Flumazenil antagonizes the effect of diazepam on negative contrast in one-way avoidance learning

The main aim of the present work was to study whether the effect of diazepam upon successive negative contrast in one-way avoidance learning-induced by shifting rats from a large reward (30s spent in the safe compartment) to a small reward (1s)-is mediated by the action of this drug on the benzodiazepine (BZ) receptor. Therefore, we studied the influence of flumazenil (FL), a BZ antagonist, on the effect of diazepam (DZ) on negative contrast. The i.p. administration of 5 and 12mg/kg, but not of 2mg/kg of FL, reliably antagonized the abolition by DZ (1mg/kg) on successive negative contrast. Moreover, FL (12mg/kg) did not affect either the avoidance response or the contrast effect. These results suggest that the GABA system is involved in the successive negative contrast effect in one-way avoidance learning, and that this experimental procedure may be useful in studies of anti-anxiety agents.     

Differential effect of buspirone and diazepam on negative contrast in one-way avoidance learning

The main aim of the present work was to investigate the effect of buspirone, a 5-HT_1A receptor agonist, on successive negative contrast in one-way avoidance learning. Successive negative contrast was induced by shifting rats from a large reward (30 s spent in the safe compartment) to a small reward (1 s). Acute administration of buspirone (0.25, 0.5, 0.75 and 1.0 mg/kg i.p.) did not attenuate the contrast effect, as opposed to that observed for diazepam (1 mg/kg i.p.). The highest dose of buspirone used, however, did interfere with the learning of the avoidance response itself. Chronic buspirone (20 days, 0.5 and 0.75 mg/kg i.p.) did not have any effect on successive negative contrast either. Overall, these results could suggest that the 5-HT_1A receptor is not involved in the negative contrast effect studied, quite different to that observed for the γ-aminobutyric acid (GABA) system. The findings are compared to results obtained with animal models selectively sensitive to some anxiolytic drugs, as are the so-called ‘conflict models’.     

One-way avoidance learning and diazepam in female roman high-avoidance and low-avoidance rats

The goal of this experiment was to study the influence of the time spent in the safe compartment (30 vs. 1 s) and of an intraperitoneal injection of diazepam (1 mg/kg vs. vehicle) on one-way avoidance learning, in inbred female roman high-avoidance and roman low-avoidance rats. Rats learned to run from a danger compartment, where they received a warning signal (88 dB tone) followed by a 1 mA electric footshock, to a safe compartment, where these stimuli were not presented. The number of trials needed to reach 10 consecutive avoidance responses was the dependent variable. Roman low-avoidance rats exposed to 1 s in the safe compartment showed poorer performance than their roman high-avoidance counterparts. These differences were not observed in rats exposed to 30 s in the safe place, and were abolished by the injection of diazepam. These results suggest the importance of fear and reinforcement in one-way avoidance learning and its usefulness for studying emotional processes underlying genetic or pharmacological manipulations.     

Differential effects of diazepam infused into the amygdala and hippocampus on negative contrast

Behavioral suppression is observed when animals shift from a high to a lower magnitude of reward in comparison to animals that continuously receive the lower magnitude reward. As previously reported, systemic administration of benzodiazepines promotes recovery from this negative contrast. This study aimed to assess where the neural substrate(s) located in the limbic areas for diazepam to induce such recovery effects on negative contrast. With food-deprived rats, the negative contrast procedure was conducted by comparing a group consuming a 32% sucrose solution which was shifted to 4% with a group consuming only 4% sucrose throughout the experiment. Represented mainly by a decreased number of licks, the negative contrast effects were clearly shown in the control groups receiving the vehicle. Systemic injection of diazepam dose-dependently reduced this contrast. Further, this negative contrast effect was significantly attenuated by local infusion of diazepam (30 microg) into the amygdala, but no such effect was confirmed when diazepam was infused into the hippocampus. Together, the present study shows that a reliable anti-contrast effect can be induced by diazepam administration peripherally or locally infused into the amygdala. These data indicate that the amygdala is involved in the recovery effects of benzodiazepines on consummatory negative contrast.     

