Fear conditioning biases olfactory sensory neuron frequencies across generations

Liff, Clara 1 ; Ayman, Yasmine1; Jaeger, Eliza1; Cardeiro, Avery1; Lee, Hudson1; Kim, Alexis1; Vina-Albarracin, Angelica1; Ferguson, Dianne-Lee1 2; Marlin, Bianca 1 2

Published Apr 08, 2026 on Dryad. https://doi.org/10.5061/dryad.80gb5mm4m

Data files

Apr 08, 2026 version files 87.35 MB

Dryad_Submission.zip

87.34 MB
README.md

11.66 KB

Abstract

We investigated changes in the cellular composition of the olfactory epithelium in response to an aversive stimulus. Using intact, optically cleared main olfactory epithelia from mice that underwent olfactory fear conditioning, we developed a dataset demonstrating that olfactory fear conditioning increases the number of odor-encoding neurons in mice that experience odor-shock conditioning (F0), as well as in their unconditioned offspring (F1). We provide evidence that the increase in F0 is due, in part, to the biasing of the stem cell layer of the main olfactory epithelium. A robust dataset of F1 behavioral metrics revealed subtle odor-specific differences between the offspring of unconditioned and conditioned parents, despite the absence of an active aversion to the conditioned odor.

Dataset DOI: 10.5061/dryad.80gb5mm4m

Description of the data and file structure

There is a CSV file containing the raw data for every data-containing subfigure in the manuscript, along with a "subject_details" CSV file containing details for every animal used to generate that data. Data files are terminated with "data.csv," while subject details files are terminated with "subject_details.csv." The number indicates the associated figure number in the manuscript, while the letter indicates the subfigure. For example, "1A_data.csv" would refer to the data file for Figure 1A. For prefixes containing an "S," the first number refers to the main figure to which the figure supplement is attached, and the second number refers to the figure supplement number. For example, "1S1" refers to Figure 1-Figure supplement 1.

Files and variables

File: Dryad_Submission.zip

Description:

Format of each "subject_details" file:

Condition:
- F0 Naïve: no conditioning
- F0 Unpaired: odor presentation and foot shock were separated by a 60 second delay period during fear conditioning
- F0 Paired: odor presentation was coterminated with a shock during fear conditioning
- F1 Unpaired: naïve offspring from F0 Unpaired fathers
- F1 Paired: naïve offspring from F0 Paired fathers
Data point: the numerical data point that was plotted in the figure (unit descriptions in data files)
Mouse_ID: unique animal identifier
Sex: male or female
Father (if F1): the mouse ID of subject's father, if second generation
Counter: experimenter who performed the count/assay/experiment

Files:

