Carisoprodol-mediated modulation of GABAA receptors: in vitro and in vivo studies.

Carisoprodol is a frequently prescribed muscle relaxant. In recent years, this drug has been increasingly abused. The effects of carisoprodol have been attributed to its metabolite, meprobamate, a controlled substance that produces sedation via GABA(A) receptors (GABA(A)Rs). Given the structural similarities between carisoprodol and meprobamate, we used electrophysiological and behavioral approaches to investigate whether carisoprodol directly affects GABA(A)R function. In whole-cell patch-clamp studies, carisoprodol allosterically modulated and directly activated human alpha1beta2gamma2 GABA(A)R function in a barbiturate-like manner. At millimolar concentrations, inhibitory effects were apparent. Similar allosteric effects were not observed for homomeric rho1 GABA or glycine alpha1 receptors. In the absence of GABA, carisoprodol produced picrotoxin-sensitive, inward currents that were significantly larger than those produced by meprobamate, suggesting carisoprodol may directly produce GABAergic effects in vivo. When administered to mice via intraperitoneal or oral routes, carisoprodol elicited locomotor depression within 8 to 12 min after injection. Intraperitoneal administration of meprobamate depressed locomotor activity in the same time frame. In drug discrimination studies with carisoprodol-trained rats, the GABAergic ligands pentobarbital, chlordiazepoxide, and meprobamate each substituted for carisoprodol in a dose-dependent manner. In accordance with findings in vitro, the discriminative stimulus effects of carisoprodol were antagonized by a barbiturate antagonist, bemegride, but not by the benzodiazepine site antagonist, flumazenil. The results of our studies in vivo and in vitro collectively suggest the barbiturate-like effects of carisoprodol may not be due solely to its metabolite, meprobamate. Furthermore, the functional traits we have identified probably contribute to the abuse potential of carisoprodol.

Meprobamate reduces accuracy of physiological detection of deception.

Normal male subjects attempted to deceive an experimenter recording electrodermal, respiratory, an cardiovascular activity. Those who had ingested a placebo or nothing were detected with statistically significant frequency on the basis of their phasic electrodermal responses, which clearly distinguished them from truthful suspects. That was not the case with deceptive subjects who had ingested 400 milligrams of meprobamate, nor did the examiner detect which subjects had received the drug.

Evaluation of CNS activities of aerial parts of Cynodon dactylon Pers. in mice.

The dried extracts of aerial parts of Cynodon dactylon Pers. (Graminae) were evaluated for CNS activities in mice. The ethanol extract of aerial parts of C. dactylon (EECD) was found to cause significant depression in general behavioral profiles in mice. EECD significantly potentiated the sleeping time in mice induced by standard hypnotics viz. pentobarbitone sodium, diazepam, and meprobamate in a dose dependant manner. EECD showed significant analgesic properties as evidenced by the significant reduction in the number of writhes and stretches induced in mice by 1.2% acetic acid solution. It also potentiated analgesia induced by morphine and pethidine in mice. EECD inhibited the onset and the incidence of convulsion in a dose dependent manner against pentylenetetrazole (PTZ)-induced convulsion. The present study indicates that EECD has significant CNS depressant activities.

Assessment of direct gating and allosteric modulatory effects of meprobamate in recombinant GABA(A) receptors.

Meprobamate is a schedule IV anxiolytic and the primary metabolite of the muscle relaxant carisoprodol. Meprobamate modulates GABAA (γ-aminobutyric acid Type A) receptors, and has barbiturate-like activity. To gain insight into its actions, we have conducted a series of studies using recombinant GABAA receptors. In αxßzγ2 GABAA receptors (where x=1-6 and z=1-3), the ability to enhance GABA-mediated current was evident for all α subunit isoforms, with the largest effect observed in α5-expressing receptors. Direct gating was present with all α subunits, although attenuated in α3-expressing receptors. Allosteric and direct effects were comparable in α1ß1γ2 and α1ß2γ2 receptors, whereas allosteric effects were enhanced in α1ß2 compared to α1ß2γ2 receptors. In "extrasynaptic" (α1ß3δ and α4ß3δ) receptors, meprobamate enhanced EC20 and saturating GABA currents, and directly activated these receptors. The barbiturate antagonist bemegride attenuated direct effects of meprobamate. Whereas pentobarbital directly gated homomeric ß3 receptors, meprobamate did not, and instead blocked the spontaneously open current present in these receptors. In wild type homomeric ρ1 receptors, pentobarbital and meprobamate were ineffective in direct gating; a mutation known to confer sensitivity to pentobarbital did not confer sensitivity to meprobamate. Our results provide insight into the actions of meprobamate and parent therapeutic agents such as carisoprodol. Whereas in general actions of meprobamate were comparable to those of carisoprodol, differential effects of meprobamate at some receptor subtypes suggest potential advantages of meprobamate may be exploited. A re-assessment of previously synthesized meprobamate-related carbamate molecules for myorelaxant and other therapeutic indications is warranted.