BPC-157,一种胃十五肽的抗焦虑作用:休克探查/埋藏试验和明暗试验(英文)

AIM: To study anxiolytic effect of a gastric pentade-capeptide, BPC-157. METHODS: In shock probe/burying test, pentadecapeptide BPC-157 (10 μg/kg, 10ng/kg, ip), diazepam (0.075, 0.0375 mg/kg, ip),and an equivolume of saline (5 mL/kg, ip) were givenat 30 min prior test. In light/dark test, the same dosageof diazepam, BPC-157, and saline were given at 45 minPrior procedure. RESULTS: Shock probe/buryingtest: rats treated with either diazepam or pentadecapeptideBPC-157 were much less afraid after the shock: almost     

Benzodiazepines reduce the tolerance to reward delay in rats

This study investigated whether benzodiazepines reduce the capacity of animals to wait for food reward. Rats trained in a T-maze were allowed to choose between two magnitudes of reward: immediate, but small (two pellets) vs delayed, but large (eight pellets). The rats learned within ten sessions to select (80–100%) the arm leading to the largest reward. Separate groups of rats were then confined for 15, 30 or 60 s in the arm associated with the largest reward before gaining access to the spacially contiguous goal-box. The choice of the other arm was not followed by a period of waiting. Under these conditions, the frequency with which the small-reward arm was chosen increased linearly as a function of the duration of the waiting period. Diazepam (2–4 mg/kg IP) dose-dependently increased the number of times the small-reward arm was chosen during the sessions for which the waiting period was fixed at 15 or 30 s. Nitrazepam (2 mg/kg IP), chlordiazepoxide (16 mg/kg IP) and clobazam (16 mg/kg IP) had similar effects. The action of diazepam was counteracted by simultaneous administration of flumazepil (Ro 15-1788, 8 mg/kg PO). In the absence of confinement, these benzodiazepines, diazepam (4 mg/kg) excepted, did not modify selection of the large-reward arm. Conversely, the serotonin uptake blockers indalpine (2–4 mg/kg IP) and zimelidine (8–16 mg/kg IP) dose-dependently increased preference for the arm leading to the delayed (25 s) but large reward.These results suggest that benzodiazepines, perhaps by increasing impulsivity, render the animals less prone than controls to tolerate delayed access to reward. It is hypothesized that serotonergic neurons play a crucial role in impulse control and in the benzodiazepines-induced shift towards the immediate reward.     

A comparison of the effects of scopolamine and diazepam on acquisition and retention of inhibitory avoidance in mice

Administration of either the muscarinic antagonist scopolamine or the benzodiazepine diazepam prior to training produced a dose-dependent impairment in the retention of one-trial inhibitory avoidance training in mice. To investigate the nature of this drug effect, the effects of scopolamine and diazepam were subsequently assessed on both acquisition and retention of inhibitory avoidance using a multiple-trial, training-to-criterion procedure. The training was conducted using either continuous trials in which the mouse was free to shuttle back and forth between shock and safe compartments or discrete trials in which the mouse was moved from the shock compartment of the safe compartment at the start of each trial. In either case, training continued until the mouse refrained from crossing into the shock compartment for a specified length of time on a single trial. Scopolamine (1.0 mg/kg) administered before training significantly increased the number of trials required to attain criterion, but did not affect retention when these mice were tested 2, 16, or 28 days later. In contrast, diazepam (1.0 mg/kg) did not significantly alter the number of trials necessary to reach criterion, but impaired retention of the inhibitory response in mice trained using discrete trials. The differences in the amnestic effects of scopolamine and diazepam revealed by this detailed analysis suggest that diazepam does not impair inhibitory avoidance performance through an effect on cholinergic function.     

