Effect of Cuminum cyminum and Rhus pentaphylla extracts on status epilepticus induced by intrahippocampal injection of kainic acid in rat
Abstract
This study aims to evaluate the anticonvulsant effect of the aqueous extract of Rhus pentaphylla and Cuminum cyminum (300 mg/kg), orally, 45 min before intrahippocampal injection of kainic acid (1 µg/µL). The scoring of seizure severity, latency of seizures and duration of total seizures were recorded in 90 min. Both extracts showed the presence of polyphenols, flavonoids, alkaloids and tannins compounds. These extracts exhibited potent antioxidant activity and showed an attenuation of severe convulsive seizures, a significant decrease in the frequency and duration of seizures and prolonged the latency of the onset seizure (p<0.001) compared to kainic acid. The aqueous extracts of R. pentaphylla and C. cyminum have a protective effect against seizures induced by kainic acid.
Introduction
Epilepsy is a chronic neurological disease characterized by unpredictable and recurrent seizures, caused by synchronous and rhythmic firing of populations of brain neurons. Antiepileptic drugs are the first line of treatment for epilepsy. One in three people still have drug-resistant epilepsy. These antiepileptic drugs are often the cause of undesirable side effects and higher rates of intolerance (Chen et al., 2017).
It is, therefore, essential to focus on traditional medicine as an alternative. Several studies reported the anticonvulsant activity of some plants on the experimental model of seizures, such as Pandanus odoratissimus (Adkar et al., 2014), Cicer arietinum (Sardari et al., 2015), Musa sapientum (Reddy et al., 2018), Anacyclus pyrethrum (Bezza et al., 2019), Artemisia persica (Daneshkhah and Setorki, 2019).
Various classes of phytochemicals identified from traditional medicinal plants have effects on convulsion, such as sesquiterpene lactone, diterpenes, triterpenes, flavonoids, coumarins (Zadali et al., 2022). The previous phytochemical analysis of Cuminum cyminum revealed that it contains alkaloid, coumarin, anthraquinone, flavonoid, glycoside, protein, resin, saponin, tannin, and steroid (Al-snafi, 2016). Another study revealed that around 16 small molecules phytoconstituents of Cuminum cyminum can cross the blood-brain barrier (Chouhan et al., 2022). In the same way, anthocyanin, (which belongs to the flavonoid family, and are natural pigments contained in Rhus pentaphylla extract) and their methylated forms reached the brain tissue (Talavéra et al., 2005). Another study shows that flavonoids are able to enter the central nervous system by crossing the blood-brain barrier (Yimer et al., 2019).
Over and above, flavonoids show considerable potential as antioxidant agents, that can contribute significantly in ROS detoxification through chemical ROS quenching, in human cells (Brunetti et al., 2013).
Here, we are interested to study C. cyminum which is a natural remedy for a diverse range of illnesses, it’s known to treat a variety of diseases due to its effective antioxidant, antimicrobial, anti-inflammatory, analgesic, anti-cancer activity, protective and central nervous effects (Al-snafi, 2016 ; Yimer et al., 2019). It’s a worthwhile herb with a rich historical basis to manage many neurological disorders. R. pentaphylla which belongs to Anacardiaceae family, the extracts of roots, leaves and seeds characterized by the presence of tannins, flavonoids, alkaloid, coumarins, and high antioxidant activity and chemoprevention, suggesting their use in medicinal and industrial fields (Mansour et al., 2011).
The objective of this research was to evaluate the effect of pretreatment by C. cyminum and R. pentaphylla in the prevention of seizures caused by kainic acid. As well as to assess the quantitative and qualitative phenolic compounds and the antioxidant activity of these extracts.
Materials and Methods
Animals
The study presented in this work was carried out on male adult Wistar rats (200-300 g) provided by the Animal Care Facility of the Faculty of Sciences Semlalia, Cadi Ayyad University, Marrakech, Morocco. The animals were kept in a stable environment with a constant temperature (22 ± 2ºC) and a 12 hour light-dark cycle (light from hour 7 to hour 19). They have free access to food and water and were allowed to acclimatize to the housing room conditions for 3 days before behavioral testing procedures were initiated. All experiments were performed between 9:00 AM and 2:00 PM.
