David Norman Stephens

Attenuation of scopolamine-induced impairment of spontaneous alternation behaviour by antagonist but not inverse agonist and agonist β-carbolines

Mice were tested in a simple automated Y-maze. Total number of arm entries and alternation behaviour were measured. The latter is thought to reflect working memory capacity at a rudimentary level. During an 8-min session, vehicle-treated mice performed 32.4±7.4 arm entries, 51.0±12.4% of which were organized in alternations (triplets). The two variables showed a negative correlation. Scopolamine (1.0 mg/kg) significantly enhanced activity, reduced alternation behaviour and diminished the correlation between the two variables. The effects of benzodiazepine receptor inverse agonist, antagonist and agonist β-carbolines on this spontaneous behaviour and on the effects of scopolamine were examined. The effects of inverse agonists and agonists on locomotor activity were complex in interaction with both vehicle and scopolamine. The scopolamine-induced reduction of alternation behaviour was significantly reversed by the antagonist ZK 93426 but not by inverse agonists; furthermore, partial agonists and agonists showed no effects. It is hypothesized that the interaction of antagonist β-carbolines with scopolamine is based on a direct GABA-ergic control of cholinergic neurotransmission, and suggests an ability of antagonist β-carbolines to antagonize amnestic properties of scopolamine.

Evaluation of the β-carboline ZK 93 426 as a benzodiazepine receptor antagonist

We describe here biochemical and pharmacological effects of the β-carboline ZK 93426, a new and potent benzodiazepine (BZ) receptor antagonist. ZK 93426 was compared with Ro 15-1788 and CGS 8216, two compounds previously described as BZ receptor antagonists. Certain effects of ZK 93426, Ro 15-1788 and CGS 8216 were quite similar (e.g., 3H-FNM displacement, “GABA ratio”, “photo-shift”). In most pharmacological tests ZK 93426 and Ro 15-1788 lacked overt effects; Ro 15-1788 was a weak agonist in some paradigms, while ZK 93426 exhibited a potent proconflict effect but also a weak anticonvulsant effect. This interesting finding with ZK 93426 suggests that BZ receptor ligands may possess differential efficacy at BZ receptor subtypes. In contrast, CGS 8216 exhibited potent proconvulsant effects in several paradigms in addition to proconflict and pentylenetetrazol generalizing effects. ZK 93426, Ro 15-1788 and CGS 8216 were almost equally potent as antagonists of the effects of BZ receptor agonists, such as diazepam and lorazepam. However, ZK 93426 was the most potent inhibitor of the convulsions produced by the BZ receptor inverse agonist DMCM.

Does the excitatory amino acid receptor antagonist 2-APH exhibit anxiolytic activity?

The activity of 2-amino-7-phosphonoheptanoic acid (2-APH), an antagonist of the NMDA subtype of glutamate receptor, was tested in several animal models of anxiolytic activity in rats and mice and compared with the activity of the standard benzodiazepine anxiolytic, diazepam. 2-APH was effective, but about 100 times less potent than diazepam in antagonising the suppressive effects of punishment on locomotor activity in the four-plate test in mice. 2-APH was also effective in enhancing exploration of the open, exposed arms of a plus maze, without altering exploration of the enclosed arms. Again 2-APH was about 100 times less effective than diazepam. In contrast to diazepam, 2-APH was ineffective in antagonising the pro-punishment properties of the anxiogenic β-carboline DMCM in a modified four-plate test, and in antagonising the discriminative stimulus provided by pentylenetetrazol. These results are discussed in the context of the equivalence of the antagonism of excitatory mechanisms and the enhancement of inhibitory systems as anxiolytic treatments.

