Identifying the neural ensembles mediating fear generalization during adolescence

PROJECT SUMMARY / ABSTRACT A core symptom observed in anxiety disorders and post-traumatic stress disorder (PTSD) is increased fear generalization, as defined by the overgeneralization of fear from a conditioned, fear-inducing stimulus to neutral stimuli. Fear generalization can lead to maladaptive responses in a safe environment. Our laboratory uses a behavioral assay called contextual fear discrimination (CFD) to assess fear generalization in mice. We shock a mouse in one context (context A), and then we can measure whether they can discriminate between this aversive, shock-paired context and a similar, but safe context (context B) over approximately 10 days of discrimination learning. In our recent publications, we show that 1) adolescent male, but not adolescent female mice overgeneralize fear; 2) adult female, but not adult male mice overgeneralize fear; and 3) aged mice of both sexes overgeneralize fear and cannot discriminate between two contexts. Therefore, there is clearly an age- and sex-dependent influence on fear overgeneralization. However, the neural ensembles mediating fear generalization across the lifespan have yet to be identified. In this grant proposal, we will use this CFD paradigm, in combination with an activity-dependent tagging genetic mouse line to identify and pharmacologically manipulate the neural ensembles mediating fear generalization. In Aim 1, we will identify and quantify the neural ensembles mediating fear generalization in adolescent and adult mice by utilizing the ArcCreERT2 x EYFP mice. This mouse line allows for the indelible labeling of cells expressing the immediate early gene (IEG) Arc/Arg3.1 and allows for a comparison between the cells that are activated during the encoding of a memory and those that are activated during the retrieval of the corresponding memory, with the overlap between the two populations being defined as an engram or memory trace. Therefore, using our activity-dependent tagging mouse line, we can determine where and how sex and age impact fear generalization neural ensembles throughout the brain. In Aim 2, we will pharmacologically manipulate adolescent and adult mice with the goal of improving behavioral fear overgeneralization and the corresponding neural circuits. In the last two decades, (R,S)- ketamine has emerged as a rapid-acting antidepressant. Interestingly, we have reported that (R,S)-ketamine decreases fear generalization. Here, male and female adolescent and adult mice will be administered a single dose of saline or (R,S)-ketamine. CFD and ex vivo whole-brain imaging will be used in order to identify the underlying neural ensembles. The outcome of this study will provide direct evidence that altered neural ensembles result in fear overgeneralization, and that (R,S)-ketamine has the potential to decrease fear overgeneralization, a core system of anxiety disorders and PTSD.