Plants extract preparation
The leaves of R. pentaphylla and the seeds of C. cyminum were collected in the Benguerir region (32°11'11"N, 7°55'19"W) and the Sidi Rahal region (31°38'20"N, 7°28'35"W) respectively. The plant material was identified by Prof. A. Ouhammou (Laboratory of Environment and Ecology) and deposited in the Herbarium of the Semlalia Faculty of Sciences, Cadi Ayyad University, Marrakech, Morocco (R. pentaphylla: MARK- 13360; C. cyminum: MARK-13291).
The extracts were prepared according to the following procedures: The plant material was crushed to obtain a powder. Then, 50 g of each powder was dissolved in 1,000 mL of distilled water and stirred at room temperature (22 ± 2°C) for 12 hours. Subsequently, the mixture was filtered, and the resulting solution was subjected to lyophilization to transform the aqueous extract into powder form.
The yield of the extraction of the two plants extracts was calculated using the following formula:
Yield (%) = (raw extract / initial extract) x 100
Phytochemical screening and phytochemical analysis
Test for flavonoids
Flavonoids are revealed by the cyanid reaction. Five milliliters of distilled water, 1 mL of concentrated hydrochloric acid, and small quantities of magnesium chips were added to 1 mL of the extract. Then, the appearance of the red color after a few minutes indicates the presence of flavonoids (Joshi et al., 2013).
Test for tannins
A few drops of ferric chloride solution (9%) were added to the extract. Then, produces a blue (gallic tannins) or dark green (catechin tannins) color in the presence of tannins (Banso and Adeyemo, 2006).
Test for alkaloids
A total of 10 g of sprayed plants are stirred in 50 mL of sulfuric acid (0.1 N) for 15 min. After filtration, the solution obtained was alkalinized with 5 mL of ammonia solution (25%) and diluted with 50 mL of distilled water. After extraction of the alkaloids with dichloromethane and vacuum evaporation, the residue obtained was taken up again with a (10%) sulfuric acid solution. Dragendorff’s reagent was added, then the presence of alkaloids causes the formation of a precipitate (Okpo et al., 2001).
Test for saponins
The presence of saponins was tested on a 20% decoction. After stirring, the extract contains the saponins when it foam was formed 1 cm high, stable for 10 min (Dohou et al., 2003).
Test for terpenes and sterols
The plant powder (4 g) was added to 50 mL of methanol and macerated for 15 min. Following filtration and evaporation, 1 mg of residue was dissolved in a few drops of acetic acid, then taken up in a watch glass where 3 mL of (acetic anhydride and sulfuric acid) mixture was added without stirring. The appearance of a blue-green color indicates the presence of sterols and terpenes (Joshi et al., 2013).
Assessment of total phenolic compounds, flavonoids and tannins
Total phenolic compounds were determined spectrophotometrically using the Folin-Ciocalteu reagent. A quantity of 50 μL of the extract was mixed with 1950 μL of distilled water and 50 μL of the freshly prepared Folin-Ciocalteu reagent. After 3 min of incubation, 500 μL of 20% sodium carbonate was added. The whole was agitated and incubated in the dark at room temperature for 60 min. The absorbance was measured at 725 nm. The results were expressed in milligrams equivalent of gallic acid per gram of dry plant material.
The flavonoid compound was quantified using the aluminum trichloride colorimetric method. 100 µL of diluted extract was mixed with 400 µL of distilled water and 30 µL of 5% sodium nitrite solution. After 5 min of incubation, 20 µL of a 10% aluminum trichloride solu-tion was added. Then, after another 5 min, 200 μL of sodium carbonate solution (1M) and 250 μL of distilled water were added. The mixture was agitated to homogenize the contents and the absorbance was measured at 510 nm. The results were expressed in milligrams equivalent to catechin per gram of dry plant material.
The determination of the tannin content was measured by the vanillin method in an acidic medium. 1 mL of vanillin solution 4% was added to 100 µL of the sample extract, the mixture was stirred using a vortex, then 500 µL of pure hydrochloric acid was added. After 15 min of incubation in the dark at room temperature, the absorbance was measured at 550 nm. Results were expressed as catechin per gram of extract.