The benzodiazepine--GABA--chloride ionophore receptor complex: common site of minor tranquilizer action.

The demonstration of specific recognition sites for benzodiazepines in the mammalian CNS has altered current thinking on the mechanisms of action of the benzodiazepines as well as the neurochemical events which are associated with anxiety. Recent studies suggest that the physiological regulation of the benzodiazepine receptor is far more complex than initially believed and includes a functional coupling to both a GABA receptor and an associated chloride ionophore. It now appears that a number of other psychopharmacologic agents, including minor tranquilizers other than the benzodiazepines as well as several convulsants and anticonvulsants, may exert their pharmacologic effects by affecting one or more regulatory sites on the benzodiazepine receptor complex. In addition to a number of drugs, at least one endogenous small molecular weight compound that has been isolated from the crude synaptosomal fraction of bovine cerebral cortex also appears to modulate this receptor complex.

Subjective and physiologic effects of morphine, pentobarbital, and meprobamate.

These studies extend previous observations on the effects of pentobarbital on subjective states and postrotational nystagmus in postaddict subjects. Pentobarbital (150 mg) induced a degree of liking and an elevation of the morphine-benzedrine group (MBG) scale score equivalent to 24 mg of morphine. The effects of pentobarbital and meprobamate on postrotational nystagmus were studied using electro-oculography. Both drugs increased the frequency and prolonged the duration of postrotational nystagmus in a dose-related manner. Meprobamate was about 1/15 as potent as pentobarbital in enhancing postrotational nystagmus and producing signs of sedation.

Persistence of drug experience in rats formerly dependent on phenobarbital or meprobamate.

The rats of groups I, II, III and IV were treated orally with phenobarbital, meprobamate, codeine and vehicle, respectively, for total 21 days, and then drugs were withdrawn. All these rats were given again orally phenobarbital for 5 days starting from 70 days after the withdrawal. In comparison with groups III and IV, groups I and II showed larger weight gain during phenobarbital re-administration and longer-lasting weight loss and an larger increase in body temperature after the termination. These results suggested that the drug experience on sedative-hypnotics persisted over two months after the withdrawal and that did not cross to that of narcotics.

Septal driving of hippocampal theta rhythm: role of gamma-aminobutyrate-benzodiazepine receptor complex in mediating effects of anxiolytics.

In free-moving male rats, when the hippocampal theta rhythm is artificially driven by stimulation in the septum at frequencies between 5 and 10 Hz, the function relating frequency to the threshold current required to drive the theta rhythm has a minimum at 7.7 Hz. This minimum is eliminated by anxiolytic drugs. Dose-response curves for this effect are reported for chlordiazepoxide, diazepam and meprobamate. The effect of meprobamate was reversed by two gamma-aminobutyrateA antagonists, picrotoxin and bicuculline, which have previously been shown to be without effects of their own. The gamma-aminobutyrateB agonist, baclofen, also without effect on its own, blocked the elimination of the 7.7-Hz minimum caused by the gamma-aminobutyrateA agonist, muscimol. The beta-carboline, ethyl-beta-carboline-3-carboxylate, had mixed agonist/antagonist properties, blocking the effects of chlordiazepoxide, diazepam and muscimol (though not sodium amylobarbitone) but itself acting like a benzodiazepine. Coupled with earlier data, these findings support a role for gamma-aminobutyrate receptors in mediating the effects of anxiolytic drugs.

Effects of general depressant drugs on the electrical responses of isolated slabs of cat's cerebral cortex.

