Lucy J. Wilks

Behavioral effects of acute and chronic administration of caffeine in the rat.

This study investigated the effects of acute and chronic caffeine treatment on behavior in the social interaction, holeboard and home-cage aggression tests and on proconvulsant actions with pentylenetetrazol. Acutely-treated rats received an IP injection of caffeine (20 or 40 mg/kg). Chronically-treated rats received caffeine in their drinking water for 21 days (50 or 100 mg/kg/day) followed by an injection of caffeine on the test day (20 or 40 mg/kg respectively). Acutely, the higher dose of caffeine (40 mg/kg) decreased levels of social interaction. In the holeboard test, 20 mg/kg of acute caffeine increased motor activity whilst 40 mg/kg reduced head-dipping behavior. In the home-cage aggression test, acute caffeine (40 mg/kg) reduced offensive aggressive behaviors. After chronic treatment with caffeine none of these behaviors differed significantly from controls. After both acute and chronic treatment, caffeine (20 and 40 mg/kg) was proconvulsant with pentylenetetrazol.

The sedative effects of CL 218,872, like those of chlordiazepoxide, are reversed by benzodiazepine antagonists

The effects of CL 218,872, initially classified as a non-sedative anxiolytic, were investigated and compared with those of chlordiazepoxide in the holeboard. The ability of two drugs that antagonise the effects of benzodiazepines, CGS 8216 and Ro 15-1788, to reverse the effects of CL 218,872 and chlordiazepoxide were also investigated, to see whether their effects might be mediated via benzodiazepine receptors. CL 218,872 (10 mg/kg) was found to be significantly sedative in both mice and rats (i.e., both locomotor activity and head-dipping were significantly decreased). In mice, the effects of CL 218,872 and of chlordiazepoxide were very similar over a range of doses, except that the stimulatory effect seen with low doses of chlordiazepoxide on head-dipping just failed to reach significance with CL 218,872. This study is in agreement with recently published results from different tests showing that sedative effects can be obtained with doses of CL 218,872 that are low and not much higher than those leading to anxiolysis. The sedative effects of both CL 218,872 (10 mg/kg) and chlordiazepoxide (20 mg/kg) were significantly reversed by RO 15-1788 (10 and 20 mg/kg) and CGS 8216 (10 mg/kg), suggesting that their effects are mediated via benzodiazepine receptors. The increase in head-dipping seen with chlordiazepoxide (2.5 mg/kg) was also reversed by RO 15-1788 and CGS 8216.

Kindling with the β-Carboline FG7142 Suggests Separation between Changes in Seizure Threshold and Anxiety-Related Behaviour

The aim of this paper was to determine whether the prolonged decrease in seizure threshold produced by chemical kindling was accompanied by behavioural changes in tests of anxiety and aggression. The responses of rats in five tests were examined after chronic treatment with the benzodiazepine inverse agonist FG7142. This treatment caused chemical kindling, so that the originally proconvulsant drug caused full seizures. This effect is very long-lasting, and our previous work with mice had suggested that it might be accompanied by an increase in anxiety-related behavior. In the present work no significant differences were found between the behaviour of FG7142-kindled rats and vehicle-treated controls in social interaction test, elevated plus maze, or the Vogel conflict test of anxiety or in tests of home cage aggression or startle responses. The results therefore show that prolonged changes in seizure threshold can occur without alterations in the apparent level of anxiety.