Antioxidant activity
DPPH assay
The antioxidant power of the aqueous extracts was evaluated by the 2,2-diphenyl-1-picrylhydrazyl (DPPH) free radical scavenging test. Each solution (50 μL) of the extracts at different concentrations was added to 2 mL of the methanolic solution of DPPH (0.004%). The prepared solutions were stirred and incubated for 20 min in the dark at room temperature. The absorbance reading of each concentration was measured at 517 nm. The negative control was prepared by adding 500 μL of methanol with 2 mL of DPPH, and gallic acid was considered as a 50% positive control. For each concentration, the test was repeated 3 times. The free radical scavenging activity was estimated using the following equation:
(I %) = [Abs control - Abs extract/ Abs control] x 100
Where (I %) of anti-free radical activity, Abs control is the absorbance of control and Abs extract is the absorbance of the extract. The IC50 values have been determined graphically by linear regression
Box I: FRAP Antioxidant Assay
Principle
The FRAP assay consists of preventing the development of Fe (II)-ferrozine complexes when samples were incubated with ferrous iron.
Requirements
Centrifuge machine; Ferric chloride solution; Phosphate buffer (0.2M, pH=6.6); Potassium ferricyanide (1%); Trichloroacetic acid (10%); Butylated hydroxytoluene; Quercetin; Spectrophotometer (model: T60UV)
Procedure
Step 1: Prepare a mixture consisting of 200 µL of each concentration of the aqueous solutions of the extracts with 500 µL of phosphate buffer solution (0.2 M, pH 6.6), then a 500 µL of potassium ferricyanide (1%) was added.
Step 2: The mixture was incubated for 20 min at 50°C.
Step 3: To stop the reaction, 500 µL mL of 10% trichloroacetic acid was added.
Step 4: The mixture was centrifuged 3,000 revolutions for 10 min.
Step 5: Then, 500 µL of the supernatant was added to 500 µL of distilled water and 100 µL of ferric chloride (0.1%) and was agitated.
Step 6: Finally, the absorbance was measured at 700 nm.
Butylated hydroxytoluene and quercetin were used as positive controls whose absorbance has been measured under the same conditions as the extracts.
Calculation
The FRAP value was calculated using the following equation:
FRAP value = [(A1 − A0)/(Ac − A0)] × 2
Where Ac is the absorbance of the positive control, A1 is the absorbance of the sample, and A0 is the absorbance of the blank
Advantages
The FRAP assay is a relatively simple, quick, and inexpensive direct method of measuring the total antioxidant activity
References
Oyaizu, 1986; Aitbaba et al., 2023
Kainic acid-induced status epilepticus
Stereotaxic surgery
Following anesthesia by intraperitoneal injection of hydrate chloral (6%), the rat was placed in a stereotaxis apparatus (Horsley Clark). A small opening in the skull was made using a dentist's drill. Then, a stainless guide cannula (1.5 cm long, 0.3 mm internal diameter and 0.4 external diameter) was lowered to the hippocampus (AP = - 2.30 mm; L = ± 1.5 mm; D = - 2.7 mm from Bregma) determined (Paxinos and Watson, 2016). The cannula was fixed with dental cement (self-curing acrylic resin) which was consolidated by 3 small screws implanted in the cranial bone on either side of the cannula (Ait laaradia et al., 2021; Ferr et al., 2012).
The microinjection system consisted of an injection cannula (27 gauge) connected to Hamilton microsyringe (10 µL) by a PE catheter. Before the injection, the mandrel was removed from the guide cannula and replaced by the injection cannula. A volume of 1 µL of kainic acid (1 µg/µL) was injected. After 2 min, the injection cannula was removed and replaced with the mandrel. The microinjections were performed in non-anesthetized rats that were held gently to avoid possible stress-related movements (Ait laaradia et al., 2021).
The rats (n=20) were divided into 4 groups of 5 rats each. Control group treated with 1 µL of 0,9% NaCl; Group microinjected with 1 µg /µL of kainic acid; Group treated with aqueous extract of C. cyminum plus kainic acid; and a group treated with the aqueous extract of R. pentaphylla plus kainic acid. Forty five minutes before the microinjection of kainic acid, the extracts of plants were administered orally by gavage. All treatments were orally administered between 9:00 AM and 12:00 AM. At the end of the experiment, the placement of the guide cannula in the hippocampus was checked in histological brain sections.
