Pain attenuating actions of vincristinet-preconditioning in chemotherapeutic agent-induced neuropathic pain: key involvement of T-type calcium channels.

Preconditioning is a well-documented strategy that induces hepatic protection, renal protection, cardioprotection, and neuroprotection but its mechanism still remains to be elucidated. Hence, the present study investigated the protective mechanism underlying pain attenuating effects of vincristine-preconditioning in chemotherapeutic agent-induced neuropathic pain. Neuropathic pain was induced by administration of vincristine (50 µg/kg, i.p.) for 10 days in rats. Vincristine-preconditioning was induced by administration of vincristine (2, 5, and 10 µg/kg, i.p) for 5 days before administration of pain-inducing dose of vincristine (50 µg/kg, i.p.). Vincristine-preconditioning (10 µg/kg, i.p) for 5 days significantly reduced vincristine (50 µg/kg, i.p.) induced pain-related behaviors including paw cold allodynia, mechanical hyperalgesia, and heat hyperalgesia. However, vincristine (2 and 5 µg/kg, i.p) did not significantly ameliorate the vincristine (50 µg/kg, i.p.) induced neuropathic pain in rats. Furthermore, to explore the involvement of calcium channels in pain attenuating mechanism of vincristine-preconditioning, T-type calcium channel blocker, ethosuximide (100 and 200 mg/kg, i.p.) and L-type calcium channel blocker, amlodipine (5 and 10 mg/kg, i.p.) were used. Pretreatment with T-type calcium channel blocker, ethosuximide significantly abolished vincristine-preconditioning-induced protective effect. However, pretreatment with L-type calcium channel blocker, amlodipine did not alter vincristine-preconditioning-induced pain-related behaviors. This indicates that vincristine-preconditioning has protective effect on pain-related parameters due to opening of calcium channels, particularly T-type calcium channels that lead to entry of small magnitude of intracellular calcium through these channels and prevent the deleterious effects of high-dose vincristine.

Interactions of tiagabine with ethosuximide in the mouse pentylenetetrazole-induced seizure model: an isobolographic analysis for non-parallel dose-response relationship curves.

The aim of this study was to characterize the interaction between tiagabine (TGB) and ethosuximide (ETS), two antiepileptic drugs, in pentylenetetrazole (PTZ)-induced clonic seizures in mice using isobolographic analysis. The nature of the interaction between the drugs administered in combination was ascertained by estimating plasma and brain concentrations of ETS and TGB using fluorescence polarization immunoassay (FPIA) and high-performance liquid chromatography (HPLC). The results indicated that both drugs produced clear anticonvulsant effects against PTZ-induced clonic seizures in mice, but that their dose-response relationship curves (DRRCs) were not parallel, consequently necessitating the isobolographic analysis for non-parallel DRRCs. The isobolographic analysis revealed that the combination of TGB with ETS at the fixed-ratio of 1:1 exerted an additive interaction against PTZ-induced clonic seizures in mice. FPIA documented that TGB significantly elevated brain ETS concentrations (by 64%), while having no effect on plasma ETS concentrations in experimental animals. In contrast, ETS had no significant impact on plasma and brain concentrations of TGB in mice, as measured by HPLC. It can be concluded that the additive interaction between TGB and ETS at the fixed-ratio of 1:1 in the PTZ test was complicated by a significant pharmacokinetic increase in total brain ETS concentrations. At present, there are no recommendations to use this drug combination in epileptic patients.

Clinical pharmacokinetics of antiepileptic drugs in paediatric patients. Part I: Phenobarbital, primidone, valproic acid, ethosuximide and mesuximide.

