Summary of funded grant The hippocampal-prefrontal circuit is implicated in many neuropsychiatric illnesses. This circuit is critically involved in cognition and emotional regulation, and is particularly vulnerable to stress, which is a key precipitating factor for these disorders. Chronic stress has deleterious effects on neuronal structure and physiological function in the hippocampus, and impairs hippocampal-dependent behavior, including processing of contextual fear. The hippocampus (HPC) and prefrontal cortex (PFC) communicate during cognitive and emotional tasks by altering the coherence of oscillatory activity between the two regions. However, the cellular and molecular events that drive changes in HPC-PFC synchrony are not well understood. Neurons projecting from the ventral CA1 region of the HPC provide the major monosynaptic input to the PFC. Many of the risk factors for neuropsychiatric disorders, including stress, affect genes that play important roles in synapse development and plasticity, and disruptions in synaptic connections of the neuronal projections between the HPC and PFC could contribute to impairments in HPC-PFC synchrony. The central hypothesis of this proposal is that cellular and molecular signaling in HPC-PFC projection neurons control their structure and function, and that these signaling pathways regulate patterns of neural activity between the HPC and PFC that influence their connectivity. The goals of this application are to 1) understand how stress drives molecular and cellular signaling in HPC-PFC projection cells to control their physiological function; and 2) determine how plasticity in HPC-PFC projection neurons impacts functional connectivity to control fear-related behavior. The funded application has three aims, designed to reveal fundamental information about molecular and cellular signaling programs in HPC-PFC projection neurons. Aim 1: Identify how stress impacts neural activity in hippocampal-prefrontal circuitry to drive enhanced fear recall; Aim 2: Determine how manipulation of plasticity in hippocampal-prefrontal projection neurons mediates functional connectivity between the HPC and PFC; Aim 3: Define how stress impacts molecular and structural correlates of plasticity in hippocampal-prefrontal projection neurons. We made substantial progress on Aims 2 and 3. Specifically, we successfully and stably overexpressed BDNF in HPC-PFC projectors, and acquired LFPs in HPC and PFC from implanted stereotrodes to assess how BDNF expression impacted connectivity in this circuit, and in parallel, assessed calcium activity in individual PFC neurons during fear recall. Towards the goals of Aim 3 we conducted single-nuclei RNA sequencing (snRNA-seq) in PFC following selective stimulation of both HPC-PFC projection neurons and a comparator projection neuron type - locus coeruleus (LC)-PFC neurons. A key, unexpected finding from these studies, which we validated at cellular resolution with RNAscope techn y, is select...