounds lack potency, the data suggest that previously inaccessible substituted analogues may be useful to segregate the proconvulsant , anticonvulsant, and antagonist actions of benzodiazepines and quinazolines.

Effects of chlordiazepoxide and of valproate on hyponeophagia in rats Evidence for a mutual antagonism between their anxiolytic properties

The effects of chlordiazepoxide (0,2.5 and 7.5 mg/kg), of valproate (0,100 and 300 mg/kg), and of combinations of these drugs, on hyponeophagia in rats, were studied using a food preference test. Chlordiazepoxide alone reduced the latency for eating, and also enhanced the time of eating and the amount of familiar food eaten at the smaller dose, and of novel food eaten at the larger dose. Valproate alone reduced the latency of eating and enhanced time of eating and the amount of novel food eaten, especially at 300 mg/kg. However, valproate antagonised the effects of chlordiazepoxide on all behavioural measures. These findings are discussed in the context of the GABA hypothesis of the actions of benzodiazepines, with the conclusion that valproate has an atypical pharmacological profile amongst putative GABA agonists, and may have partial agonist activity at the proposed benzodiazepine/GABA receptor complex.

Effect of diazepam on plasma corticosterone levels.

In light of the numerous but rather conflicting reports on the action of benzodiazepines upon the hypothalamo-hypophyseal-adrenal (HHA) axis activity, the effect of different doses of diazepam (0.1, 1.0 and 10.0 mg/kg) administered 15, 30, 60, 120 and 240 min before decapitation on plasma corticosterone level was studied in rats. While 0.1 mg/kg diazepam had no effect, 1.0 mg/kg diazepam decreased plasma corticosterone levels 30 and 60 min following drug administration. On the other hand, treatment with 10.0 mg/kg diazepam produced an increase in plasma corticosterone levels from 15-120 min following drug administration.

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.

Actions of benzodiazepines on the neuroendocrine system

The administration of benzodiazepines to rats is associated with alterations in the secretion of anterior pituitary hormones. Although basal secretion of prolactin and thyroid stimulating hormone (TSH) was not changed by benzodiazepines, the secretory response to physiological and pharmacological stimuli was attenuated. Diazepam reduced the rise in secretion of prolactin during proestrus and the increase in release induced by stress, administration of fluoxetine plus 5-hydroxytryptophan and haloperidol. Diazepam also blocked the rise in release of TSH induced by exposure to cold. To establish the site at which benzodiazepines act, anterior pituitary binding for [ 3H]diazepam was characterized. Scatchard analysis indicated a K d of 11 nM and a B max of 27 fmol/10 6 cells. In cultures of anterior pituitary cells diazepam enhanced the inhibition of prolactin secretion produced by γ-aminobutyric acid (GABA) and dopamine. The potency of benzodiazepine analogues established that the benzodiazepine binding site in the pituitary belonged to the peripheral subtype of benzodiazepine receptors. However, the potency of analogues for inhibition of the release of prolactin and TSH following systemic administration indicates that benzodiazepines with affinity for the central subtype of receptors were the most potent. These data are interpreted to suggest that the actions of benzodiazepines on the neuroendocrine system result from their actions within the CNS on the neurons involved in the regulation of pituitary function. In addition, peripheral benzodiazepine receptors are present on cells of the anterior pituitary and these binding sites can modulate secretion of prolactin in response to its inhibitory regulators.

Adenosine receptor interactions and anxiolytics

[ 3 H]-N 6- cyclohexyladenosine and [ 3H]-1,3-diethyl-8-phenylxanthine label the A 1 subtype of adenosine receptor in brain membranes. The affinities of methylxanthines in competing for A 1 adenosine receptors parallel their potencies as locomotor stimulants. The adenosine agonist N 6-(-phenylisopropyl) adenosine is a potent locomotor depressant. Both diazepam and N 6-( l- phenylisopropyl) adenosine cause locomotor stimulation in a narrow range of subdepressant doses. Combined stimulant doses of the two agents depress motor activity, as do larger doses of either one, given separately. Evidence supporting and against the hypothesis that some of the actions of benzodiazepines are mediated via the adenosine system is reviewed. A number of compounds interact with both systems, probably because of physico-chemical similarities between adenosine and diazepam. It is concluded that of the four classic actions of benzodiazepines, the sedative and muscle relaxant (but not anxiolytic or anticonvulsant) actions could possibly be mediated by adenosine.