1G_1S1_1S2_subject_details.csv
- Subject details for each mouse in Figures 1G, 1-Figure supplement 1, and 1-Figure supplement 2 (refer to above for format)
1G_data.csv
- M71 (olfactory sensory neuron) cell counts from iDISCO-cleared main olfactory epithelia on day 21 following olfactory fear conditioning, grouped by condition (column name)
1I_1S1_1S2_subject_details.csv
- Subject details for each mouse in Figures 1I, 1-Figure supplement 1, and 1-Figure supplement 2 (refer to above for format)
1I_data.csv
- MOR23 (olfactory sensory neuron) cell counts from iDISCO-cleared main olfactory epithelia on day 21 following olfactory fear conditioning, grouped by condition (column name)
2D_data.csv
- M71 (olfactory sensory neuron) cell counts from iDISCO-cleared main olfactory epithelia on day 21 following olfactory fear conditioning with a control odor, grouped by condition (column name)
2D_subject_details.csv
- Subject details for each mouse in Figure 2D (refer to above for format)
2E_data.csv
- Ratio of MOR23 to M71 (olfactory sensory neuron) cells, grouped by condition (column name)
2E_subject_details.csv
- Subject details for each mouse in Figure 2E (refer to above for format)
3D_data.csv
- Percent EdU-positive M71 (olfactory sensory neuron) cells, grouped by condition (column name)
3D_subject_details.csv
- Subject details for each mouse in Figure 3D (refer to above for format)
3E_data.csv
- Percent EdU-positive MOR23 (olfactory sensory neuron) cells, grouped by condition (column name)
3E_subject_details.csv
- Subject details for each mouse in Figure 3E (refer to above for format)
4C_4S1_data.csv
- M71 (olfactory sensory neuron) cell counts from iDISCO-cleared main olfactory epithelia on days 42 and 63 following olfactory fear conditioning, grouped by condition (column name)
4C_4S1_subject_details
- Subject details for each mouse in Figures 4C, 4-Figure supplement 1 (refer to above for format)
4E_4S2_data.csv
- Acetophenone avoidance indices [(time spent in acetophenone chamber-time spent in control odor chamber)/time spent in either chamber] at day 42, grouped by condition (column name)
4E_4S2_subject_details.csv
- Subject details for each mouse in Figures 4E, 4-Figure supplement 2 (refer to above for format)
4F_4S2_data.csv
- Acetophenone avoidance indices [(time spent in acetophenone chamber-time spent in control odor chamber)/time spent in either chamber] at day 63, grouped by condition (column name)
4F_4S2_subject_details.csv
- Subject details for each mouse in Figures 4F, 4-Figure supplement 2 (refer to above for format)
5C_5S1_5S2_5S3_subject_details.csv
- Subject details for each mouse in Figures 5C, 5-Figure supplement 1, 5-Figure supplement 2, 5-Figure supplement 3 (refer to above for format)
5C_5S1_5S2_data.csv
- Acetophenone avoidance indices [(time spent in acetophenone chamber-time spent in control odor chamber)/time spent in either chamber], grouped by condition (column name)
5E_5S1_data.csv
- Distance traveled (m) during acetophenone trichamber assay, grouped by condition (column name)
5F_5S1_data.csv
- Time freezing (s) during acetophenone trichamber assay, grouped by condition (column name)
5G_5S1_5S2_5S3_subject_details.csv
- Subject details for each mouse in Figures 5G, 5-Figure supplement 1, 5-Figure supplement 2, 5-Figure supplement 3 (refer to above for format)
5G_5S1_5S2_data.csv
- Lyral avoidance indices [(time spent in lyral chamber-time spent in control odor chamber)/time spent in either chamber], grouped by condition (column name)
5I_5S1_data.csv
- Distance traveled (m) during lyral trichamber assay, grouped by condition (column name)
5J_5S1_data.csv
- Time freezing (s) during lyral trichamber assay, grouped by condition (column name)
5K_5S1_5S2_5S3_subject_details.csv
- Subject details for each mouse in Figures 5K, 5-Figure supplement 1, 5-Figure supplement 2, 5-Figure supplement 3 (refer to above for format)
5K_5S1_5S2_data.csv
- Propanol avoidance indices [(time spent in propanol chamber-time spent in control odor chamber)/time spent in either chamber], grouped by condition (column name)
5M_5S1_data.csv
- Distance traveled (m) during propanol trichamber assay, grouped by condition (column name)
5N_5S1_data.csv
- Time freezing (s) during propanol trichamber assay, grouped by condition (column name)
5S3a_data.csv
- Mean speed (m/s) during acetophenone trichamber assay, grouped by condition (column name)
5S3B_data.csv
- Number of entries in control odor and conditioned odor sides during acetophenone trichamber assay, grouped by condition (column name)
5S3C_data.csv
- Time spent (s) in control odor and conditioned odor sides during acetophenone trichamber assay, grouped by condition (column name)
5S3D_data.csv
- Mean speed (m/s) during lyral trichamber assay, grouped by condition (column name)