1. In the neuronally isolated cortex of the cat, local application of diphenhydramine, promethazine, gammahydroxybutyrate, gammabutyrolactone, gamma aminobutyric acid, hyoscine and pethidine, and the intravenous injection of diazepam and meprobamate depressed or abolished the surface negative and surface positive response to direct stimulation and raised the stimulus threshold of the positive burst response. These effects were the same as previously demonstrated for general and local anaesthetics on the same preparation.2. Chlorpromazine produced a similar depression in small concentrations but caused spontaneous activity in higher concentrations.3. In contrast to local application, pethidine when given by intravenous injection in a high dose produced convulsant activity in the isolated cortical slab. The possibility was suggested that the convulsant activity was produced by a metabolite of pethidine.4. The results of this investigation suggest that the central depression produced by a number of structurally unrelated drugs is indicative of an anaesthetic-like property of these drugs.

A sequential screning test based on the running component of audiogenic seizures in mice, including reference compound PD50 values.

1. The running component of audiogenic seizures in mice has been used as the basis of a sequential screening test for the detection of a variety of centrally acting drugs.2. For acceptance by the test, an active compound must completely suppress the running component in a total of sixteen mice at a dose of 1/5 LD50 intraperitoneally.3. Considerable economies in the numbers of animals required for screening have been achieved, the mean number of mice required to reject an inactive compound being 2.0.4. The running component is highly sensitive to anticonvulsants and general central depressants, and insensitive to phenothiazine tranquillizers and morphine. Reserpine caused an increase in the severity of the running component.5. The statistical model used in this test is of general application to screening test situations which use quantal data.

Effects of drugs acting alone and in combination on the motor activity of intact mice.

1. When administered to intact white mice, the central depressants-diphenhydramine, promethazine, chlorpromazine, gammahydroxybutyrate, gammabutyrolactone, hyoscine, and pethidine-produced sedation in small doses, but excitement and convulsions in higher doses. When given to mice pretreated with subanaesthetic doses of phenobarbitone these drugs abolished the righting reflex both in convulsant doses (hyoscine excepted) and in non-convulsant doses. These effects are similar to the effects previously observed with local anaesthetics.2. Meprobamate, diazepam and chlorpromazine produced a loss of righting reflex both when given alone and following phenobarbitone. When given alone in higher doses, chlorpromazine induced convulsions.3. The central stimulants bemegride and picrotoxin antagonized the loss of righting reflex produced by phenobarbitone, but nikethamide, caffeine and strychnine did not alter the depressant effects of phenobarbitone.4. On the basis of these and previous studies with intact white mice a tentative classification of drugs having generalized depressant and stimulant effects on the central nervous system was proposed and discussed.

[Effects of antianxiety drugs on the water intake in trained and untrained rats and mice (author's transl)]. / Effets des anxiolytiques sur la prise de boisson en situation nouvelle et familière

In water-deprived rats and mice, animals trained to the test situation spent more time in drinking than naive animals (first exposure to the test situation). The time spent in drinking, either during 5 min or during 10 min was recorded. As compared to controls, benzodiazepines, phenobarbital, meprobamate, and mecloqualone increased drinking time whether the experiments were run on naive or on experienced animals [5 or 10 (in mice) and 9 (in rats) exposures in the test situation]. All drugs were injected i.p. 30 min before testing. This release of the drinking behavior was more pronounced during the last 5 min than during the first 5 min of the 10 min test session. These results suggest that: 1. The inhibition of water intake of naive animals as compared to trained rats and mice, could be related to some emotional factors elicited by the first exposure to an unknown situation. 2. The increase in drinking time induced by the antianxiety drugs in a novel and in a familiar situation seems difficult to correlate only with the antianxiety action of these compounds. 3. Antianxiety drugs could interfere with the regulatory mechanism of thirst.

The Geller-Seifter conflict paradigm with incremental shock.

The typical Geller-Seifter conflict paradigm for predicting clinical efficacy of anxiolytics is a mult VI/CRF schedule in which response rates in the CRF (conflict) portion are depressed by response-contingent electric shock. In 1-h sessions, anxiolytics raise the depressed conflict rates. Recently it was shown that replacing the single shock level with an arrangement whereby shock begins at zero and is increased with each response in the conflict portion produced more orderly data and facilitated training and maintenance of experimental subjects; chlordiazepoxide was the test drug. In the present study, those results are replicated in 30-min sessions, and the incremental paradigm is demonstrated to be as specific for anxiolytics as the standard Geller-Seifter paradigm. The possibility of very short sessions is suggested.