Anxiogenic action of a convulsant benzodiazepine: reversal by chlordiazepoxide

Ro5-3663, a convulsant benzodiazepine and a ligand for CNS picrotoxin sites, but not for benzodiazepine sites, had a significant dose-related (2 and 4 mg/kg) anxiogenic action in the social interaction test of anxiety. This anxiogenic effect was reversed by chlordiazepoxide (5 mg/kg).

Intrinsic and antagonistic effects of β-carboline FG 7142 on behavioral and EEG actions of benzodiazepines and pentobarbital in cats

The methyl-β-carboline-3-carboxamide FG 7142 (10 mg/kg i.p.) produced a pronounced state of alertness, attentive behavior and fearfulness in the cat. Electroencephalographic (EEG) desynchronization in the cortex and theta rhythms in the hippocampus occurred concomitantly. Behavioral and hippocampal EEG actions of FG 7142 were reduced by Ro 15-1788 (10 mg/kg i.p.). The benzodiazepines diazepam and quazepam produced behavioral and EEG effects which were completely antagonized by FG 7142. The pentobarbital effects also were partially antagonized. These findings suggest that the cat provides an interesting model to study the anxiogenic properties of β-carbolines.

Multiple benzodiazepine receptors: autoradiographic localization in normal human amygdala

Because previous studies have emphasized the importance of the amygdala in the therapeutic actions of benzodiazepines and described differences in benzodiazepine receptor distribution between human and rat, this study examined the distribution of multiple benzodiazepine receptors in normal human amygdala by light microscopic autoradiography. Benzodiazepine receptors were labeled with [ 3H]flunitrazepam at 5 representative rostro-caudal levels. Type 1 and Type 2 receptors were differentiated by the addition of 200 nM CL218,872 to serial sections and subsequent analysis based on the differential occupancy at Type 1 and Type 2 receptors by this drug. Results demonstrated that like the rat amygdala, human amygdala contains a higher density of benzodiazepine receptors in the basolateral nuclear complex compared to the corticomedial complex, and more Type 2 than Type 1 receptors overall. However, while rat amygdala is enriched rostrally in Type 1 receptors, this subclass in humans is elevated caudally. Pathways including the amygdala and limbic and cortical targets of its efferents may be preferential loci of benzodiazepine anxiolytic activity.

Structure-activity relationships in kynurenine, diazepam and some putative endogenous ligands of the benzodiazepine receptors

Kynurenine, an endogenous cerebral and peripheral neuroactive metabolite of l-tryptophan, exerts stimulant and convulsant effects in mice, rats and frogs. In mice it (intracerebroventricularly, ICV) antagonized the anticaffeine effect of diazepam and in smaller doses potentiated its sedative action. In rats l-kynurenine (ICV) potentiated the convulsant action of caffeine. The effect of pentylenetetrazol was not altered in either species. The convulsant effect of l-kynurenine is the most resistant among various convulsants towards the protective action of diazepam. The structure of l-kynurenine is similar to benzophenones, metabolites of diazepam, and has four structural fragments common with diazepam. Putative endogenous and non-endogenous ligands of the benzodiazepine receptors have from one to three of these common fragments. Among the antagonists of diazepam exhibiting stimulant and convulsant action ethyl-β-carbonline-3-carboxylate has the same four fragments, Ro 5–3663 and Ro 15–1788 have three and caffeine two. The most striking dissimilarity is a diazo-moiety (NCCN or NCCC N) absent in the structure of l-kynurenine. This moiety seems to be the most important for the binding to the benzodiazepine receptors. A role of each fragment and their combinations as well as the stereoconfiguration for the pharmacological activity is considered. It is suggested that l-kynurenine is a putative endogenous modulator or, less probably, ligand of the benzodiazepine receptor of either type (most probably that which mediates anxiolytic action of benzodiazepines) or a part of this receptor. The benzodiazepine receptor might be a phylogenetically transformed kynurenine receptor. Highly selective antagonism of purines to l-kynurenine suggests that it can modulate the function of the benzodiazepine receptors via purinergic mechanisms. Stimulant and convulsant action of l-kynurenine can be related to a moiety of succinic acid (OCCCCO) which is typical of quinolinic acid, the strongest endogenous convulsant among kynurenines, and aspartic acid, an excitatory amino acid. l-Kynurenine is suggested to be an anxiogenic and convulsigenic endogenous factor.

