The Ontogeny of GABAergic Enhancement of the γ-Hydroxybutyrate Model of Generalized Absence Seizures

The ontogeny of GABAergic enhancement of generalized absence seizures was examined in the gamma-hydroxybutyrate (GHB) model of generalized absence seizures. The GHB seizure was quantitated in developing and adult rats in the presence of varying doses of the GABAa agonist muscimol or intracerebroventricularly (i.c.v.) administered GABA. Both GABA and muscimol potentiated GHB-induced seizures in an age-dependent fashion. The adult dose of 1 mg/kg muscimol was extremely potent in rats less than 28 days of age and resulted in the death of all younger animals tested secondary to profound hypothermia. A dose of 0.1 mg/kg muscimol was associated with a significant prolongation of GHB seizure in rats less than 35 days of age, but had no effect on older animals. The response to GHB was also age dependent, with the greatest sensitivity occurring during the fourth and fifth week of life. The developmental sensitivity of the rat to GHB seizure correlated with enhancement of the seizure by muscimol and GABA, and both phenomena parallel the maturation of thalamocortical recruiting mechanisms thought to play a role in the pathogenesis of the bilaterally synchronous spike wave discharges that characterize generalized absence seizures.     

Effect of gamma-crotonolactone in experimental models of generalized absence seizures

The effect of the unsaturated lactone, gamma-crotonolactone (GCL), against spike wave discharges (SWD) in two pharmacological models of generalized absence seizures in rat was investigated. The models used were the gamma-hydroxybutyrate (GHB) model and the low dose pentylenetetrazole model. GCL pretreatment resulted in a significant decrease in duration of SWD in both models. In addition, this compound reduced the ability of the agonist, muscimol, to enhance SWD duration in the GHB model and was effective in developing animals as well as in adult animals with GHB-induced SWD. These data suggest that GCL may be a useful tool in the study of basic mechanisms of SWD generation in experimental absence seizures.     

gamma-Hydroxybutyric acid-induced seizures bear no relation to core temperature

The hypothesis that the absencelike seizures induced by gamma-hydroxybutyrate (GHB) are secondary to the effect of this drug on body temperature was tested using the prodrug of GHB, gamma-butyrolactone (GBL). Dosages of GBL less than 400 mg/kg produced a consistent profound hypothermia associated with bilaterally synchronous spike-wave discharges (SWD), whereas higher doses were associated with a more complex effect on core temperature associated with an EEG pattern of burst suppression. The threshold dose for the hypothermia and SWD was the same, but the temperature changes occurred later and lasted longer than the SWD induced by GHB. Rats aged less than 28 days were less sensitive to the hypothermia but more sensitive to the SWD produced by GHB than adult animals. The antiepileptic drug (AED) ethosuximide (ESM), known to attenuate GHB-induced SWD did so, but had no effect on the hypothermia, whereas GHB-induced hypothermia, but not SWD, was blocked by raising the ambient temperature from 26 degrees to 32 degrees C. These data do not support the hypothesis that GHB-induced absencelike seizure activity is a result of the hypothermia produced by this drug. Rather they suggest that the SWD and hypothermia are caused by separate, independent mechanisms.     

Gamma-hydroxybutyric acid-induced absence seizures in GluR2 null mutant mice

In this electrophysiological study, we examined the susceptibility of GluR2 mutant null mice to absence seizures in comparison with wild-type controls. The prodrug of (GHB), gamma-butyrolactone (GBL) was given systemically to induce the absence seizures. We also tested the severity and duration of the seizure activity in this model. The results showed that the latency from GBL administration to onset of seizure was significantly prolonged in GluR2(-/-) mice when compared to GluR2(+/+) mice. The duration of spike-and-wave discharges (SWD) was also significantly decreased in the GluR2(-/-) mice. Ninety minutes following GBL administration, wild-type animals continued to exhibit intermittent SWD bursts while GluR2(-/-) mice had returned to baseline. These data suggest that the GluR2 subunit may be involved in the initiation and maintenance of absence seizures induced by GBL.     