The staircase test: some evidence of nonspecificity for anxiolytics.

In the staircase test, a naive mouse is placed in a Plexiglas chamber containing a five-step staircase, and the number of rearings and steps climbed are recorded for 3 min. A claim for drug-class specificity has been made because conventional anxiolytics reduced rearings at doses that did not reduce steps climbed, while non-anxiolytics affected both measures in parallel. In the present study chlordiazepoxide, meprobamate, and ethanol registered the expected true positive effect by reducing rearings at doses that did not reduce steps climbed. Nicotine, which has some clinical anxiolytic action, registered a small true positive. The benzodiazepine anxiolytic alprazolam reduced both measures, a false negative, although it reduced rearings more than steps climbed. The putative novel anxiolytics CGS 9896, ketanserine, and tracazolate registered negatives, as did the known clinical anxiolytic buspirone. The non-anxiolytics phencyclidine and phenacetin registered true negatives, but morphine registered a clear false positive. The anxiogenics FG 7142 and pentylenetetrazol produced no significant effects. Because of the equivocal false negative for alprazolam, the clear false negative for buspirone, and the clear false positive for morphine, we concluded that the test lacks the degree of therapeutic-class specificity previously proposed but may still be useful in basic research.

Benzodiazepines and behavioral effects of reward (water) omission in the rat.

Two behaviors related to nonreward (omission of water in an enclosure where the rats were habituated to drink) were studied. The time spent licking the bottles during water omission and the time spent drinking during a subsequent 5-min drinking session (water available) were recorded. The drinking session was performed 30 min after the water-omission session. Rats subjected to water omission showed an enhanced drinking time that varied with the length of the water omission session, with the motivational state of the animals, and with the previous number of water-omission sessions. Diazepam, chlordiazepoxide, lorazepam, and meprobamate (i.p., 30 min before water omission), increased the time spent licking the empty bottles, but failed to abolish subsequently enhanced drinking. However, some of our data suggested that minor tranquilizers weakly reduced the increased drinking induced by nonreward, despite their direct stimulation on water drinking. It is proposed that either minor tranquilizers are devoid of general antifrustration activity or nonreward-induced frustration and nonreward-induced drive enhancement may not be correlated.

Diazepam-induced place preference conditioning: appetitive and antiaversive properties.

The place conditioning paradigm was used to examine the reinforcing properties of diazepam. Rats were injected with diazepam (0.5-5.0 mg/kg, IP) and 30 min later were confined for 30 min to one side of a shuttle box, in which each of the two compartments had distinctive features. On alternate (control) days they received vehicle injections and were confined for 30 min to the opposite side. At almost all doses tested, diazepam produced place preference for the distinctive compartment that had been previously associated with the drug. Preference for the drug side developed regardless of whether diazepam was paired or unpaired with the least-preferred side, and regardless of whether testing was carried out in the undrugged or in the drugged state. The rats preferred the drug side over a novel compartment, but they did not change their initial preference for the side when diazepam was given after removal from the training box. Animals injected with meprobamate (70 mg/kg, PO), a non-benzodiazepine anxiolytic, also developed conditioned preference for the drug side, comparable to that seen following cocaine hydrochloride (10 mg/kg, IP). The diazepam (2.5 mg/kg)-induced place preference was antagonized by CGS 8216 (3 mg/kg, IP), picrotoxin (2 mg/kg, IP) and naloxone (0.8 mg/kg, SC), injected 3 min before and 15 and 20 min after diazepam respectively. Sodium valproate (200 mg/kg, IP) did not influence diazepam (1 mg/kg)-induced place preference. Sodium valproate by itself had marginal effects on place conditioning. Picrotoxin and naloxone, but not CGS 8816, produced place aversion which, in the case of picrotoxin, was due to state dependent learning.(ABSTRACT TRUNCATED AT 250 WORDS)

Stimulant and relaxant drugs combined with stimulant and relaxant information: a study of active placebo.

