Bidirectional effects of benzodiazepine receptor ligands against picrotoxin- and pentylenetetrazol-induced seizures

Summary The dose response curves of picrotoxin-induced seizures and pentylenetetrazol-induced seizures were shifted to the right by the benzodiazepine (BZ) receptor agonist lorazepam, and to the left by the inverse agonists, DMCM, ZK 90886, FG 7142 and CGS 8216. The BZ receptor antagonists ZK 93426 and Ro 15-1788 had no effect on the dose response curves. The anticonvulsive action of lorazepam and the proconvulsive action of DMCM against picrotoxin-induced seizures and against pentylenetetrazol-induced seizures was inhibited by low doses of ZK 93426 and Ro 15-1788.These results indicate that the bidirectional effects of benzodiazepine receptor ligands on picrotoxin and pentylenetetrazol induced seizures is actually mediated through benzodiazepine receptors.     

Enhanced sensitivity to β-carboline inverse agonists in rats chronically treated with FG 7142

The biochemical and behavioural effects of the chronic administration of the beta-carboline inverse agonist FG 7142 were studied in the rat. Repeated administration of FG 7142 (15 mg/kg IP, twice daily for 10 consecutive days) induced sensitization to the effects of this drug, which from proconvulsant became a full convulsant. Thus, myoclonic seizures were observed in 30% and 80% of the animals by the third and the eighth day of treatment, respectively. The sensitization to the convulsant effect of FG 7142 persisted for up to 50 days after withdrawal and was completely prevented by the concurrent administration of the benzodiazepine receptor antagonist Ro15-1788 (15 mg/kg IP, twice a day for 10 days). Moreover, four to twelve days after withdrawal from chronic treatment with FG 7142, an increased sensitivity to the proconvulsant beta CCE and to the convulsant DMCM was observed. In addition, convulsions induced by isoniazid (350 mg/kg, SC) were potentiated in rats chronically treated with FG 7142 at 5 and 20 days after withdrawal. These pharmacological effects were paralleled by a decrease in the density of low affinity GABA receptors in the cerebral cortex and cerebellum. These results are consistent with the view that repeated administration of FG 7142 induces a long-lasting down-regulation of the GABAergic function which results in an increased sensitivity to beta-carboline inverse agonists and isoniazid. The possibility that a concomitant decrease in the responsiveness to benzodiazepines and Ro15-1788 takes place after chronic treatment with FG 7142 is also discussed.     

Selective susceptibility to inhibitors of GABA synthesis and antagonists of GABA(A) receptor in rats with genetic absence epilepsy

Thalamocortical spike-and-wave discharges characterize the nonconvulsive absence seizures that occur spontaneously in genetic absence epilepsy rats from Strasbourg (GAERS), a selected strain of Wistar rats. GABA is crucial in the generation of absence seizures. The susceptibility to convulsions induced by threshold doses of various GABA receptor antagonists and inhibitors of GABA synthesis, kainic acid and strychnine, was compared in GAERS and in nonepileptic rats from a selected control strain (NE). The brain structures involved in the drug-elicited convulsive seizures were mapped by c-Fos immunohistochemistry. Injection of various antagonists of the GABA(A) receptor, bicuculline and picrotoxin, and inverse agonists of the benzodiazepine site (FG 7142 and DMCM) induced myoclonic spike-and-wave discharges followed by clonic or tonic-clonic seizures with high paroxysmal activity on the cortical EEG. The incidence of the convulsions was dose-dependent and was higher in GAERS than in NE rats. Mapping of c-Fos expression showed that the frontoparietal cortex was constantly involved in the convulsive seizures elicited by a threshold convulsant dose, whereas limbic participation was variable. In contrast, GAERS were less susceptible than NE rats to the tonic-clonic convulsions induced by the inhibitors of glutamate decarboxylase, isoniazide and 3-mercaptopropionic acid. The GABA(B) receptor antagonist CGP 56999 and kainic acid induced a similar incidence of seizures in GAERS and NE rats and predominantly activated the hippocampus. No difference in the tonic seizures elicited by strychnine could be evidenced between the strains. These results suggest that an abnormal cortical GABAergic activity may underlie absence seizures in GAERS.     

