The nucleus accumbens (NAc) regulates reward-motivated behavior, but the temporal dynamics of NAc neurons that enable "free-willed" animals to obtain rewards remain elusive. Here, we recorded Ca2+ activity from individual NAc neurons when mice performed self-paced lever-presses for sucrose. NAc neurons exhibited three temporally-sequenced clusters, defined by times at which they exhibited increased Ca2+ activity: approximately 0, -2.5 or -5 sec relative to the lever-pressing. Dopamine D1 receptor (D1)-expressing neurons and D2-neurons formed the majority of the -5-sec versus -2.5-sec clusters, respectively, while both neuronal subtypes were represented in the 0-sec cluster. We found that pre-press activity patterns of D1- or D2-neurons could predict subsequent lever-presses. Inhibiting D1-neurons at -5 sec or D2-neurons at -2.5 sec, but not at other timepoints, reduced sucrose-motivated lever-pressing. We propose that the time-specific activity of D1- and D2-neurons mediate key temporal features of the NAc through which reward motivation initiates reward-seeking behavior. Volitional reward taking emerges through a sequence of preluding events. Yet, the underlying neural mechanism is not fully understood. Here authors show a series of temporal dynamics of nucleus accumbens neurons that may substantiate such preluding events to commit "free-willed" animals to reward taking.

BACKGROUND: Over the course of chronic drug use, brain transcriptional neuroadaptation is thought to contribute to a change in drug use behavior over time. The function of the transcription factor CREB (cAMP response element binding protein) within the nucleus accumbens (NAc) has been well documented in opposing the rewarding properties of many classes of drugs, yet the gene targets through which CREB causally manifests these lasting neuro-adaptations remain unknown. Here, we identify zinc finger protein 189 (Zfp189) as a CREB target gene that is transcriptionally responsive to acute and chronic cocaine use within the NAc of mice.METHODS: To investigate the role of the CREB-Zfp189 interaction in cocaine use, we virally delivered modified clustered regularly interspaced short palindromic repeats (CRISPR)/dCas9 constructs capable of selectively localizing CREB to the Zfp189 gene promoter in the NAc of mice.RESULTS: We observed that CREB binding to the Zfp189 promoter increased Zfp189 expression and diminished the reinforcing responses to cocaine. Furthermore, we showed that NAc Zfp189 expression increased within D1 medium spiny neurons in response to acute cocaine but increased in both D1-and D2-expressing medium spiny neurons in response to chronic cocaine. CREB-mediated induction of Zfp189 potentiated electrophysiological activity of D1-and D2-expressing medium spiny neurons, recapitulating the known effect of CREB on these neurons. Finally, targeting CREB to the Zfp189 promoter within NAc Drd2-expressing neurons, but not Drd1-expressing neurons, was sufficient to diminish cocaine-conditioned behaviors.CONCLUSIONS: Together, these findings point to the CREB-Zfp189 interaction within the NAc Drd21 neurons as a molecular signature of chronic cocaine use that is causal in counteracting the reinforcing effects of cocaine.

Glutamatergic synapses are key cellular sites where cocaine experience creates memory traces that subsequently promote cocaine craving and seeking. In addition to making across-the-board synaptic adaptations, cocaine experience also generates a discrete population of new synapses that selectively encode cocaine memories. These new synapses are glutamatergic synapses that lack functionally stable AMPARs, often referred to as AMPAR-silent synapses or, simply, silent synapses. They are generated de novo in the NAc by cocaine experience. After drug withdrawal, some of these synapses mature by recruiting AMPARs, contributing to the consolidation of cocaine-associated memory. After cue-induced retrieval of cocaine memories, matured silent synapses alternate between two dynamic states (AMPAR-absent vs AMPAR-containing) that correspond with the behavioral manifestations of destabilization and reconsolidation of these memories. Here, we review the molecular mechanisms underlying silent synapse dynamics during behavior, discuss their contributions to circuit remodeling, and analyze their role in cocaine-memory-driven behaviors. We also propose several mechanisms through which silent synapses can form neuronal ensembles as well as cross-region circuit engrams for cocaine-specific behaviors. These perspectives lead to our hypothesis that cocaine-generated silent synapses stand as a distinct set of synaptic substrates encoding key aspects of cocaine memory that drive cocaine relapse.

Carrying different aspects of emotional and motivational signals, glutamatergic synaptic projections from multiple limbic and paralimbic brain regions converge to the nucleus accumbens (NAc), in which these arousing signals are processed and prioritized for behavioral output. In animal models of drug addiction, some key drug-induced alterations at NAc glutamatergic synapses underlie important cellular and circuit mechanisms that promote subsequent drug taking, seeking, and relapse. With the focus of cocaine, we review changes at NAc glutamatergic synapses that occur after different drug procedures and abstinence durations, and the behavioral impact of these changes.

The development of drug addiction is associated with functional adaptations within the reward circuitry, within which the nucleus accumbens (NAc) is anatomically positioned as an interface between motivational salience and behavioral output. The functional output of NAc is profoundly altered after exposure to drugs of abuse, and some of the functional changes continue to evolve during drug abstinence, contributing to numerous emotional and motivational alterations related drug taking, seeking, and relapse. As in most brain regions, the functional output of NAc is critically dependent on the dynamic interaction between excitation and inhibition. One of the most prominent sources of inhibition within the NAc arises from fast-spiking interneurons (FSIs). Each NAc FSI innervates hundreds of principal neurons, and orchestrates population activity through its powerful and sustained feedforward inhibition. While the role of NAc FSIs in the context of drug addiction remains poorly understood, emerging evidence suggests that FSIs and FSI-mediated local circuits are key targets for drugs of abuse to tilt the functional output of NAc toward a motivational state favoring drug seeking and relapse. In this review, we discuss recent findings and our conceptualization about NAc FSI-mediated regulation of motivated and cocaine-induced behaviors. We hope that the conceptual framework proposed in this review may provide a useful guidance for ongoing and future studies to determine how FSIs influence the function of NAc and related reward circuits, ultimately leading to addictive behaviors.