This article reviews 119 papers published since 1964 on the pharmacokinetics of phenobarbital, primidone, valproic acid, ethosuximide and mesuximide (methsuximide) in paediatric patients. Particular attention has been paid to the role of age in determining the variability of pharmacokinetic parameters, but the effect of other factors, such as different formulations and routes of administration, concomitant treatments, gender and pathological conditions other than epilepsy, have also been considered. Mean phenobarbital terminal half-life (t1/2z) is very long in neonates (45 to 409 hours) and decreases with age. Therefore, a low dose per kilogram (dose/kg) is recommended during the neonatal period. The dose requirement decreases with increasing age, especially in children also taking valproic acid, which inhibits phenobarbital metabolism. Primidone is metabolised to phenobarbital and phenylethylmalonilamide; the metabolic conversion rate is increased by enzyme-inducing drugs and inversely correlated with age, being virtually absent in neonates. Valproic acid is extensively bound to plasma proteins, but there is a high interindividual and intraindividual diurnal variability in the binding, which depends on the concentration of binding proteins (i.e. albumin) and binding modulators (e.g. free fatty acids) but not on age (at least in those patients aged between 3 months and 65 years). The clearance (CL/F) of valproic acid positively correlates with the unbound concentrations and is strongly age-dependent, being low in neonates and high at the end of the first postnatal month, and progressively decreasing from 2 months to 14 years. The combination of these factors leads to a very poor correlation between plasma concentrations and dose/kg (C/D) and between plasma concentrations of total valproic acid and efficacy. Children also taking enzyme-inducing antiepileptic drugs require a larger valproic acid dose/kg, whereas the coadministration of aspirin (acetylsalicylic acid) may decrease the clearance of unbound drug (CLu/F), and thus require a decrease in the daily dose of valproic acid. Ethosuximide is well absorbed, minimally protein bound and slowly eliminated. Lower C/D ratios are reported in children younger than 10 years old than in older children and in individuals also taking enzyme-inducing drugs (i.e. primidone). According to the only available paper on mesuximide in paediatric patients, the C/D ratio is less sensitive to both age and associated therapy.

Clinically significant pharmacokinetic drug interactions with carbamazepine. An update.

Carbamazepine is one of the most commonly prescribed antiepileptic drugs and is also used in the treatment of trigeminal neuralgia and psychiatric disorders, particularly bipolar depression. Because of its widespread and long term use, carbamazepine is frequently prescribed in combination with other drugs, leading to the possibility of drug interactions. The most important interactions affecting carbamazepine pharmacokinetics are those resulting in induction or inhibition of its metabolism. Phenytoin, phenobarbital (phenobarbitone) and primidone accelerate the elimination of carbamazepine, probably by stimulating cytochrome P450 (CYP) 3A4, and reduce plasma carbamazepine concentrations to a clinically important extent. Inhibition of carbamazepine metabolism and elevation of plasma carbamazepine to potentially toxic concentrations can be caused by stiripentol, remacemide, acetazolamide, macrolide antibiotics, isoniazid, metronidazole, certain antidepressants, verapamil, diltiazem, cimetidine, danazol and (dextropropoxyphene) propoxyphene. In other cases, toxic symptoms may result from elevated plasma concentrations of the active metabolite carbamazepine-10,11-epoxide, due to the inhibition of epoxide hydrolase by valproic acid (sodium valproate), valpromide, valnoctamide and progabide. Carbamazepine is a potent inducer of CYP3A4 and other oxidative enzyme system in the liver, and it may also increase glucuronyltransferase activity. This results in the acceleration of the metabolism of concurrently prescribed anticonvulsants, particularly valproic acid, clonazepam, ethosuximide, lamotrigine, topiramate, tiagabine and remacemide. The metabolism of many other drugs such as tricyclic antidepressants, antipsychotics, steroid oral contraceptives, glucocorticoids, oral anticoagulants, cyclosporin, theophylline, chemotherapeutic agents and cardiovascular drugs can also be induced, leading to a number of clinically relevant drug interactions. Interactions with carbamazepine can usually be predicted on the basis of the pharmacological properties of the combined drug, particularly with respect to its therapeutic index, site of metabolism and ability to affect specific drug metabolising isoenzymes. Avoidance of unnecessary polypharmacy, selection of alternative agents with lower interaction potential, and careful dosage adjustments based on serum drug concentration monitoring and clinical observation represent the mainstays for the minimisation of risks associated with these interactions.

Influence of some novel N-substituted azoles and pyridines on rat hepatic CYP3A activity.