Bidirectional effects of beta-carbolines and benzodiazepines on cognitive processes

Experiments with benzodiazepine receptor ligands in two paradigms involving cognitive processing were performed in order to test whether the concept of bidirectional effects of benzodiazepine receptor ligands could also be applied to cognitive functions. Benzodiazepine receptor agonists like chlordiazepoxide, lorazepam, ZK 93423 and ZK 91296 induced amnesia in a passive avoidance paradigm. Mice treated with the benzodiazepine receptor antagonist, ZK 93426, reached a learning criterion after fewer foot-shocks than saline treated mice both in naive animals and in scopolamine pre-treated animals. Furthermore, ZK 93246, attenuated the amnesic effect of corneal electroshock. The inverse agonists FG 7142 and DMCM decreased the detrimental effect of scopolamine on retrieval. In a signal detection paradigm, chlordiazepoxide impaired signal detection. In aged rats ZK 93426, ZK 90886 and FG 7142 had no effect on signal detection but ZK 93426 and FG 7142 attenuated the impairment of signal detection induced by scopolamine. These effects of benzodiazepine receptor ligands may reflect changes in arousal/vigilance, suggesting that BZ inverse agonists may have useful properties in enhancing vigilance.     

Differential sensitivity to inverse agonists of GABA(A)/benzodiazepine receptors in rats with genetic absence-epilepsy.

A strain of Wistar rats, genetic absence epilepsy rats from Strasbourg (GAERS), was selected and inbred over 40 generations for occurrence of spontaneous spike-wave discharges characteristic of absence seizures, simultaneously with a strain of non-epileptic rats (NER). GAERS demonstrate an excessive sensitivity to antagonists of the GABA(A) receptor. The sensitivity to convulsions induced by various inverse agonists of the GABA(A)/benzodiazepine receptor was compared in GAERS and NERs. The beta-carbolines FG 7142 and DMCM, and the imidazobenzodiazepines RO 19-4603 and the alpha 5-selective RY 024 were several times more convulsant in GAERS than in NERs. The largest differences were found with the non-selective RO 19-4603- and FG 7142. The proconvulsant imidazobenzodiazepine RO 15-4513, binding also to diazepam-insensitive receptors, had low efficacy. The high affinity binding of GABA(A)/BZD receptors with (3H) RO 15-1788 in the brain of naive rats and after administration of FG 7142 did not differ in GAERS and NERs. The data indicate that the hypersensitivity of GAERS to various inverse agonists of the GABA(A)/benzodiazepine receptor involves cortical GABA(A) receptors and is not related to differential activity of a subunit-selective receptor.     

Behavioral neurobiology of inverse agonist FG 7142 induced anxiety syndrome in rats

1. Neurobehavioral survey of inverse agonist FG 7142 was performed employing a novel anxiety paradigm namely FR-2 Two-Way crossover in a shuttle box. 2. FG syndrome was found to be similar to learned helplessness following shock treatment. Significant increase in mean latency to escape was observed from 0 to 25th trial. Effect of FG 7142 on the behavioral and neurological profile did not deviate significantly from controls. However, a general increase in arousal, darting and sideway movement (weaving) of the head were noted. 3. Drugs with specificity at benzodiazepine (BDZ) receptor site were employed as pretreatments in order to study their influence on FG 7142 induced anxiety syndrome. Diazepam and ZK 91296, significantly blocked the inverse agonist response. Head weaving as well as darting movements were completely abolished although rats demonstrated arousal and vigilance. 4. A dose dependent inhibition of FG response was observed with RO 15-1788, a specific BDZ receptor blocker. ZK 93423 at low doses (1 mg/kg) failed to reverse the escape deficit induced by FG. Higher doses of ZK 93423 could not be tested as significant sedation and ataxia were noted. 5. It is suggested that a careful combination of a BDZ agonist and inverse agonist may be beneficial in the treatment of generalised anxiety disorders.     

