Evidence that the increased anxiety detected in the elevated plus-maze during chlordiazepoxide withdrawal is not due to enhanced noradrenergic activity

Rats displayed a reduction in the percentage of time spent on the open arms of the elevated plus-maze 24-30 hours after withdrawal from chronic chlordiazepoxide treatment (10 mg/kg/day IP for 4 weeks). This indicated an anxiogenic response in this test. This anxiogenic response was not significantly reversed by DL-propranolol (5 and 10 mg/kg IP) or clonidine (0.02 and 0.04 mg/kg IP). These results provide no evidence to suggest that the anxiogenic effects of chlordiazepoxide withdrawal are mediated by an increase in noradrenergic activity. The possible involvement of multiple transmitter systems in benzodiazepine withdrawal symptomology is discussed.     

The effects of sertraline and fluoxetine on anxiety in the elevated plus-maze test in mice

The aim of this study was to evaluate the acute and chronic effects of two SSRIs (sertraline and fluoxetine) on anxiety by the elevated plus-maze test. Diazepam increased the time spent in open arms significantly whereas the anxiogenic m-chlorophenylpiperazine (m-CPP) decreased the time significantly. Acute sertraline (10 mg x kg(-1)) and fluoxetine (20 mg x kg(-1)) treatment significantly decreased the time spent in open arms. Acute fluoxetine (20 mg x kg(-1)) treatment also decreased the total number of enclosed arm entries. Seven days sertraline treatment decreased the time spent in open arms, whereas 14 days fluoxetine treatment increased the time spent in open arms. These results show that acute administration of SSRIs may produce anxiogenic effects in the elevated plus-maze test.     

Effect of valepotriates on the behavior of rats in the elevated plus-maze during diazepam withdrawal

The effect of a mixture of valepotriates on the elevated plus-maze performance of diazepam withdrawn rats was evaluated. The rats were chronically (28 days) treated with diazepam (doses increased up to 5.0 mg/kg) and then treated with control solution for 3 days to induce a withdrawal syndrome. Chronically vehicle-treated rats were used as control.The abstinent animals treated with vehicle showed a significant decrease in the percentage of time spent in the open arms when compared with the control animals. Diazepam and valerian 12.0 mg/kg reversed this anxiogenic effect. Valerian 6.0 mg/kg did not show any difference in relation to the others group.     

The effects of histaminergic agents in the central amygdala of rats in the elevated plus-maze test of anxiety

Reports indicate that histamine and histaminergic agents can change anxiety-related behaviours in both animals and humans. The amygdala is an important brain site in the modulation of fear or anxiety. In the present study, we investigated the effects of intracentral amygdala microinjection of histaminergic agents on anxiety-related behaviours in rats, using the elevated plus-maze test of anxiety. Intracentral amygdala administration of histamine (0.01-0.5 microg/0.5 microl bilateral) decreased %open armtime and % open arm entries, but not locomotor activity, showing an anxiogenic response. Intracentral amygdala microinjection of pyrilamine (H1 receptor antagonist) and ranitidine (H2 receptor antagonist) (both at 1-20 microg/0.5 microl bilateral) did not change anxiety-related parameters in our experiments. In another series of experiments, histamine (0.5 microg/0.5 microl bilateral) was coadministrated with pyrilamine and ranitidine (both at 1-20 mg/0.5 microl bilateral). The results showed that pyrilamine but not ranitidine could significantly reverse the anxiogenic effect of histamine at doses of 10 and 20 microg/0.5 microl bilateral. The results suggest that histamine may modulate anxiety via H1 but not H2 receptors in the rat central amygdala.     