Evidence of strain differences in GABA-benzodiazepine coupling

The effects of a single dose of oxazepam on seizure threshold, receptor occupancy and brain oxazepam concentration were investigated at several time points after drug treatment in two inbred strains of mice (NIH and C3H/HE). The C3H/HE strain showed a greater sensitivity to the effects of both pentylenetetrazol and oxazepam. Furthermore, the C3H/HE strain showed decreases in both receptor occupancy and seizure threshold across time, whereas the NIH strain showed no change in either measure. Although within-strain correlations were observed between seizure threshold and receptor occupancy, the C3H/HE strain had similar seizure thresholds to the NIH strain throughout but lower percentage receptor occupancies, thus a between-strain correlation was not observed. The C3H/HE strain had a higher number of specific benzodiazepine binding sites and these results may reflect a strain difference in GABA-benzodiazepine receptor coupling. In a further experiment, the development of acute tolerance to the effects of oxazepam was investigated. Brain concentrations of oxazepam and receptor occupancies were determined for each strain of mouse, at two different time points (1.5 and 7.5 h after drug treatment), at which equivalent seizure thresholds were obtained by manipulating the starting dose of oxazepam. For each strain, when equivalent seizure thresholds were observed at different time points, equivalent receptor occupancies were also observed. However, a trend towards higher brain concentrations of oxazepam at the later time point was detected for the two strains, suggesting that there may be some decrease across time in the affinity of the receptor for oxazepam.

Evidence for Simultaneous Anxiolytic and Aversive Effects Several Hours after Administration of Sodium Phenobarbitone to the Rat

Using a place conditioning paradigm, it was shown that the state experienced by rats 8 h after a 50-mg/kg dose of sodium phenobarbitone had aversive properties. However, at this same time, rats showed an increase in the percentage of time spent on the open arms of an elevated plus-maze, and an increase in the number of punished licks in a conflict test. These two effects are indicative of an anxiolytic action. It was therefore possible to demonstrate, simultaneously, an anxiolytic effect of sodium phenobarbitone and that it induced an aversive state. Conditioned place aversion could also be demonstrated 1 h after a 20-mg/kg dose of sodium phenobarbitone, but this dose was ineffective in the elevated plus-maze test of anxiety. These results provide further evidence that drugs that reduce anxiety do not necessarily have secondary positive reinforcing properties.

Does the behavioural activation detected after a single dose of a benzodiazepine reflect a withdrawal response

The effects, in mice, of a single dose of lorazepam or oxazepam were determined, in the holeboard, 24, 48 and 72 hours after treatment. Lorazepam produced significant increases in both spontaneous locomotor activity and in rearing 48 hours after treatment and oxazepam produced a significant overall increase in rearing over the three time points. There was no detectable in vivo receptor occupancy for either drug at the 48 hour time point, so that these effects were not due to residual concentrations of drug in the brain. We therefore suggest that we were detecting a spontaneous withdrawal response to a single dose of benzodiazepine. The increases in both locomotor activity and rearing, detected 48 hours after lorazepam, could be reversed by treating simultaneously with Ro 15-1788 (a benzodiazepine antagonist). When Ro 15-1788 was injected 20 minutes prior to testing, the mice that had been treated 48 hours previously with lorazepam still showed increased locomotor activity and rearing. We conclude that the hyperactivity was not caused by any change in the levels of endogenous substances acting at the benzodiazepine receptor.

The effects of benzodiazepines in newborn rats suggest a function for type 2 receptors

In day 4 female rats lorazepam, diazepam and clonazepam produced dose-related increases in forward walking and loss of righting and diazepam produced a dose-related increase in paddling. Lorazepam and diazepam increased jerks of the fore- and hind-limbs and the whole body, and clonazepam increased the latter two; these increases were not dose-related. Some doses of lorazepam and the lowest dose of diazepam increased tonic-clonic movements. Thus the benzodiazepines were observed to have two kinds of stimulant effect in day 4 rats. One is to cause hyperactivity and this effect is dose-related. The other is to cause a type of seizure-like behavior, although this action is not dose-related and the responses can be distinguished from those caused by convulsant compounds. The effects of the benzodiazepine antagonist Ro 15-1788 resembled those of the benzodiazepines. It increased hind-limb and whole body jerks, forward walking, paddling, loss of righting and tonic-clonic movements. CL 218,872, which is selective for type 1 benzodiazepine receptors, was devoid of significant effects. This suggests that the behavioral changes observed with the other compounds were mediated by the type 2 receptors.