Ethanol induces conditioned place preference in genetically selected alcohol-preferring rats

The study of the biological mechanisms of ethanol reward has greatly suffered from problems to obtain ethanol-induced conditioned place preference (CPP) in rats. In the present study, CPP was obtained in genetically selected Marchigian Sardinian alcohol-preferring (msP) rats, derived from Sardinian alcohol-preferring rats, following intragastric (IG) ethanol administration by means of a permanent IG catheter, but not after intraperitoneal (IP) injection or IG gavage. Rats with permanent IG catheter, received IG administration of 0.35, 0.7, 1.5 or 2.8 g/kg ethanol, as a 10% v/v solution. In ethanol-experienced rats 0.7 or 1.5, but not 0.35 or 2.8 g/kg ethanol significantly increased in comparison to controls the time spent in the ethanol-associated previously non-preferred compartment, which became preferred in the post-conditioning test. In ethanol-naive rats, only 0.7 g/kg ethanol significantly increased the time spent in the ethanol-associated compartment. On the other hand, no effect was observed in alcohol-experienced rats following IG gavage, or IP injection of 0.35, 0.7 or 1.5 g/kg ethanol. The present results provide evidence that ethanol possesses postingestive rewarding properties in msP rats, and that it can reliably induce CPP in them, provided that an appropriate method of administration is adopted. Received: 26 October 1997/Final version: 26 May 1998     

Enhancement of a diazepam withdrawal symptom by bicuculline and yohimbine

The role of the GABA system in producing a pentylenetetrazol-like interoceptive discriminative stimulus during withdrawal from diazepam was investigated in rats by determining the sensitivity of this system to GABAergic drugs before and after chronic treatment with diazepam. Food-restricted rats were trained to obtain a reward of food by responding on one lever following an injection of pentylenetetrazol (PTZ; 20 mg/kg) and the other lever following an injection of saline (1 ml/kg). After rats had acquired this discrimination, the effectiveness of Ro 15–1788, bicuculline and yohimbine to substitute for pentylenetetrazol was determined. Prior to chronic treatment with diazepam, rats selected the appropriate lever for saline after Ro 15–1788 and the appropriate lever for pentylenetetrazol after bicuculline (0.04–2.5mg/kg) or yohimbine (0.16–5.0mg/kg). Although the selection of the appropriate lever for pentylenetetrazol was dose-dependent, full substitution for pentylenetetrazol was not obtained with either drug as larger doses of bicuculline produced convulsions while the rats began to select the appropriate lever for saline after larger doses of yohimbine (bell-shaped curve). Diazepam blocked the pentylenetetrazol-like interoceptive discriminative stimulus for bicuculline. The rats were then injected with diazepam (80mg/kg/8 hr) for 24 days. Upon termination of the administration of diazepam, the animals were tested for lever-selection following the administration of saline, Ro 15–1788 (10mg/kg), bicuculline (0.32, 0.64 and 1.25mg/kg) or yohimbine (0.16, 0.64 and 2.5mg/kg). After saline, 33% of the rats selected the appropriate lever for pentylenetetrazol whereas selection of this lever was enhanced after Ro 15–1788, bicuculline or yohimbine. Thus, either displacement of diazepam or further reduction of GABAergic activity increased the subjective effect of the withdrawal from diazepam. These data support the hypothesis that the chronic administration of diazepam produces a reduction in the activity of the GABA/benzodiazepine complex, which is manifested during withdrawal as a pentylenetetrazol-like stimulus.     

Haloperidol and clozapine affect social behaviour in rats postnatally lesioned in the ventral hippocampus

Neonatal ibotenic acid lesion of the ventral hippocampus results in altered patterns of social behaviour. After puberty, lesioned animals spent less time in social interaction and the nonaggressive/aggressive behaviour ratio shifted towards increased aggressiveness. In this study, the effects on social behaviour of the neuroleptic drugs haloperidol (HAL) and clozapine (CLO) after acute and subchronic treatment were studied. Seven-day-old rats were lesioned and social behaviour was tested at the age of 13 weeks. Drug effects were tested after acute (HAL 0.025 mg/kg, CLO 1.0 mg/kg) and subchronic (10 injections, HAL 0.075 mg/kg, CLO 5.0 mg/kg) administration. For comparison, diazepam (DZP, 0.5 mg/kg) was used in the acute experiment. After acute administration, DZP had no effect on social behaviour in sham-lesioned rats, but nonaggressive behaviour increased significantly in lesioned animals. CLO and HAL did reduce the time sham-lesioned rats spent in social contact, and CLO also increased % nonaggressive behaviour in lesioned rats. Here, HAL had no effect. Subchronic administration did not alter social behaviour in sham-lesioned animals. However, CLO increased the time lesioned animals spent in social interaction, whereas HAL had an effect on nonaggressive behaviour. The results of this study indicate that the lesion model is sensitive to differentiated effects of classical neuroleptic drugs such as HAL and atypical neuroleptic drugs like CLO. It might be a useful tool in the search for potential neuroleptic drugs.     