A specific inhibitory action of certain benzothiazepines and benzodiazepines on the sodium-calcium exchange process of heart and brain mitochondria

A specific inhibitory action of certain benzothiazepines and benzodiazepines on the sodium-calcium exchange process of heart and brain mitochondria

Beta-Carbolines enhance shock-induced suppression of drinking in rats.

By using Vogel's method to test the anxiolytic action of benzodiazepines and reducing the intensity of the current delivered to the drinking tube, it is possible to distinguish the pharmacological activity of three types of ligands for the benzodiazepine recognition site. An anticonflict action typical of anxiolytic benzodiazepines, a proconflict action typical of many beta-carbolines, including FG 7142 (beta-carboline-3-carboxylic acid ethyl ester methyl amide), and an antagonistic action of the proconflict and anticonflict actions typical of RO 15-1788 (ethyl-8-fluoro-5, 6-dihydro-5-methyl-6-oxo-4H-imidazol[1,5-alpha]-[1, 4]-benzodiazepine-3-carboxylate) and CGS 8216 (2-phenylpyrazolo[4,3-c]quinolin-3-(5H)-one). Pentylenetetrazole, which causes convulsions by interacting with a subunit of the gamma-aminobutyric acid receptor that is different from the benzodiazepine recognition site, also induces a proconflict action that is antagonized by anxiolytic benzodiazepines but not by RO 15-1788.

Diethylpyrocarbonate modification of benzodiazepine receptors from calf cerebral cortex.

Diethylpyrocarbonate (DEP), an amino acid modifying reagent, causes complete inactivation of particulate and deoxycholate-solubilized benzodiazepine-receptors from calf cerebral cortex. No heterogeneity was observed in DEP-sensitivity of the receptors. Protection from DEP-induced inactivation was provided by the centrally active benzodiazepines, diazepam and nitrazepam, but not by the peripherally active Ro5-4864, suggesting that DEP modifies a residue which is essential for the central actions of benzodiazepines. GABA did not protect against inactivation or influence the protection afforded by diazepam, indicating that the DEP-modifiable residue is independent of GABA binding sites, or that GABA binding sites are also sensitive to DEP. DEP-induced inactivation of benzodiazepine-receptors proceeds much faster at pH 10.1 than at pH 8.1 or 6.0, indicating the modification of a high pKa side group, possibly the phenol of a tyrosyl residue. This postulation is in accord with our previous findings with the modifying reagents tetranitromethane and N-acetylimidazole.

A benzodiazepine receptor antagonist decreases sleep and reverses the hypnotic actions of flurazepam.

The benzodiazepine receptor antagonist 3-hydroxymethyl-beta-carboline, which blocks several of the pharmacological actions of benzodiazepines, induces a dose-dependent increase in sleep latency in the rat. Furthermore, at a low dose that by itself does not affect sleep, 3-hydroxymethyl-beta-carboline blocks sleep induction by a large dose of flurazepam. The benzodiazepine receptor may play a role in both the physiological regulation and pharmacological induction of sleep.

The biological basis of benzodiazepine actions.

In spite of intensive research during the decade following the introduction of chlordiazepoxide into clinical medicine, the mechanism(s) of actions of benzodiazepines remained obscure. An important breakthrough occurred with the discovery of the role of gamma -amino-butyric acid (GABA) as the main inhibitory neurotransmitter in the mammalian central nervous system (CNS). Demonstration that there was a close association between benzodiazepine actions and GABAergic mechanisms (Polc, Mohler & Haefely, 1974; Costa, Guidotti & Mao, 1975; Haefelyet at., 1975) led to massive amounts of research which clarified the relationship between GABA and benzodiazepines and resulted in a number of fundamental neurochemical discoveries. This chapter reviews current understanding of the biological basis of benzodiazepine actions and builds on earlier comprehensive reviews (Haefely, 1977, 1978, 1980, 1983b; Haefelyet al., 1975, 1978, 1981, 1983a).