The effects of histaminergic agents in the dorsal hippocampus of rats in the elevated plus-maze test of anxiety

High levels of histamine are found in the hippocampus. The central histamine system is involved in many physiological behavioural processes including anxiety-related behaviours both in animals and humans. In the present study, we investigated the effects of intra-hippocampal CA1 (intra-CA1) microinjection of histaminergic agents on anxiety-related behaviours in rats, using elevated plus-maze test of anxiety. Intra-CA1 administration of histamine (at the dose of 10 microg/rat) increased open arm time (%OAT) and open arm entry (%OAE) but not locomotor activity, thus showing an anxiolytic response. Intra-CA1 microinjection of pyrilamine (H1 receptor antagonist; at the doses of 10, 20 and 40 microg/rat) in combination with histamine (10 microg/rat) showed a decrease in the %OAT and %OAE. Higher dose of the antagonist (40 microg/rat) by itself increased both %OAT and %OAE, but not locomotor activity, indicating an anxiolytic effect. Intra-CA1 microinjection of ranitidine (H2 receptor antagonist), at the doses of 10, 20 and 40 microg/rat, also reduced the histamine response. Furthermore, the H2 receptor antagonist by itself reduced %OAT and %OAE without affecting locomotor activity. The results may indicate an anxiogenic effect for the antagonist. Our results showed that histamine may modulate anxiety via H1 and H2 receptors in the CA1 region of hippocampus of the rat.     

Social isolation modifies nicotine's effects in animal tests of anxiety

1. These experiments determined whether the housing conditions of rats influenced the effects of nicotine in two animal tests of anxiety, social interaction and elevated plus-maze tests. 2. In animals housed singly for 7 days, (-)nicotine (0.025 mg kg(-1) s.c.) was ineffective, but 0.05, 0.1 and 0.25 mg kg(-1) (s.c.) significantly increased the time spent in social interaction, without changing locomotor activity, thus indicating anxiolytic actions. (-)Nicotine (0.45 mg kg(-1) s.c.) significantly reduced social interaction, indicating an anxiogenic effect. 3. However, in group-housed animals, (-)nicotine (0.025 mg kg(-1) s.c.) had a significant anxiolytic effect in the social interaction test, but 0.01, 0.05, 0.1, 0.25 and 0.45 mg kg(-1) were ineffective. (-)Nicotine (1 mg kg(-1)) reduced motor activity and social interaction in the group-housed animals. 4. In the elevated plus-maze, the time-course and the dose-response curve to nicotine were investigated. In both singly- and group-housed rats, (-) nicotine (0.1 - 0.45 mg kg(-1) s.c.) decreased the per cent entries into, and per cent time spent on, the open arms, indicating anxiogenic effects. 5. The housing condition influenced the time course, with significant effects at 5 and 30 min after injection in group-housed rats, and significant effects at 30 and 60 min in singly-housed rats. 6. In the social interaction test there was no difference in the scores of the first and last rats removed from group cages, whereas the order of removal from the cages did affect the scores in the elevated plus-maze. 7. These results provide further evidence that the two animal tests model distinct states of anxiety, and show how social isolation powerfully modifies both anxiolytic and anxiogenic effects of nicotine.     

Flumazenil but not nitrendipine reverses the increased anxiety during ethanol withdrawal in the rat

After 7 day's gradual introduction of ethanol, rats were maintained for a further 4 weeks on a liquid diet containing 10% ethanol (mean daily dose 11.8±0.2 g/kg/day). Control-treated rats received liquid diet alone. Pairs of rats were tested in the social interaction test of anxiety 8 h after withdrawal. Withdrawal from ethanol significantly reduced the time spent in social interaction compared with controls, indicating an anxiogenic withdrawal response. Nitrendipine (50 mg/kg) had no effect on, whereas flumazenil (4 mg/kg) significantly reversed, this withdrawal response. This reversal appeared to be long-lasting as there was still no evidence of increased anxiety when rats were again withdrawn after 3 more days of ethanol diet.     