Lorazepam and FG 7142 induce tolerance to the DMCM antagonistic effect of benzodiazepine receptor ligands

Mice were given chronic treatment with lorazepam 10 mg/kg PO or FG 7142 40 mg/kg IP once a day for 14 days. The pretreatments with lorazepam and FG 7142 did not change the sensitivity of the mice to the convulsant effect of DMCM. Lorazepam pretreated mice showed a significantly lower sensitivity to the anticonvulsant effects of the benzodiazepine (BZ) receptor ligands lorazepam, ZK 93423, ZK 91296, Ro 15-1788 and ZK 93426 administered acutely by the IP route when challenged with DMCM 24 hr after the last dose of lorazepam. FG 7142 pretreated mice showed a significantly lower sensitivity to the anticonvulsant effect of the two agonists lorazepam and ZK 93423 and to the antagonist Ro 15-1788, whereas the effects of ZK 91296 and ZK 93426 were left unchanged. The reduced DMCM antagonistic effects of the BZ receptor ligands may indicate that these ligands may either have lost some of their affinity to those BZ receptors being responsible for the DMCM-induced seizures or they may have lost some efficacy in allosterically inhibiting DMCM binding or as a third possibility may have lost efficacy at a BZ receptor site downstream to the seizure-inducing center in the brain.     

Suppression of the immune response by benzodiazepine receptor inverse agonists

24 h after administration of a single dose of the benzodiazepine receptor inverse agonists N'-methyl-beta-carboline-3-carboxamide (FG 7142) and 3-carbomethoxy-4-ethyl-6,7-dimethoxy-beta-carboline (DMCM), a profound suppression of the immune response was observed in rodents. This immunosuppression was manifest as a decrease in phytohemagglutinin (PHA) and concanavalin-A (Con-A) stimulated T cell proliferation in rats and mice administered FG 7142 and a decrease in allogeneic cytotoxic T lymphocyte activity in mice administered either FG 7142 or DMCM. The effects of FG 7142 were antagonized by the prior administration of Ro 15-1788, a benzodiazepine receptor antagonist. These findings demonstrate that the neural pathways subserved by benzodiazepine receptors can modulate immune function, and suggest that these receptors may be involved in the stress-induced modulation of immune function.     

Beta-carbolines characterized as benzodiazepine receptor agonists and inverse agonists produce bi-directional changes in palatable food consumption

Drugs which bind to specific benzodiazepine recognition sites fall into three categories: agonists, antagonists, and inverse agonists. A set of biochemical parameters is available which distinguishes between the three. In addition, actions of the drugs result in physiological and behavioural effects which are distinguishable. beta-Carboline derivatives provide a group of compounds which show high affinity for the benzodiazepine sites, and which contains examples belonging to each of the three categories. Evidence is reviewed which shows that beta-carboline benzodiazepine receptor agonists (ZK 93423, ZK 91296) produce increases in the consumption of a palatable diet by non-deprived rats, that beta-carboline inverse agonists (FG 7142, DMCM) produce an anorectic effect, and that the beta-carboline ZK 93426 acts as a benzodiazepine receptor antagonist. The results support the proposal of bi-directional control of feeding responses through the action of drugs at a common benzodiazepine receptor. Furthermore, benzodiazepine receptor inverse agonists provide a novel class of anorectic agents. Evidence is also reviewed which is suggestive of the modulation of food-related reward by drug actions at benzodiazepine receptors.     

Blockade of FG 7142 kindling by anticonvulsants acting at sites distant from the benzodiazepine receptor

Repeated administration of the beta-carboline FG 7142 results in sensitisation to its convulsant effects (chemical kindling); acutely FG 7142 is not convulsant, but following several treatments full seizures develop. It has been suggested that the increased sensitivity results from changes in benzodiazepine (BZ)/GABA receptor function. The present experiments studied the ability of BZ receptor ligands and anticonvulsant drugs with diverse mechanisms of action to block the expression and development of kindling to once daily injection of FG 7142 (40 mg/kg, i.p.) in mice. In fully kindled mice, the BZ receptor agonists clonazepam, ZK 93,423 and CL 218,872, and the antagonists flumazenil and ZK 93,426 prevented FG 7142 convulsions, as did 2 anticonvulsants, sodium valproate, possibly acting by influencing GABAergic transmission, and ethosuximide. A further two substances, MK 801 and 2-chloradenosine which act respectively via glutamatergic and purinergic mechanisms were also effective. When administered concomitantly with repeated FG 7142, all of these substances prevented or strongly reduced the development of kindling. Phenytoin and carbamazepine were ineffective in protecting against FG 7142 convulsions in kindled mice, and in preventing the development of kindling when administered repeatedly together with FG 7142. Since MK 801 and 2-chloradenosine prevented kindling, these results suggest that an interaction of FG 7142 with BZ receptors is not sufficient to induce kindling, which may instead result from secondary changes in sites distant from BZ/GABA receptors.     