The lateral vestibular nucleus is a very important site for depressant actions of benzodiazepines on the crossed extensor reflex

We have reported that the crossed extensor reflex (CER) in Wistar male chloralose-anesthetized rats is very susceptible to the depressive action of benzodiazepine derivatives. The present study was designed to elucidate a possible site for this action in the central nervous system. Since the reflex contraction of the femoral quadriceps muscle elicited by an electrical stimulation of contralateral sciatic nerve was little influenced by diazepam (DZ) (0.05-2 mg/kg i.v.) in the chronic spinal rat, it is very probable that DZ may act first on supraspinal structures. DZ (2.5 μg) injected into the 4th ventricle produced much more potent and rapid inhibition of the CER than injection into the lateral ventricle. To find the most sensitive site involved in the area close to the 4th ventricle, microinjections of DZ in doses of 1 and 25 ng into the lateral vestibular nucleus (LVN) ipsilateral to the testing muscle produced a dose-dependent reflex depression which was clearly antagonized by Ro 15-1788, a benzodiazepine antagonist. Injections of 100 ng of DZ into other areas, i.e., the contralateral LVN, other subcerebellar nuclei, the red nucleus and the substantia nigra, produced no effects on the CER. Furthermore, electrolytic lesions of the ipsilateral LVN decreased the CER, while an electrical conditioning stimulation of the same nucleus increased it.

Gaba and endorphln mechanisms in relation to the effects of benzodiazepines on feeding and drinking

1. 1. Behavioural actions of benzodiazepines have a number of significant characteristics. Anxiolytic effects are demonstrable both clinically and experimentally; in addition, there is excellent evidence for a reinforcing effect of these compounds, and a direct involvement in ingestional responses. This review focusses on the effects of benzodiazepines on the latter feeding and drinking responses. 2. 2. A necessary mediator of benzodiazepine action in the central nervous system appears to be the facilitation of inhibitory GABAergic neurotransmission. It follows, therefore, that behavioural consequences of benzodiazepine action may depend crucially on enhanced GABAergic activity in the brain. Evidence for some involvement of GABAergic mechanisms in the control of feeding and drinking responses is reviewed. Only a few data are so far available to link benzodiazepines effects on ingestional behaviour directly to GABAergic transmission. 3. 3. A major current theme in the psychopharmacology of feeding and drinking behaviour is the possible involvement of endogenous opioid peptides. There is a strong suggestion in the experimental data that there are links between benzodiazepine and endorphinergic mechanisms in relation to ingestional responses. A promising future line of approach appears to be a delineation of benzodiazepine-GABA-endorphin interrelations in the control of food and water consumption.

An erythrocyte deformability improving action of midazolam.

A water soluble imidazobenzodiazepine, midazolam, [8-chloro-6-(fluorophenyl)-1-methyl-4H-imidazo-1,5-A)(1,4)benzodiazepine], is now under clinical trials as an anesthesia inducer in surgical operation. In addition to its typical benzodiazepine actions, we now report its new action on erythrocyte deformability. Erythrocyte filterability as a measure of erythrocyte de-formability was measured in vitro and ex vivo as described before (Experientia 37: 890, 1981). The decrease in deformability was induced by doubling the osmolality. Midazolam was found to significantly prevent the decrease in filterability at 10−4 M in vitro and at 100 mg/kg p.o. and 10 mg/ kg i.v. ex vivo. This erythrocyte deformability improving action of midazolam was attributed to its preventive effect on an echinocytic morphological change of the erythrocyte from the normal discocyte cell shape under scanning electronmicroscopy ; under hyperosmorality,65% of erythrocytes was changed to echinocytes, whereas in the presence of 10−4 M of midazolam, only 32% of erythrocytes was changed (p<0.001).