ASSESSMENT OF ANTI-EPILEPTIC EFFICACY OF HERBAL FORMULATIONS IN A RAT MODEL OF KANIC ACID-INDUCED EPILEPSY

ASSESSMENT OF ANTI-EPILEPTIC EFFICACY OF HERBAL FORMULATIONS IN A RAT MODEL OF KAINIC ACID-INDUCED EPILEPSY

1.0 TEST SYSTEM DETAILS:

Species : Rattus norvegicus (Rats)

Strain : Sprague Dawley

Age : 6-7 weeks

Sex : Male

No. of animals : 10 /Group

Total animals : 60

2.0 TEST ARTICLES DETAILS

HB1: Herbal formulation -1

3.0 ALLOCATION OF GROUPS:

Group No.	Group Description	Disease disease-inducing agent administered	Treatment administered	Dose Volume and Route
G1	Normal Control	Normal Saline administered by intraperitoneal route (i.p.),	0.5% MC, p.o., b.i.d.	5 ml/kg, p.o.
G2	Disease Control	Kanic acid 12 mg/kg (dissolved in Normal Saline), i.p., single dose	0.5% MC, p.o., b.i.d.
G3	Reference Control		Carbamazepine 80 mg/kg, p.o. + 0.5 % MC, p.o.
G4	Treated with a low dose of HB1		HB1: 100 mg/kg, b.i.d.  in 0.5% MC
G5	Treated with an intermediate  dose of HB1		HB1: 300 mg/kg, mg/kg, b.i.d. in 0.5% MC
G6	Treated with high dose 2 of HB1		HB1: 1000 mg/kg,   b.i.d. in 0.5% MC

Abbreviations: MC-Methyl Cellulose, p.o.-per os. q.d.: quaque die; bid: bis in die, i.p.-Intraperitoneal

4.0 METHOD:

Healthy animals will be selected for the study, randomized based on body weight, and assigned to 6 groups of 10 animals each.

• Animals of Group G1 will be designated as normal-control and administered 0.5% MC, p.o., b.i.d.

• Disease control animals (assigned to group G2) will receive 0.5% MC, p.o., b.i.d.

• Animals of group G3 will be treated with reference drug Carbamazepine at the dose of 80 mg/kg, p.o., and will be additionally administered 0.5% MC in the evening.

• Animals of group G4-G6 will be treated with HB1 at different dose levels ranging from 100-1000 mg/kg, b.i.d.

• Compound administration will be initiated 14 days before Kainic acid injection.

• The normal control group (G1) will be administered an injection of Normal Saline by intraperitoneal route. In contrast, animals allocated to groups G2 – G6 will be injected with Kainic acid (12 mg/kg, i.p.) dissolved in Normal Saline.

• Seizure activity will be observed in the animals immediately after the administration of Kainic acid.

• Rats will be subjected to the Morris Water Maze test to evaluate the cognitive deficits on the third day and 10th day post-kainic acid administration. During this period the animals will continue to receive the test articles.

• On Day 11, post-kainic acid administration, animals will be sacrificed under an overdose of thiopentone anesthesia. After suitable anesthesia but before the animal dies, blood will be collected from the retro-orbital plexus to estimate biochemical parameters. Immediately after the animal dies, the brain will be weighed, and half will be fixed in 10% neutral buffered formalin for histopathology. In contrast, the other half will be stored at -80°C for the various biochemical and molecular evaluations.

5.0 PARAMETERS TO BE EVALUATED:

• Body weight: Twice a week

• Mortality post-kainic-acid administration

• Seizure scoring

• Cognitive function

• Brain weight and its relative organ weight.

• Cytokine estimation in serum: IL-1β, TNF-α, and IL-6 (anti-inflammatory)

• Oxidative stress parameters in the brain tissue: Reduced Glutathione, Malondialdehyde, Catalase, Total nitrate, superoxide dismutase Total antioxidant capacity (TAC).

• Histopathological analysis of pancreas (Hematoxylin & Eosin-stained).

• Immunofluorescence staining of the brain tissue

6.0 REFERENCE(S):

1. Friederike Kienzler-Norwood., et al. A novel animal model of acquired human temporal lobe epilepsy based on the simultaneous administration of kainic acid and lorazepam. Epilepsia. Feb;58(2):222-230 (2017).

2. Kuan Ming Chiu., et al. Protective Effects of Bupivacaine against Kainic Acid-Induced Seizure and Neuronal Cell Death in the Rat Hippocampus. Biol. Pharm. Bull. 38, 522–530 (2015).

3. Yogendra K. gupta., et al. Protective effect of curcumin against kainic acid-induced seizures and oxidative stress in rats. Indian J Physiol Pharmacol; 53 (1): 39–46 (2009).

                               

                                               END OF THE DOCUMENT

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