β-Carbolines characterized as benzodiazepine receptor agonists and inverse agonists produce bi-directional changes in palatable food consumption

Drugs which bind to specific benzodiazepine recognition sites fall into three categories: agonists, antagonists and inverse agonists. A set of biochemical parameters is available which distinguishes between the three. In addition, actions of the drugs result in physiological and behavioural effects which are distinguishable. β-Carboline derivatives provide a group of compounds which show high affinity for the benzodiazepine sites and which contains examples belonging to each of the three categories. Evidence is reviewed which shows that β-carboline benzodiazepine receptor agonists (ZK 93423, ZK 91296) produce increases in the consumption of a palatable diet by non-deprived rats, that β-carboline inverse agonists (FG 7142, DMCM) produce an anorectic effect and that the β-carboline ZK 93426 acts as a benzodiazepine receptor antagonist. The resuks support the proposal of bi-directional control of feeding responses through the action of drugs at a common benzodiazepine receptor. Furthermore, benzodiazepine receptor inverse agonists provide a novel class of anorectic agents. Evidence is also reviewed which is suggestive of the modulation of food-related reward by drug actions at benzodiazepine receptors.     

Low and high doses of benzodiazepine receptor inverse agonists respectively improve and impair performance in passive avoidance but do not affect habituation

The benzodiazepine receptor inverse agonists, methyl 6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate (DMCM) and N-methyl-beta-carboline-3-carboxamide (FG 7142), were given to rats at various stages of a passive avoidance task. When the drugs were given before trial 1, low doses enhanced, and high doses impaired, performance as assessed 24 h later. A group given drugs on both trials showed that the impairment was not due to state-dependent effects. When the drugs were given immediately after trial 1, or before trial 2, they were without effect, except for the low dose of DMCM which impaired consolidation. It is discussed whether the changes in passive avoidance performance are due to direct or indirect effects. Between-trial habituation of exploratory head-dipping was measured in a holeboard. When FG 7142 was given before trial 1, the high dose impaired between-trial response decrement; but this was because it decreased the level of head-dipping on trial 1. When FG 7142 was given immediately after trial 1, or before trial 2, it was without effect on between-trial habituation.     

Changes in local cerebral glucose utilization induced by the beta-carbolines FG 7142 and DMCM reveal brain structures involved in the control of anxiety and seizure activity

The brain regions that may be functionally involved in the control of anxiety and the development of seizures were examined using quantitative 1-14C-deoxyglucose autoradiography. For this purpose, beta-carbolines FG 7142 and DMCM were employed. They exert their effects via the benzodiazepine receptor, and whereas both possess anxiogenic properties, FG 7142 is a proconvulsant and DMCM a potent convulsant. The pattern of increases of local cerebral glucose utilization (LCGU) induced by FG 7142 was mainly restricted to limbic structures, such as the lateral septal nucleus, the anterior thalamic nuclei, and the mamillary nuclei. However, structures involved in motor regulation were also affected. A pronounced increase in LCGU was observed in the posterior part of the substantia nigra, pars reticulata. Further, the LCGU of the globus pallidus, the ventral thalamic nucleus, and the cerebellum was increased. DMCM likewise increased LCGU of the mamillary body and the lateral septal nucleus. In contrast to FG 7142, the hippocampal formation displayed an increase in LCGU, while LCGU of the anterior thalamic nuclei was unchanged. A pronounced increase in LCGU was seen in the substantia nigra, pars reticulata in addition to other structures functionally involved in central motor regulation. The specific benzodiazepine antagonist Ro 15-1788 antagonized the effects of both FG 7142 and DMCM. It is concluded that the beta-carbolines FG 7142 and DMCM produce selective effects upon LCGU that are mediated by benzodiazepine receptors.(ABSTRACT TRUNCATED AT 250 WORDS)     

Immediate effects of 14 non MAOI antidepressants in rats with spontaneous petit mal-like seizures