Seizure behavior scoring
After intra-hippocampal injection of kainic acid, each rat was placed in the center of the glass and its behavior was observed and recorded for 90 min. The following seizure parameters were noted: latency (min), Latency of the tonic-clonic seizure (sec), duration of the tonic-clonic seizure (sec), duration of seizures (min), percentage of protection against tonic-clonic seizures.
A seizure severity scale was determined according to Racine scale. The scoring was as follows: 0: normal activity; 1: immobility and /or fixed gaze; 2: stiffness, tail lengthening, head twitching; 3: repetitive movements, bilateral tremors of the hind limbs; 4: minor seizure or shaking, jumping, falling; 5: tonic-clonic convulsion or multiple convulsions; 6: severe tonic-clonic seizure and 7: Death.
The cage was cleaned at the end of each test to avoid the presence of odor which may result in behavioral change. The assessment of the severity of seizures was evaluated as the sum of cumulative scores given to each seizure behavioral category. The latency of seizures, the time between the moment of the microinjection of kainic acid and the onset of the first seizure, were also recorded. The total duration of seizures was considered as the sum of the elementary durations of episodes of seizures during the 90 min observation time.
Evaluation of protection against seizures
The percentage of protection was calculated using the equation:
%Protection = 100 – (Number of animals presenting tonic-clonic convulsions/Total number of animals) × 100
Statistical analysis
Results are presented as mean ± standard error of the mean (SEM). Sigma Plot 12.5 software was used for statistical analysis. Comparison of the different variables between groups was performed by one-way analysis of variance (ANOVA) test, followed by a Tukey post hoc test. The results with p<0.05 were considered statistically significant.
Results
Yield of plant extracts
C. cyminum extract yielded 26.8% whereas R. pentaphylla yielded 22%.
Phytochemical screening
The preliminary phytochemical screening of R. pentaphylla and C. cyminum extracts had shown the presence of bioactive chemicals such as, alkaloids, flavonoids, steroids, terpenes, and tannins (Table I).
Table I: Phytochemical constituents
| Cuminum cyminum | Rhus pentaphylla | |
| Qualitative data | ||
| Alkaloides | +++ | ++ |
| Saponines | - | - |
| Sterols/Terpenes | ++ | +++ |
| Quantitative data | ||
| Polyphenols (mg EAG/g MS) | 14.8 ± 0.2 | 21.6 ± 0.1 |
| Flavonoids (mg EC/g MS) | 9.7 ± 0.4 | 14.4 ± 0.7 |
| Tannins (mg EC/g MS) | 4.9 ± 0.5 | 10.7 ± 0.2 |
Assessment of total polyphenol, flavonoids and tannins content
Based on the absorbance values of the extract solutions, and compared to the corresponding standard solution, the aqueous extracts of C. cyminum and R. pentaphylla had shown a moderate content of polyphenols, flavonoids and tannins. R. pentaphylla microsyringe seizure severity evaluated by scorings (Table I).
Antioxidant activity
The antioxidant activity was tested in vitro using two complementing tests: reducing power and DPPH free radical scavenging. The inhibitory concentration (IC50), is represented in Table II. The antioxidant properties were compared to those of quercetin and butylatedhydroxytoluene (BHT). Stronger antioxidant activity was indicated by lower IC50 values. Both C. cyminum and R. pentaphylla showed significant antioxidant activity, with the DPPH test the lowest IC50 was obtained for R. pentaphylla (IC50 = 47.2 ± 0.2 µg/mL). However, the IC50 values was less effective than those of synthetic antioxidant agent butylhydroxytoluene and quercetin (IC50 values of 3.8 ± 0.1 µg/mL to 1.3 ± 0.0 µg/mL).