A series of N-substituted heteroaromatic compounds structurally related to clotrimazole was synthesized, and the effects of these compounds on ethosuximide clearance in rats were determined as a measure of their abilities to induce cytochrome P4503A (CYP3A) activity. Ethosuximide clearance and in vitro erythromycin N-demethylase activity were shown to correlate. In this series, imidazole or other related heteroaromatic "head groups" were linked to triphenylmethane or other phenylmethane derivatives. Within the series, it was found that 1-triphenylmethane-substituted imidazoles elicited the greatest increase in CYP3A activity, and that among the triphenylmethyl-substituted imidazoles, the highest activities were achieved by the substitution of F- or Cl- in either the meta or para position of one of the phenyl rings. Diphenylmethyl-substituted pyridine was effectively devoid of activity. Compounds eliciting the largest increase in CYP3A activity (viz. 1-[(3-fluorophenyl)diphenylmethyl]imidazole, 1-[(4-fluorophenyl)diphenylmethyl]imidazole, and 1-[tri-(4-fluorophenyl)methyl]imidazole) produced little or no increase in ethoxyresorufin O-dealkylase (EROD) activity (i.e. CYP1A), whereas benzylimidazole, which elicited only a small increase in CYP3A activity, produced an almost 9-fold increase in CYP1A activity. For a series of eleven compounds exhibiting a wide range of influence on CYP3A activity, a positive correlation was found between ethosuximide clearance and hepatic CYP3A mRNA levels.

Effect of positively and negatively charged liposomes on skin permeation of drugs.

To clarify the effect of the surface charge of liposomes on percutaneous absorption, the permeation of liposomal drugs through rat skin was investigated in vitro and in vivo. Liposomes were prepared using egg yolk lecithin (EPC, phase transition temperature, -15 to -17 degrees C), cholesterol and dicetylphosphate (DP) or stearylamine (SA) (10:1:1, mol/mol). Also examined was the penetration behavior of positively and negatively charged liposomes, using a fluorescent probe (Nile Red). The in vitro penetration rate of melatonin (MT) entrapped in negatively charged liposomes was higher than that of positively charged ones (p<0.05). When the percutaneous absorption of ethosuximide (ES) encapsulated was estimated in vivo, the absorption of ES from negatively charged liposomes was slightly higher than that from positively charged liposomes. Additionally, the absorption of ES from both types of liposomes was superior to that from the lipid mixtures consisting of the same composition as the vesicles. The percutaneous absorption of betahistine (BH) from a gel formulation containing negatively charged liposomes of BH was much more than that from the formulation with positively charged ones, with 2-fold higher AUC (p<0.05). Histological studies revealed that the negatively charged liposomes diffused to the dermis and the lower portion of hair follicles through the stratum corneum and the follicles much faster than the positive vesicles at the initial time stage after application. Thus, the rapid penetration of negatively charged liposomes would contribute to the increased permeation of drugs through the skin.

Interspecies scaling: predicting pharmacokinetic parameters of antiepileptic drugs in humans from animals with special emphasis on clearance.

The objective of this study was to test the interspecies-scaling approach in a series of antiepileptic drugs. Clearance, volume of distribution, and elimination half-life were scaled up from animal data obtained from literature. Four different methods were utilized to generate plots to scale up the clearance values: (i) clearance vs body weight (simple allometric equation); (ii) the product of clearance and maximum life-span potential (MLP) vs body weight (an approach recommended in literature); (iii) the two-term power equation which incorporates both body weight and brain weight suggested by Boxenbaum; and (iv) the product of clearance and brain weight vs body weight (a new approach being introduced in this study). When the predicted values for clearance were qualitatively compared with the observed values in humans, it was found that our proposed method predicted the clearance better than the other three methods. Using the simple allometric equation, the prediction of volume of distribution as a function of body weight was found to be satisfactory. The elimination half-life could not be predicted from simple allometric equations for any of the drugs studied; however, utilizing the equation CL = VK, prediction for half-life was feasible. The results of this study indicate that it is possible to predict reliably the pharmacokinetic parameters of these antiepileptic drugs in humans from animal data using an allometric approach.

Once-daily ethosuximide in the treatment of absence epilepsy.

