Lack of effects of GHB precursors GBL and 1,4-BD following i.c.v. administration in rats

Gamma-hydroxybutyrate (GHB) is used therapeutically and recreationally worldwide. Since the scheduling of GHB by the USA and the United Nations in 2000-2001, the recreational use of GHB precursors has reportedly increased. The aim of this study was to examine if potency differences of GHB and GHB-like compounds are due to their blood-brain barrier permeability. The effects of peripheral and central administration of GHB, GHB precursors gamma-butyrolactone (GBL) and 1,4-butanediol (1,4-BD), and the gamma-aminobutyric acid (GABA)(B) receptor agonist baclofen on schedule-controlled responding were examined in rats. GHB and baclofen were 276- and 253-fold more potent, respectively, after intracerebroventricular (i.c.v.) administration than after intraperitoneal (i.p.) administration, whereas GBL and 1,4-BD, up to a dose of 1780 microg were without effect after i.c.v. administration. These data suggest that GBL and 1,4-BD are not metabolically converted to GHB in the brain, that enhanced brain penetration cannot account for potency differences between compounds, and that baclofen, like GHB, can readily cross the blood-brain barrier.     

Different sensitivity to the motor incoordinating effects of gamma-hydroxybutyric acid (GHB) and baclofen in GHB-sensitive and GHB-resistant rats

The present study investigated whether the differential sensitivity of selectively bred gamma-hydroxybutyric acid (GHB)-sensitive (GHB-S) and GHB-resistant (GHB-R) rats to GHB- and baclofen-induced sedation/hypnosis generalized to the motor incoordinating effect of the two drugs. To this aim, GHB-S and GHB-R rats were tested on a Rota-Rod after the acute administration of GHB (100-500 mg/kg, i.p.) and baclofen (1.25-5 mg/kg, i.p.). Significant line differences were observed in the dose-response curves for both GHB and baclofen, with GHB-S rats displaying a greater sensitivity to the motor incoordinating effects of both drugs than GHB-R rats. No line difference was observed in diazepam (1.25-5 mg/kg, i.p.), pentobarbital (5-15 mg/kg, i.p.), and ethanol (1-1.5 g/kg, i.p.) dose-response curves. These results suggest that the differential sensitivity of GHB-S and GHB-R rats to GHB and baclofen extends to the effects produced by doses of the two drugs which are 5-10 times lower than those for which rats have been selectively bred. These results are discussed in terms of the GABA(B) receptor being the likely neural substrate on which the differential sensitivity of GHB-S and GHB-R rats resides.     

GHB-C rats: the control line of GHB-sensitive (GHB-S) and GHB-resistant (GHB-R) rats

Gamma-hydroxybutyric acid (GHB)-sensitive (GHB-S) and GHB-resistant (GHB-R) rats have been selectively bred for their opposite sensitivity to the sedative/hypnotic effect of GHB. This opposite sensitivity has been found to generalize to the GABA(B) receptor agonist, baclofen. A control line [named GHB-control (GHB-C)] has been derived from the foundation stock of GHB-S and GHB-R rats. GHB-C rats have been bred without any evaluation of their sensitivity to GHB. The experiments described here were designed to evaluate the sensitivity of GHB-C rats, from the 13th generation, to the sedative/hypnotic effect of GHB (1 g/kg, i.p.) and baclofen (20 mg/kg, i.p.). All measures (onset, sleep time and r = sleep time/onset) of sensitivity to GHB- and baclofen-induced sedation/hypnosis in GHB-C rats were significantly different from and intermediate to those recorded in GHB-S and GHB-R rats. Furthermore, these values were similar to those recorded in the foundation stock. These results suggest that GHB-C rats may constitute a valid control line for GHB-S and GHB-R rats, representing the "general population" from which GHB-S and GHB-R rats were derived. Furthermore, the relative equidistance of sensitivity to GHB- and baclofen-induced sedation/hypnosis of GHB-C rats from those of GHB-S and GHB-R rats suggests that genetic factors contributes to the development of both sensitivity in GHB-S rats and resistance in GHB-R rats.     

