Heptanone

5XFAD mice (3.5 or 7.5 months) were orally gavaged for 13 days with 100 mg/kg/day bexarotene. Odorants (heptanone, isoamyl acetate, limonene, and ethyl valerate; Sigma Aldrich, St. Louis, MO) were diluted 1 × 10⁻³ in mineral oil and applied to a cotton-applicator stick enclosed in a piece of plastic tubing32 (link). Odors were delivered for 4 successive trials (1 block), 20 seconds each, separated by 30 second inter-trial intervals, by inserting the odor stick into a port on the side of the animal’s home cage at day 13 of bexarotene treatment32 (link). Testing took place during the light phase of the animals’ (12:12) light:dark cycle. The duration of time spent investigating was defined as snout-oriented sniffing within 1 cm of the odor presentation port, and was recorded across all trials by a single observer blinded to genotype and treatment.

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.