Effects of Tempol on Epileptic Activity in Picrotoxin-Induced Epilepsy in Rats
Correspondence Address: Source of Support: None, Conflict of Interest: None DOI: 10.4103/0028-3886.314542
Source of Support: None, Conflict of Interest: None
Keywords: Epilepsy, picrotoxin, tempolKey Message: Epilepsy is a common neurological disease, the etiology and pathophysiology of which are not yet fully understood. Oxidative stress plays a role in the pathogenesis of epilepsy. Pretreatment with Tempol reduces Ptx induced tonic-clonic seizures through its anti-inflammatory and antioxidant effects. Further studies are needed to investigate the mechanisms behind the antiepileptic effect of tempol.
The World Health Organization (WHO) estimates that epilepsy affects 0.8% of the global population. Although epilepsy is a common neurological disease, its etiology and pathophysiology are not yet fully understood. Epilepsy is a disease that occurs as a result of abnormal simultaneous discharges of neurons and is characterized by recurrent seizures. Many factors have been found to be effective in the onset of epilepsy, including genetic factors, glutamatergic cell toxicity increased by intracellular electrolyte metabolism disorders, mitochondrial dysfunction, oxidative stress, decreased growth factor and increased cytokine concentrations.
Studies have shown that free radicals are associated with the pathogenesis of epilepsy, and oxidative stress results from increased production of free oxygen radicals. Oxidative stress plays a role in the pathogenesis of many neurological diseases, including epilepsy, and leads to the emergence of not only status epilepsy, but also its chronic form. Reactive oxygen species (ROS) may affect neuronal excitability and seizure susceptibility.,
A variety of experimental epilepsy models have been applied to understand the basic mechanisms of epilepsy, and more efficient drugs have duly been discovered for treatment. The formation of oxidative stress in the central nervous system has been demonstrated through the administration of penicillin, pilocarpine, and pentylenetetrazol (PTZ) in different animal models of epilepsy.,, There have also been many studies demonstrating that antiepileptic medicines with neuroprotective and antioxidant activity inhibit free oxygen radicals.,
PTZ is the most preferred substance for the formation of experimental epilepsy models. Although the mechanism of action for PTZ is unknown, it is believed to selectively block the GABA A receptors associated with chloride channels and facilitate the depolarization of neurons by reducing the effect of GABA and the other inhibitor neurotransmitters., On the other hand, Ptx is an agent that may lead to generalized seizures in systemic applications by blocking the chloride channels associated with GABA.
The tempol (4-hydroxy220.127.116.11-tetramethylpiperidine-N-oxyl) nitroxide compound is a membrane-permeable radical scavenger similar to superoxide dismutase (SOD) that can provide protection against oxidative stress. Tempol antioxidants scavenge superoxide anions (O2-) with low molecular weight.
This study evaluates the effects of tempol on epileptic activity through behavioral parameters in acute Ptx models in awake rats.
This experimental study was conducted on 42 adult male Wistar Albino rats weighing 450–500 grams. Following a council decision of the Mustafa Kemal University Experimental Research Application and Research Centre Ethics Committee (2016/2-4), the rats were divided into three groups and held in cages containing six rats each until the end of the experiment. Standard pellet feed and city drinking water were used for animal nutrition.
Ptx was purchased from Sigma and was dissolved in Dimethyl sulfoxide (DMSO). Tempol was purchased from Merck Manual and dissolved in saline.
Establishing an acute picrotoxin model
Ptx 2.5 mg/kg was injected as a single i.p dose. Following injection, the animals were observed for 30 min in glass observation cages measuring 35 cm × 35 cm × 35 cm. The seizure behavior of the animals was scored based on the method in the study by Fisher and Kittner.
The study of rats was terminated after scoring.
IBM SPSS Statistics (version 21.0. Armonk, New York: IBM Corp.) was used for the statistical analysis. Numerical variables were expressed as mean ± standard error. P < 0.05 was considered sufficient for statistical significance. A one-way variance analysis (one-way ANOVA) was used for the comparison of the groups, followed by a Tukey test for a post hoc analysis.
The results of the Ptx-induced epilepsy were as follows.
The total number of seizures significantly decreased in the Ptx + tempol 100 and Ptx + tempol 150 groups when compared to the Ptx group (P < 0.05). The total seizure time significantly decreased in the Ptx + tempol 100 and Ptx + tempol 150 groups when compared to the Ptx group (P < 0.05). Phases of seizure significantly decreased in the Ptx + tempol 150 group when compared to the Ptx group (P < 0.05) [Table 1].
This study was carried out to identify the relationship between the epileptic seizure model and oxidative stress, in which the results of tempol administration at doses of 100 and 150 mg/kg in experimental epilepsy models established with Ptx 2.5 mg/kg were recorded.