Failure of gamma-hydroxybutyric acid both to increase neuroactive steroid concentrations in adrenalectomized-orchiectomized rats and to induce tolerance to its steroidogenic effect in intact animals

Gamma-Hydroxybutyric acid (GHB), a drug proposed in the treatment of alcohol withdrawal syndrome, increases the cerebrocortical and plasma concentrations of the neuroactive steroids allopregnanolone and allotetrahydrodeoxycorticosterone (THDOC). In the present study, we examined the role of hypothalamic-pituitary-adrenal (HPA) axis in the effect of GHB by measuring the concentrations of these steroids in the brain and plasma of adrenalectomized-orchiectomized (Adx-Orx) rats. The acute administration of GHB (500 mg/kg, i.p.) induced in 30 min an increase in the concentrations of allopregnanolone, THDOC and their precursors pregnenolone and progesterone in different brain areas (cerebral cortex, hypothalamus and cerebellum) and plasma of sham-operated rats but had no effect on the concentrations of these compounds in Adx-Orx rats, suggesting that activation of the HPA axis mediates the effect of GHB on brain and plasma concentrations of neuroactive steroids. Moreover, we evaluated whether repeated exposure of GHB induces tolerance to its steroidogenic effects. Chronic administration of GHB (500 mg/kg, i.p., twice a day for 10 days) to intact animals failed to affect the levels of progesterone, allopregnanolone, or THDOC measured 3 or 48 h after the last drug administration, whereas a challenge injection of GHB or ethanol was still able to increase the concentrations of these steroids in brain and plasma. These results indicate that repeated exposure to GHB fails to induce tolerance or cross-tolerance to the steroidogenic action of GHB or ethanol, respectively.     

GABAB receptor activation exacerbates spontaneous spike-and-wave discharges in DBA/2J mice

Rich evidence has highlighted that stimulation of γ-amino-butyric acid (GABA)_B receptors increases the occurrence of spike-and-wave discharges (SWDs), the electroencephalographic (EEG) landmark of absence epilepsy (AE). Recent findings suggest that the outcomes of GABA_B activation in vivo are contingent on the chemical characteristics of the agonist. In particular, the endogenous ligand γ-hydroxybutyrate (GHB) and its precursor γ-butyro-lactone (GBL) have been shown to elicit different effects than the prototypical GABA_B agonist baclofen. In view of these premises, the present study was aimed at the characterization of the effects of baclofen (0.5–10 mg/kg, i.p.) and GBL (5–100 mg/kg, i.p.) on the spontaneous SWDs and locomotor activity of DBA/2J mice.While both baclofen and GBL dose-dependently increased SWDs episodes, high doses of the latter (100 mg/kg, i.p.) reduced the occurrence of these phenomena and increased the number of isolated spikes. Interestingly, both compounds elicited a dose-dependent reduction of locomotor activity, in comparison with their vehicle-treated controls. The GABA_B selective antagonist, SCH50911 (50 mg/kg, i.p.), reversed the changes in SWD occurrence and locomotion induced by baclofen and GBL, but failed to elicit intrinsic effects on either paradigm. These results indicate that GABA_B receptor signaling might exert differential effects on SWDs in DBA/2J mice.     

Decreased catalepsy response to haloperidol in the genetically dystonic (dt) rat

The ontogeny of petit mal-like seizures induced by gamma-hydroxybutyrate (GHB) was investigated. The prodrug of GHB, gamma-butyrolactone (GBL) was administered in varying dosages under continuous EEG monitoring from cortical and depth electrodes to rats varying in postnatal age from 1 to 85 days. The brain pharmacokinetics of GHB were determined at various ages as was the effect of ethosuximide on GBL-induced EEG changes. In adult rats, GBL produced a predictable sequence of electrical events beginning with spike bursts and progressing to polyspiking separated by low voltage activity. In 1-day-old rats, GBL produced voltage suppression with stupor. Poorly organized spiking appeared at postnatal day 3 and by day 9 marked burst suppression with polyspiking separated by low voltage activity was noted. However, the full array of electrical events seen in adult rats did not appear until day 28. Ethosuximide was ineffective against GHB seizure until the third postnatal week of life. GHB had a longer half-life in brain in the first week of postnatal life. These data suggest that in the rodent, petit mal-like seizure activity may require a fully mature brain and raise the possibility of different mechanisms being responsible for the various stages of EEG changes induced by GBL.     

Low doses of gamma-hydroxybutyric acid stimulate the firing rate of dopaminergic neurons in unanesthetized rats.

