Hole-Board Test

The hole-board apparatus was used for the assessment of anxiolytic and CNS depressant activity. The apparatus contains sixteen equidistant holes on a black sheet that was placed on a transparent poly acrylic rectangular box. The floor of the box was placed 15 cm above the ground. The mice under study were orally treated with vehicle and diazepam 1 mg/kg, and each extract of L. serriola at a dose of 300, 400, and 500 mg/kg was administered 1 h before recording the number of head dips. Each mouse was placed at the center of the apparatus and was allowed to explore the apparatus liberally for 5 min, and the number of head dips was counted [23]. A head dip was counted only when both eyes of the mouse disappeared in the hole [24]. The apparatus was washed completely with 10% ethanol solution for the next mice.

The elevated plus maze (EPM) test was used for the assessment of anxiety, stress, motor coordination, and exploratory behavior of rodents. The apparatus contains 4 arms, two open and two closed, 49 cm in length and 10 cm broad, arranged in such a way that both arms are opposite to each other in the arrangement and the maze was placed 50 cm above the floor. The mice under study were administered with vehicle, diazepam 1 mg/kg, and with each extract of L. serriola at dosses of 300, 400, and 500 mg/kg orally just 1 h before the test. Each mouse was placed on the platform of EPM and the time spent and the number of entries in open arms was noted for five minutes. An entry was noted when the mouse placed all four paws in the arm [23]. The EPM apparatus was cleaned with 10% ethanol solution after the performance of the test, for the next mice.

The light–dark test was also employed for the assessment of anxiolytic activity of investigational substances. The apparatus consists of a box divided into two compartments, a bright compartment painted white and a dark compartment painted dark. The two compartments are linked through a small opening at the center of both compartments. The mice were administered with vehicle, diazepam 1 mg/kg, and each extract of L. serriola dried seeds at dosses of 300, 400, and 500 mg/kg just 1 h before the test. Each mouse after treatment was placed in the center of the light box and observation was made for five minutes. The time spent by each mouse in the light compartment was recorded [25]. The mouse was returned to the cage after being removed from the box. The apparatus was washed thoroughly with 10% ethanol solution for the next mice.