ZK 91296, a partial agonist at benzodiazepine receptors

ZK 91296 (ethyl 5-benzyloxy-4-methoxymethyl-β-carboline-3-carboxylate) is a potent and selective ligand for benzodiazepine (BZ) receptors. Biochemical investigations indicate that ZK 91296 may be a partial agonist at BZ receptors. Such partial agonism may explain to some extent why ZK 91296 needs higher BZ receptor occupancy than diazepam for the same effect against chemical convulsants and for behavioural effects. The lack of sedatiye effects, and the very potent inhibition of reflex epilepsy, spontaneous epilepsy and DMCM-induced seizures suggest, furthermore, that ZK 91296 may possess pharmacological selectivity for a particular type of BZ receptor interaction, perhaps including topographic as well as receptor subtype differentiation.

Bidirectional effects of β-carbolines and benzodiazepines on cognitive processes

Abstract 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 93426, 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.

Treatment strategies for senile dementia: antagonist β-carbolines

Abstract An effective pharmacological treatment for senile dementia is not yet available and even the therapeutic potency of muscarinic agonists has been rather disappointing up to now. Different lines of evidence and a hypothesis are presented below suggesting that β-carbolines with antagonist or partial inverse agonist properties at the GABA-benzodiazepine receptor complex may offer a treatment for senile dementia primarily by disinhibiting the remaining cholinergic neurons of basal forebrain. Preclinical behavioral and biochemical data, as well as studies with human volunteers, support the idea that such β-carbolines exert nootropic effects in general.

β-carbolines can enhance or antagonize the effects of punishment in mice

Six β-carboline ligands at central benzodiazepine (BZ) receptors were tested for their anxiolytic or anxiogenic properties in mice in the four-plate test. ZK 93 423 and ZK 91 296 increased activity which had been suppressed by punishment (1 mA, 60 ms footshock) at doses which exerted no effect on unpunished locomotion. ZK 93 426, ZK 90 886, FG 7142, and DMCM exerted no antipunishment activity themselves, and antagonized the ability of diazepam to increase both punished and unpunished locomotor activity. DMCM, FG 7142, and ZK 90 886, but not ZK 93 426, also enhanced the ability of a reduced level of footshock (0.3 mA) to suppress activity. This propunishment activity of DMCM and ZK 90 886 took place at doses which had no effect on unpunished locomotion. The nature of the effect of the individual β-carbolines on punishment was related to the nature of their interaction with the BZ/GABA receptor/chloride channel complex (GBC complex). Thus the antipunishment properties of ZK 93 423 and ZK 91 296 were associated with their ability to increase binding of 35S-t-butylbicyclophosphorothionate (TBPS) to its binding site associated with the chloride channel, whereas DMCM, FG 7142 and ZK 90 886, which exerted propunishment effects, reduced TBPS binding. ZK 93 426, which was neutral with respect to punished activity, had the weakest effect on TBPS binding. These results are discussed in the context of a possible role of the GBC complex in anxiety.

Modulation of anxiety by β-carbolines and other benzodiazepine receptor ligands: Relationship of pharmacological to biochemical measures of efficacy

Several beta-carbolines and other benzodiazepines (BZ) receptor ligands have been investigated for anxiolytic or anxiogenic action in 4 unrelated animal models of anxiety using rats. The substances could be grouped into essentially 2 groups. The first, anxiolytics, exhibited antipunishment activity in a lick-suppression test, antagonised the discriminative stimulus provided by pentylenetetrazol, resembled chlordiazepoxide (CDP) in a drug discrimination test, and reduced the rise in plasma corticosterone levels following swim stress. Such substances included several benzodiazepines, the beta-carboline ZK 93 423, and the triazolapyridazine CL 218 872. A subgroup of anxiolytics were active in only some of these tests. They included two beta-carbolines, ZK 91 296 and ZK 95 962, and the pyrazoloquinoline CGS 9896, and these 3 substances were also distinguishable in not producing rate-decreasing effects in any of the 3 operant tests. The second group were anxiogenic in that they produced a discriminative stimulus resembling that of PTZ, they antagonised the CDP cue, exhibited propunishment effects in the lick-suppression test, and themselves caused increases in plasma corticosterone in otherwise unstressed animals. Such substances included the beta-carbolines DMCM, FG 7142 and ZK 90 886, and the pyrazoloquinoline CGS 8216. Two substances, Ro 15-1788 and ZK 93 426 had little or only weak activity in any test. The classification of these substances into anxiolytics or anxiogenics could be predicted qualitatively both by their ability to enhance (anxiolytics) or decrease the binding of 35S-TBPS to rat brain membranes and by whether their own binding was increased (anxiolytics) by adding the GABA agonist muscimol to the in vitro incubation medium. For the limited number of substances for which full data was available, there was also a quantitative relationship between the degree of enhancement of 35S-TBPS binding by a substance and its potency in the CDP cue test when such potency was expressed as numbers of BZ receptors occupied at the ED50 value in the pharmacological test. Furthermore, for the anxiolytics, activity in the CDP cue correlated significantly with potency in 2 other tests. Otherwise, surprisingly weak correlations existed between potencies in the different tests. In particular, the beta-carboline ZK 95 962 was highly potent in antagonising the PTZ cue but inactive in both a conflict test and in protecting against stress. These results are discussed in terms of differences in the neuropharmacologies of the 4 tests and in selectivity of the BZ receptor ligands for subtypes of BZ receptor.