Table II: IC50 (µg/mL) values
| DPPH | FRAP | |
|---|---|---|
| Cuminum cyminum | 180.4 ± 0.3 | 252.0 ± 0.3 |
| Rhus pentaphylla | 21.1 ± 0.9 | 106.1 ± 0.3 |
| Butylatedhydroxytoluene | 3.8 ± 0.1 | 1.0 ± 0.0 |
| Quercetin | 1.3 ± 0.0 | 1.5 ± 0.0 |
Epileptic behavior testing
Mortality
In the kainic acid group, two rats died during the establishment of the model. The remaining 4 rats were successfully modeled, but died in the following, two or three days after. No rats died in the treated groups.
Effects of C. cyminum and R. pentaphylla on the score of kainic acid induced status epilepticus
The total score of status epilepticus was high in kainic acid group (Figure 1). C. cyminum and R. pentaphylla groups presented similar levels of scores but a two- thirds reduction compared to kainic acid group score (p<0.001).
Figure 1: Different phases of tonico-clonic seizures caused by administration of kainic acid (upper row); Effect of oral pretreatment with total extract of C. cyminum (Cc) (300 mg / kg) and R. pentaphylla (300 mg / kg) on the score of seizures induced by intrahippocampal kainic acid injection (lower row). Mean ± S.E.M. (n = 5 per group). ***p<0.001 compared to KA; +++ p<0.001 compared to control
Latency of the first seizure
Microinjection of kainic acid into the hippocampus elicited seizures with a latency of 1 min, while a pretreatment with either C. cyminum or R. pentaphylla delayed this latency to 5 and 7 min respectively. These observed increases in the latency of the first seizure in the C. cyminum and R. pentaphylla groups were statistically significant (p <0.001) (Figure 1).
Total duration of seizures
The mean total duration of seizures induced by kainic acid microinjections outlasted 28 min (1700s) during the 90 min observation period. Pretreatment with either C. cyminum (300 mg/kg) or R. pentaphylla (300 mg/kg) significantly reduced kainic acid-induced total seizure duration (p<0.001) (Figure 1).
Percentage of tonic-clonic seizures
In the kainic acid group, the percentage of tonic-clonic seizures was 31% of the total observation time However, in the C. cyminum group or the R. pentaphylla group tonic-clonic seizures percentage was near zero. This result confirmed the reduced seizure severity evaluated by scorings.
Discussion
In the present study, the antiepileptic activity of C. cyminum and R. pentaphylla was assessed using the kainic acid model, which is characterized by status epilepticus. Kainic acid is a potent and selective agonist of glutamatergic receptors (Coyle, 2007). It is well-established that the mechanism of kainic acid-induced epilepsy involves increased production of reactive oxygen species (ROS), mitochondrial dysfunction, and neuronal apoptosis in various brain regions, particu-larly the hippocampus (Zhang and Zhu, 2011).
These results demonstrated that pretreatment with C. cyminum or R. pentaphylla significantly reduced the seizure parameters, including seizure duration and intensity, prolonged the time to onset of seizures, and decreased the mortality rate compared to the group pretreated with kainic acid alone. The antiepileptic effect of R. pentaphylla can be attributed, at least in part, to the pharmacological effects of polyphenols, which possess neuroprotective and antiepileptic properties. Additionally, R. pentaphylla leaves exhibited acetylcholinesterase inhibitor activity, as reported in previous studies (Mansour et al., 2011). acetylcholinesterase inhibitors have been shown to exert anticonvulsant activity by enhancing cortical GABA transmission, as demonstrated by Gersner et al. (2015). This suggests a potential upregulation of acetylcholinesterase that facilitates the release of GABAergic signaling, normalizing cortical excitation/inhibition ratio, and implying a possible role for acetylcholinesterase in status epilepticus treatment.
The findings regarding the effects of C. cyminum extract align with a study conducted by Hosseinzadeh et al. (2002), which confirms the anticonvulsant properties of C. cyminum's aqueous extracts, ethanol, and essential oil in a rat model of epilepsy induced by pentylenetetrazole. Several studies have identified phytochemical constituents in C. cyminum extract, including flavonoids, alkaloids, terpenoids, and tannins (Kabuto et al., 1992). These compounds may explain the protective effect against severe and prolonged tonic-clonic seizures induced by kainic acid. Flavonoids exert antiepileptic activity by acting on the GABA receptor and potentiating its effect (Diniz et al., 2015). Among these compounds, particular attention has been given to 6-bromoflavone and 6-bromo-3'-nitroflavone, which exhibit specific affinity for benzodiazepine receptors (receptors coupled to GABAA receptors) (Medina et al., 1997). These receptors are targeted by effective therapeutic agents widely prescribed for their anxiolytic, muscle relaxant, sedative-hypnotic, and anticonvulsant effects. Thus, C. cyminum suppresses the convulsive effects of kainic acid through GABAergic processes (Khatibi et al., 2008).