Tempol is a membrane-permeable free radical scavenger. In a previous study, tempol was shown to reduce lipid peroxidation by capturing free radicals in ischemic brain damage, thus reducing ischemic brain damage. Tempol has been shown in many studies to have a neuroprotective effect by reducing oxidative stress in the brain.,
In previous studies, oxidative stress due to the increased production of reactive oxygen types has been shown to play a role in the pathogenesis of several neurological diseases, including epilepsy.,, The increased production of oxygen radicals causes neuronal death by triggering signaling pathways. Oxidative stress causes an increase in membrane phospholipase activity and reveals free fatty acids by breaking the phospholipids in the phospholipase membrane in epilepsy models. Increased fatty acids and free radicals lead to cellular dysfunction and lipid peroxidation in membranes. Oxidative stress causes an increase in malondialdehyde (MDA) levels an important marker of lipid peroxidation and a decrease in glutathione (GSH), SOD, and catalase levels. Increased oxidative stress in the brain results in seizure activity.
The antioxidant defense system consists of diverse endogenous antioxidative enzymes, such as SOD, glutathione peroxidase (GPx), and GSH. These antioxidative enzymes provide significant cellular defense against oxidative stress by extinguishing free radicals, thereby reducing elevated ROS levels. As reported in one study, tempol is a cell membrane-permeable amphibolite that can catalyze to facilitate the hydrogen peroxide metabolism, which increases endogenous SOD levels. As such, tempol can be considered to have a significant antioxidant effect on these ROS, and so can act as a breakdown in the antioxidant defense system, and thus alleviate the convulsions caused by PTZ.
In a further study, tempol was shown to be an anticonvulsant due to its SOD-like effect and to significantly reduce or completely prevent epileptic seizures when administered to immature animals in a 150 mg/kg dose.
Studies have shown that epilepsy, an important neurological disease, can cause oxidative stress and mitochondrial dysfunction, among the other forms of damage, to the brain. In this regard, the use of antioxidant compounds can provide neuroprotection, and can help to reduce the damage caused by epileptic seizures and the use of anticonvulsant drugs. Rosmarinic acid (RA) prevents the cell damage caused by free radicals by acting as an antioxidant and can reduce the cell damage generated by Ptx seizures.
In this study, after administering Ptx 2.5 mg/kg, tempol at 150 mg/kg significantly extended the seizure onset time when compared to the Ptx group, while tempol 100 mg/kg significantly shortened the total duration of the seizure when compared to the Ptx group.
Ptx causes convulsions through the inhibition of the GABA receptor/chloride channel complex, and is the most commonly used chemoconvulsant in studies into the effects of antiepileptic drugs.
In another study, a significant increase was observed in MDA levels, as an important indicator of free radical production resulting from lipid peroxidation, in the brain cortex tissue, as well as a significant decrease in antioxidant enzymes, such as catalase and SOD, in an epilepsy model involving rats aimed at identifying the relationship between epilepsy and oxidative stress. We have seen that oxidative stress increases in epilepsy, and that antioxidant drugs such as tempol prevent the lipid peroxidation caused by free radicals in the body by capturing the free radicals that have increased as a result of oxidative stress. We have further observed a decrease in the MDA levels that increase as a result of oxidative stress and an increase in antioxidant enzymes.,, Antioxidant treatments can be benefitted from in the prevention of oxidative stress in epilepsy.
In this study, tempol 100 and 150 mg/kg were administered in an experimental epilepsy model established with Ptx 2.5 mg/kg. Ptx was administered after being dissolved in DMSO, which studies have shown has an antiepileptic effect on temporal lobe epilepsy, while another study found that low dose DMSO reduced epileptic activity while high dose DMSO increased epileptic activity.,
Ptx was found to be effective in epilepsy when administered at doses of 2, 3, 3.5, and 4 mg/kg. In this study, the total number of seizures and the total seizure time decreased in the tempol 100 and tempol 150 mg/kg groups, respectively, in an epilepsy model induced with Ptx 2.5 mg/kg. Furthermore, a decrease in the phases of seizure was observed in the tempol 150 mg/kg group.
In this study, the administration of tempol 100 and 150 mg/kg to an experimental epilepsy model induced with Ptx 2.5 mg/kg, the onset of seizure was extended by tempol 150 mg/kg; the total number of seizures was decreased by tempol 100 and 150 mg/kg; and the total seizure time was shortened in the tempol 100 mg/kg group. Furthermore, the phase of seizures was decreased in the tempol 150 group; the total number of seizures and the total seizure time were decreased in the tempol 100 and tempol 150 mg/kg groups in the epilepsy model induced with Ptx 2.5 mg/kg; and a decrease was noted in the phases of seizure in the tempol 150 mg/kg group. In another study, Tempol (100 and 200 mg/kg) pretreatment provided significant protection against PTZ-induced epilepsy.
Tempol 100 mg/kg and tempol 150 mg/kg can be considered effective in epilepsy models induced with Ptx, with tempol 150 mg/kg in particular being found to be more effective. Pretreatment with Tempol reduces Ptx-induced tonic-clonic seizures via its anti-inflammatory and antioxidant effects. Further studies are needed to investigate the antiepileptic effect mechanisms of tempol.
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