Effect of Vortioxetine on Maximal electroshock (MES) induced seizures in Sprague Dawley rats

Sharath Kumar C¹, Navin Patil², Amrita Parida*¹, Kabirdev¹, Prachi Priyadarshini¹, Shanti Gurung²,

Karthik Rao N³, Manju V⁴

¹Department of Pharmacology, Kasturba Medical College, Manipal, Manipal Academy of Higher Education, Manipal - 576104, Karnataka, India.

²Department of Pharmacology, UCMS, Bhairahawa, Rupandehi, Nepal.

³Department of Medicine, KS Hegde Medical Academy, Mangaluru.

⁴Department of Paediatrics, Melaka-Manipal Medical College, Manipal Campus, Manipal Academy of Higher Education, Manipal, Karnataka, India – 576104.

*Corresponding Author E-mail: amrita_parida@yahoo.com

ABSTRACT:

Aim: To investigate the antiepileptic activity of vortioxetine. Materials and method: Vortioxetine was screened for its antiepileptic activity in Sprague-Dawley rats using maximal electroshock model. 4 groups of rats (each 6 rats) were used. First group was administered distilled water, second group diazepam and the third and fourth groups were given vortioxetine 10mg/kg and 20mg/kg respectively. These drug were given 30 minutes before the animal was subjected to electroshock (150mA, 50Hz for 0.2 seconds). Results: Vortioxetine effectively reduced tonic hind limb extension (THLE) in the rats and the effect was statistically significant compared to the control group. The seizure duration was also significantly lower compared to the control. There was no substantial difference in the duration of seizures and THLE between the diazepam and vortioxetine group. However, the total seizure score of the vortioxetine group was not statistically significant compared to the control group. Conclusion: Vortioxetine has the potential to be clinically useful in treating epilepsy. Further detailed studies using other animal models and in humans are required to prove its efficacy in epilepsy.

KEYWORDS: Anti-seizure, antidepressant, serotonin reuptake inhibitors, anti-epileptic, maximal electroshock.

INTRODUCTION:

Epilepsy affects millions of people around the globe. Epileptic attacks can cause impairment of consciousness which might lead to bodily harm. It also frequently interferes with performance of day to day activities including education and occupation. Most of the antiepileptic drugs needs to be taken lifelong, which adds to the economic burden on the patient. Numerous side effects and lack of compliance are other major issues associated with antiepileptic drugs. Though the drugs that are available for treatment of epilepsy inhibit seizures, there is no agent which treats the cause of epilepsy (epileptogenesis).¹ Hence, there is no existing therapy for complete cure of epilepsy.

Most of the antiepileptic drugs, that we know today act by one of the three broad mechanisms:² 1. Prolonging the inactivated state of the sodium channels. 2. Facilitating the GABA mediated opening of chloride channels and lastly 3. Inhibiting the “T” type calcium current. The quest for an ideal anti-epileptic agent is still on and many compounds are being tested for their antiepileptic potential. One such group of drugs are the antidepressants. Since a long time it was believed that antidepressants have pro-convulsive property and can lead to precipitation of seizures. However, recent studies have suggested that, the newer drugs like selective serotonin/norepinephrine reuptake inhibitors do not lower seizure threshold. Moreover, it was found that, these antidepressants might actually have anticonvulsant property. The exact mechanism of how these drugs regulate the neurogenic signals is still unknown. The postulated hypothesis is that, most probably the SSRIs and SNRIs modulate the GABAergic/glutamatergic transmission in brain and this action is site specific.³ There are also studies which have shown that there is a link between monoamines and epilepsy. These studies which show a relation between serotonin (5-HT) and epilepsy are based on various neuroimaging studies in epileptic patients and in experimental animal models of epilepsy.⁴

Vortioxetine, a selective serotonin reuptake inhibitor (SSRI), is a newer drug approved for use in major depressive disorder (MDD). It is 5-HT_1D, 5-HT₃ and 5-HT₇ receptors blocker. It acts as a partial agonist at the 5-HT_1B receptor. It is also a complete agonist at the 5-HT_1A receptor. In addition to modulating these serotonin receptors, vortioxetine is also a serotonin transporter (SERT) inhibitor.⁵ The exact mechanism of action of vortioxetine is still unknown. Due to its direct modulatory effects on the various serotonin receptors, its pharmacological actions are different from the other SSRIs. The older antidepressants cause desensitization of 5-HT_1A receptor by increasing the serotonin levels as a result of inhibiting its uptake. The 5-HT_1A receptors are located presynaptically, hence its inactivation creates a negative feedback and is possibly responsible for their reduced their antidepressive effects. With vortioxetine, this desensitization is not seen as it is an agonist of 5-HT_1Areceptor.⁵ Since the way vortioxetine acts is novel, its role in epilepsy needs to be evaluated. There are limited studies on action of vortioxetine on epilepsy hence we decided to investigate the effects of this drug on rodent model of epilepsy. Hence the current study was to screen for antiepileptic activity of vortioxetine using the Maximal Electroshock (MES) model in Sprague Dawley rats.