In unanesthetized rats the intravenous (i.v.) administration of gamma-hydroxybutyric acid (GHB) at the doses of 50-400 mg/kg produced a dose-related stimulation (10-56%) of the firing rate of dopaminergic (DA) neurons in the pars compacta of the substantia nigra. Doses of 1000 and 1500 mg/kg inhibited the firing rate almost completely. In unanesthetized rats the intraperitoneal injection of GHB at the dose of 750 mg/kg produced a brief initial stimulation (23%) followed by a modest reduction in the firing rate (29%). On the other hand, in chloral hydrate-anesthetized rats the i.v. administration of GHB at cumulative doses of up to 200 mg/kg failed to modify the firing rate of DA neurons, while a cumulative dose of 400 mg/kg suppressed neuronal firing. The results indicate that sub-anesthetic doses of GHB stimulate the firing rate of DA neurons in unanesthetized rats.     

Gamma-butyrolactone (GBL) disruption of passive avoidance learning in the day-old chick appears to be due to its effect on GABAB not gamma-hydroxybutyric [corrected] acid (GHB) receptors

Gamma-butyrolactone (GBL) is a prodrug to gamma-hydroxybutyric acid (GHB) and metabolises to GHB when ingested. Discrimination stimulus studies report generalisation of effects of GHB to GBL. While amnesia is one of the most commonly reported symptoms of GHB's ingestion in human users, as yet few studies have examined this effect. Although an endogenous GHB specific receptor is present in the brain, several studies have indicated that the clinical effects of exogenous doses of GBL/GHB are due to its action on GABA(B) receptors rather than on the GHB receptor. In this series of studies, New Hampshire x White leghorn cockerels were trained using a modified version of the passive avoidance learning task. Subcutaneous injections of GBL induced a memory deficit by 10 min post-training, which persisted for at least 24 h. No effect on memory was seen with administration of the specific GHB agonist NCS-356 (gamma-p-chlorophenyl-trans-4-hydroxycrotonate). The GBL-induced memory deficit appeared similar to the deficit produced by baclofen, where the antagonist facilitated learning. Additionally, GBL-induced memory deficit was ameliorated by application of a GABA(B) antagonist. The results support the hypothesis that GBL exerts its influence on memory via the GABA(B) receptor rather than by the specific GHB receptor.     

Specific gamma-hydroxybutyrate-binding sites but loss of pharmacological effects of gamma-hydroxybutyrate in GABA(B)(1)-deficient mice

gamma-Hydroxybutyrate (GHB), a metabolite of gamma-aminobutyric acid (GABA), is proposed to function as a neurotransmitter or neuromodulator. gamma-Hydroxybutyrate and its prodrug, gamma-butyrolactone (GBL), recently received increased public attention as they emerged as popular drugs of abuse. The actions of GHB/GBL are believed to be mediated by GABAB and/or specific GHB receptors, the latter corresponding to high-affinity [3H]GHB-binding sites coupled to G-proteins. To investigate the contribution of GABAB receptors to GHB actions we studied the effects of GHB in GABAB(1)-/- mice, which lack functional GABAB receptors. Autoradiography reveals a similar spatial distribution of [3H]GHB-binding sites in brains of GABAB(1)-/- and wild-type mice. The maximal number of binding sites and the KD values for the putative GHB antagonist [3H]6,7,8,9-tetrahydro-5-hydroxy-5H-benzocyclohept-6-ylidene acetic acid (NCS-382) appear unchanged in GABAB(1)-/- compared with wild-type mice, demonstrating that GHB- are distinct from GABAB-binding sites. In the presence of the GABAB receptor positive modulator 2,6-di-tert-butyl-4-(3-hydroxy-2,2-dimethyl-propyl)-phenol GHB induced functional GTPgamma[35S] responses in brain membrane preparations from wild-type but not GABAB(1)-/- mice. The GTPgamma[35S] responses in wild-type mice were blocked by the GABAB antagonist [3-[[1-(S)-(3,4dichlorophenyl)ethyl]amino]-2-(S)-hydroxy-propyl]-cyclohexylmethyl phosphinic acid hydrochloride (CGP54626) but not by NCS-382. Altogether, these findings suggest that the GHB-induced GTPgamma[35S] responses are mediated by GABAB receptors. Following GHB or GBL application, GABAB(1)-/- mice showed neither the hypolocomotion, hypothermia, increase in striatal dopamine synthesis nor electroencephalogram delta-wave induction seen in wild-type mice. It, therefore, appears that all studied GHB effects are GABAB receptor dependent. The molecular nature and the signalling properties of the specific [3H]GHB-binding sites remain elusive.     

