Page 88 - AN-4-1

Advanced Neurology Tetrapleura tetraptera protects the hippocampus

body weight was administered intraperitoneally to the twitching; stage 2 – Myoclonic jerks; stage 3 – Myoclonic
rats. Sodium valproate (Sanofi-Aventis, United Kingdom) jerks with rearing; stage 4 – Forelimb clonus; stage
45
tablets (200 mg) were dissolved in distilled water, and a 5 – Seizures with rearing, rolling over to the side, and
dose of 200 mg/kg body weight was administered orally. generalized clonic-tonic seizures; stage 6 – Death. The rats
were monitored for 24 h after the last administration to
2.4. Phytochemical screening of T. tetraptera fruit check for mortality. The percentage of quantal protection

Qualitative and quantitative analyses of the phytochemicals against seizures and death was calculated. Quantal
present in the ground T. tetraptera fruits were conducted protection was determined as the number of live rats per
following standard methods. 47 group divided by the total number of rats in the group. 45
2.5. Determination of median lethal dose of 2.8. Spontaneous alternation behavior test
T. tetraptera On day 22, a spontaneous alternation behavior test was
The ethical approval for the lethal dose testing on mice conducted in a T-maze. Briefly, the rats were acclimated
was also granted by the Faculty of Basic Medical Sciences in the test room for one h before the test commenced. The
Research and Ethical Committee. To assess the toxicity of T-shaped maze consisted of two short, goal arms (left and
the T. tetraptera fruit extract, an oral median lethal dose was right) and one long, start arm. Gates were placed at the
carried out on 25 CD-1 mice, following the Organization junctions of the arms and were opened when the rat was
for Economic Cooperation guidance document. The placed in the start arm. The rats were allowed to choose
up-and-down procedure was used to determine the median between entering either of the short arms.
lethal dose. Briefly, the 25 CD-1 mice were assigned to Each animal underwent five successive trials, each
43
five groups (n = 5). After overnight fasting, the mice were lasting 60 s. When all four paws of the rat entered one of
administered the extract, with water provided ad libitum. the short arms, the gate was closed, and an arm entry was
Two hours after administration, the mice were allowed recorded before the rat was returned to the holding cage.
access to food. In Group 1, T. tetraptera was administered If a rat did not enter a goal arm within the allotted time,
43
at 1,000 mg/kg, and the mice were observed for signs of the trial was scored as blank, and the rat was returned to
toxicity. After 48 h, with no deaths recorded, the procedure the holding cage. After each trial, the T-maze arena was
was repeated for the second through fifth groups at doses cleaned with 70% ethanol. The frequency with which the
of 2,000, 3,000, 4,000, and 5,000 mg/kg, respectively. As rats alternated between the short arms and the total trial
no signs of toxicity or death were recorded during the duration were recorded. The percentage alternation was
experiment, the dosing procedure was discontinued, and then calculated. 11,48
the lethal dose was estimated.
2.9. Termination of the experiment
2.6. Administration of T. tetraptera, sodium Ketamine hydrochloride (Rotex Medica, Germany)
valproate, and PTZ
was administered intraperitoneally as an anesthetic
The T. tetraptera fruit extract (2 g) was dissolved in 20 mL agent at a dose of 50 mg/kg. The thoracic cavity of each
distilled water. Calculated doses of 250 (5%), 500 (10%), rat was opened, and a cannula was inserted into the left
and 1,000 (20%) mg/kg of the median lethal dosage ventricle of the heart for perfusion fixation using 10%
(LD ) of T. tetraptera fruit extract were administered. In buffered formalin. The entire brain was removed and
50
addition, 200 mg/kg sodium valproate was administered post-fixed in 10% buffered formalin for 48 h, followed by
orally, 1 h before the PTZ intraperitoneal injections, which processing for paraffin wax sectioning for histological and
were given on alternate days (every 48 h) until day 21. The immunohistochemical analysis.
administration schedule is shown in Table 1.
2.10. Nissl substance and immunohistochemistry
2.7. Kindling induction and determination of seizure protocols
scores
Briefly, the left dorsal hippocampal brain region of the
To induce kindling, 40 mg/kg PTZ was administered rats was routinely dehydrated using a graded series of
intraperitoneally every 48 h. Kindling was achieved ethanol, cleared in xylene, and embedded in paraffin wax.
on alternate day 21. Following the administration of Serial sections (10 μm thick) were mounted on slides
PTZ, seizure scores, and mortality rates were observed and were rehydrated for both Nissl substance staining
and recorded for up to 30 min. The seizure stage and and immunohistochemistry. For the Nissl substance
accompanying behaviors were noted using the following study, the sections were stained with 1% cresyl violet,
Racine scale: Stage 0 – No response; stage 1 – Ear and facial supplemented with a few drops of acetic acid, for 10 min.

Volume 4 Issue 1 (2025) 82 doi: 10.36922/an.6862