5S3E_data.csv
- Number of entries in control odor and conditioned odor sides during lyral trichamber assay, grouped by condition (column name)
5S3F_data.csv
- Time spent (s) in control odor and conditioned odor sides during lyral trichamber assay, grouped by condition (column name)
5S3G_data.csv
- Mean speed (m/s) during propanol trichamber assay, grouped by condition (column name)
5S3H_data.csv
- Number of entries in control odor and conditioned odor sides during propanol trichamber assay, grouped by condition (column name)
5S3I_data.csv
- Time spent (s) in control odor and conditioned odor sides during propanol trichamber assay, grouped by condition (column name)
6_l_r_xcutoffs.csv
- x cutoff points (pixels) to delinate entries to left and right chamber entrances, per video, for analyses in Figures 6E-I
  - filename: name of video file
  - left cutoff: x coordinate (in pixels) delineating the boundary between the left chamber and the center chamber
  - right cutoff: x coordinate (in pixels) delineating the boundary between the center chamber and the right chamber
6_moseq_df.csv
- moseq_df for all Figure 6 data, generated by kpMoSeq software (see code/software section below for citation and software access). This file contains syllable labels and kinematic information for each frame of each video analyzed.
  - name: video filename
  - centroid_x: x-coordinate of centroid of subject
  - centroid_y: y-cooridnate of centroid of subject
  - heading: heading (in radians) of subject
  - angular_velocity: angular velocity (in radians/second) of subject
  - velocity_px_s: velocity (pixels/second) of subject
  - syllable: syllable label for frame
  - frame_index: video frame index
  - group:
    - f0_up: F0 (parental generation) unpaired
    - f0_p: F0 paired
    - f1_up: F1 unpaired (naïve offspring of unpaired fathers)
    - f1_p: F1 paired (naïve offspring of paired fathers)
  - onset: true (first frame of syllable) or false (not first frame of syllable)
6_stats_df.csv
- stats_df for all Figure 6 data, generated by kpMoSeq software (see code/software section below for citation and software access). This file contains summary statistics for each syllable in each video.
  - group: f0_up, f0_p, f1_up, or f1_p (see description of 6_moseq_df.csv)
  - name: video file
  - syllable: behavioral syllable (number index)
  - heading_mean: average heading (in radians) of every frame of the syllable in the video
  - heading_std: heading standard deviation of every frame of the syllable in the video
  - heading_min: minimum heading of the syllable in the video
  - heading_max: maximum heading of the syllable in the video
  - angular_velocity_mean: average angular velocity (in radians/second) of every frame of the syllable in the video
  - angular_velocity_std: angular velocity standard deviation of every frame of the syllable in the video
  - angular_velocity_min: minimum angular velocity of the syllable in the video
  - angular_velocity_max: maximum angular velocity of the syllable in the video
  - velocity_px_s_mean: average velocity (in pixels/second) of every frame of the syllable in the video
  - velocity_px_s_std: velocity standard deviation of every frame of the syllable in the video
  - velocity_px_s_min: minimum velocity of the syllable in the video
  - velocity_px_s_max: maximum velocity of the syllable in the video
  - frequency: frequency of the syllable in the video (relative to all other syllables)
  - duration: average duration of the syllable in the video
6_subject_details.csv
- Subject details for each mouse in Figure 6 and 6-Figure supplement 1
  - file: video filename, with the number indicating the subject ID (e.g. "processed_395.analysis" refers to the video file for subject CWL395)
  - odor: the odor used in olfactory fear conditioning
  - p/up: condition
    - p=paired. Odor presentation was coterminated with a shock during fear conditioning
    - up=unpaired. Odor presentation and foot shock were separated by a 60 second delay period during fear conditioning
  - c_o_side: conditioned odor side
    - R: the conditioned odor was presented in the right chamber
    - L: the conditioned odor was presented in the left chamber
  - sex: subject sex
  - gen: generation (F0 or F1)
  - ai: avoidance index. Values range from -1 to 1, with negative values indicating avoidance of the conditioned odor and positive values indicating approach of the conditioned odor.
    - ai = (time spent in conditioned odor chamber-time spent in control odor chamber)/time spent in either chamber

Code/software

kpMoSeq software (Weinreb et al., 2023. doi: 10.1038/s41592-024-02318-2) was used for Figure 6 data. Custom analysis code for Figure 6 is available on Github.