Nimodipine restores the altered hippocampal phenytoin pharmacokinetics in a refractory epileptic model

doi:10.1016/j.neulet.2006.11.075

Neuroscience Letters

Volume 413, Issue 2, 14 February 2007, Pages 168-172

https://doi.org/10.1016/j.neulet.2006.11.075 Get rights and content

Abstract

The present work was undertaken to examine the central pharmacokinetics of phenytoin (PHT) in an experimental model of epilepsy, induced by administration of 3-mercaptopropionic acid (MP), and possible participation of P-glycoprotein in this model of epilepsy. Repeated seizures were induced in male Wistar rats by injection of 3-MP (45 mg kg⁻¹, i.p.) during 10 days. Control rats (C) were injected with saline solution. In order to monitor extracellular PHT levels, either a shunt microdialysis probe or a concentric probe was inserted into carotid artery or hippocampus, respectively. All animals were administered with PHT (30 mg kg⁻¹, i.v.) 30 min after intraperitoneal administration of vehicle (V) or nimodipine (NIMO, 2 mg kg⁻¹). No differences were found in PHT plasma levels comparing all experimental groups. In pre-treated rats with V, hippocampal PHT concentrations were lower in MP (maximal concentration, C_max: 2.7 ± 0.3 μg ml⁻¹, p < 0.05 versus C rats) than in C animals (C_max: 5.3 ± 0.9 μg ml⁻¹). Control rats pre-treated with NIMO showed similar results (C_max: 4.5 ± 0.8 μg ml⁻¹) than those pre-treated with V. NIMO pre-treatment of MP rats showed higher PHT concentrations (C_max: 6.8 ± 1.0 μg ml⁻¹, p < 0.05) when compared with V pre-treated MP group. Our results indicate that central pharmacokinetics of PHT is altered in MP epileptic rats. The effect of NIMO on hippocampal concentrations of PHT suggests that P-glycoprotein has a role in reduced central bioavailability of PHT in our epileptic refractory model.

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Acknowledgements

This work was supported by grants of UBACYT M033 from Universidad de Buenos Aires, and CONICET (PIP No. 02267 and PIP 5798) from the Consejo Nacional de Investigaciones Científicas y Técnicas.