Alteration of GLUR2 expression in the rat brain following absence seizures induced by gamma-hydroxybutyric acid

We explored the involvement of the glutamate receptor subunit B (GluR2) in the mechanism of absence seizures induced by gamma-hydroxybutyric acid (GHB). The expression and distribution of GluR2 protein in rat brain were examined during and after GHB-induced absence seizures. The data indicate that GluR2 protein expression significantly decreases following the onset of absence seizures. The suppression of GluR2 expression was prolonged and it outlasted the duration of the continuous absence seizure activity. The alteration of GluR2 protein levels was accompanied by a re-distribution of GluR2 expression from laminae V to IV in cerebral cortex. We also analyzed the duration and latency of absence seizures induced by GHB 72 h following an initial GHB-induced absence seizure, a time when suppression of GluR2 protein was maximal. The second absence seizure was significantly more prolonged than the first. These data may indicate that the putative down-regulation of GluR2 following GHB-induced absence seizure could have contributed to the potentiation of subsequent seizures in animals. A related hypothesis posed by the data is that down-regulation of GluR2 is involved in the mechanisms of the maintenance of recurrent absence seizure activity once it is initiated and therefore, may contribute to the chronicity of seizures in absence epilepsy.     

Gamma hydroxybutyrate in the monkey. III. Effect of intravenous anticonvulsant drugs.

Monkeys were treated intravenously with various anticonvulsant drugs before and after the intravenous administration of gamma hydroxybutyrate (GHB). Continuous electroencephalographic (EEG) and temperature monitoring was performed throughout all experiments. The GHB-induced EEG changes were abolished by ethosuximide and clonazepam, marginally improved by diazepam, and unaffected by phenobarbital. The GHB-induced myoclonic jerks were abolished by ethosuximide, significantly improved by diazepam, and worsened by clonazepam. Phenobarbital was effective in diminishing the frequency of GHB-induced myoclonic jerks only when given prior to administration of GHB. The GHB-induced stupor was improved only by ethosuximide. The GHB model of petit mal seizures is quite specific for drugs used in this disorder. GHB may play a role in the pathogenesis of absence seizures in children.     

Ganaxolone,a selective,high-affinity steroid modulator of the γ-aminobutyric acid-A receptor,exacerbates seizures in animal models of absence

Ganaxolone (3α-hydroxy-3β-methyl-5α-pregnan-20-one) is a novel neurosteroid which has anticonvulsant properties in a number of seizure models as well as the ability to enhance function of the γ-aminobutyric acid-A (GABA_A) receptor complex via a neurosteroid binding site. The object of these experiments was to ascertain the efficacy of ganaxolone against absence seizures. Ganaxolone was assessed in the low-dose pentylenetetrazol (PTZ) and the γ-hydroxybutyric acid (GHB) model of absence seizures in rats. Ganaxolone pretreatment resulted in a significant prolongation of absence seizure in both the PTZ and GHB models. Further, ganaxolone in doses above 20 mg/kg alone produced bilaterally synchronous spike wave discharges (SWDs) associated with behavioral arrest. These data suggest that augmentation of GABA_A receptor complex function by neurosteroids has the potential to result in or exacerbate absence seizures.     

γ-Hydroxybutyric acid-induced absence seizures in GluR2 null mutant mice

In this electrophysiological study, we examined the susceptibility of GluR2 mutant null mice to absence seizures in comparison with wild-type controls. The prodrug of (GHB), γ-butyrolactone (GBL) was given systemically to induce the absence seizures. We also tested the severity and duration of the seizure activity in this model. The results showed that the latency from GBL administration to onset of seizure was significantly prolonged in GluR2^−/− mice when compared to GluR2^+/+ mice. The duration of spike-and-wave discharges (SWD) was also significantly decreased in the GluR2^−/− mice. Ninety minutes following GBL administration, wild-type animals continued to exhibit intermittent SWD bursts while GluR2^−/− mice had returned to baseline. These data suggest that the GluR2 subunit may be involved in the initiation and maintenance of absence seizures induced by GBL.     

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.     

Withdrawal syndrome from gamma-hydroxybutyric acid (GHB) and 1,4-butanediol (1,4-BD) in Sardinian alcohol-preferring rats

Gamma-hydroxybutyric acid (GHB) and its precursors, 1,4-butanediol (1,4-BD) and gamma-butyrolactone (GBL), are recreational drugs widely abused in the US, Europe and Australasia. A severe withdrawal syndrome from GHB, 1,4-BD and GBL has been increasingly documented over the last years, necessitating the development of a reliable animal model for investigations of potential therapeutic approaches. The present study describes the induction and occurrence of audiogenic seizures as a sign of withdrawal from GHB and 1,4-BD in selectively bred Sardinian alcohol-preferring (sP) rats, treated with escalating doses of GHB (1.5-3.5 g/kg, twice daily; i.g.) or 1,4-BD (500-1000 mg/kg, twice daily; i.g.) for 9 consecutive days. Acute administration of the selective GABA(B) receptor antagonist, SCH 50911, dramatically increased seizure occurrence. We propose that the inherent sensitivity of sP rats to different GHB-associated responses may have contributed to the unraveling of a phenomenon which was otherwise not recognizable in other rat strains.     

