2023 Undergraduate Research Symposium: Mahmoud Teran The development of memories, including those throughout extinction learning, require neuroplasticity for encoding and retrieval to occur. As environmental factors can affect gene expression, epigenetic changes can play a critical role in the development of memory formation and updating and ultimately manifesting in behavioral changes reflective of the learning. The purpose of the current project is to show the utility of Positron Emission Tomography (PET), a functional, molecular imaging technique that employs radioactive substances to measure specific markers in the brain. As part of our efforts to understand the neurobiological mechanisms of fear learning and the role of epigenetic changes using PET, images of the brains were collected before and after extinction learning of acquired long-term fear (LTF) memories induced by shock tone pairings made every 30 seconds over 38 min. PET images were collected by injecting the radiotracer [18F]TFAHA, a substrate for class IIa histone deacetylases (HDACs). The resulting accumulation of the breakdown product of [18F]TFAHA could be considered as a proxy for class IIa HDAC expression changes before and after LTF memory. HDAC activity is directly related to gene expression changes through deacetylation of histone lysine residues. This results in a closed chromatin conformation and prevents the binding of RNA polymerase II and ultimate gene transcription and protein synthesis. This can then be manifested and seen as behavioral change. Our preliminary analysis of PET images results indicate a significant interaction effect between pre-to-post class IIa HDAC expression-activity, sex, and shock condition. Additional PET image analysis is anticipated to show the extent of HDAC activity in both the sexes following LTF.

1. Utilization of In vivo Longitudinal PET
Imaging to Assess Class IIa HDAC Activity
following Long-Term Extinction Fear Memory
in Female and Male Rats
Department of Psychiatry & Behavioral
Neurosciences
Presented by: Mahmoud Teran

2. Background
● Development of memories, including those throughout extinction
learning, require neuroplasticity for encoding and retrieval to occur
● Environmental factors can affect gene expression, epigenetic
changes can play a critical role in the development of memory
formation and updating
● Epigenetic changes related to learning ultimately manifest in
behavioral changes reflective of the learning

3. Background - Epigenetics & class IIa HDACs
Park and Kim Experimental & Molecular Medicine (2020) 52:204–212
● Environmental factors can affect
gene expression, repressing or
expressing genes. This is
regulated by HDACs and HATs
● Changes in activity levels of
HDACs is indicative of learning,
specifically related to acquired
long-term fear (LTF) memories

4. Purpose
To assess changes in class IIa HDACs before and after
extinction learning via PET imaging to understand the
neurobiological mechanisms of extinction learning of
acquired long-term fear memories in female and male
rodents

5. Methods - Subjects
● Species: Wild type Wistar Rat
● Radiotracer: [18F] TFAHA
● Imaging conducted: PET/CT
● Co-registration rat brain atlas:
Px. Rat W. Schiffer
Courtesy of Morgan M. Glover

6. Methods - Experimental Design
Figures are courtesy of Morgan M. Glover
Day 1: Baseline PET imaging
Day 11: Habituation – Context A, 10m, no additional stimuli
Day 12: Acquisition – Context A, 8m50s, five 10s tones co-terminating with 1s 1mA
footshock
Days 13 – 32: Consolidation Period – Pair-housed, handled every few days
Day 33: Fear conditioning recall or Extinction Learning– Context B, 38m, thirty 10s
tones
Day 33: PET imaging – Post-behavior

7. [18F]-TFAHA-used as HDAC PET Tracer
● A substrate for class IIa histone deacetylases (HDACs)
● Resulting accumulation of the breakdown product of [18F]TFAHA could be
considered as a proxy for class IIa HDAC expression changes before and
after long-term fear memory
Tang et al. American Journal of Nuclear Medicine and Molecular
Imaging (2014) 4. 324-332.

8. Rats subjected to shock show increased freezing behavior
during acquisition
Three-way ANOVA
Sex (F vs M): F(1, 10) = 1.50, p = 0.249
Group (NS vs S): F(1, 10) = 16.68, p = 0.002**
ITI x Group: F(4, 40) = 5.032, p = 0.002**
Signif. Codes: ‘^’ 0.1, ‘*’< 0.05, ‘**’< 0.01
Three-way ANOVA
Sex (F vs M): F(1, 10) = 0.03554, p = 0.854
Group (NS vs S): F(1, 10) = 6.374, p = 0.03*
ITI (binned) x Group: F(3, 30) = 1.594, p < 0.001**
ITI x Group: F(19, 190) = 5.025, p < 0.001**
Signif. Codes: ‘^’ 0.1, ‘*’< 0.05, ‘**’< 0.01
Figures are courtesy of Morgan M. Glover
Acquisition Extinction Learning

9. PET imaging allows for visualization of changes in brain activity
Show the utility of Positron Emission Tomography (PET), a functional,
molecular imaging technique that employs radioactive substances to
measure specific markers in the brain.
Anterior Dorsal Hippocampus
Cerebellum
D) Courtesy of Morgan M.
Glover
A) Coronal view
C) Sagittal View
B) Axial view
D)
(A, B, and C) Source:

10. Decreasing expression-activity levels from pre-to-post PET scans for
males and no decrease for shock females indicating that different
mechanisms between sexes regulate fear extinction
Three-way ANOVA
Sex (F vs M): F(1, 4) = 113.5, p < 0.001**
Group (NS vs S): F(1, 4) = 0.00283, p = 0.963
pre/post x Group: F(1, 4) = 109.7, p < 0.001**
pre/post x Sex: F(1, 4) = 124.0, 0.001**
pre/post x Group x Sex: F(1, 4) = 41.21, p < 0.003**
Signif. Codes: ‘^’ 0.1, ‘*’< 0.05, ‘**’< 0.01
Figures are courtesy of Morgan M. Glover

11. Conclusion
● Behavioral results indicate female and male shock subjects acquired and
extinguished fear behavior, but no sex difference
● Preliminary analysis of PET imaging results indicate a significant interaction effect
between pre-to-post class IIa HDAC expression-activity, sex, and shock condition
● With decreasing expression-activity levels from pre-to-post PET scans for males
and no decrease for shock females, this could indicate that different mechanisms
between sexes regulate fear extinction
● Additional PET image analysis is anticipated to show the extent of HDAC activity in
both the sexes following LTF.

12. Acknowledgments
I would like to express my special thanks of gratitude to my lab P.I. Dr.
Shane Perrine for giving me this opportunity. I would also like to thank Dr.
Srinivasu Kallakuri, Dr. Cameron Davidson, Mr. Todd Sasser, and Morgan M.
Glover for all of their help and assistance.
Thank You!

13. Sources
Park, SY., Kim, JS. A short guide to histone deacetylases including recent progress on class II enzymes. Exp Mol Med 52,
204–212 (2020). https://doi.org/10.1038/s12276-020-0382-4
Tang, Wayland & Kuruvilla, Sharon & Galitovskiy, Valentin & Pan, Min-Liang & Grando, Sergei & Mukherjee, Jogeshwar.
(2014). Targeting histone deacetylase in lung cancer for early diagnosis: (18)F-FAHA PET/CT imaging of NNK-treated A/J
mice model. American Journal of Nuclear Medicine and Molecular Imaging. 4. 324-332.