Moreover, several studies have shown that natural flavonoids such as rutin, quercetin, and isoquercitrin, which are commonly found in edible plants including C. cyminum and R. pentaphylla (Shamsiev et al., 2021; Fadhil et al., 2021), possess neuroprotective and anti-convulsant properties by attenuating kainic acid-induced oxidative stress in experimental models of epilepsy (Choudhary et al., 2011; Nassiri et al., 2013). According to the present results, pretreatment with C. cyminum and R. pentaphylla reduced the effects of kainic acid, indicating their potential in seizure management and providing partial validation of their use in traditional medicine.
Furthermore, in status epilepticus, ROS plays a significant role in mitochondrial dysfunction and neuronal death (Wu et al., 2019). Antioxidant therapies have a long history of effectively treating epilepsy by reducing oxidative stress (Forman and Zhang, 2021). Numerous animal studies have also confirmed the efficacy of antioxidant treatments, indicating their antiepileptic effects in animal models of epilepsy (Golechha et al., 2011; Ambrogini et al., 2014). The present results further demonstrate the high antioxidant activity of the studied extracts in vitro, suggesting another potential pathway for protecting against epileptic seizures.
Based on the present findings, pretreatment with C. cyminum and R. pentaphylla attenuated the effects of kainic acid, providing evidence of their efficacy against seizures. It can be concluded that the anticonvulsant effect of C. cyminum and R. pentaphylla is likely mediated by multiple mechanisms and molecules. These findings suggest that a high dietary or supplemental intake of these plant extracts may reduce the risk of seizures in epileptic patients. However, further studies are necessary to identify the active compounds and determine their exact mechanisms of action to minimize potential side effects.
Conclusion
The pretreatment with C. cyminum or R. pentaphylla extracts offers attenuation of the severe seizures induced by kainic acid. This observed anticonvulsant effect might be due, at least in part, to the protective antioxidant effects of extracts against oxidative stress induced by kainic acid.
Ethical Issue
All tests were carried out in compliance with the European Community Guidelines (EEC directive 86/609/EEC, dated November 24, 1986). All attempts were made to keep animal suffering to a minimum and to decrease the number of animals used.
Acknowledgement
Authors are grateful to Mr. Abderrazzak Regragi expert in animal laboratory handling for his assistance in this study.
References
Adkar P, Jadhavn P, Ambavade S, Shelke T, Bhaskar V. Protective effect of leaf extract of Pandanus odoratissimus Linn on experimental model of epilepsy. Int J Nutr Pharmacol. 2014; 4: 81-87.
Aitbaba A, Sokar Z, Chait A. Analgesic and anti-inflammatory effects of hydroalcoholic extract of Astragalus ibrahimianus. Bangladesh J Pharmacol. 2023; 18: 41-48.
Ait Laaradia M, Oufquir S, El Hidan MA, Marhoume F, Laadraoui J, Bezza K, El Gabbas Z, Aboufatima R, Boumezzough A, Chait A. Assessment of the relationship between the amount of scorpion venom in the central nervous system and the severity of scorpion envenomation in rats. Toxin Rev. 2021; 40: 179-88.
Al-snafi AE. The pharmacological activities of Cuminum cyminum: A review. J Pharm. 2016; 6: 46-65.
Ambrogini P, Minelli A, Galati C, Betti M, Lattanzi D, Ciffolilli S, Piroddi M, Galli F, Cuppini R. Post-seizure α-tocopherol treatment decreases neuroinflammation and neuronal degeneration induced by status epilepticus in rat hippocampus. Mol Neurobiol. 2014; 50: 246-56.
