Olfactory Function Assessment in Mice

To assess the olfactory function of mice after different treatments, an odor cross-habituation test was performed using a previously described method to detect olfactory defects in mice [36 (link)]. Monomolecular odorants including heptanone, isoamyl acetate, ( +) enantiomer of limonene, and ethyl valerate (Sigma Aldrich, China) were diluted to 1 ×10⁻³ mol/L in mineral oil and coated on a cotton ball. The cotton ball was then wrapped in an odorless plastic bag with small holes and placed in the bottle cap to prevent the odor liquid from reaching the cage or animals, but allow the volatile odor fumes to diffuse. Odor cross-habituation was tested in four 20 s trials by placing a cotton ball on the side of the animal's cage with a 30 s interval between them. During sniffing, the time recording was terminated when the mouse's nose was within 1 cm of the odor of the cotton ball.