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Drug-resistant epilepsy affects approximately one-third of the patients with epilepsy. The pharmacoresistant condition in epilepsy is mainly explained by six hypotheses. In addition, several experimental models have been used to understand the mechanisms involved in pharmacoresistant epilepsy and to identify novel therapies to control this condition. However, the global prevalence of this disease persists without changes. Several factors can explain this situation. First of all, the pharmacoresistant epilepsy is explained by different and independent hypotheses. Each hypothesis indicates specific mechanisms to explain the drug-resistant condition in epilepsy. However, there are different findings suggesting common mechanisms between the different hypotheses. Other important situation is that the experimental models designed for the screening of drugs with potential anticonvulsant effect do not consider factors such as age, gender, type of epilepsy, and comorbid disorders. The present review focuses on indicating the limitations for each hypothesis and the relationships among them. The relevance to consider central and peripheral phenomena associated with the drug-resistant condition in different types of epilepsy is also indicated. The necessity to establish a global hypothesis that integrates all the phenomena associated with the pharmacoresistant epilepsy is proposed.
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Despite the approval of a considerable number of last generation antiepileptic drugs (AEDs) (only in the last decade, six drugs have gained Food and Drug Administration approval), the global figures of seizure control have seemingly not improved, and available AED can still be regarded as symptomatic treatments. Fresh thinking in AEDs drug discovery, including the development of drugs with novel mechanisms of action, is required to achieve truly innovative antiepileptic medications.
The transporter hypothesis proposes that inadequate penetration of AEDs across the blood–brain barrier, caused by increased expression of efflux transporters such as P-glycoprotein (P-gp), contributes to drug-resistant epilepsy. Neuroinflammation due to high levels of glutamate has been identified as one of the causes of P-gp upregulation, and several studies in animal models of epilepsy suggest that antiinflammatory drugs might prevent P-gp overexpression and, thus, avoid the development of refractory epilepsy.
We have applied ligand-based in silico screening to select compounds that exert dual anticonvulsant and antiinflammatory effects. Five of the hits were tested in animal models of seizure, with protective effects. Later, two of them (sebacic acid (SA) and gamma-decanolactone) were submitted to the recently described MP23 model of drug-resistant seizures. All in all, SA displayed the best profile, showing activity in the maximal electroshock seizure (MES) and pentylenetetrazol (PTZ) seizure models, and reversing resistance to phenytoin (PHT) and decreasing the P-gp upregulation in the MP23 model. Furthermore, pretreatment with SA in the pilocarpine status epilepticus (SE) model resulted in decreased histamine release in comparison with nontreated animals.
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Citation Excerpt :
However, the repetitive induction of generalized seizures with MPA results in Pgp overexpression and reduces brain bioavailability and anticonvulsant effects of phenytoin [18,27]. This PHT-resistant phenotype is reversed by the pharmacological inhibition of Pgp [19,20,22,23,26]. This experiment was designed to test the hypothesis that TFS can inhibit the drug resistance condition induced by the repetitive MPA administration, thus enhancing the anticonvulsant effects of PHT.
Transcranial focal stimulation (TFS) is a noninvasive neuromodulation strategy that reduces seizure activity in different experimental models. Nevertheless, there is no information about the effects of TFS in the drug-resistant phenotype associated with P-glycoprotein (Pgp) overexpression. The present study focused on determining the effects of TFS on Pgp expression after an acute seizure induced by 3-mercaptopropionic acid (MPA). P-glycoprotein expression was analyzed by western blot in the cerebral cortex and hippocampus of rats receiving 5 min of TFS (300 Hz, 50 mA, 200 μs, biphasic charge-balanced squared pulses) using a tripolar concentric ring electrode (TCRE) prior to administration of a single dose of MPA. An acute administration of MPA induced Pgp overexpression in cortex (68 ± 13.4%, p < 0.05 vs the control group) and hippocampus (48.5 ± 14%, p < 0.05, vs the control group). This effect was avoided when TFS was applied prior to MPA. We also investigated if TFS augments the effects of phenytoin in an experimental model of drug-resistant seizures induced by repetitive MPA administration. Animals with MPA-induced drug-resistant seizures received TFS alone or associated with phenytoin (75 mg/kg, i.p.). TFS alone did not modify the expression of the drug-resistant seizures. However, TFS combined with phenytoin reduced seizure intensity, an effect associated with a lower prevalence of major seizures (50%, p = 0.03 vs phenytoin alone). Our experiments demonstrated that TFS avoids the Pgp overexpression induced after an acute convulsive seizure. In addition, TFS augments the phenytoin effects in an experimental model of drug-resistant seizures. According with these results, it is indicated that TFS may represent a new neuromodulatory strategy to revert the drug-resistant phenotype.
ABC transporters in drug-resistant epilepsy: mechanisms of upregulation and therapeutic approaches
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Animal models are often employed not only to investigate ABCB1 activity and substrates brain penetration, but also to assess the brain penetration of AEDs and the associated variation in seizure frequency and severity. In order to explore this question, Höcht et al. investigated the influence of nimodipine (an ABCB1 inhibitor) on the brain uptake of phenytoin in epileptic rats [72]. The experimental and control groups were either injected with nimodipine or vehicle.
Drug-resistant epilepsy (DRE) affects approximately one third of epileptic patients. Among various theories that try to explain multidrug resistance, the transporter hypothesis is the most extensively studied. Accordingly, the overexpression of efflux transporters in the blood-brain barrier (BBB), mainly from the ATP binding cassette (ABC) superfamily, may be responsible for hampering the access of antiepileptic drugs into the brain. P-glycoprotein and other efflux transporters are known to be upregulated in endothelial cells, astrocytes and neurons of the neurovascular unit, a functional barrier critically involved in the brain penetration of drugs. Inflammation and oxidative stress involved in the pathophysiology of epilepsy together with uncontrolled recurrent seizures, drug-associated induction and genetic polymorphisms are among the possible causes of ABC transporters overexpression in DRE.