Unusual presentation of a patient with GBL withdrawal: a case report

GBL (gamma-butyro-lactone) is converted to Gamma hydroxyl butyrate (GHB) in the body. GBL and GHB are available in liquid form and powder form. Once categorised under "legal highs", these two are not associated with any dependence or withdrawal in animal studies. But there are case reports indicating their high dependence potential in humans. We here present a case of a 29 year old who came to the attention of psychiatric services with very bizarre presentation and needed a host of investigations and expert views from various medical disciplines. He was treated mainly symptomatically followed by a sudden dramatic recovery on the 11th day after presentation. GBL is getting popular as a recreational drug and its withdrawal should be seriously considered in the list of medical causes leading to Delirium.     

Enhanced Anticonvulsant Activity of Neuroactive Steroids in a Rat Model of Catamenial Epilepsy

PURPOSE: Perimenstrual catamenial epilepsy may in part be due to withdrawal of the endogenous progesterone-derived neurosteroid allopregnanolone that potentiates gamma-aminobutyric acidA (GABA(A)) receptor-mediated inhibition. Here we sought to determine whether the anticonvulsant potencies of neuroactive steroids, benzodiazepines, phenobarbital (PB), and valproate (VPA) are altered during the heightened seizure susceptibility accompanying neurosteroid withdrawal in a rat model of perimenstrual catamenial epilepsy. METHODS: Test drugs were evaluated for their ability to alter the convulsant activity of pentylenetetrazol (PTZ) in young adult female rats, in pseudopregnant rats with prolonged exposure to high levels of progesterone (and its neurosteroid metabolites), and in pseudopregnant rats 24 h after acute withdrawal of neurosteroids by treatment with the 5alpha-reductase inhibitor finasteride. Test drugs were administered at doses equivalent to twice their ED50 values for protection against PTZ-induced clonic seizures in naive young adult female rats. RESULTS: The anticonvulsant activity of allopregnanolone (5 mg/kg, s.c.), pregnanolone (5 mg/kg, s.c.), allotetrahydrodeoxycorticosterone (15 mg/kg, s.c.), and tetrahydrodeoxycorticosterone (10 mg/kg, s.c.) were enhanced by 34-127% after neurosteroid withdrawal. The anticonvulsant activity of PB (65 mg/kg, i.p.) was also enhanced by 24% in neurosteroid-withdrawn animals. In contrast, the anticonvulsant activity of diazepam (4 mg/kg, i.p.), bretazenil (0.106 mg/kg, i.p.), and VPA (560 mg/kg, i.p.) were reduced or unchanged in neurosteroid-withdrawn animals. CONCLUSIONS: The anticonvulsant activity of neuroactive steroids is potentiated after neurosteroid withdrawal, supporting the use of such agents in the treatment of perimenstrual catamenial epilepsy.     

Effect of the synthetic polyamine N,N'-bis-(3-aminopropyl) cyclohexane-1,4-diamine (DCD) on rat spinal cord nociceptive transmission

In rats submitted to a C-fiber reflex response paradigm, intravenous (i.v.) administration of 2.5, 5 and 10 mg/kg of the synthetic polyamine N,N'-bis-(3-aminopropyl) cyclohexane-1,4-diamine (DCD) dose-dependently reduced both the integrated C reflex responses and wind-up activity. Inhibitory effects of the polyamine on spinal cord nociceptive transmission are likely to be consequence of blockade by extracellular DCD of NMDA receptor channels localized in dorsal horn neurons, although modulatory actions at supraspinal level and at other ion channels could also be possible.     

Variety of symptoms after drug use of gamma-hydroxybutyric acid (GHB)

Gamma-hydroxybutyric acid (GHB, "liquid ecstasy") and its legal prodrugs gamma-butyrolactone and 1,4-butanediol are gaining importance as recreational drugs in Germany. Because of the wide availability of GHB and its prodrugs physicians are increasingly being confronted with cases of intoxication. The effect of GHB intoxication is comparable with those of alcohol and/or benzodiazepines. Likewise, symptoms of withdrawal may occur. In this review, we summarise current data regarding the history, pharmacodynamics and pharmacokinetics of the drug as well as the relevant symptoms of intoxication or withdrawal as they pertain to neurology and psychiatry.     