Human studies on the benzodiazepine receptor antagonist β-carboline ZK 93 426: preliminary observations on psychotropic activity

The β-carboline ZK 93 426, a benzodiazepine receptor antagonist, was administered intravenously to human volunteers at two different doses (0.01 mg/kg, 0.04 mg/kg) according to a double-blind, placebo controlled design. Vital functions (i.e. blood pressure, heart rate, ECG, EEG), peripheral (finger) skin temperature and performance in psychometric tests for psychotropic and cognitive effects were evaluated. Blood samples were collected in addition and certain pharmacokinetic parameters were estimated. ZK 93 426 in both doses was well tolerated and exhibited no side effects. A decrease in peripheral skin temperature and heart rate was observed. In a general estimation of behavioural changes, volunteers experienced a stimulant and activating effect of the drug. An improvement in performance was observed in two cognitive tasks, the “logical reasoning task” and “pictures differences task” which estimated concentration and attention, respectively. No effects were found in time estimation. Plasma levels 5 min after intravenous administration of ZK 93 426 were 16±10 ng/ml and 52±31 ng/ml for 0.01 mg/kg and 0.04 mg/kg, respectively. Total clearance was calculated as 46±22 ml/min/kg (0.04 mg/kg).

Contrasting effects of the competitive NMDA antagonist CPP and the non-competitive NMDA antagonist MK 801 on performance of an operant delayed matching to position task in rats

The effects of the competitive NMDA antagonist CPP and the non-competitive NMDA antagonist MK 801 (dizolcipine) on short term working memory in the rat were investigated. The behavioural paradigm used was discrete trial, operant delayed matching to position, as originally described by Dunnett (1985), with delays of 0, 5, 15 and 30 s. These delays generated an orderly “forgetting” curve in control rats, with matching accuracy decreasing from approximately 100% at 0-s delay to approximately 75% at 30-s delay. Intraperitoneal (IP) administration of CPP (10 mg/kg) produced a markeddelay dependent impairment in performance, suggesting a specific effect on short term working memory. This effect was accompanied by a minor decrease in the speed of responding, and a slight increase in the number of missed trials. Lower doses of CPP had no significant effects on either matching accuracy or sedation. In contrast, IP administration of MK 801 (0.1 and 0.2 mg/kg) caused a markeddelay independent impairment in the accuracy of delayed matching performance, suggesting a non-specific disruption of performance. A lower dose (0.05 mg/kg) of MK 801 had no significant effect on matching accuracy. The two lower doses of MK 801 increased the number of nose pokes made during the delays and tended to increase the speed of responding, suggesting a stimulant-like action. The highest dose of MK 801 had the opposite effects and also decreased the number of trials completed. The results with CPP therefore support the hypothesized role of NMDA receptors in learning and memory, and the contrasting effects of these two NMDA antagonists support previous suggestions of different behavioural effects resulting from administration of competitive and non-competitive NMDA antagonists.