MATERIALS AND METHODS:

Animals:

Twenty-four rats (Sprague Dawley strain) were used. They were divided into four groups. Both male and female rats were included in the study. Weight of the animals ranging 150-300g were used. Animals were housed in standard environmental conditions (temperature of 22±3˚C and a 12-hour alternate light and dark cycle). The animals were provided with regular rodent diet and water ad libitum. The protocol was approved (letter no-IAEC/KMC/114/2019) by the Institutional Animal Ethics Committee, KMC, MAHE, Manipal.

Drugs:

Vortioxetine 20mg tablets, (brand name – Brintellix) Takeda Pharmaceuticals America, Inc. were indented from hospital pharmacy. The tablets were first powdered and the powder was mixed with distilled water. The drug was administered through intraperitoneal (i.p.) route.

Diazepam injection 10mg/2mL (brand - Calmpose) was also indented from the hospital pharmacy and the calculated dose was given by i.p. route.

Experimental procedure:

All the animals were subjected to electric shock 48 hours prior to the experiment and were found to show positive THLE and hence all of them were taken up for the main experiment. The rats were divided into four groups with 6 rats in each. Diazepam (5mg/kg) was the standard drug used. It is known to prevent tonic hind limb extension (THLE) in the MES model. The test drug vortioxetine was given in two different doses of 10mg/kg and 20mg/kg. These drug were given 30 minutes before the animal was subjected to electroshock. Distilled water was injected to the negative control group. All the drugs were given through i.p. route

Table 1: Different groups and the treatment given.

Group 1 (Negative control)	Distilled water
Group 2 (Standard positive control)	Diazepam (5 mg/kg)
Group 3 (Test drug)	Vortioxetine (10 mg/ kg)
Group 4 (Test drug)	Vortioxetine (20 mg/ kg)

An electro-convulsiometer was used to deliver the shock using ear electrodes. The intensity of the stimulus employed was 150mA, 50Hz for 0.2 seconds.

The following stages are seen in MES induced seizures:

a) Flexion

b) Extension

c) Clonic convulsions

d) Stupor

e) Recovery or death

After administration of shock, all the animals were observed for 2 minutes. Absence/ reduction of the extensor phase of the MES convulsions was taken as the criteria of protection for treatment groups.

The following parameters were observed for:

1) Onset of seizures

2) Time duration of tonic hind limb extension

3) Total time period of seizures

4) Recovery or death

The seizure score⁶ for each group was calculated based on the observations. The scoring is done accordingly: 0 = no seizure; 1 = only forelimb extension and no hind limb extension. 2 = complete forelimb extension with partial hind limb extension, 3 = complete tonic hind limb extension (THLE) and 4 = post ictal depression.

Percentage inhibition of seizures as compared to controls is calculated as follows:⁶

Percentage protection = (Number of animals with hind limb extension absent / Total number of animals) x 100

Statistical Analysis:

One-way ANOVA followed by Tukey’s post-hoc test was used to analyse the results. SPSS package version 17 was used for the statistical analysis. p value < 0.05 was considered statistically significant.

RESULTS:

Any substance that inhibits/ reduces the tonic hind limb extension is said to possess anti-epileptic activity. In this study, the different parameters that were noted for different groups were as follows:

1) Onset of seizures

2) Time duration of hind limb extension

3) Total time duration of seizures

4) Recovery or death

5) Total seizure score

The above data for the different groups is tabulated below. There was no death in any of the groups.

One-way ANOVA followed by Tukey’s post-hoc test was used to compare the means of different groups.

DISCUSSION:

The dysfunctioning of neuronal mechanisms in brain can lead to numerous diseases. Sometimes, a common derangement can cause multiple problems. The neurotransmitter serotonin is known to be involved in various CNS disorders. Variations in the monoamine activity in mood disorders is well established. Most of the effective drugs used in major depressive disorders act by modulating the 5-HT and norepinephrine levels in brain. Studies have proved that patients with mood disorders like anxiety and depression are at increased risk of developing epilepsy.⁷ And the vive-versa is also true, i.e., epileptic patients present with mood disorders more commonly compared to the general population.⁸ This similarity strengthens the evidence that the underlying neuronal mechanism could be similar in both the cases. Some of the studies have suggested that the depression that is seen in epileptic patients could be an intra or post ictal effect of the seizure.⁹ The hypothesis of a common pathway being involved in epilepsy and mood disorders is further supported by numerous animal and human studies. It is seen that in epileptic patients, there is reduction in the serotonin levels in brain as well as reduced binding of serotonin to the 5-HT^1A receptor.^10,11 The 5-HT_1A receptor is responsible for hyperpolarization of neuronal membranes, hence its activation, reduces excessive neuronal firing. Stimulation of 5-HT also increases GABA levels in brain.^12-14 Another clue to the common link between epilepsy and mood disorders is that, many of the anticonvulsants currently used (sodium valproate, carbamazepine) are also effective in mood disorders. It is noted that these drugs increase serotonin levels in brain.^11,15,16