Once-daily ethosuximide was used to treat 10 consecutive children with typical absence seizures. Three patients had gastrointestinal side effects which resolved when the same total daily dose was divided into 2 doses. Two other patients continued to have seizures on ethosuximide, whether given once or twice daily. Five patients had complete seizure control without adverse effects on once-daily ethosuximide.

Synergistic anticonvulsant effect of valproic acid and ethosuximide on pentylenetetrazole-induced epileptic phenomena in rats.

The possible synergistic effect of valproic acid and ethosuximide in combination on pentylenetetrazole-induced epilepsy was investigated in rats. Valproic acid and ethosuximide administered intraperitoneally both showed dose-dependent anti-epileptic activity towards pentylenetetrazole-induced myoclonias and tonic-clonic seizures. The valproic acid-ethosuximide combination had a synergistic pharmacological effect. Against myoclonias combined valproic acid-ethosuximide produced a non-significant decrease in the effective dose of both drugs compared with treatment with either drug alone. In the case of tonic-clonic seizures the protective effect against the seizures was significantly increased by combined treatment compared with treatment with either drug alone. Neither plasma concentrations nor any other pharmacokinetic parameters were significantly changed when the same doses of valproic acid and ethosuximide were given, singly or in combination.

Tissue distribution of ethosuximide and clobazam in a seizure related fatality.

The case of a six-year-old male who died in a hospital while receiving several anticonvulsant drugs is described. Phenytoin, desmethyldiazepam, clobazam (an experimental 1,5 benzodiazepine), and desmethylclobazam were quantitated in serum, liver, and brain tissue by high performance liquid chromatography. Ethosuximide was quantitated by gas chromatography. To our knowledge, this is one of few reports describing tissue concentrations of ethosuximide collected at autopsy and the first report of clobazam/desmethylclobazam tissue distribution in man.

Release and rectal absorption of ethosuximide from suppositories in rabbits.

Ethosuximide was formulated in different suppository bases. In vitro release experiment demonstrated more rapid and higher release of the drug from water soluble polyethylene glycol (PEG) bases than from Witepsol fatty bases. Rectal administration in PEG 400:4000 and 400:6000 to rabbits gave high plasma levels where Cmax was 45.66 and 42.66 micrograms.ml-1, respectively; while, in the presence of Witepsol E76 and W35 it was 34.00 and 28.33 micrograms.ml-1, respectively. The systemic availability was 89.39%, 82.72%, 58.80% and 47.45% when the bases were PEG 400:4000, PEG 400:6000, Witepsol E76 and Witepsol W35, respectively.

The metabolism of ethosuximide.

The metabolism of the antiepileptic drug ethosuximide (3-ethyl-3-methylpyrollidine-2,5-dione) (I) in animals and humans is reviewed. Chiral aspects of the metabolism of ethosuximide are discussed. Clarification of the precise nature of the hydroxymetabolites of ethosuximide is presented.

Evaluation of longevity enhancing compounds against transactive response DNA-binding protein-43 neuronal toxicity.

In simple systems, lifespan can be extended by various methods including dietary restriction, mutations in the insulin/insulin-like growth factor (IGF) pathway or mitochondria among other processes. It is widely held that the mechanisms that extend lifespan may be adapted for diminishing age-associated pathologies. We tested whether a number of compounds reported to extend lifespan in C. elegans could reduce age-dependent toxicity caused by mutant TAR DNA-binding protein-43 in C. elegans motor neurons. Only half of the compounds tested show protective properties against neurodegeneration, suggesting that extended lifespan is not a strong predictor for neuroprotective properties. We report here that resveratrol, rolipram, reserpine, trolox, propyl gallate, and ethosuximide protect against mutant TAR DNA-binding protein-43 neuronal toxicity. Finally, of all the compounds tested, only resveratrol required daf-16 and sir-2.1 for protection, and ethosuximide showed dependence on daf-16 for its activity.

Ethosuximide and phenytoin dose-dependently attenuate acute nonconvulsive seizures after traumatic brain injury in rats.

