A major challenge in treating addiction is that vulnerability to cue-induced craving and relapse persist even after long periods of abstinence. There is presently no FDA-approved medication to lessen psychostimulant craving. We study persistent relapse vulnerability using the `incubation of craving' model, in which rats exhibit withdrawal- dependent intensification (incubation) of cue-induced craving over the first weeks of withdrawal, with high levels of craving then persisting for months. Incubation of craving also occurs in humans. Previously, we showed that incubation depends on strengthening of synapses in nucleus accumbens (NAc) core via synaptic incorporation of Ca2+-permeable AMPARs (CP-AMPARs). We then showed that metabotropic glutamate receptor 1 (mGlu1) positive allosteric modulators (PAMs) internalize these CP-AMPARs (a form of LTD) and thereby reduce cue- induced craving for cocaine and methamphetamine. The goal of this proposal is to take the first steps towards advancing the translation of this basic science discovery. Specifically, we hypothesize that mGlu1 PAMs may be developed as anti-craving drugs to enable recovering psychostimulant users to maintain abstinence by avoiding cue-induced relapse. Barriers to this goal include less than optimal pharmacokinetic properties of existing PAMs, but efforts are underway to develop improved drugs for another indication (schizophrenia). Therefore, this application will address other barriers, with the goal of making mGlu1 PAMs more attractive to potential partners and thus accelerate their translation as anti-craving drugs. Aim 1 will use the incubation of craving model to further validate mGlu1 as a target for psychostimulant (cocaine and methamphetamine) craving reduction by replicating our effects with an additional mGlu1 PAM, conducting dose-response studies, and studying female rats. While studies in the incubation model have high translational value because we can study craving after prolonged abstinence from drug self-administration, they are very time-consuming (6-8 weeks) and labor- intensive. Aim 2 will develop higher throughput screens for the ability of mGlu1 PAMs to internalize CP-AMPARs. Work from other groups and our preliminary data indicate that mGlu1-induced CP-AMPAR internalization depends on mGlu1-induced translation of the “LTD protein” oligophrenin-1 (OPHN1). We propose to develop an in vitro screen that is precisely focused on this target mechanism, i.e., mammalian cells expressing mGlu1 will be exposed to mGlu1 PAMs (with glutamate) and OPHN1 expression will be measured. As an alternate strategy, we will evaluate an in vivo screen that is significantly faster than the incubation model. This is based on extensive work showing that a single i.p. cocaine injection increases CP-AMPAR levels in excitatory synapses on VTA dopamine neurons within hours. mGlu1 PAMs internalize CP-AMPARs through the same mechanism in the VTA and the NAc, so their effects in this screen shoul...