Project Summary: Many patients suffer from chronic pain in the absence of identifiable injury. Such pains are termed “functional” and include irritable bowel syndrome, temporomandibular joint disorder, fibromyalgia, migraine and others. For reasons that are not understood, almost all functional pain syndromes (FPS) are female prevalent. FPS patients experience pain-free interictal periods punctuated by attacks of pain. The frequency of attacks is predictive of risk of chronification. Pain episodes thus produce a priming effect, establishing a state of increased vulnerability to future attacks, likely reflecting peripheral and central sensitization. FPS patients commonly identify stress as a key trigger of pain. Repeated stress may thus promote vulnerability and pain in a sexually dimorphic fashion. We have developed an injury-free rodent model of FPS based on hyperalgesic priming with repeated stress. Hyperalgesic priming produces a pain-free state of increased vulnerability that has been termed “latent sensitization” (LS). Following induction of LS, normally subthreshold triggers can produce pain attacks, modeling the interictal and ictal periods of FPS. We will use this model to test the novel hypothesis that repeated stress activates kappa opioid receptor (KOR) signaling in the hypothalamus resulting in release of prolactin (PRL) and dysregulation of prolactin receptor (PRLR) isoform expression selectively in female nociceptors. PRL signals through homodimers of PRLR long and short (i.e., PRLR-L and PRLR-S) isoforms that respectively regulate transcription and pain. Repeated stress down-regulates PRLR-L promoting female-selective pain through stress-induced PRL/PRLR-S signaling. The balance of PRLR isoforms may therefore “tune” female nociceptors to promote LS and pain from normally subthreshold stimuli. We will use genetic and chemogenetic manipulations along with anatomical, neurochemical, electrophysiological, pharmacological and behavioral studies in male and female mice to evaluate the role of dorsal root ganglion (DRG) PRLR-L down-regulation and stress-related hypothalamic KOR activation as essential mechanisms of LS and stress-related pain in females. Aim 1 will establish the effects of repeated stress on hypothalamic KOR signaling and PRL release. Aim 2 will establish a potential causal relationship of repeated stress or hypothalamic KOR activation on DRG PRLR isoform expression, neural excitability, LS and stress-related pain. Aim 3 will determine if KOR antagonists, DA agonists or a PRL antibody will prevent LS and FPS-like pain selectively in females. The proposed studies will characterize a previously unknown stress-related neuroendocrine link between hypothalamic KOR and PRL/PRLR signaling to promote female selective functional pain. Importantly, these studies will advance knowledge about previously unknown biological mechanisms and may unravel mechanisms for therapeutic interventions allowing improved therapy of FPS in women.