Prolonged effects of repeated cocaine on medial prefrontal cortex dopamine response to cocaine and a stressful predatory odor challenge in rats

The present study examined the effects of seven daily saline (1 ml/kg, i.p.) or cocaine injections (15 mg/kg, i.p.) on extracellular dopamine levels in the medial prefrontal cortex (mPFC) after challenge with cocaine or stressful predatory odor presentation given 1 week (early withdrawal) or 3 weeks later (late withdrawal). Cocaine challenge at early withdrawal produced an increase in dopamine levels that was temporally shifted so that maximal levels of dopamine were significantly higher and attained 20 min earlier in the cocaine-pretreated group (maximal levels of saline controls=378% increase, cocaine=494% increase above baseline). Cocaine challenge at late withdrawal produced a similar effect on the temporal shift of maximal dopamine levels, with a significantly higher maximal percent increase of dopamine in cocaine-pretreated rats (saline-pretreated=420% increase, cocaine-pretreated=515% increase). Challenge with TMT, a predatory odor from fox that produces a stress response in rats, produced a maximal 75-200% increase in basal dopamine levels in both groups at both early and late withdrawal times. As with cocaine challenge, daily cocaine produced a leftward shift in the time at which maximal dopamine levels were attained in response to TMT. Cocaine-pretreated animals demonstrated maximal dopamine levels 40-80 min after TMT removal, while saline-pretreated rats showed maximal levels 100-140 min after TMT removal. These results suggest that there are long-term changes in the mPFC dopamine response to subsequent challenge with cocaine as well as a stressful predatory odor. The altered response of mPFC dopamine after repeated daily cocaine may impact relapse to drug-seeking or drug-taking behavior.     

Psychiatric aspects of acute withdrawal from gamma-hydroxybutyrate (GHB) and its analogue gamma-butyrolactone (GBL): implications for psychiatry services in the general hospital

Abstract

Objective. The objective of this study was to describe the psychiatric symptoms, management and outcomes in a consecutive series of patients being managed medically for symptoms of withdrawal from gamma-hydroxybutyrate (GHB) and its analogue gamma-butyrolactone (GBL) in a general hospital setting. Methods. A toxicology database was used to identify patients presenting with a history suggestive of withdrawal from GHB and analogues. Electronic and paper medical records were searched for demographic features, neuropsychiatric symptoms, psychiatric management while in hospital and overall outcome. Results. There were 31 presentations with withdrawal from the drugs involving 20 patients. Of these 17 (54%) were referred to and seen by the liaison psychiatry team. Anxiety (61.3%) and agitation (48.4%) were the most common symptoms. Of the 17 cases seen by the liaison psychiatry team, 52.9% required close constant observation by a mental health nurse and 29.4% required to be detained in hospital under mental health legislation. Conclusions. The significant proportion of patients presenting with neuropsychiatric symptoms and requiring intensive input from the liaison psychiatry team during withdrawal from GHB and its analogues points to the importance of close liaison between medical and psychiatric teams in managing these patients in the general hospital.     

Consequences of amygdala kindling and repeated withdrawal from ethanol on amphetamine-induced behaviours

It has been shown previously that chronic ethanol treatment in mice leads to accelerated behavioural sensitization to psychomotor stimulants [Manley & Little (1997) J. Pharmacol. Exp. Ther., 281, 1330-1339], whilst repeated experience of ethanol withdrawal sensitizes pathways underlying seizure activity (Becker & Hale (1993) Alcohol Clin. Exp. Res., 17, 94-98]. The aim of the current experiment was to investigate the consequences of repeated withdrawal from ethanol on amphetamine-induced behaviours in the rat and compare this with animals with electrical kindling of the amygdala, a procedure that has been shown to enhance alcohol withdrawal seizures [Pinel et al. (1975) Can. J. Neurol. Sci., 2, 467-475]. For the kindling experiments, electrodes were surgically implanted in the left basolateral amygdala and were stimulated daily at the afterdischarge threshold until a criterion of three consecutive stage 5 seizures was reached. Fully kindled rats showed a marginally significant reduction in sensitivity to the locomotor stimulant effects of acute amphetamine compared with sham and partially kindled rats which had experienced subthreshold stimulation of the amygdala. Sham and partially kindled rats sensitized readily to the locomotor activating effects of amphetamine (0.125 mg/kg) following repeated treatments, but the fully kindled rats did not. Fully kindled rats also failed to show place preference conditioning to amphetamine (0.5 mg/kg). Rats, withdrawn three times from chronic ethanol (liquid-diet), kindled more quickly to PTZ (30 mg/kg, i.p.) than rats with the same overall exposure to ethanol (24 days) followed by a single withdrawal or control animals. However, there was no difference in the locomotor stimulating effects of acute amphetamine (0.25-1 mg/kg, i.p.), the rate of sensitization to amphetamine (0.125 mg/kg, i.p.) or amphetamine induced conditioned place preference (1 mg/kg, i.p.). These observations suggest that, in rats, repeated withdrawal from a relatively mild chronic ethanol treatment modulates neuronal systems that may also be involved in PTZ-induced kindling but not those involved in either the acute stimulant effects of amphetamine or behavioural sensitization or appetitive conditioning following repeated amphetamine administration. Behavioural changes following amygdala kindling differed from those following repeated ethanol withdrawal, suggesting that withdrawal kindling from a mild ethanol treatment differs in its effects from amygdala kindling.     