1. Wistar rats of a strain presenting spontaneous petit mal-like seizures were injected intraperitoneally with graded doses of 14 non-monoamine oxidase inhibitor antidepressants and the immediate effects on behavior and the EEG were recorded. 2. Amineptine and nomifensine, the two drugs interacting with dopaminergic neurotransmission, reduced the duration of spontaneous spike-wave discharges (SWD) and were thus potentially antiepileptic. 3. Trazodone increased SWD duration. 4. The antidepressants, imipramine-like (imipramine, chlorimipramine, desipramine, metapramine and amitriptyline) and non-imipraminic (minaprine, maprotiline, viloxazine, mianserin, fluvoxamine and indalpine), and the 3 noted above, had potentially convulsive effects.     

Stimulation of cortical acetylcholine efflux by FG 7142 measured with repeated microdialysis sampling

The effects of the benzodiazepine receptor partial inverse agonist β-carboline FG 7142 on cortical ACh efflux were determined using in vivo microdialysis in freely-moving rats. Additionally, a within-subjects, repeated-dialysis experimental design (four microdialysis sessions; removable dialysis probe) was evaluated as a method for measuring changes in basal and FG 7142-stimulated ACh efflux in the frontoparietal cortex. FG 7142 (4.0, 8.0, and 16.0 mg/kg) produced a 150–470% increase in cortical ACh efflux, with a dose-dependent effect on the duration of the increase in efflux. Basal cortical ACh efflux was lower in session 4 than in session 1. However, the ability of FG 7142 to stimulate efflux was unchanged by repeated dialysis testing. The ability of tetrodotoxin (1.0 μM) to suppress both basal and FG 7142-stimulated ACh efflux was also unaffected by repeated dialysis testing. These results demonstrate that systemically administered benzodiazepine receptor inverse agonists stimulate cortical ACh efflux, and that repeated-measures experimental designs can be valid for determining certain changes in cortical ACh efflux with in vivo microdialysis. © 1995 Wiley-Liss, Inc.     

Cardiovascular and somatic startle and defense: Concordant and discordant actions of benzodiazepine receptor agonists and inverse agonists

Benzodiazepine receptor (BZR) agonists and inverse agonists yield generally opposing effects on GABAergic transmission, and the functional consequences of these ligands are often bidirectional. BZR agonists exert anxiolytic effects, whereas the BZR partial inverse agonist FG 7142 has been reported to have anxiogenic actions in a variety of paradigms. In keeping with this literature, we found that the cardioacceleratory defensive response is enhanced by FG 7142, and attenuated by the BZR agonist chlordiazepoxide. In contrast, both compounds attenuated basal and fear-potentiated somatic startle responses. This did not appear to reflect a global reduction of startle reactivity, however, as the cardiac startle response was not significantly altered. These findings support the view that multiple substrates underlie distinct aspects or features of fear and anxiety. The results are consistent with the suggestion that FG 7142 may selectively enhance those aspects of anxiety that depend on cortical-cognitive processing.     

Beta-carboline interactions at the BZ-GABA receptor chloride-ionophore complex in the rat cerebral cortex

beta-Carboline congeners can act at the benzodiazepine (BZ) recognition site of the BZ-GABA receptor complex to increase GABA-stimulated chloride conductance (agonist effect), inhibit this conductance (inverse agonist effect) or block the actions of agonists and inverse agonists (antagonist effect). In this communication we describe the effects of several beta-carbolines (ZK 93423, ZK 91296, ZK 93426, and DMCM) on GABA-stimulated chloride influx into vesicles prepared from rat cerebral cortex. ZK 93423 produces an approximate 2-fold left-shift of the GABA dose-response curve at a concentration of 1.0 microM consistent with its full agonist activity (positive intrinsic efficacy), while the same concentration of ZK 91296 produces over a 1-fold left-shift consistent with its partial agonist activity. At higher concentrations (0.1 mM), ZK 91296 inhibits GABA-stimulated chloride influx which appears to be mediated through a non-BZ receptor mechanism since this effect is not reversed by the BZ antagonist ZK 93426. The augmenting effect of both ZK 93423 and ZK 91296 on GABA-stimulated chloride flux was reduced by the antagonist ZK 93426 in a dose-dependent manner and reached GABA-stimulated control levels at a ZK 93426 concentration of 1.0 microM. Interestingly, there was a further inhibition of the GABA-stimulated chloride influx at higher concentrations of ZK 93426 which is not seen when ZK 93426 is used in the absence of BZ agonists. The inverse agonist activity of DMCM was incompletely blocked by the antagonist ZK 93426. These data show that ZK 93426 can antagonize the effects of the full agonist ZK 93423 and partial agonist ZK 91296 at the BZ receptor. Furthermore, the interaction of the agonists with ZK 93426 results in the appearance of inverse agonist-like activity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Behavioral and biochemical evidence for a long-lasting decrease in GABAergic function elicited by chronic administration of FG 7142

