Presented by Justin P. Smith AKT/GSK-3 β / β -CATENIN SIGNALLING WITHIN HIPPOCAMPUS AND AMYGDALA REFLECTS GENETICALLY DETERMINED DIFFERENCES IN POSTTRAUMATIC STRESS DISORDER LIKE SYMPTOMS Presented by Justin P. Smith

PTSD Review Traumatic Event Persistent exaggerated fear responses, avoidance behavior, hyper-arousal, and emotional numbing .33% of PTSD patients have symptoms for 10+ years despite therapeutic interventions

Background Evidence for genetic & environmental factors β-catenin required in amygdala for normal consolidation, but not acquisition of fear memories* Suggesting β-catenin modulates synaptic remodeling and stabilization of long-term memory* *Maguschak & Ressler Nat Neurosci 2008

Background cont. Inescapable foot shock- their model of PTSD Maternal inexperience as potential risk factor B6N – PTSD susceptible, increased contextual and sensitized fear response B6JOIa – PTSD resilient Maguschak and Ressler paper suggests that the transcription factor β-catenin plays a similarly important role in consolidation of fear memories at the level of the basolateral amygdala

Mechanism GSK-3 β (glycogen synthase kinase -3 β) controls β-catenin “Normal” conditions: GSK-3 β phosphorylates (℗) β-catenin to decrease activity AKT ℗ inactivates GSK-3 β ℗ which stabilizes β-catenin Stabilized levels of β-catenin then mediates gene transcription

Wada 2009

Why-What Evidence for role of kinases and transcription factors, little info about activity changes that last more than a couple of hours (exempli gratia PTSD) This study looks at AKT/GSK-3 β / β -catenin-cascades within the dorsal hippocampus and basolateral amygdala (BLA) in the long-term aftermath of exposure to an inescapable foot shock

Animals Male B6N (N) & B6JOIa (J) mice Embryo-transfers (Donor/Recipients) (Embryo/Mom) B6N donors and B6N recipients (N/N), B6N donors and B6JOla recipients (N/J), B6JOla donors and B6JOla recipients (J/J), B6JOla donors and B6N recipients (J/N)

Experiments Experiment 1. Male B6N Experiment 2. Male B6N and B6JOla Groups: foot shock, unshocked (cage control). tested for sensitized and contextual fear (28 days after shock) before brain removal (42 days after shock) Experiment 2. Male B6N and B6JOla Within-strain (N/N, J/J) and between-strain (N/J, J/N) embryo transfers Shocked, tested for sensitized and contextual fear (28 days after shock), Western blot analysis hippocampus and amygdala Limited yield of the transfers did not allow inclusion of unshocked controls (no cage controls!)

Box plot Maximum Upper quartile (75th percentile) Median Dr. John Tukey Maximum Upper quartile (75th percentile) Median Lower quartile (25th percentile) Minimum

Fig 1. B6N mice purchased from commercial breeders Fig 1. B6N mice purchased from commercial breeders. Western blot, shock/no shock. Relative to mean levels of non-shock controls. PTSD-like symptoms coincide with changes in kinase and transcription factor activities at remote time points Amygdala

-within-strain (N/N, J/J) -between-strain (N/J, J/N) -N- PTSD mice Fig.2 Fear behavior in mice originating from commercial breeders or within-strain and between-strain embryo transfers. 28 days after shock. Back in box (Morning) Embryo transfers -within-strain (N/N, J/J) -between-strain (N/J, J/N) -N- PTSD mice -J- Resilient mice Maternal effect Tone (Afternoon) Maternal effect Maternal effect

Fig 3 Fig 4 Embryo Mother Donor Effect

Fig 5 Fig 6 Recipient Effect

Fig. 7. Changes in signaling cascades within the hippocampus.

Fig. 8. Changes in signaling cascades within the amygdala.

Discussion Behavior influenced by both genetic and maternal factors Molecular level results influenced by embryo genotype, not maternal genotype Unclear as to when after foot shock changes occur (different waves of kinase activity?)

Discussion cont. It is tempting to speculate that the changes in kinase activity observed in the present study contribute to this phenotype, similarly to the role of PKA/adenylate cyclase 1, NMDA receptor NR1 subunits and CaMKII in development and/or maintenance of remote contextual fear memory. Are they speculating???

Discussion cont. Fig 2C- _/J had higher sensitized fear than _/N Not statistically secured were separately analyzed (Fig 5C & 6C) AKT/GSK-3 β /β -catenin pathway decreased rather than increased GSK-3 β activity within the amygdala coincides with sustained PTSD-like symptoms the same pathway appears to be regulated in the opposite manner in the amygdala and hippocampus

Take home Identifies lasting changes in AKT/GSK-3 β / β -catenin-cascades in the amygdala and hippocampus B6N’s showed long-term contextual & sensitized fear dHippocampus: increased phosphorylated AKT In BLA: higher levels of phosphorylated AKT and GSK-3 β Increased β -catenin levels Embryo transfer did not alter effect (mother’s genotype had no effect) Shocked B6N levels of phosphorylated GSK-3 β and β -catenin levels were decreased in dHippocampus but increased in BLA compared to shocked B6JOIa

Thank you