Despite their dangers, the number of opioid prescriptions written for veterans has increased sharply since 2000, and veterans are more prone than the general population to both suicide and the development of use disorders following opioid treatment. Target receptors for opioids are widely expressed throughout the brain and periphery, but their reinforcing properties are largely mediated by their action in mesocorticolimbic areas such as the ventral tegmental area (VTA) and the nucleus accumbens (NAc). In the VTA, activation of presynaptic mu-opioid receptors is known to blunt release of the inhibitory neurotransmitter GABA, thus “disinhibiting” dopamine neuron activity. Preliminary work has established that the modulatory peptide neurotensin can activate presynaptic neurotensin 1 receptors (NtsR1) to enhance GABA release in the VTA. While this novel form of synaptic plasticity would be expected to directly counteract the effects of opioids, it is not known how repeated opioid exposure interacts with neurotensin effects on GABA signaling. Opioids and neurotensin are both known to modulate pain; however, there are significant gaps in our knowledge of how these compounds interact at the synaptic and circuit level in the VTA to affect drug reinforcement. Improved treatments for opioid use disorders are desperately needed, both for the general population but also for aging veterans that will increasingly develop painful conditions that require long-term treatment. The proposed studies are necessary to determine the feasibility of targeting the neurotensin system to modulate reinforcement and relapse in individuals that no longer can control their opioid intake. We will combine brain slice electrophysiology and cell type-specific molecular techniques with self- administration of the opioid remifentanil in mice to explore these issues. The use of operant self-administration in mice offers several key advantages: mice are able to titrate their intake based on individual sensitivity, and using mice instead of rats opens up the powerful tools of mouse genetics (i.e., Cre-lox technology) to experimental manipulations. The hypothesis to be tested is that a history of remifentanil self-administration decreases neurotensin-induced enhancement of GABA release in the NAc  VTA circuit, removing a critical break on dopamine neuron excitability during drug intake to increase reinforcement. Experiments in Aim 1 will identify the sensitivity of individual GABA inputs in the VTA to neurotensin, and determine how plasticity is affected by remifentanil self-administration as well as following a forced abstinence. Experiments in Aim 2 will use chemogenetics to activate specific GABA inputs to determine their effect on remifentanil self-administration behavior and cue responding following a forced abstinence. A novel cell type-specific neurotensin receptor knockout will provide additional information on the role of specific cell types on opioid self-administration. Experiments in Aim 3 will use a discovery approach to determine transcriptomic and epigenomic alterations following remifentanil self-administration in single cell types of the VTA. This will be done with several novel NuTRAP (Nuclear Tagging and Translating Ribosome Affinity Purification) mouse lines under the control of Cre recombinase that allow for labeling and isolation of both DNA and RNA from specific cell types. Improved strategies are desperately needed to improve the quality of life for veterans at risk of adverse consequences following opioid treatment. Data obtained will delineate the behavioral and physiological interactions between GABA input to the VTA, neurotensin signaling, and opioid exposure, and identify novel gene and receptor targets for exploration as treatments for opioid use disorders.

The public health crisis known as “the opioid epidemic” continues to take the lives of veterans, not only due to the development of opioid use disorders and overdose but also from an enhanced risk of suicide. Improved treatments for opioid use disorders are desperately needed, but the neurophysiological adaptations that drive increased reinforcement in these individuals are not adequately understood. The proposed studies will combine electrophysiological, behavioral, molecular, and cell type-specific techniques in mice to elucidate the relationship between the modulatory peptide neurotensin, inhibitory synaptic transmission, and opioid self- administration. The mechanistic understanding gleaned from this work will help identify novel therapeutic targets to treat opioid use disorders in veterans and the general population.