The aforementioned pathological mechanisms will be herein discussed together with the multiple strategies to overcome the activity of efflux transporters in the BBB – from direct transporters inhibition to down-regulation of gene expression resorting to RNA interference (RNAi), or by targeting key modulators of inflammation and seizure-mediated signalling.
High incidence of persistent subtherapeutic levels of the most common AEDs in children with epilepsy receiving polytherapy
2018, Epilepsy Research
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In our experience, in some individualized cases treatment with nimodipine 2 mg/kg was able to reverse the pharmaco-resistant phenotype (Lazarowski et al., 2006). This drug, at equivalent doses restored the normal phenytoin pharmacokinetics in the hippocampus of a refractory epilepsy model and avoided new seizures (Hocht et al., 2007). Furthermore, the pharmacokinetic hypothesis suggests that decreases in AED plasma levels observed in several cases of patients with refractory epilepsy could be secondary to an overexpression of efflux transporters in peripheral organs, such as the intestine, liver, and kidney, thereby reducing the access of AEDs to the epileptic focus in the brain (Lazarowski et al., 2007).
Low levels of AEDs can be secondary drug–drug interactions or related to irregular intake due to poor treatment adherence. This latter behavior is highly suspected in ambulatory pediatric epileptic patients when controls of AEDs are subtherapeutic. However, it cannot be considered for inpatients during long periods of hospitalization. A few isolated case reports have documented persistent low levels (PLL) of AEDs in hospitalized epileptic children, but no population study has currently been reported.
The aim of this study was to document the incidence of PLL of the most common AEDs – phenytoin (PHT), phenobarbital (PHB), valproic acid (VA), and carbamazepine (CBZ) – in pediatric epileptic in- and outpatients (PEP).
21,040 plasma levels of the aforementioned AEDs from 3279 PEP were retrospectively analyzed. Plasma levels of AEDs were measured by an automated method using trademarked commercial kits with their corresponding quality control programs. Randomized samples were also controlled by HPLC methods. Only cases with more than 3 controls were included in the study.
A high rate of PLL of PHT was detected in in- (71.7%) and outpatients (74.1%), while PLL of PHB, VA, and CBZ were detected in a lower proportion. Rates of PLL of PHT were similar in in- and outpatients. PLL of PHB was more commonly observed in outpatients while PLL of VA and CBZ were more frequently seen in inpatients. In some hospitalized patients receiving polytherapy, PLL of at least one AED were documented during a long time.
Treatment non-adherence could be present in part of the outpatients, but cannot explain the PLL observed in a group of inpatients as described here. The recently described “pharmacokinetic hypothesis” of pharmaco-resistant epilepsy should be addressed in cases with AEDs-PLL, particularly in hospitalized cases. Perhaps, instead of stopping the subtherapeutic medication, the increasing doses of this AED and/or administration of inhibitors of CYP and P-glycoprotein, could help to achieve its therapeutic range, allowing a better pharmacologic effect and avoiding the development of more severe complications, such as status epilepticus or SUDEP.
Flavonoid compounds as reversing agents of the P-glycoprotein-mediated multidrug resistance: An in vitro evaluation with focus on antiepileptic drugs
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Citation Excerpt :
Hence, aware that the P-gp overexpression can be induced by seizures activity and chronic AED therapy, together with the fact that several AEDs are P-gp substrates, is of paramount importance the evaluation of the effect of multiple flavonoids on the activity of P-gp efflux pump and their influence on AEDs transport. In fact, the number of studies focusing on the effect of flavonoids in the disposition of central nervous system-acting drugs, including AEDs, is scarce; nevertheless, the inhibition of P-gp has previously demonstrated to restore the AEDs anticonvulsant activity in several animal models of refractory epilepsy (Brandt, Bethmann, Gastens, & Löscher, 2006; Clinckers, Smolders, Meurs, Ebinger, & Michotte, 2005; Höcht et al., 2007). Therefore, for this purpose, Madin-Darby canine kidney cell line transfected with the human multidrug resistance-1 (MDR1; ABCB1) gene encoding P-gp (MDCK-MDR1) and its respective wild-type Madin-Darby canine kidney II (MDCK II) cells were used to identify P-gp inhibitors among several flavonoids and evaluate their effects on the cell uptake of AEDs and their active metabolites.
The pharmacoresistance to antiepileptic drugs (AEDs) remains a major unsolved therapeutic need. The overexpression of multidrug transporters, as the P-glycoprotein (P-gp), at the level of the blood-brain barrier of epileptic patients has been suggested as a key mechanism underlying the refractory epilepsy. Thus, efforts have been made to search for therapeutically useful P-gp inhibitors. Herein, the strategy of flavonoid/AED combined therapy was exploited as a possible approach to overcome the P-gp-mediated pharmacoresistance. For this purpose, several in vitro studies were performed using Madin-Darby canine kidney II (MDCK II) cells and those transfected with the human multidrug resistance-1 (MDR1) gene, overexpressing the P-gp (MDCK-MDR1). Overall, the results showed that baicalein, (−)-epigallocatechin gallate, kaempferol, quercetin and silymarin, at 200 μM, produced a marked increase on the intracellular accumulation of rhodamine 123 in MDCK-MDR1 cells, potentially through inhibiting the P-gp activity. In addition, with the exception of lamotrigine, all other AEDs tested (phenytoin, carbamazepine and oxcarbazepine) and their active metabolites (carbamazepine-10,11-epoxide and licarbazepine) demonstrated to be P-gp substrates. Furthermore, the most promising flavonoids as P-gp inhibitors promoted a significant increase on the intracellular accumulation of the AEDs (excluding lamotrigine) and their active metabolites in MDCK-MDR1 cells, evidencing to be important drug candidates to reverse the AED-resistance. Thus, the co-administration of AEDs with baicalein, (−)-epigallocatechin gallate, kaempferol, quercetin and silymarin should continue to be explored as adjuvant therapy for refractory epilepsy.
List of chemical compounds studied in this article:
Baicalein (PubChem CID: 5,281,605); Carbamazepine (PubChem CID: 2554); Carbamazepine 10,11-epoxide (PubChem CID: 2555); (−)-Epigallocatechin gallate (PubChem CID: 65064); Kaempferol (PubChem CID: 5280863); Lamotrigine (PubChem CID: 3878); Licarbazepine (PubChem CID: 114709); Oxcarbazepine (PubChem CID: 34312); Phenytoin (PubChem CID: 1775); Silymarin (PubChem CID: 7073228); Quercetin (PubChem CID: 5280343); Verapamil (PubChem CID: 2520).