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.     

Muscarinic cholinergic mediation of the GH response to gamma-hydroxybutyric acid: neuroendocrine evidence in normal and parkinsonian subjects

We have recently reported that parkinsonian patients show a significant GH response to gamma-hydroxybutyric acid (GHB), but not to gamma-aminobutyric acid (GABA)-ergic drug administration. In order to establish whether muscarinic cholinergic receptors mediate the GH secretion induced by GHB, normal men and parkinsonian patients were tested with GHB both in the absence and in the presence of the anticholinergic agent, pirenzepine. Both normal controls and parkinsonian patients showed a significant serum GH rise in response to GHB (25 mg/kg body weight p.o.) even though a slightly, but significantly lower response was observed in parkinsonian patients. Pretreatment with pirenzepine (100 mg p.o. 2 h before GHB) completely suppressed the GHB-induced GH release in both normal controls and parkinsonian patients. These data indicate that a cholinergic mechanism mediates the GH response to GHB in normal men. In addition the data indicate that this pathway is preserved in the parkinsonian brain.     

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.     

Effects of alkyl-substituted gamma-butyrolactones and succinimides on the evoked and spontaneous activity of hippocampal slices in vitro

Alkyl-substituted and unsubstituted derivatives of gamma-butyrolactone (GBL), gamma-hydroxybutyrate (GHB), and succinimide were tested for their effects on the evoked potential (EP) and spontaneous activity of hippocampal slices incubated in vitro. When tested alone, only beta-ethyl-beta-methyl-GBL (beta-EMGBL) and tetramethylsuccinimide (TMSM) had any effect, producing greatly augmented EPs and spontaneous epileptiform discharges. The nonexcitatory drugs were also tested for the ability to block the excitation produced by beta-EMGBL, TMSM, and penicillin. The unsubstituted compounds had no effect on the augmented EP induced by any of the convulsants. The alpha-ethyl-alpha-methyl-substituted GBL (alpha-EMGBL) and ethosuximide (ESM), blocked the augmented EP induced by beta-EMGBL and TMSM but not that induced by penicillin. The spontaneous activity induced by all three excitatory agents was blocked by GHB. Unsubstituted succinimide had no effect on spontaneous activity. The spontaneous activity produced by beta-EMGBL and TMSM was blocked by alpha-EMGBL, but ESM blocked only that induced by TMSM. Neither alpha-EMGBL nor ESM blocked the spontaneous activity produced by penicillin. We conclude that the ability to block the EP more closely parallels in vivo anticonvulsant activity whereas effects on the spontaneous activity may be more related to neuronal depressant effects. The alkyl-substituted GBLs and succinimides had very similar, albeit not identical, effects on incubated hippocampal slices. That may indicate that they have a similar but not a common mechanism of action.     

The effects of coenzyme Q10 on seizures in mice: The involvement of nitric oxide

Coenzyme Q10 is a potent antioxidant in both mitochondria and lipid membranes. It has also been recognized to have an effect on gene expression. This study was designed to investigate whether acute or subchronic treatment with coenzyme Q10 altered the seizures induced by pentylenetetrazole or electroshock in mice. We also evaluated the involvement of nitric oxide in the effects of coenzyme Q10 in pentylenetetrazole-induced seizure models. Acute oral treatment with different doses of coenzyme Q10 did not affect the seizure in intraperitoneal pentylenetetrazole, intravenous pentylenetetrazole, and electroshock models in mice. Subchronic oral administration of coenzyme Q10 (100 mg/kg or more) increased time latencies to the onset of myoclonic jerks and clonic seizures induced by intraperitoneal pentylenetetrazole and at the doses of 25 mg/kg or more increased the seizure threshold induced by intravenous infusion of pentylenetetrazole. Subchronic doses of coenzyme Q10 (50 mg/kg or more) also decreased the incidence of tonic seizures in the electroshock-induced seizure model. Moreover, acute treatment with the precursor of nitric oxide synthesis, l-arginine (60 mg/kg), led to a significant potentiation of the antiseizure effects of subchronic administration of coenzyme Q10 (400 mg/kg in intraperitoneal and 6.25 mg/kg in intravenous pentylenetetrazole tests). Acute treatment with l-NAME (5 mg/kg), a nonspecific nitric oxide synthase inhibitor, significantly attenuated the antiseizure effects of subchronic doses of coenzyme Q10 in both seizure models induced by pentylenetetrazole. On the other hand, acute administration of aminoguanidine (100 mg/kg), a specific inducible nitric oxide synthase inhibitor, did not affect the seizures in mice treated with subchronic doses of coenzyme Q10 in both intraperitoneal and intravenous pentylenetetrazole tests. In conclusion, only subchronic and not acute administration of coenzyme Q10 attenuated seizures induced by pentylenetetrazole or electroshock. We also demonstrated, for the first time, the interaction between nitric oxide and coenzyme Q10 in antiseizure activity probably through the induction of constitutive nitric oxide synthase.     