Comparative effects of valproate,anxiolytic, or anxiogenic drugs on the light/dark aversion test

The potential anxiolytic-like effects of 200, 300, and 400 mg/kg i.p. of sodium valproate were evaluated in the light/dark aversion test in mice. For comparison, we included the reference anxiolytic drug, diazepam (2.5, 5.0, 10 mg/kg i.p.) as well as putative anxiolytics, i.e., clonidine (0.015, 0.030, 0.060 mg/kg i.p.), verapamil (5, 10, 20 mg/kg i.p.), or anxiogenic drugs, pentylenetetrazol (5, 10, 15 mg/kg i.p.). Results showed that control mice preferred the dark compartment where they spent approximately 70% of their time. After administration of 5 and 10 mg/kg i.p. of diazepam or 400 mg/kg i.p. of valproate, the increase in exploratory behavior in the lit area was associated with an increase both in the number of transitions and in the time spent in the lit area. Pretreatment with 0.030 and 0.060 mg/kg i.p. clonidine caused an increase in the time spent in the lit area. Verapamil had no significant effect on both measures of this test. Pentylenetetrazol, in doses which did not cause convulsions, was anxiogenic. In conclusion, these results in the light/dark aversion test are consistent with clinical data showing that valproate, a GABA-agonist, and clonidine, an α₂ adrenoceptor agonist, possess anxiolytic properties. © 1992 Wiley-Liss, Inc.     

Effects of chronic desipramine on waiting behaviour for a food reward in olfactory bulbectomized rats

The effects of olfactory bulbectomy (OB) on the 'waiting behaviour' of rats for a food reward in a T-maze, as well as the effect of chronic treatment with desipramine (7.5 mg/kg, i.p. x 14 days) prior to and throughout testing was assessed. In the T-maze, the time spent in locating a reward was longer in OB rats during the first phase of the food reward test. However, all groups chose the larger reward in at least 80% of trials by day 5 of test. Upon imposition of a delay in acquisition of the larger reward (phase 2), sham-operated rats chose the larger reward in correspondingly fewer trials until, by day 5, only 13% (control sham group) and 29% (desipramine-treated sham group) of trials were for the larger but delayed reward. By contrast, both OB rat groups continued to choose the larger but delayed reward throughout phase 2 in the majority of trials. The typical behavioural hyperactivity of OB rats in the 'open field' apparatus was attenuated by chronic administration of DMI. The results suggest that the OB rat has a deficit in food-motivated behaviour and appears to have reduced adaptation of this learned response when the conditions of the test are altered. Chronic desipramine treatment did not attenuate this effect.     

Endogenous opioids are necessary for benzodiazepine palatability enhancement: naltrexone blocks diazepam-induced increase of sucrose-'liking'

Opioid agonists and benzodiazepine agonists each increase food intake. Both also increase hedonic 'liking' reactions to sweet tastes in rats. Do opioids and benzodiazepines share overlapping mechanisms of hedonic impact? Or are benzodiazepine and opioid effects on hedonic impact mediated by independent mechanisms? The present study examined whether blockade of opioid receptors prevents benzodiazepine-induced enhancement of taste palatability, as assessed by the affective taste reactivity test. Rats were implanted with oral cannulae, and prior to an oral infusion of bittersweet quinine-sucrose solution, all received i.p. injections of either vehicle, or diazepam alone (5 mg/kg diazepam+0 mg/kg naltrexone), naltrexone alone (1 mg/kg naltrexone+0 mg diazepam), or both diazepam plus naltrexone (5 mg/kg diazepam+1mg/kg naltrexone). Videotaped hedonic ('liking') and aversive ('disliking') orofacial reactions elicited by sucrose/quinine taste were compared across drug conditions. Diazepam administration alone more than doubled hedonic 'liking' reactions to the bittersweet taste, while reducing 'disliking' in half, compared to vehicle levels. Naltrexone by itself had little effect on taste-elicited affective reactions, and only marginally increased aversive gapes. However, naltrexone completely blocked diazepam's enhancement of positive hedonic 'liking' reactions, and naltrexone similarly disrupted diazepam-reduction of aversive 'disliking' taste reactions. These results indicate that endogenous opioid neurotransmission may be crucial to benzodiazepine enhancement of hedonic 'liking' for natural taste reward.     