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Nimodipine restores the altered hippocampal phenytoin pharmacokinetics in a refractory epileptic model

Abstract

Section snippets

Acknowledgements

Brain Res.

Pharmacol. Biochem. Behav.

Neurosci. Lett.

Epilepsy Res.

Epilepsy Behav.

Epilepsy

N. Engl. J. Med.

P-glycoprotein: a defense mechanism limiting oral bioavailability and CNS accumulation of drugs

Int. J. Clin. Pharmacol. Ther.

Modification of 3H-MK801 binding to rat brain NMDA receptors after the administration of a convulsant drug and an adenosine analogue. A quantitative autoradiographic study

Neurochem. Res.

Refractory phenotype reversion by nimodipine administration in a model of epilepsy resistant to phenytoin treatment

Epilepsia

Astrocytic response in hippocampus and cerebral cortex in an experimental epilepsy model in rat

Neurochem. Res.

Effect of nimodipine, a dihydropyridine calcium channel antagonist on the pharmacokinetics of carbamazepine in rhesus monkeys

Indian J. Physiol. Pharmacol.

Validation of a new intraarterial microdialysis shunt probe for the estimation of pharmacokinetic parameters

J. Pharm. Biomed. Anal.

Microdialysis in drug discovery

Curr. Tech. Drug Discov.

Stereoisomers of calcium antagonists which differ markedly in their potencies as calcium blockers are equally effective in modulating drug transport by P-glycoprotein

Biochem. Pharmacol.

Calcium modulation in epilepsy

Pol. J. Pharmacol.

Multidrug-resistance (MDR) proteins develops refractory epilepsy phenotype: clinical and experimental evidences

Curr. Drug Ther.