Vigabatrin has an anxiolytic effect in the elevated plus-maze test of anxiety.

tau-Vinyl GABA (vigabatrin, GVG) is a novel antiepileptic drug that irreversibly inhibits GABA transaminase and elevates GABA levels in all parts of the brain. In the present study, we investigated the anxiolytic and behavioral effects of GVG in the elevated plus-maze and the hole board compared to diazepam. Doses of 500 and 1,000 mg/kg GVG were injected IP to different groups of male Wistar rats and animals were tested either 4 or 24 h after injection. Animals administered diazepam (1.5 mg/kg, IP) and saline (1 ml) were tested 20 min after injection. GVG and diazepam were found to decrease significantly the number of squares visited and rearing; both had a suppressant effect on locomotor activity. Neither drug had an effect on exploration (head dipping). GVG at a dose of 1,000 mg/kg was shown to have a similar anxiolytic activity either after 4 or 24 h as diazepam, while GVG at 500 mg/kg did not show any significant anxiolytic effect.     

Effects of scopolamine and its quaternary analogue in the murine elevated plus-maze test of anxiety

Several lines of evidence suggest that muscarinic cholinergic receptors may play a role in fear/anxiety reactions in animals. In the present study, the behavioural effects of the muscarinic receptor antagonist scopolamine (0.125-1.0mg/kg) were examined in male mice exposed to the elevated plus-maze. In contrast to scopolamine methyl bromide, which was behaviourally inert under present test conditions, scopolamine hydrobromide produced behavioural changes indicative of enhanced anxiety. The effects included a reduction in percentage of open arm entries and a marked stimulation of risk assessment measures (i.e. stretched attend postures and closed arm returns), as well as more subjective signs of enhanced fear/anxiety such as vocalisation, struggling and escape-oriented behaviour. Although scopolamine also enhanced total arm entries, perhaps suggesting a general stimulant action, this effect was specific to the closed arms and was not accompanied by systematic increases in either rearing or head-dipping. Data are discussed in relation to the possible involvement of central muscarinic substrates in risk assessment behaviour in animals and hypervigilance states in humans.     

A robust animal model of state anxiety: fear-potentiated behaviour in the elevated plus-maze

Fear (i.e., decreased percentage time spent on open-arm exploration) in the elevated plus-maze can be potentiated by prior inescapable stressor exposure, but not by escapable stress. The use of fear-potentiated plus-maze behaviour has several advantages as compared to more traditional animal models of anxiety. (a) In contrast to the traditional (spontaneous) elevated plus-maze, which measures innate fear of open spaces, fear-potentiated plus-maze behaviour reflects an enhanced anxiety state (allostatic state). This "state anxiety" can be defined as an unpleasant emotional arousal in face of threatening demands or dangers. A cognitive appraisal of threat is a prerequisite for the experience of this type of emotion. (b) Depending on the stressor used (e.g., fear of shock, predator odour, swim stress, restraint, social defeat, predator stress (cat)), this enhanced anxiety state can last from 90 min to 3 weeks. Stress effects are more severe when rats are isolated in comparison to group housing. (c) Drugs can be administered in the absence of the original stressor and after stressor exposure. As a consequence, retrieval mechanisms are not affected by drug treatment. (d) Fear-potentiated plus-maze behaviour is sensitive to proven/putative anxiolytics and anxiogenics which act via mechanisms related to the benzodiazepine-gamma-aminobutyric acid receptor, but it is also sensitive to corticotropin-releasing receptor antagonists and glucocorticoid receptor antagonists and serotonin receptor agonists/antagonists complex (high predictive validity). (e) Fear-potentiated plus-maze behaviour is very robust, and experiments can easily be replicated in other labs. (f) Fear-potentiated plus-maze behaviour can be measured both in males and females. (g) Neural mechanisms involved in contextual fear conditioning, fear potentiation and state anxiety can be studied.Thus, fear-potentiated plus-maze behaviour may be a valuable measure in the understanding of neural mechanisms involved in the development of anxiety disorders and in the search for novel anxiolytics. Finally, the involvement of corticotropin-releasing factor and corticosteroid-corticotropin-releasing factor interactions in the production of fear-potentiated plus-maze behaviour are discussed.     