Acute seizures frequently occur following severe traumatic brain injury (TBI) and have been associated with poor patient prognosis. Silent or nonconvulsive seizures (NCS) manifest in the absence of motor convulsion, can only be detected via continuous electroencephalographic (EEG) recordings, and are often unidentified and untreated. Identification of effective anti-epileptic drugs (AED) against post-traumatic NCS remains crucial to improve neurological outcome. Here, we assessed the anti-seizure profile of ethosuximide (ETX, 12.5-187.5 mg/kg) and phenytoin (PHT, 5-30 mg/kg) in a spontaneously occurring NCS model associated with penetrating ballistic-like brain injury (PBBI). Rats were divided between two drug cohorts, PHT or ETX, and randomly assigned to one of four doses or vehicle within each cohort. Following PBBI, NCS were detected by continuous EEG monitoring for 72 h post-injury. Drug efficacy was evaluated on NCS parameters of incidence, frequency, episode duration, total duration, and onset latency. Both PHT and ETX attenuated NCS in a dose-dependent manner. In vehicle-treated animals, 69-73% experienced NCS (averaging 9-10 episodes/rat) with average onset of NCS occurring at 30 h post-injury. Compared with control treatment, the two highest PHT and ETX doses significantly reduced NCS incidence to 13-40%, reduced NCS frequency (1.8-6.2 episodes/rat), and delayed seizure onset: <20% of treated animals exhibited NCS within the first 48 h. NCS durations were also dose-dependently mitigated. For the first time, we demonstrate that ETX and PHT are effective against spontaneously occurring NCS following PBBI, and suggest that these AEDs may be effective at treating post-traumatic NCS.

Effects of fluoxetine on the anticonvulsant action of valproate and ethosuximide in mouse model of myoclonic convulsions.

Depression is becoming a growing problem in rural areas. This psychiatric disorder often accompanies epilepsy. The aim of this study was to assess the influence of fluoxetine (FXT), a commonly used antidepressant, on the protective action of two conventional antiepileptic drugs: ethosuximide (ETX) and valproate (VPA), against pentylenetetrazole (PTZ)-induced convulsions in mice. Motor coordination and long-term memory deficits induced by FXT, antiepileptic drugs alone and in combinations with FXT were assessed in the chimney test and passive-avoidance task, respectively. Brain concentrations of ETX and VPA were measured by immunofluorescence. Obtained results indicate that FXT at the dose of 15 mg/kg (i.p., 30 min before the test) significantly increased the threshold for clonic convulsions. The antidepressant drug at lower doses remained ineffective in this respect. Moreover, FXT at the highest subprotective dose (10 mg/kg, i.p.) markedly enhanced the anticonvulsant effects of VPA, but not of ETX, against PTZ-induced seizures. The interaction between FXT and VPA seems to be pharmacodynamic because the antidepressant drug did not alter the brain concentration of VPA. With regard to adverse effects, FXT, VPA, ETX, and the combinations of FXT with antiepileptic drugs, did not impair motor coordination and long-term memory in mice. In conclusion, the combination of FXT with VPA may be advantageous in the treatment of myoclonic epilepsy, and therefore it should be recommended for further study in clinical conditions.

Effects of WIN 55,212-2 mesylate (a synthetic cannabinoid) on the protective action of clonazepam, ethosuximide, phenobarbital and valproate against pentylenetetrazole-induced clonic seizures in mice.