The psychoactive ingredient of marijuana induces behavioural sensitization

Here we describe, for the first time, the occurrence of behavioural sensitization after chronic exposure to Delta9-tetrahydrocannabinol. Rats were treated twice a day, for five days, with increasing doses (5, 10, 20, 40, 40 mg/kg i.p.) of Delta9-tetrahydrocannabinol or its vehicle and after 20 days of withdrawal, animals were challenged with 5 mg/kg (i.p.) of the drug and their behaviour was assessed. Contrary to the motor inhibition induced in control rats, challenge with Delta9-tetrahydrocannabinol in pre-exposed animals elicited a complex behavioural syndrome mainly characterized by oral stereotyped items. Due to the relevance of behavioural sensitization in drug-seeking behaviour that persists long after discontinuation of drug use, our findings suggest that cannabinoids could trigger neurobiological alteration not dissimilar from those observed with more harmful abused drugs.     

Early administration of entacapone prevents levodopa-induced motor fluctuations in hemiparkinsonian rats

The purpose of this study was to investigate the effect of the catechol-O-methyltransferase (COMT) inhibitor, entacapone, in the reversal and prevention of "wearing-off" phenomena in hemiparkinsonian rats. Catechol-O-methyltransferase (COMT) inhibitors increase the half-life and bioavailability of levodopa, providing more continuous dopamine receptor stimulation. This raises the possibility of using levodopa and a COMT inhibitor not only to treat motor complications, but also to prevent their development. Male Sprague-Dawley rats received a unilateral 6-hydroxydopamine (6-OHDA) administration in the nigrostriatal pathway. Two sets of experiments were performed. First, animals were treated with levodopa (50 mg/kg/day with benserazide 12.5 mg/kg/day, twice daily (b.i.d.), intraperitoneally (i.p.) for 22 days. On day 23, animals received either entacapone (30 mg/kg, i.p.) or vehicle with each levodopa dose. In the second set, animals were treated either with levodopa (50 mg/kg/day, i.p.) plus entacapone (30 mg/kg/day, i.p.) or levodopa (50 mg/kg/day, i.p.) plus vehicle, administered two or three times daily [b.i.d. or thrice daily (t.i.d.), respectively] for 22 consecutive days. Entacapone both reversed and prevented the shortening of the motor response duration that defines "wearing-off" motor fluctuations. Entacapone also decreased the frequency of failures to levodopa. The combination of levodopa and entacapone may reduce the likelihood of motor fluctuation development and may thus become a valuable approach to treat Parkinson disease whenever levodopa is needed.     

Tolerance to the antinociceptive effects of ethanol during ethanol withdrawal

Prior research has indicated that tolerance develops to the antinociceptive effects of ethanol and continues even during withdrawal. Three potential pharmacological mechanisms for this tolerance are examined, using nitrendipine (L-type calcium channel blocker), theophylline (adenosine A1/A2 antagonist) and flumazenil (benzodiazepine antagonist). Rats received 10 days of exposure to an ethanol-containing liquid diet (6.5% w/v). A radiant heat tail-flick assay was used to assess hyperalgesia at 12 h after removal of the liquid diet, as well as tolerance to the effects of cumulative doses of ethanol (0.5-2 g/kg). Co-administration of flumazenil (10 mg/kg, i.p., b.i.d.), nitrendipine (5 mg/kg, i.p., b.i.d.) or theophylline (1 mg/kg, i.p., b.i.d.) with chronic ethanol prevented development of the hyperalgesia produced by ethanol withdrawal, but only theophylline reduced tolerance to the antinociceptive effects of ethanol administered during ethanol withdrawal. In contrast, when administered during ethanol withdrawal, theophylline (1-10 mg/kg) blocked the anti-hyperalgesic effects of ethanol during ethanol withdrawal, whereas nitrendipine (5-25 mg/kg) enabled ethanol to produce levels of antinociception comparable to non-dependent rats. These findings indicate that L-type calcium channels and adenosine receptors play important, but differing roles in the development of hyperalgesia during withdrawal, and to tolerance to the antinociceptive effects of ethanol.