Prior maze experience required to alter midazolam effects in rats submitted to the elevated plus-maze

In rodents, prior maze experience increases open arm avoidance (OAA) and compromises the anxiolytic effects of benzodiazepines in a subsequent exposure to the elevated plus-maze (EPM), a phenomenon referred to as "one trial tolerance" (OTT). Nevertheless, a possible correlation between these intriguing events remains unclear. Using maze-naive and maze-experienced (free exploration of the EPM for 5 min) rats, Experiment 1 confirmed the anxiolytic effects of midazolam (MDZ; 0.125-1.0 mg/kg) in maze-naive rats, while both increased OAA and OTT to the MDZ anxiolytic effects were observed in maze-experienced rats. However, our results from Experiment 2, designed to assess whether open, enclosed or both arms experience is involved in increased OAA and OTT, showed that MDZ retained its efficacy in rats confined either to an open or enclosed arm, where no significant changes in open arm exploration were observed when compared to the maze-naive group, therefore suggesting that prior experience in the whole apparatus may be involved in the loss of the anxiolytic MDZ effects. Results are discussed in terms of a possible correlation between increased OAA and the OTT phenomenon elicited in a subsequent exposure to the EPM.     

Histaminergic H1 receptors mediate L-histidine-induced anxiety in elevated plus-maze test in mice

The central histaminergic system is reported to mediate behavioural, hormonal and physiological homeostasis of living organisms. Recent reports indicate its prominent role in various neurobehavioural disorders such as depression and psychosis. This study evaluated the effect of activation of the central histaminergic system in anxiety-like conditions, using the elevated plus-maze test in mice, and elucidated the role of different histaminergic receptors mediating such effects. Peripheral administration of L-histidine (L-His), in a dose-dependent manner, significantly decreased the exploration time in open arms and number of entries into open arms without modifying the number of entries into closed arms of the elevated plus-maze, indicating anxiogenesis. Further, such effects of central histamine were significantly attenuated, in a dose-dependent manner, by pretreatment with pyrilamine (H1 receptor antagonist). Pretreatment with either zolantidine (H2 receptor antagonist) or thioperamide (H3 receptor antagonist), however, failed to attenuate the L-His-induced anxiogenesis. Our results indicate that anxiogenic effects of central histaminergic system appear to be mediated prominently by activation of H1 receptors.     

Anxiolytic and anxiogenic drug effects on exploratory activity in an elevated plus-maze: a novel test of anxiety in the rat

The current studies further investigated the effects, in animal models of anxiety, of novel putative anxiolytic and anxiogenic compounds believed to induce their effects by actions at the GABA-benzodiazepine receptor complex. It was expected that the results would also provide further validation for a novel test of anxiety based on the ratio of open to closed arm entries in an elevated plus maze in the rat. The novel putative anxiolytics CL 218,872 (10-20 mg/kg) and tracazolate (5 mg/kg) significantly elevated the percentage of time spent on the open arms of an elevated plus-maze, consistent with their anxiolytic activity in several other animal tests. Also consistent with results from other animal tests, no anxiolytic activity was observed for the phenylquinoline PK 8165 (10-25 mg/kg), the 3,4-benzodiazepine tofisopam (25-50 mg/kg), or buspirone (0.5-20 mg/kg). The benzodiazepine receptor inverse agonists FG 7142 (1-5 mg/kg) and CGS 8216 (3-10 mg/kg) had anxiogenic activity in this test, as did the atypical benzodiazepine Ro 5-4864 (1-5 mg/kg). Interestingly, however, the benzodiazepine receptor antagonists Ro 15-1788 (10-20 mg/kg) and ZK 93426 (5-10 mg/kg) had no anxiogenic activity in this test.     