Banso A, Adeyemo S. Phytochemical screening and antimalarial assessment of Abutilon mauritianum, Bacopa monnifera and Datura stramonium. Biokemistri 2006; 18: 39-44.
Bezza K, El Gabbas Z, Laadraoui J, Laaradia MA, Oufquir S, Chait A. Ameliorative potential of Anacyclus pyrethrum extract in generalized seizures in rat: Possible cholinergic mediated mechanism. Bangladesh J Pharmacol. 2019; 14: 188-195.
Brunetti C, DI Ferdinando M, Fini A, Pollastri S, & Tattini M. Flavonoids as antioxidants and developmental regulators: Relative significance in plants and humans. Int J Mol Sci. 2013; 14: 3540-55.
Chen B, Choi H, Hirsch LJ, Katz A, Legge A, Buchsbaum R, Detyniecki K. Psychiatric and behavioral side effects of antiepileptic drugs in adults with epilepsy. Epilepsy Behav. 2017; 76: 24-31.
Choudhary N, Bijjem KR, Kalia AN. Antiepileptic potential of flavonoids fraction from the leaves of Anisomeles malabarica. J Ethnopharmacol. 2011; 135: 238-42.
Chouhan H, Purohit A, Ram H, Chowdhury S, Kashyap P, Panwar A, Kumar A. The interaction capabilities of phytoconstituents of ethanolic seed extract of cumin (Cuminum cyminum L.) with HMGâ€CoA reductase to subside the hypercholesterolemia: A mechanistic approach. Food Frontiers. 2022; 3: 300-15.
Coyle JT. Kainic acid: Insights into excitatory mechanisms causing selective neuronal degeneration. In: Ciba Foundation Symposium 126â€Selective Neuronal Death: Selective neuronal death: Ciba Foundation Symposium. 2007; 126: 186-203.
Daneshkhah M, Setorki M. Effect of Artemisia persica on seizure severity and memory and learning disorders in pentylenetetrazole-kindled mice. Bangladesh J Pharmacol. 2019; 14: 36-44.
Diniz TC, Silva JC, Lima-Saraiva SRGD, Ribeiro FPRDA, Pacheco AGM, de Freitas RM, Almeida JRGDS. The role of flavonoids on oxidative stress in epilepsy. Oxid Med Cell Longev. 2015; 2015.
Dohou N, Yamni K, Tahrouch S, Massani LMI, Badoc A, Gmira N.. Screening phytochimique d’une endémique Libéro-Marocaine, Thymelaea luthroïdes. Bull Soc Pharm Bordx. 2003; 142: 61-78.
Fadhil H, Mraihi F, Zouied D, Ayadi MT, Cherif JK. Anticorrosion inhibition behavior of Rhus pentaphylla fruit extracts in (1 M) HCl against carbon steel and their chemical characterization using HPLC-MS-ESI. Chem Select. 2021; 6: 5281-89.
Ferr B, Gervasoni D, Vogt C. PreÌcis de neurochirurgie steÌreÌotaxique appliqueÌe aux rongeurs de laboratoire. Paris, Lavoisier, 2012, pp 122-27.
Forman HJ, Zhang H. Targeting oxidative stress in disease: Promise and limitations of antioxidant therapy. Nat Rev Drug Discov. 2021; 20: 689-709.
Gersner R, Ekstein D, Dhamne SC, Schachter SC, Rotenberg A. Huperzine A prophylaxis against pentylenetetrazole-induced seizures in rats is associated with increased cortical inhibition. Epilepsy Res. 2015; 117: 97-103.
Golechha M, Chaudhry U, Bhatia J, Saluja D, Singh Arya D. Naringin protects against kainic acid-induced status epilepticus in rats: Evidence for an antioxidant, anti-inflammatory and neuroprotective intervention. Biol Pharm Bull. 2011; 34: 360-65.
Hosseinzadeh H, Ramezani M, Fadishei M, Basirat M. Anticonvulsant effects of Cuminium cyminum L. seeds extracts and essential oil in mice. J Med Plant. 2002; 1: 9-14.
Joshi A, Bhobe M, Saatarkar A. Phytochemical investigation of the roots of Grewia microcos Linn. J Chem Pharm Res. 2013; 5: 80-87.