Chronic treatment with the beta-carboline derivative FG 7142 (15 mg/kg i.p. twice a day for 10 consecutive days) produced a long-lasting enhancement of shock-induced suppression of drinking in rats, without affecting unpunished behaviour. This proconflict effect was observed up to 15 days after withdrawal from FG 7142. A significant sensitization to seizures induced by isoniazid, a drug known to inhibit GABAergic transmission, was also found to occur after long-term (25 days) withdrawal. Moreover, the density of low-affinity GABA receptors was decreased by 30% in the cerebral cortex of rats repeatedly injected with FG 7142 at 5 and 15 days after withdrawal. The capacity of high-affinity GABA receptors, as well as the apparent dissociation constants for both high- and low-affinity GABA receptors were unchanged. Similar modifications in [3H]GABA binding were also observed in the cerebellum. The enhancement of punishment suppressed behaviour, the sensitization to isoniazid-induced convulsions and the decrease in the density of low-affinity GABA receptors suggest that chronic administration of FG 7142 induces a persistent down-regulation of GABAergic transmission in the central nervous system.     

Chronic treatment with diazepam or the inverse benzodiazepine receptor agonist FG 7142 causes differential changes in the effects of GABA receptor stimulation

Daily treatment of mice with diazepam leads to the development of tolerance to the anxiolytic and anticonvulsant effect of the benzodiazepine, while daily treatment with the proconvulsant benzodiazepine receptor inverse agonist FG 7142 produces sensitization to its effects in that seizures develop (chemical kindling). In the present study, the effects of GABA receptor stimulation were studied 2 days after termination of 13 days treatment with diazepam, 20 mg/kg i.p./day, and FG 7142, 40 mg/kg i.p./day. For GABA receptor stimulation, the GABA agonist progabide was chosen because among several GABA receptor stimulants tested it was the only compound that induced increases in seizure threshold in non-toxic doses. Using the threshold for maximal (tonic extension) electroconvulsions as a measure for anticonvulsant efficacy, the anticonvulsant effect of progabide (100 mg/kg i.p.) was unchanged after chronic treatment with diazepam but was lost in FG 7142 kindled animals. Conversely, the hypothermic effect of progabide was reduced after treatment with diazepam but not with FG 7142. Baseline seizure threshold was unchanged 2 days after chronic administration of diazepam but increased in the FG 7142 pretreated mice. The data indicate that tolerance to benzodiazepines and kindling by FG 7142 are associated with different changes in GABA receptor function.     

Bidirectional effects of chronic treatment with agonists and inverse agonists at the benzodiazepine receptor

We have studied in rodents the effects of beta-carboline inverse agonists on chronic treatment and after repeated administration of benzodiazepine agonists. Chronically, the inverse agonist FG 7142 caused chemical kindling, i.e., a decrease in the threshold to the convulsive effects of the drug. This change was accompanied by decreases in the effects of beta-carboline but not benzodiazepine agonists. In addition the effects of GABA receptor agonists were decreased and the effects of GABA antagonists marginally increased. The GABA stimulated benzodiazepine binding was lower after FG 7142 kindling. Some evidence was found in mice to suggest that these changes were accompanied by behavioural alterations, but studies in rats did not show any changes. Repeated administration of benzodiazepine agonists, sufficient to cause tolerance to their pharmacological actions and to those of beta-carboline agonists, increased all of the effects of the partial inverse agonists and some of the actions of the full inverse agonists. We suggest that this is due not to precipitation of withdrawal but to a "withdrawal shift" in the coupling at the receptor inophore. This would increase the intrinsic properties of inverse agonists and decrease those of agonists. Evidence for this hypothesis is summarised.