Anxiolytic-like effects of ginsenosides on the elevated plus-maze model in mice

In a previous study, we reported that ginseng extract has anxiolytic-like effects in the elevated plus-maze model and that the ginseng saponin fraction plays an important role. This experiment was performed to investigate the anxiolytic-like effects of ginsenosides Rb1, Rg1, Rg3-R, and Rg3-S, and the Rg5 and Rk mixture isolated from the ginseng saponin fraction in the elevated plus-maze. Furthermore, the anxiolytic-effects of Rb1, Rg1, Rg3-R, Rg3-S, and the Rg5 and Rk mixture were compared with those of a well-known active anxiolytic drug (diazepam). The oral administration of ginsenoside Rb1 significantly increased the number of open arm entries and the time spent on the open arm compared with those in the vehicle-treated group. Ginsenoside Rg1 and the Rg5 and Rk mixture also significantly increased the number of open arm entries and the time spent on the open arm. However, ginsenosides Rg3-R and Rg3-S did not increase the number of open arm entries or the time spent on the open arm. On the other hand, ginsenoside Rb1 and the Rg5 and Rk mixture decreased locomotor activity in a manner similar to diazepam. These data indicate that ginsenosides Rb1, Rg1, and the Rg5 and Rk mixture have anxiolytic-like effects, but ginsenosides Rg3-R and Rg3-S do not in this model. We provide evidence that some ginsenosides may be useful for the treatment of anxiety.     

Lack of effects of 5HT3 receptor antagonists in the social interaction and elevated plus-maze tests of anxiety in the rat

The effects of three 5HT₃ receptor antagonists: BRL 43964 (0.1 and 1 mg/kg, oral), GR 38032F (0.1 and 1 mg/kg, oral), and zacopride (0.01, 0.1 and 1 mg/kg, IP) were examined in low light test conditions of the social interaction test. None of the three 5HT₃ receptor antagonists had a significant effect on social interaction. In contrast, in two experiments chlordiazepoxide (7.5 mg/kg) significantly increased social interaction and this effect was greatest in the unfamiliar test condition. In a third experiment, the effects of GR 38032F (0.1 and 1 mg/kg, oral) and zacopride (0.01, 0.1 and 1 mg/kg, oral) were investigated in the high light test conditions of the social interaction test; neither compound had a significant effect. In the elevated plus-maze, chlordiazepoxide (7.5 mg/kg oral or IP) significantly increased both the per cent number of entries made onto open arms and the per cent of time spent on the open arms, indicating an anxiolytic action. Zacopride (0.01, 0.1 and 1 mg/kg, oral or IP) had no significant effect in this test. The effect of the baseline rate of responding in the social interaction test on the effects of 5-HT₃ antagonists is discussed. The results from the present experiment and those from other animal tests of anxiety caution against the conclusion that 5HT₃ receptor antagonists are anxiolytic.     

The effects of corticotrophin releasing factor (CRF) and handling stress on behavior in the elevated plus-maze test of anxiety

The effects on rodent anxiety of corticotrophin releasing factor (CRF), common experimental stressors and the CRF receptor blocker, α-helical CRF, were measured using the hole board and elevated plus-maze tests. Centrally administered (intracerebroventricular, i.c.v.) CRF increased anxiety in an anxioselective manner. α-Helical CRF (i.c.v) antagonized the effects of CRF, implicating central CRF receptors. Common experimental stressors, such as surgical implantation of cannulas and intraperitoneal injections of saline also selectively increased anxiety in the plus maze. Endogenous CRF binding to central CRF receptors probably mediates the anxiogenic effects of stressors, since α-helical CRF reversed the increased anxiety following surgery. Finally, repeated gentle handling seemed to blunt the anxiogenic effect of CRF. Handling also altered the effect of CRF on behavior, creating an apparently CRF-mediated suppression of rearing and exploration which was not present in rats not stressed with repeated handling. Together the data suggest a modifiable modulatory role of CRF in rodent anxiety. The findings also suggest that careful attention should be paid to stress history when examining the role of CRF in rodent behavior.     