Reduction of submissive behavior in rats: a test for antidepressant drug activity.

Randomly paired rats were food deprived overnight and placed in an apparatus compelling them to compete for a food reward. About half of these pairs developed a dominant-submissive relationship measured as a significant difference in time spent on the feeder by each rat. This relationship developed over a 2-week period and remained stable for at least the next 5 weeks. Treatment of the submissive subjects, for at least 2 weeks, with imipramine, desipramine, or fluoxetine (10 mg/kg) significantly reduced submissive behavior. The effect faded after cessation of treatment with desipramine. Fluoxetine was further tested at 2.5- and 5-mg/kg doses and showed a dose-dependent reduction of submissive behavior. Treatment of submissive rats with the anxiolytic diazepam (1 mg/kg) was ineffective. The prevalence of dominant-submissive relationships and the effect of desipramine and imipramine on submissive behavior were gender independent. The predictive, face, and construct validity of the behavioral test is discussed.     

Prevention of diazepam withdrawal syndrome by nifedipine-behavioural and neurochemical studies

Our studies aimed at investigating whether the dihydropyridine calcium antagonist, nifedipine, could prevent anxiogenic-like consequences of diazepam withdrawal in rats. Animals withdrawn from chronic diazepam (2 mg/kg/day i.p. for 2 weeks) drank significantly less water than did control rats in the unfamiliar arm of a Y maze. This anxiogenic-like effect could be prevented by acute administration of nifedipine (at 10 mg/kg i.p., but not at lower doses), which, on its own, did not change water intake in naive rats. Given chronically in combination with diazepam for the second half of a 2-week treatment with this drug, nifedipine (at the daily dose of 5 mg/kg i.p.) also suppressed the reduction of water intake normally observed on diazepam withdrawal. Biochemical measurements showed that acutely, as well as chronically, administered nifedipine increased 5-HT turnover in the hippocampus of diazepam-treated rats, thereby suggesting that the prevention of diazepam withdrawal-induced anxiogenic behaviour by the calcium antagonist might be underlain by serotoninergic mechanisms.     

Unbiased cocaine conditioned place preferences (CPP) obscures conditioned locomotion, and nimodipine blockade of cocaine CPP is due to conditioned place aversions

The effect of nimodipine (0, 0.1, 1.0 and 10 mg/kg, SC), a dihydropyridine L-type Ca²⁺ channel antagonist, on the establishment of cocaine-(10 mg/kg IP) conditioned place preferences (CPP) was investigated. Nimodipine produced conditioned place aversions (CPA) on its own; reductions in cocaine CPP are apparently due to this CPA. There is a high negative correlation between time spent in the CS+ compartment and the difference in locomotion rates between the CS+ and the non-drug (CS−) compartments, independent of drug effects. This relationship is responsible for an increased rate of locomotion observed in the CS− compartment in cocaine-conditioned rats. Analysis of covariance indicated that cocaine CPP occurred independently of cocaine’s effects on locomotion. Furthermore, cocaine produces an increase in the rate of locomotion in the CS+ compartment when time spent in this compartment is equated with time spent in the CS− compartment. This suggests that cocaine’s effects on CPP and “conditioned” locomotion are due to separate mechanisms of action. On the other hand, nimodipine-induced place aversions and locomotor rates are not independent of each other, indicating a common mechanism of action, or that one is a consequence of the other. It is concluded that place preferences and place aversions can sometimes be secondary to compartment-specific locomotor changes, and locomotion effects can be confounded by differential times spent in each compartment. The relationships between these two behaviours must be controlled for before conclusions of CPP or CPA can be drawn in drug conditioning studies. Received: 25 January 1996 / Final version: 7 November 1996     

