Neurological severity score (NSS)

The NSS was determined and recorded using the following four parameters.

The Zea-longa score was determined by observing movements of the mice in and out of the cage: 0, mice with no neurological deficits; 1, mice could not fully extend the left forepaw; 2, mice circled to the left while walking; 3, mice fell to the left or limped; and 4, mice were unable to walk spontaneously; 5, death [12].

We determined the ability of the mice to grab and hold onto a 0.5 cm diameter wire elevated at a height of 70 cm from the surface. Performance was evaluated using a four-grade score: 0, mice could hang on the wire for over 5 s with hind legs placed on the wire; 1, mice could hang on the wire for 5 s; 2, mice could hang on the wire for 3~4 s; and 3, mice could hang on the wire for 0~2 s. For scores 1~3, the mice were unable to place their hind legs on the wire [13].

Circling behavior was determined by observing and scoring the ability of the mice to remain in contact with the floor with their forelegs: 0, mice had no observable deficits; 1, mice exhibited flexion of the contralateral torso; 2, mice circled clockwise; 3, mice continuously spun clockwise; and 4, no movements or reactions [14].

The mice were held by their tail, and the bending of the torso or forelimbs in the air was observed and scored: 0, normal reaction; 1, mice occasionally exhibited flexion of the asymmetric forelimb; 2, asymmetric forelimb flexion; 3, mice exhibited flexion of the asymmetric forelimb and torso; and 4, no motor activity or reaction [14].