The rostral ventromedial medulla (RVM) is an important site of opioid action and a critical relay in the bulbo- spinal inhibitory and facilitatory pathways that modulate pain. Persistent pain leads to adaptive changes in the RVM that change the balance of these pathways and that increase the potency of opioid analgesics. Investigation of the mechanisms by which 5 opioid receptor (MOPr) agonists suppress acute nociception and relieve hyperalgesia in persistent pain states will provide important insights into these adaptive changes. The overall goal of this proposal is to investigate the role of postsynaptic MOPr, via G protein inwardly rectifying K+ channels (GIRKs), and presynaptic MOPr, via inhibition of GABA release, in antinociception and in the adaptive changes that occur in the RVM during persistent inflammatory pain induced by injection of complete Freund's adjuvant (CFA) in the hindpaw. My hypotheses are: (1) presynaptic inhibition of GABA release and disinhibition of pain inhibitory neurons is the predominant mechanism for the production of antinociception in the uninjured state and (2) postsynaptic inhibition of pain facilitatory neurons, whose activity increases after injury, is the predominant mechanism for the enhanced antinociception and the anti-hyperalgesic effects of MOPr agonists under conditions of persistent pain. Behavioral, neurochemical, and molecular experiments are proposed. The first aim will confirm the role of Gi/o proteins in the antinociceptive effects of MOPr and 4 opioid receptor (DOPr) agonist agonists in the RVM, and establish that they play a more important role in enhanced antinociception and anti-hyperalgesia in persistent inflammatory pain. Behavioral pharmacological experiments will determine whether inactivation of Gi/o proteins by pertussis toxin reduces the antinociceptive and anti-hyperalgesic effects of the MOPr agonist DAMGO or the DOPr agonist DELT in the RVM of saline- and CFA-treated rats. The second aim will target GIRK channels to examine the role of postsynaptic MOPr and DOPr in opioid antinociception and anti-hyperalgesia. These experiments will determine the extent to which microinjection of the GIRK channel inhibitor tertiapin in the RVM reduces the antinociceptive or anti-hyperalgesic effects of DAMGO or DELT in saline and CFA-treated rats. qRT-PCR and Western blotting will determine if persistent pain changes the levels or types of GIRK channels in the RVM. The third aim will examine the role of presynaptic MOPr and DOPr by measuring basal and K+ evoked release of GABA using microdialysis of the RVM in saline- and CFA- treated rats. These experiments will determine if antinociceptive and anti-hyperalgesic concentrations of DAMGO and DELT inhibit GABA release in uninjured and CFA-treated rats, and whether inhibition of GABA release is enhanced in CFA-treated rats. These studies will increase our understanding of the mechanisms that subserve the enhanced potency of opioid agonists and how persistent pain alters the physiology and pharmacology of CNS pain modulatory pathways. They will also provide important insights into new non- opioid pharmacological approaches to the relief of persistent pain that lack the adverse effects of opioids. PUBLIC HEALTH RELEVANCE: Chronic pain is a significant health care problem for which effective pharmacotherapies continue to elude us. A better understanding of how chronic pain alters the physiology and pharmacology of the central nervous system pain modulatory pathways through which these drugs act is required. These studies will increase our understanding of the mechanisms that subserve the enhanced potency of opioid drugs and provide important insights into new non-opioid pharmacological approaches that can be used for the relief of chronic pain.