The aim of this study was to determine the effect of WIN 55,212-2 mesylate (WIN - a non-selective cannabinoid CB1 and CB2 receptor agonist) on the protective action of four classical antiepileptic drugs (AEDs: clonazepam [CZP], ethosuximide [ETS], phenobarbital [PB], and valproate [VPA]) in the mouse pentylenetetrazole (PTZ)-induced clonic seizure model. WIN (15 mg/kg, i.p.) significantly enhanced the anticonvulsant action of ETS, PB and VPA, but not that of CZP against PTZ-induced clonic seizures. The ED(50) values of ETS, PB and VPA were reduced from 148.0, 13.9 and 137.1mg/kg to 104.0, 8.3 and 85.6 mg/kg, respectively (P<0.05). WIN (5 and 10mg/kg, i.p.) had no impact on the anticonvulsant action of all studied AEDs against PTZ-induced clonic seizures. WIN (15 mg/kg, i.p.) significantly elevated total brain concentrations of ETS and VPA, but not those of CZP and PB in mice. Moreover, WIN combined with CZP, ETS, PB and VPA significantly impaired motor performance, long-term memory and muscular strength in mice subjected to the chimney, passive avoidance and grip-strength tests, respectively. Pharmacodynamic enhancement of the anticonvulsant action of PB by WIN against PTZ-induced clonic seizures is favorable from a preclinical viewpoint. Advantageous effects of WIN in combination with ETS and VPA against PTZ-induced seizures were pharmacokinetic in nature. However, WIN combined with CZP, ETS, PB and VPA impaired motor coordination and long-term memory as well as reduced skeletal muscular strength in the experimental animals.

In vitro concentration dependent transport of phenytoin and phenobarbital, but not ethosuximide, by human P-glycoprotein.

AIMS: One possible mechanism for epilepsy drug resistance is overexpression of P-glycoprotein in the blood-brain barrier, but whether (or which) antiepileptic drugs (AEDs) are transported by P-gp remains unclear. We evaluated AEDs as P-gp substrates using cell monolayers. MAIN METHODS: Bi-directional transport assays and concentration equilibrium transport assays (CETAs) were performed for phenytoin (PHT), phenobarbital (PB), and ethosuximide (ESM) using wildtype Madin-Darby Canine Kidney II cell line MDCKII and porcine renal endothelial cell line LLC-PK1 cells and these cells transfected with human MDR1 cDNA to express P-gp. KEY FINDINGS: Wildtype cells demonstrated no efflux transport of PHT, PB, or ESM. In CETAs, both MDR1-transfected cell lines transported PHT from basolateral to apical when PHT loading concentrations were 5 or 10, but not 20microg/ml. MDCK-MDR1 cells transported PB when initial concentrations were 10 or 20, but not 5microg/ml. LLC-MDR1 did not transport PB. P-gp inhibitor verapamil blocked efflux transport. MDR1-transfected cells did not transport ESM at 5.6 or 56microg/ml. Bi-directional transport assays demonstrated weak transport for PHT but not PB or ESM. SIGNIFICANCE: Human P-gp transports PHT and PB, but not ESM, in a concentration dependent manner. CETA may be more sensitive than bi-directional assays to detect transport of drugs with high passive diffusion. Potential P-gp substrates should be tested at clinically relevant concentration ranges.

Ethosuximide: from bench to bedside.

Ethosuximide, 2-ethyl-2-methylsuccinimide, has been used extensively for "petit mal" seizures and it is a valuable agent in studies of absence epilepsy. In the treatment of epilepsy, ethosuximide has a narrow therapeutic profile. It is the drug of choice in the monotherapy or combination therapy of children with generalized absence (petit mal) epilepsy. Commonly observed side effects of ethosuximide are dose dependent and involve the gastrointestinal tract and central nervous system. Ethosuximide has been associated with a wide variety of idiosyncratic reactions and with hematopoietic adverse effects. Typical absence seizures are generated as a result of complex interactions between the thalamus and the cerebral cortex. This thalamocortical circuitry is under the control of several specific inhibitory and excitatory systems arising from the forebrain and brainstem. Corticothalamic rhythms are believed to be involved in the generation of spike-and-wave discharges that are the characteristic electroencephalographic signs of absence seizures. The spontaneous pacemaker oscillatory activity of thalamocortical circuitry involves low threshold T-type Ca2+ currents in the thalamus, and ethosuximide is presumed to reduce these low threshold T-type Ca2+ currents in thalamic neurons. Ethosuximide also decreases the persistent Na+ and Ca2+ -activated K+ currents in thalamic and layer V cortical pyramidal neurons. In addition, there is evidence that in a genetic absence epilepsy rat model ethosuximide reduces cortical gamma-aminobutyric acid (GABA) levels. Also, elevated glutamate levels in the primary motor cortex of rats with absence epilepsy (but not in normal animals) are reduced by ethosuximide.