In human populations, cigarettes and alcohol generally serve as gateway drugs, which people use first before progressing to marijuana, cocaine or other illicit substances. To understand the biological basis of the gateway sequence of drug use, we developed an animal model in mice and focused on the effects of nicotine on subsequent responses to cocaine. We found that pretreatment of mice with nicotine increased the response to cocaine as assessed by both addiction-related behaviors and synaptic plasticity in the striatum, a brain region critical for addiction-related reward. Locomotor sensitization was increased by 98%, conditioned place preference was increased by 78%, and cocaine-induced reduction in long-term potentiation (LTP) was enhanced by 24%. The responses to cocaine were altered only when nicotine was administered first, and nicotine and cocaine were then administered concurrently. Reversing the order of drug administration was ineffective. Cocaine had no effect on nicotine induced behaviors and synaptic plasticity. Nicotine primed the response to cocaine by enhancing its ability to induce transcriptional activation of the FosB gene through inhibiting histone deacetylase, causing global histone acetylation in the striatum. We tested this conclusion further with a histone deacetylase inhibitor and found that it similarly simulated the actions of nicotine on cocaine by priming the response to cocaine, and enhancing FosB gene expression and LTP depression in the nucleus accumbens. Conversely, in a genetic mouse model of Rubinstein Taybi’s syndrome, characterized by reduced histone acetylation, the effects of cocaine on LTP were diminished. We achieved a similar effect pharmacologically by infusing a low-dose of theophylline, an activator of histone deacetylase, into the nucleus accumbens. These data from mice prompted an analysis of epidemiological data, which indicated that most cocaine users initiate cocaine use after the onset of smoking while actively smoking and that initiating cocaine use after smoking increases the risk of becoming dependent on cocaine, consistent with our data in mice. If our findings in mice apply to humans, a decrease in smoking rates in young people could also lead to a decrease in cocaine addiction.

Consolidation of long-term memories depends on de novo protein synthesis. Several translational regulators have been identified, and their contribution to the formation of memory has been assessed in the mouse hippocampus. None of them, however, has been implicated in the persistence of memory. Although persistence is a key feature of long-term memory, how this occurs, despite the rapid turnover of its molecular substrates, is poorly understood. Here we find that both memory storage and its underlying synaptic plasticity are mediated by the increase in level and in the aggregation of the prion-like translational regulator CPEB3 (cytoplasmic polyadenylation element-binding protein). Genetic ablation of CPEB3 impairs the maintenance of both hippocampal long-term potentiation and hippocampus-dependent spatial memory. We propose a model whereby persistence of long-term memory results from the assembly of CPEB3 into aggregates. These aggregates serve as functional prions and regulate local protein synthesis necessary for the maintenance of long-term memory.

The cellular mechanisms by which prions cause neurological dysfunction are poorly understood. To address this issue, we have been using cultured cells to analyze the localization, biosynthesis, and metabolism of PrP molecules carrying mutations associated with familial prion diseases. We report here that mutant PrP molecules are delayed in their maturation to an endoglycosidase H-resistant form after biosynthetic labeling, suggesting that they are impaired in their exit from the endoplasmic reticulum (ER). However, we find that proteasome inhibitors have no effect on the maturation or turnover of either mutant or wild-type PrP molecules. Thus, in contrast to recent studies from other laboratories, our work indicates that PrP is not subject to retrotranslocation from the ER into the cytoplasm prior to degradation by the proteasome. We find that in transfected cells, but not in cultured neurons, proteasome inhibitors cause accumulation of an unglycosylated, signal peptide-bearing form of PrP on the cytoplasmic face of the ER membrane. Thus, under conditions of elevated expression, a small fraction of PrP chains is not translocated into the ER lumen during synthesis, and is rapidly degraded in the cytoplasm by the proteasome. Finally, we report a previously unappreciated artifact caused by treatment of cells with proteasome inhibitors: an increase in PrP mRNA level and synthetic rate when the protein is expressed from a vector containing a viral promoter. We suggest that this phenomenon may explain some of the dramatic effects of proteasome inhibitors observed in other studies. Our results clarify the role of the proteasome in the cell biology of PrP, and suggest reasonable hypotheses for the molecular pathology of inherited prion diseases.

The cellular prion protein, PrPC, is neuroprotective in a number of settings and in particular prevents cerebellar degeneration mediated by CNS‐expressed Doppel or internally deleted PrP (‘ΔPrP’). This paradigm has facilitated mapping of activity determinants in PrPC and implicated a cryptic PrPC‐like protein, ‘π’. Shadoo (Sho) is a hypothetical GPI‐anchored protein encoded by the Sprn gene, exhibiting homology and domain organization similar to the N‐terminus of PrP. Here we demonstrate Sprn expression and Sho protein in the adult CNS. Sho expression overlaps PrPC, but is low in cerebellar granular neurons (CGNs) containing PrPC and high in PrPC‐deficient dendritic processes. In Prnp0/0 CGNs, Sho transgenes were PrPC‐like in their ability to counteract neurotoxic effects of either Doppel or ΔPrP. Additionally, prion‐infected mice exhibit a dramatic reduction in endogenous Sho protein. Sho is a candidate for π, and since it engenders a PrPC‐like neuroprotective activity, compromised neuroprotective activity resulting from reduced levels may exacerbate damage in prion infections. Sho may prove useful in deciphering several unresolved facets of prion biology.