Earlier there was a misconception that the antidepressants have proconvulsant activity and were to be avoided in epileptic patients. However recent animal and human studies have proved that the newer antidepressants like SSRI/SNRIs do not cause seizure and in fact, have anticonvulsant property. By increasing the serotonin levels in brain and modulating the various serotonin receptors, they have been able to supress epileptic seizures in animal models of epilepsy.⁴ Though there are no published controlled human clinical trials results related to direct use of antidepressants in epilepsy, data from other trials shows patients of major depressive disorder (MDD) who were on SSRI/SNRIs had lesser incidences of seizure attacks compared to placebo.¹⁷ Serotonin reuptake inhibitors have also enhanced the activity of other antiepileptic drugs in the MES animal models.¹⁸

Vortioxetine is a newer serotonin reuptake inhibitor, approved for use in MDD. It has a complex mechanism of action on the serotonin receptors and is therefore different from the earlier SSRIs. This drug is a 5-HT_1D, 5-HT₃ and 5-HT₇ receptor blocker. It acts as a partial agonist at the 5-HT_1B and as a full agonist at the 5-HT_1A receptor. In addition to modulating these serotonin receptors, vortioxetine is also a serotonin transporter (SERT) inhibitor. In addition to targeting the serotonergic pathway, it also affects the glutamate and GABA levels in the brain and this action is site specific. As stated earlier, role of 5-HT_1A receptor is vital in epilepsy. The other SSRIs in due course of treatment cause desensitization of 5-HT_1A receptor located presynaptically, and this can lead to their decreased efficacy in both depression as well as epilepsy. Vortioxetine is devoid of this action and since it is agonistic at 5-HT_1A, it is found to be more efficacious in MDD.⁵ The antiepileptic activity of other SSRIs/SNRIs have been proved in many animal models, but there is only one published data on the role of vortioxetine in epilepsy.³

In the current study, vortioxetine was tried at a dose of 10mg/kg and 20mg/kg for epilepsy in rats using the MES model. The doses were determined based on a previous study³. The MES model is simple yet very effective model for screening drugs for epilepsy.¹⁹ Drugs those are found to be effective in abolishing/ reducing the hind limb extension in MES model are said to be effective in GTCS clinically.^20-22 In our study, vortioxetine was found to effective reduce THLE in the rats and the results were statistically significant compared to the distilled water control. The seizure duration was also significantly lower compared to the distilled water group. There was no substantial difference in the duration of seizures and THLE between the diazepam and vortioxetine group. However, the total seizure score of the vortioxetine group was not statistically significant compared to the control group. Similar results were seen in the study conducted by Ogun MN et al.³ In their study, Vortioxetine was found to significantly reduce the epileptic discharges induced by penicillin in rats compared to controls and there were no significant differences in the spike activity between vortioxetine and the standard treatment drug (diazepam) group.

Since the results are encouraging, further detailed studies need to be done to find out the exact effects of vortioxetine in epilepsy. The effect of vortioxetine as an add on drug to the current anticonvulsants also needs to be investigated.

Limitations of the study:

Major limitation of the study is a low sample size due to lesser number of animals used because of ethical issues. Secondly, a single model was used for screening the antiepileptic effect of vortioxetine due to which the results might be inconclusive. Extensive studies are required to find out the exact effect of vortioxetine in epilepsy.

CONCLUSION:

Vortioxetine significantly reduced seizure activity and the duration of hind limb extension phase in MES model of epilepsy in rats compared to the control group. There was no significant difference in the antiepileptic activity between vortioxetine and diazepam group. Further studies in other animal models as well as humans are required to substantiate the antiepileptic activity of vortioxetine.

CONFLICT OF INTEREST:

None

FUNDING SOURCE:

None

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Received on 25.08.2020 Modified on 29.09.2020

Research J. Pharm. and Tech. 2021; 14(8):4337-4340.

DOI: 10.52711/0974-360X.2021.00753

Groups (n = 6 in each group)	Duration of THLE (Mean± SEM) in seconds	Duration of seizure (Mean± SEM) in seconds	Mean seizure score	Percentage protection (%)
Group 1 (Distilled water) control	64.83 ± 5.9	134.3 ± 10.47	3 ± 0.63	Zero
Group 2 (Diazepam)	0*	33.3 ± 6.29*	0.83 ± 0.26*	100
Group 3 (Vortioxetine 10mg/kg)	14.5±4.9*	57.83 ± 2.4*	2.33 ± 1.03	11.1
Group 4 (Vortioxetine 20mg/kg)	9.33 ± 4.0*	46.66 ± 6.77*	2.5 ± 0.83	33.3