Research indicates that olfactory dysfunction is a notable phenotype of neurodegenerative diseases (e.g., Parkinson’s disease, Alzheimer’s disease) and neurodevelopmental disorders (e.g., autism, epilepsy)^[1,2]. These findings provide critical insights for early disease diagnosis and open new avenues for exploring therapeutic mechanisms^[3].

Olfactory training is a systematic approach that uses diverse odor stimuli to improve olfactory function. It has been widely applied in studies addressing olfactory dysfunction. Regular exposure to a variety of odors has been shown to promote the remodeling of olfactory receptors and associated neural pathways, aiding the recovery of olfactory function and even alleviating symptoms of diseases related to olfactory impairment (e.g., Alzheimer’s disease, autism, and depression)^[4].

Next, we will introduce several common olfactory behavioral experiment methods and demonstrate how the RWD Olfactometer facilitates olfactory discrimination testing, olfactory sensitivity detection, and olfactory training.

Common Olfactory Behavioral Experiment Methods

1. Buried Food Test

Objective: To assess olfactory sensitivity.
Method: Mice are fasted for 24 hours before the experiment. Food pellets are buried in sand or another substrate within a testing chamber. The mouse is then placed in the chamber, and the time it takes to locate the food is recorded. By comparing the time required for different mouse models to find the food, their olfactory function can be evaluated.

2. Habituation/Dishabituation Test

Objective: To assess olfactory discrimination ability.
Method: Mice are first habituated to a specific odor. Afterward, a new odor is presented, and the number of explorations and the time spent investigating the new odor are recorded. If the mouse is able to distinguish the two odors, it will spend more time exploring the novel odor. By comparing the exploration time and frequency of different mouse models, their olfactory discrimination ability can be evaluated.

3. Hole Board Test

Objective: To assess olfactory recognition ability.
Method: Mice are placed on a board with multiple holes, each containing a different odor. The number of times the mouse sniffs each hole is recorded. By comparing the sniffing frequency across different mouse models, the olfactory recognition ability can be evaluated.

4. Odor Preference Test

Objective: To assess the animal’s preference for different odors.
Method: Mice are placed in a test chamber with two odor sources. The time spent near each odor source is recorded. By comparing the time across different mouse models, their odor preference is evaluated.

Although these traditional experimental methods are simple to perform, mice can be influenced by external environmental factors and human intervention during the experiment, which affects the reproducibility, operational efficiency, and compatibility with other research techniques (such as optogenetics, fiber photometry, electrophysiology, etc.).

To address these challenges, RWD has developed the Olfactometer. With precise experimental design, automated data recording, and highly compatible device configurations, this system is not only a reliable tool for olfactory research but also provides a novel behavioral approach for studies in learning, memory, decision-making, and other areas.

RWD Olfactometer Experimental Examples

1.Olfactory Discrimination Test

Method:

Mice are presented with two different odors: a reward odor and a non-reward odor. Licking the water port in response to the reward odor results in a water reward, while licking the port after detecting the non-reward odor yields no reward. The software automatically tracks the correct rate to assess the mice’s olfactory discrimination ability.

System Advantages:

• Customizable random trials with up to 32 different reward and non-reward odor pairs, accommodating complex training needs.
• Fast odor switching and continuous background airflow effectively prevent odor cross-contamination.
• Precise control of odor release time and flow rate, with a water port sensor that accurately detects licking behavior, improving experiment reproducibility.

2.Olfactory Sensitivity Test

Method:

The test odor is diluted in gradient steps, with the solvent used as the non-reward odor. Multiple Go/No-Go programs are created and run, exposing the mice to various concentrations of the odor. The mice’s response accuracy to each concentration is recorded to assess their olfactory sensitivity. The learning time can also be used as a reference to evaluate early disease models, such as Alzheimer’s disease.

System Advantages:

• Supports multiple program combinations, enabling flexible design of concentration gradient stimuli.
• Mice are exposed to odors in a fixed, awake state, minimizing interference from instinctive exploratory behavior and enhancing experimental accuracy and efficiency.
• Event and result data are automatically saved in CSV format, providing clear, quantifiable results.

3.Olfactory Training

Method:

A set of diverse odors (e.g., floral, fruity, and spicy scents) is selected, and the Olfactometer is used to set a sequence of odor release rules. The system precisely controls the odor release time and flow rate, ensuring that each mouse receives standardized odor stimulation. After the training period, behavioral tests and olfactory function assessments are conducted to evaluate whether the mice’s olfactory function and associated disease symptoms have improved.

System Advantages:

• The single-channel olfactometer can precisely control the release of up to 16 different odors in sequence, meeting diverse training needs.
• The specialized software automates the experimental process, reducing manual intervention and ensuring high reproducibility for each training session.
• The system reserves 8 TTL signal ports, supporting integration with optogenetics, fiber photometry, electrophysiology, and two-photon imaging systems to monitor neural signal changes during odor stimulation. This facilitates the exploration of complex mechanisms in conjunction with other technologies.

The Olfactometer is designed to provide more diverse, efficient and precise solutions for neuroscience research. If you’re interested in the system’s features or experimental methods, feel free to contact us to explore more potential applications!

References
[1] Hornix, Betty E., Robbert Havekes, and Martien JH Kas. “Multisensory cortical processing and dysfunction across the neuropsychiatric spectrum.” Neuroscience & Biobehavioral Reviews 97 (2019): 138-151.
[2] Dan, Xiuli, et al. “Olfactory dysfunction in aging and neurodegenerative diseases.” Ageing research reviews 70 (2021): 101416.
[3] Lyons-Warren, Ariel M., et al. “A systematic-review of olfactory deficits in neurodevelopmental disorders: From mouse to human.” Neuroscience & Biobehavioral Reviews 125 (2021): 110-121.
[4] Vance, David E., et al. “Does olfactory training improve brain function and cognition? A systematic review.” Neuropsychology review 34.1 (2024): 155-191.