Project Summary/Abstract Chronic stress is comorbid with a wide spectrum of conditions, such as anxiety, depression, sleep disturbances, and pain. Chronic stress plays a crucial role in exacerbating pain and the loss of quality of life. The mechanisms by which stress exacerbate pain, and how these mechanisms contribute to nervous system pathology, are poorly understood. The ultimate goal of this proposal is to rectify this deficiency. In this application, we study how interoceptive and somatic pain inputs interact within lateral parabrachial (PBl) circuits to affect behavioral responses to pain. We focus on interoceptive inputs from catecholaminergic neurons with the caudal mediodoral nucleus of the solitary tract (NTS) because these neurons play a pivotal role in facilitating behavioral responses to stress and emotional stimuli. We Hypothesize that chronic stress potentiates catecholaminergic NTS (NTScat) inputs to PBl and amplifies nociceptive signaling within PBl, resulting in enhanced behavioral responses to pain. We propose 3 aims: Aim 1: Noxious stimuli activate the NTScat èPBl circuit in naïve animals. We predict that: (1A) Noxious stimuli enhances NTScat neuronal activity; (1B) Optogenetic activation of NTScat neurons causes NA release in PBl; and (1C) Reversible inactivation of NTScat neurons results in diminished release of NA in PBl in response to noxious stimulation. Aim 2: Chronic stress alters dynamics of NA release from NTScat terminals in PBl. In an animal model of chronic stress we predict: (2A) NA release in PBl evoked by noxious stimulation is enhanced by chronic stress; (2B) PBl neuron responses to noxious stimuli are enhanced by chronic stress; inactivation of NTScat inputs to PBl attenuates these responses; and (2C) Suppression of NTScat inputs to PBl attenuates the effects of chronic stress on pain-related behaviors. Aim 3: NA released by NTScat terminals presynaptically regulates sensory and affective pain inputs to PBl and increases its excitability. We predict that NA released from NTScat afferents: (3A) Has a prolonged faciliatory effect on spontaneous synaptic activity in PBl; (3B) Suppresses inhibitory inputs from the central nucleus of the amygdala to PBl; and (3C) Facilitates excitatory inputs to PBl neurons from superficial dorsal horn afferents.