Prosapip1- a novel mediator of alcohol abuse disorder

We previously reported that excessive consumption of alcohol activates the H-Ras/PI3K/AKT signaling in the nucleus accumbens (NAc) of rodents, which ultimately leads to the activation of the mechanistic target of rapamycin complex 1 (mTORC1) [1-3]. mTORC1 is localized in dendrites and plays an important role in synaptic plasticity by promoting the translation of synaptic proteins [4]. We therefore hypothesized that mTORC1-dependent mRNA translation contributes to neuroadaptations that underlie excessive alcohol consumption. To do so, we utilized the high throughput RNA sequencing (RNA seq) approach to identify novel gene products whose mTORC1-dependent translation is induced in the NAc in response to excessive alcohol consumption. Specifically, mice underwent an intermittent access to 20% alcohol or water only using 2-bottle choice paradigm for 8 weeks, and were then treated with rapamycin or vehicle. The NAc was dissected, polysomes (RNA undergoing translation) were isolated, and RNA seq was performed. Among the 12 identified candidates whose translation was dependent of mTORC1 was ProSAP-interacting protein 1 (Prosapip1), a synaptic protein [5] that we found to be highly expressed in the striatum, and whose function is not well understood. First, we confirmed the RNAseq data and showed that the translation of Prosapip1 but not its transcription was induced in response to excessive alcohol drinking in an mTORC1-dependent manner. The corresponding increase in Prosapip1 protein in the NAc of mice in response to a binge drinking session was localized to the synaptic fraction and was maintained even after 24 hours of withdrawal. Prosapip1 levels were not altered in other brain regions where mTORC1 is not activated by alcohol. To test for potential cellular consequences of Prosapip1 increase by alcohol, we infused a lentivirus overexpressing Prosapip1 in NAc and observed an increased F-actin content. Interestingly, similar result was found in NAc of mice after excessive alcohol drinking. Conversely, knocking down the endogenous protein by intra-NAc infusion of lentivirus delivering shRNA against Prosapip1 reduces the amount of actin filaments. We are currently analyzing the consequences of these actin cytoskeleton modifications on the number and types of dendritic spines in medium spiny neurons (MSNs). Next, we tested the behavioral outcomes of Prosapip1 knock-down and we observed a decreased operant self-administration of 20% alcohol when a fixed-ratio 2 schedule (i.e. two presses on the active lever resulted in the delivery of one drink of alcohol) was applied. The decreased alcohol intake by Prosapip1 knockdown was not due to non-specific locomotion or cognitive changes. We furthermore showed that decreasing Prosapip1 levels in the NAc reduces the conditioned place preference (CPP) for alcohol.Together, our data suggest that alcohol-induced activation of mTORC1 in the NAc leads to translation of the synaptic protein Prosapip1 that in turn drives the motivation to obtain and consume alcohol.