Effects of acute and chronic treatment on the pro‐and anti‐convulsant actions of CL 218, 872, PK 8165 and PK 9084, putative ligands for the benzodiazepine receptor

CL 218, 872 is a triazolopyridazine that acts at the benzodiazepine binding site. At low doses (0.5−7.5 mg kg−1) it is proconvulsant when combined with subconvulsant doses of picrotoxin but not when combined with pentetrazol (leptazol, pentylenetetrazol). At high doses (20−60 mg kg−1) CL 218, 872 counteracted seizures caused by pentetrazol but not those caused by picrotoxin. There was tolerance to the proconvulsant effects after five days of treatment and to the anticonvulsant effects after 15–20 days. Two phenylquinolines, PK 8165 and PK 9084, that also act at the GABA‐benzodiazepine receptor complex have proconvulsant actions in combination with picrotoxin. Significant tolerance to these effects had not developed even after 20 days of treatment. It is concluded that three different sites on the GABA‐benzodiazepine complex mediate the pro‐ and anti‐convulsant actions of CL 218,872 and the proconvulsant actions of PK 8165 and PK 9084.

The role of the benzodiazepine receptor in mediating long-lasting anticonvulsant effects and the late-appearing reductions in motor activity and exploration.

The number of head-dips, the time spent head-dipping, the number of rears and the locomotor activity of mice placed in a holeboard was reduced by lorazepam (0.25 mg/kg) 1 and 1.5 h after oral administration and these reductions were reversed by the benzodiazepine antagonist flumazenil (1 mg/kg). Activity returned to control levels at 3 and 4.5 h for head-dipping, and between 3 and 12 h for rearing and locomotor activity. However, significant late-appearing reductions were found for the number of head-dips and the time spent head-dipping (6 h) and rearing and locomotor activity (15 h) and these decreases could not be reversed by flumazenil. Similar results were found after oral administration of oxazepam (7 mg/kg). Oxazepam reduced the number of head-dips and time spent head-dipping at 1, 1.5 and 3 h and these reductions were reversed by flumazenil (1 mg/kg). Head-dipping activity returned to normal at 4.5 h. Significant reductions were also found for both measures at 1, 6 and 7.5 h and these late reductions could not be reversed by flumazenil. This suggests that the late-appearing reductions in holeboard behaviours, resulting from lorazepam or oxazepam administration to mice, is not mediated by the benzodiazepine receptor. This conclusion was supported by the results from in vivo binding, which showed no change in the % receptor occupancy 3–15 h after administration of lorazepam or oxazepam. In contrast to the holeboard behaviours, the anticonvulsant effects of the two drugs showed good correlations with receptor occupancy. The anticonvulsant effect of oxazepam (7 mg/kg) significantly decreased 1–3 h after oral administration, but thereafter a steady anticonvulsant effect was retained for up to 24 h. The anticonvulsant effect of lorazepam (0.25 mg/kg) also significantly decreased 1–3 h after administration, and thereafter remained steady for up to 15 h. At all the time-points tested, oxazepam (7 mg/kg) had a significantly greater anticonvulsant effect than lorazepam (0.25 mg/kg). This was mirrored by higher percentage receptor occupancies in cortex, hippocampus and cerebellum, from 3 to 15 h after administration.

Anticonvulsant effects of chronic treatment with phenytoin against pentylenetetrazole-induced seizures in the mouse

Acute administration of phenytoin (40 and 75 mg/kg) was unable to counteract seizures induced by pentylenetetrazole and even prolonged them in some cases. These prolonged seizures remained with chronic (10 days) treatment but an anticonvulsant effect of phenytoin (40 and 75 mg/kg) also emerged, shown by a decrease in the incidence of seizures. This latter effect could be detected even 24 hr after the last dose. Both acute and chronic treatment with phenytoin (40 or 75 mg/kg) were able to enhance anticonvulsant efficacy of diazepam against pentylenetetrazole-induced seizures. The mechanisms underlying these three independent effects of phenytoin and their clinical relevance are discussed.