Prior exposure to the elevated plus-maze sensitizes mice to the acute behavioral effects of fluoxetine and phenelzine

A single undrugged experience of the elevated plus-maze modifies future drug responses in the test. The present study investigated the effects of maze-experience on the acute behavioral effects of the monoamine oxidase inhibitor phenelzine and the serotonin reuptake inhibitor fluoxetine. Phenelzine (2.5-12.5 mg/kg) had no clear effect on plus-maze behavior in test-naive Swiss Webster mice, but dose-dependently increased anxiety-like behavior in maze-experienced subjects. Similarly, fluoxetine (5-20 mg/kg) produced non-significant trends for increased anxiety-like behavior in maze-naive mice, but significantly and dose-dependently increased anxiety-like behavior and suppressed locomotor activity in maze-experienced mice. The anxiogenic effects of the benzodiazepine receptor inverse agonist N-methyl-beta-carboline-3-carboxamide (FG 7142) (20 mg/kg) was abolished by prior test experience, suggesting an alteration in gamma-aminobutyric acid (GABA)/benzodiazepine receptor function with maze-experience. However, the benzodiazepine receptor antagonist flumazenil (5-20 mg/kg) produced a silent profile regardless of maze-experience. Present findings provide further evidence demonstrating that prior test history is a critical consideration in mouse studies of anxiety-related behavior.     

On the effects of lindane on the plus-maze model of anxiety

The behavioral effects of an acute subconvulsant dose of the insecticide and ectoparasiticide γ-hexachlorocyclohexane, lindane, were compared in the plus-maze (+-maze) animal model of anxiety to those elicited by the anxiogenic GABA_A-antagonist pentylenetetrazole (PTZ) and the anxiolytic benzodiazepine diazepam. The effects of the coadministration of diazepam and lindane, and of diazepam and PTZ were also studied. The results from the analysis of the data with the multivariate statistical technique principal component analysis indicated that the effects of subconvulsant doses of lindane and PTZ on the behavior of rats in the +-maze are similar, and opposite to those elicited by diazepam, and that the administration of the convulsants would antagonize the effects of the benzodiazepine. The putative anxiogenic properties of the insecticide and the utility of the +-maze model in neurotoxicological research are discussed.     

Galanin GAL-R1 receptor null mutant mice display increased anxiety-like behavior specific to the elevated plus-maze.

The neuropeptide galanin coexists with norepinephrine and serotonin in neural systems mediating emotion. Previous findings suggested that galanin modulates anxiety-related behaviors in rodents. Three galanin receptor subtypes have been cloned; however, understanding their functions has been limited by the lack of galanin receptor subtype-selective ligands. To study the role of the galanin GAL-R1 receptor subtype in mediating anxiety-related behavior, we generated mice with a null mutation in the Galr1 gene. GAL-R1 -/- are viable and show no abnormalities in health, neurological reflexes, motoric functions, or sensory abilities. On a battery of tests for anxiety-like behavior, GAL-R1 -/- showed increased anxiety-like behavior on the elevated plus-maze test. Anxiety-related behaviors on the light/dark exploration, emergence, and open field tests were normal in GAL-R1 -/-. This test-specific anxiety-like phenotype was confirmed in a second, independent cohort of GAL-R1 null mutant mice and +/+ controls. Principal components factor analysis of behavioral scores from 279 mice suggested that anxiety-like behavior on the elevated plus-maze was qualitatively distinct from behavior on other tests in the battery. In addition, exposure to the elevated plus-maze produced a significantly greater neuroendocrine response than exposure to the light/dark exploration test, as analyzed in normal C57BL/6J mice. These behavioral findings in the first galanin receptor null mutant mouse are consistent with the hypothesis that galanin exerts anxiolytic actions via the GAL-R1 receptor under conditions of relatively high stress.