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.     

Differences Between Two Feline Epilepsy Models in Sleep and Waking State Disorders, State Dependency of Seizures and Seizure Susceptibility: Amygdala Kindling Interferes with Systemic Penicillin Epilepsy

The objective of the study was to determine whether contemporary feline models of petit mal (systemic penicillin epilepsy) or temporal lobe epilepsy (amygdala kindling) resemble human seizure disorders with respect to disturbances of sleep and waking states, the state dependency of seizures, and transference of seizure susceptibility. These variables were examined in 6-h polygraphic recordings before and during exposure to both seizure models in 24 cats; 12 cats had intramuscular (i.m.) injections of 300,000 or 400,000 IU/kg of penicillin prior to kindling, and 12 were kindled before penicillin challenge. Results were as follows. First, penicillin increased light slow wave sleep (SWS) and drowsiness, during which spike-wave (SW) activity was maximal. Generalized tonic-clonic convulsions (GTCs) occurred predominantly in drowsiness after awakening from SWS. Second, kindling produced more deep SWS than did penicillin; susceptibility to kindled GTCs peaked during deep SWS, especially in transition to rapid eye movement sleep (REM). Third, penicillin did not influence subsequent sleep disorders or seizure susceptibility during kindling; kindling interfered with penicillin-induced GTCs, SW activity, and sleep disorders. Collectively, the findings suggest distinct state disorders and state-dependent seizure profiles in the two models. These differences parallel human analogues and may have contributed to the transference results. Kindling is a chronic model with persistent sleep and seizure abnormalities that differ from and may have discouraged penicillin epilepsy. Penicillin is an acute model with transient state and seizure disorders, a fact that may account for the absence of penicillin transference to kindling.     

The effect of acute aripiprazole treatment on chemically and electrically induced seizures in mice: The role of nitric oxide

Aripiprazole is an antipsychotic drug which acts through dopamine and serotonin receptors. Aripiprazole was noted to have antiseizure effects in a study on mice, while it induced seizures in a few human case reports. Dopaminergic and serotonergic systems relate to nitric oxide, and aripiprazole also has effects on dopamine and serotonin receptors. This study investigated the effects of aripiprazole on seizures and the potential role of nitric oxide in the process. The following three models were examined to explore the role of aripiprazole on seizures in mice: 1 — pentylenetetrazole administered intravenously, 2 — pentylenetetrazole administered intraperitoneally, and 3 — electroshock. Aripiprazole administration delayed clonic seizure in intravenous and intraperitoneal pentylenetetrazole models. In the electroshock-induced seizure model, tonic seizure and mortality protection percent were increased after aripiprazole administration. In intraperitoneal administration of pentylenetetrazole, aripiprazole effects on clonic seizure latency were significantly decreased when l-NAME — a nonselective nitric oxide synthase (NOS) inhibitor, 7-nitroindazole — a selective neuronal NOS (nNOS) inhibitor, or aminoguanidine — a selective inducible NOS (iNOS) inhibitor was injected before aripiprazole administration. In the intravenous pentylenetetrazole method, administration of l-NAME or aminoguanidine inhibited aripiprazole effects on clonic seizure threshold. Aminoguanidine or l-NAME administration decreased aripiprazole-induced protection against tonic seizures and death in the electroshock model. In both intravenous and intraperitoneal seizure models, aripiprazole and l-arginine coadministration delayed the onset of clonic seizures. Moreover, it increased protection against tonic seizures and death in intraperitoneal pentylenetetrazole and electroshock models. In conclusion, the release of nitric oxide via iNOS or nNOS may be involved in anticonvulsant properties of aripiprazole.     

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.