Project Summary/Abstract Post-traumatic stress disorder (PTSD) and opioid use disorder (OUD) are highly comorbid, resulting in worse symptom severity and treatment outcomes than either disorder alone. Individuals with OUD appear to be particularly susceptible to PTSD. Studying each disorder in isolation fails to capture the complexity of the interrelationships between PTSD and OUD. While prior trauma tends to occur before the development of substance use disorders, little is known about the causal mechanism of this relationship. Both PTSD and OUD can be conceptualized as learning disorders, in which opioid- or fear-associated cues gain a strong control over behaviors. In order to advance our understanding of PTSD and OUD comorbidities, the goal of this proposed research is to shed light on the stress-induced neuroadaptations that may promote vulnerability for opioid learning and reward. The Fanselow laboratory has developed a stressor model that recapitulates some of the symptomology of PTSD. In this stressor model, severe stress sensitizes future fear learning. I demonstrated that severe stress additionally enhances future opioid-context learning. Both this stressor model and opioid-context learning are dependent on basolateral amygdala (BLA) activity. Stress enhances the excitability and neural plasticity in the BLA, leading to the intriguing possibility that severe stress fundamentally changes learning processes in the BLA, making individuals more susceptible to opioid reward learning. Using a novel activity-dependent labeling technique in a rat model, the first aim of this proposal will examine whether there is significant overlap in neurons activated by severe stress and morphine-context pairing. Such findings would provide evidence that neurons responsive to severe stress directly engage in future opioid-reward cue learning. The second aim in this proposal will examine if stress-responsive neurons are necessary for stress enhanced opioid learning. This work will promote an interdisciplinary training experience involving a combination of sophisticated behavioral approaches paired with in vivo chemogenetic, in vivo activity-dependent labeling, and ex vivo analysis techniques that will ultimately broaden our understanding of how traumatic stress can lead to development of OUD. These experiments will be conducted at UCLA, an excellent training environment that prioritizes productivity, mentorship, and collaboration, ensuring the success of the project.