Kabuto H, Yokoi I, and Mori A. Monoamine metabolites, iron induced seizures, and the anticonvulsant effect of tannins. Neurochem Res. 1992; 17: 585-90.
Khatibi A, Haghparast A, Shams J, Dianati E, Komaki A, Kamalinejad M. Effects of the fruit essential oil of Cuminum cyminum L. on the acquisition and expression of morphine-induced conditioned place preference in mice. Neurosci. Lett. 2008; 448: 94-98.
Mansour HB, Yatouji S, Mbarek S, Houas I, Delai A, Dridi D. Correlation between antibutyrylcholinesterasic and antioxidant activities of three aqueous extracts from Tunisian Rhus pentaphyllum. Ann Clin Microbiol Antimicrob. 2011; 10: 1-9.
Medina JH, Viola H, Wolfman C, Marder M, Wasowski C, Calvo D, Paladini AC. Overview– Flavonoids: A new family of benzodiazepine receptor ligands. Neurochem Res. 1997; 22: 419-25.
Nassiri-Asl M, Farivar TN, Abbasi E, Sadeghnia HR, Sheikhi M, Lotfizadeh MPB. Effects of rutin on oxidative stress in mice with kainic acid-induced seizure. J Integr Med. 2013; 11: 337-42.
Okpo SO, Fatokun F, Adeyemi OO. Analgesic and anti-inflammatory activity of Crinum glaucum aqueous extracts. J Ethnopharmacol. 2001; 78: 207-11.
Oyaizu M. Studies on products of browning reaction anti-oxidative activities of products of browning reaction prepared from glucosamine. Jpn J Nutr. 1986; 44: 307-15.
Paxinos G, Watson C. The rat brain in stereotaxic coordinates. London. Elsevier, 2006.
Racine RJ. Modification of seizure activity by electrical stimulation: II. Motor seizure. Electroencephalogr Clin Neurophysiol. 1972; 32: 281-94.
Reddy AJ, Dubey AK, Handu SS, Sharma P, Mediratta PK, Ahmed QM, Jain S. Anticonvulsant and antioxidant effects of Musa sapientum stem extract on acute and chronic experimental models of epilepsy. Pharmacog Res. 2018; 10: 49.
Sardari S, Amiri M, Rahimi H, Kamalinejad M, Narenjkar J, Sayyah M. Anticonvulsant effect of Cicer arietinum seed in animal models of epilepsy: Introduction of an active molecule with novel chemical structure. Iran Biomed J. 2015; 19: 45.
Shamsiev A, Park J, Olawuyi IF, Odey G, Lee W. Optimization of ultrasonic-assisted extraction of polyphenols and anti-oxidants from cumin (Cuminum cyminum L.). Korean J Food Preserv. 2021: 28: 510-21.
Talavéra S, Felgines C, Texier O, Besson C, Gil-Izquierdo A, Lamaison JL, Rémésy C. Anthocyanin metabolism in rats and their distribution to digestive area, kidney, and brain. J Agric Food Chem. 2005; 53: 3902-08.
Wu Y, Chen M, Jiang J. Mitochondrial dysfunction in neurodegenerative diseases and drug targets via apoptotic signaling. Mitochondrion 2019; 49: 35-45.
Yimer EM, Tuem KB, Karim A, Ur-Rehman N, Anwar F. Nigella sativa L. (black cumin): A promising natural remedy for wide range of illnesses. Evid Based Complement Altern Med. 2019; 2019.
Youdim KA, Shukitt-Hale B, Joseph JA. Serial Review: Flavonoids and isoflavones (Phytoestrogens: Absorption, metabolism, and bioactivity). Free Radic Biol Med. 2004; 36: 1683-93.
Zadali R, Baghery M, Abbasi M, Yavari N, Miran M, Ebrahimi SN. Anticonvulsant activity of Iranian medicinal plants and molecular docking studies of isolated phytochemicals. S Afr J Bot. 2022; 149: 646-57.
Zhang XM, and Zhu J. Kainic acid-induced neurotoxicity: Targeting glial responses and glia-derived cytokines. Curr Neuropharmacol. 2011; 9: 388-98.