RATIONALE: The active placebo hypothesis states that placebo effects are potentiated when an active drug is administered. OBJECTIVE: This hypothesis was tested in an experiment where information about the effect of a drug was combined with administration of an active drug or placebo. METHODS: Information that a drug acted as a relaxant, a stimulant, or as a placebo was crossed with oral administration of a relaxant drug (700 mg carisoprodol), a stimulant drug (400 mg caffeine) or placebo (lactose) in healthy volunteers ( n=94). Dependent variables were subjective and physiological measures of arousal, as well as serum carisoprodol and caffeine levels. Data were collected from 15 to 280 min after administration of drug or placebo. RESULTS: Caffeine increased alertness, systolic and diastolic blood pressure, startle blink reflexes, and skin conductance responses. Subjects were calmer after carisoprodol, and heart rate was increased. There was a positive correlation between increased arousal and carisoprodol levels when stimulant information had been provided. However, this was only seen when carisoprodol levels were very low. There was no evidence that caffeine modulated the placebo response. CONCLUSIONS: Active placebo responses were seen only transiently when carisoprodol levels were low, and only in the subjective arousal data. Caffeine did not support active placebo responses. The overall findings did not favour the active placebo hypothesis.

The use of social interaction as a method for detecting anxiolytic activity of chlordiazepoxide-like drugs.

The social interaction test in rats provides a method for detecting anxiolytic activity that does not use food or water deprivation, or electric shock, and therefore obviates difficulties of interpretation that might arise from drug-induced changes in motivation. Since social interaction is measured under more than one test condition any overall increase or decrease in social behaviour can be detected independently from the drug x test condition interaction that characterizes an anxiolytic drug. The Geller-Seifter conflict test was designed with two schedules of reinforcement for the same reasons. Any candidate test for anxiolytic action that examines drug effects under only one experimental condition is open to misinterpretation and may also prove unreliable if the critical experimental factors ( e.g. the level of food deprivation or the shock intensity) are changed. The testing procedure in the social interaction test is relatively time consuming in terms of observer-hours, but no lengthy pretraining of the animals is required. There is no way of fully automating the scoring and therefore it is important that the observers do not know the experimental group of the rats that they are scoring, and that tape recordings are made so that the scores can be checked. It has not so far been fruitful to analyze drug effects on every individual social behaviour, but this method does allow changes in individual behaviours to be detected. By entering the data directly into a computer we are now able to store the frequency and duration of each behaviour as well as the sequence of behaviours. It will then be possible to determine whether a detailed analysis of drug effects on the patterning of social behaviours will prove a useful addition to the social interaction test

Uropharmacology: X. Central nervous system stimulants and depressants.

Several drugs that are utilized primarily for their effects on the central nervous system also affect lower urinary tract function. Most of these effects are produced by the action of these drugs on adrenergic and cholinergic receptors or by direct action of lower urinary tract musculature. Central nervous system stimulants and depressants which are known to affect the storage or evacuation role of the lower urinary tract are discussed.

Influence of meprobamate and phenobarbital upon local cerebral glucose utilization: parallelism with effects of the anxiolytic diazepam.

The [1-14C]2-deoxyglucose technique was employed for an evaluation of the regional pattern of alteration of brain metabolism induced by the anxiolytics phenobarbital (which is described in small doses as anxiolytic agent) and meprobamate. Their effects were compared with those produced by the anxiolytic benzodiazepine diazepam which we have described in a previous study. In low doses, both meprobamate (30 mg/kg i.v.) and phenobarbital (5 mg/kg i.v.) elicited a regional pattern of changes similar to those seen with diazepam. Thus, the local cerebral glucose utilization (LCGU) of the mammillary nuclei, the lateral and ventral thalamic nuclei, the anterior thalamic nuclei and the geniculate nuclei was significantly decreased. A doubling of the dose (meprobamate 60 mg/kg i.v.; phenobarbital 10 mg/kg i.v.), however, resulted in a decrement in LCGU in virtually every brain region examined. Further, at this higher dose, phenobarbital significantly increased LCGU in the interpeduncular nucleus. These data demonstrate that both meprobamate and phenobarbital, in moderate doses induce selective alterations in LCGU in particular brain regions with the pattern of changes similar to that induced by diazepam. The structures affected may be of general importance for the expression of the anxiolytic actions of each of those classes of minor tranquilizers in clinical use.