Long-term diazepam treatment produces changes in cholecystokinin receptor binding in rat brain

This study examined the effect of chronic diazepam administration on central benzodiazepine and CCK-8 receptor binding in rat brain. After a two-week treatment with diazepam (5 mg/kg per day) tolerance developed towards the sedative but not towards the anxiolytic action of this drug as determined using elevated plus-maze and open field tests. The % entries the rats made onto open arms and % time the rats spent in open arms were markedly decreased 24 h after the last dose of diazepam, probably indicating withdrawal anxiety. There were no changes in [3H]flunitrazepam binding either 30 min or 24 h after the last diazepam dose. However, 30 min after the last diazepam administration the apparent number of sulphated [3H]CCK-8 binding sites was significantly increased in the primary olfactory cortex. Acute diazepam treatment (5 mg/kg) had no influence on [3H]flunitrazepam or sulphated [3H]CCK-8 binding in any brain region studied. Cessation of chronic diazepam treatment was followed after 24 h by an increase in the number of CCK-8 receptors in frontal cortex and hippocampus as compared to the vehicle group. These results demonstrate that certain alterations in CCK-8 receptor characteristics may be important in the anti-anxiety effect, tolerance, and withdrawal reaction reaction after benzodiazepine administration.     

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.     

A role for the mesolimbic dopamine system in the reinforcing properties of diazepam

The conditioned place preference paradigm was used to investigate the neurochemical and neuroanatomical substrates which mediate the rewarding properties of diazepam. The results confirmed that diazepam (1 and 2.5 mg/kg, IP) produced place preference for a distinctive environment that had previously been paired with injections of the drug. Pretreatment with haloperidol (0.1 mg/kg) antagonised the place preference induced by diazepam (1 mg/kg). Pretreatment with domperidone (2 mg/kg) failed to influence this effect of diazepam. Haloperidol (0.1 mg/kg) and domperidone (2 mg/kg) alone did not produce place aversion. In separate experiments the diazepam-induced place preference was examined in rats having 6-hydroxydopamine (6-OHDA) lesions of the nucleus accumbens. These animals did not show preference for the compartment associated with diazepam. Depletion of central noradrenaline produced by systemic injections of DSP4 did not affect diazepam-induced place preference conditioning. These findings suggest that dopamine-containing neurons of the mesolimbic system are a component of the neural circuitry that mediates the reinforcing properties of diazepam.     

Enhancement of acetylcholine release by flumazenil in the hippocampus of rats chronically treated with diazepam but not with imidazenil or abecarnil

The effects of long-term treatment (three times a day for 3 weeks) with pharmacologically active doses of the novel anxiolytics and anticovulsants abecarnil (0.5 mg/kg, IP) and imidazenil (0.5 mg/kg, IP) on basal hippocampal acetylcholine release in freely moving rats were compared with those of diazepam (3 mg/kg, IP). Challenge doses of diazepam, abecarnil, and imidazenil decreased the extracellular acetylcholine concentration in the hippocampus by the same extent in animals chronically treated with the respective drug or vehicle. Moreover, the abrupt discontinuation of long-term treatment with diazepam, abecarnil, or imidazenil failed to affect hippocampal acetylcholine release during the first 5 days of withdrawal. In contrast, the acute administration of the benzodiazepine receptor antagonist flumazenil (1 mg/kg, IP) 2 days after diazepam withdrawal elicited a marked increase (65%) in acetylcholine release in the hippocampus. Flumazenil failed to induce the same effect 5 days after diazepam withdrawal or 2 or 5 days after discontinuation of long-term treatment with abecarnil or imidazenil. These results indicate that (i) the inhibitory effects of full (diazepam), partial (imidazenil), and selective (abecarnil) benzodiazepine receptor agonists on acetylcoholine output in rat hippocampus are not affected by repeated drug administration; (ii) discontinuation of long-term treatment with each type of agonist does not affect hippocampal cholinergic mechanisms; and (iii) flumazenil increases acetylcholine release only in the hippocampus of rats chronically treated with diazepam. Together, these data further differentiate the pharmacology of benzodiazepine receptor full agonists from that of partial and selective agonists.