Cocaine-associated memories are persistent, but, on retrieval, become temporarily destabilized and vulnerable to disruptions, followed by reconsolidation. To explore the synaptic underpinnings for these memory dynamics, we studied AMPA receptor (AMPAR)-silent excitatory synapses, which are generated in the nucleus accumbens by cocaine self-administration, and subsequently mature after prolonged withdrawal by recruiting AMPARs, echoing acquisition and consolidation of cocaine memories. We show that, on memory retrieval after prolonged withdrawal, the matured silent synapses become AMPAR-silent again, followed by re-maturation ~6 h later, defining the onset and termination of a destabilization window of cocaine memories. These synaptic dynamics are timed by Rac1, with decreased and increased Rac1 activities opening and closing, respectively, the silent synapse-mediated destabilization window. Preventing silent synapse re-maturation within the destabilization window decreases cue-induced cocaine seeking. Thus, cocaine-generated silent synapses constitute a discrete synaptic ensemble dictating the dynamics of cocaine-associated memories and can be targeted for memory disruption.

Understanding the transcriptional changes that are engaged in stress resilience may reveal novel antidepressant targets. Here we use gene co-expression analysis of RNA-sequencing data from brains of resilient mice to identify a gene network that is unique to resilience. Zfp189, which encodes a previously unstudied zinc finger protein, is the highest-ranked key driver gene in the network, and overexpression of Zfp189 in prefrontal cortical neurons preferentially activates this network and promotes behavioral resilience. The transcription factor CREB is a predicted upstream regulator of this network and binds to the Zfp189 promoter. To probe CREB-Zfp189 interactions, we employ CRISPR-mediated locus-specific transcriptional reprogramming to direct CREB or G9a (a repressive histone methyltransferase) to the Zfp189 promoter in prefrontal cortex neurons. Induction of Zfp189 with site-specific CREB is pro-resilient, whereas suppressing Zfp189 expression with G9a increases susceptibility. These findings reveal an essential role for Zfp189 and CREB-Zfp189 interactions in mediating a central transcriptional network of resilience.

Stress resilience involves numerous brain-wide transcriptional changes. Determining the organization and orchestration of these transcriptional events may reveal novel antidepressant targets, but this remains unexplored. Here, we characterize the resilient transcriptome with co-expression analysis and identify a single transcriptionally-active uniquely-resilient gene network. Zfp189, a previously unstudied zinc finger protein, is the top network key driver and its overexpression in prefrontal cortical (PFC) neurons preferentially activates this network, alters neuronal activity and promotes behavioral resilience. CREB, which binds Zfp189, is the top upstream regulator of this network. To probe CREB-Zfp189 interactions as a network regulatory mechanism, we employ CRISPR-mediated locus-specific transcriptional reprogramming to direct CREB selectively to the Zfp189 promoter. This single molecular interaction in PFC neurons recapitulates the pro-resilient Zfp189-dependent downstream effects on gene network activity, electrophysiology and behavior. These findings reveal an essential role for Zfp189 and a CREB-Zfp189 regulatory axis in mediating a central transcriptional network of resilience.

Despite diverse acute pharmacological actions, all drugs of abuse produce many similar behaviors, such as psychomotor sensitization, conditioned drug taking, drug seeking, and relapse ( 1 Nestler E.J. Is there a common molecular pathway for addiction?. Nat Neurosci. 2005; 8: 1445-1449 Crossref PubMed Scopus (1025) Google Scholar ). The development of these addiction-related behaviors is thought to arise from diverse molecular and cellular adaptations, which collectively result in convergent long-term functional alterations of the striatum. However, at the cellular level, on the one hand, exposure to different classes of drugs, such as stimulants versus opioids ( 2 Russo S.J. Dietz D.M. Dumitriu D. Morrison J.H. Malenka R.C. Nestler E.J. The addicted synapse: Mechanisms of synaptic and structural plasticity in nucleus accumbens. Trends Neurosci. 2010; 33: 267-276 Abstract Full Text Full Text PDF PubMed Scopus (490) Google Scholar ), induces distinct and often opposing forms of adaptations in striatal medium spiny neurons (MSNs). On the other hand, exposure to alcohol promotes alcohol consumption by simultaneously activating both excitatory and inhibitory synaptic signaling, two seemingly opposing mechanisms, in the striatum ( 3 Wang J. Lanfranco M.F. Gibb S.L. Yowell Q.V. Carnicella S. Ron D. Long-lasting adaptations of the NR2B-containing NMDA receptors in the dorsomedial striatum play a crucial role in alcohol consumption and relapse. J Neurosci. 2010; 30: 10187-10198 Crossref PubMed Scopus (145) Google Scholar , 4 Nie H. Rewal M. Gill T.M. Ron D. Janak P.H. Extrasynaptic delta-containing GABAA receptors in the nucleus accumbens dorsomedial shell contribute to alcohol intake. Proc Natl Acad Sci U S A. 2011; 108: 4459-4464 Crossref PubMed Scopus (76) Google Scholar ). It remains underexplored how such opposing synaptic mechanisms contribute to the same behavioral states induced by drugs of abuse. Distinct Synaptic Strengthening of the Striatal Direct and Indirect Pathways Drives Alcohol ConsumptionBiological PsychiatryVol. 81Issue 11PreviewRepeated exposure to addictive drugs or alcohol triggers glutamatergic and gamma-aminobutyric acidergic (GABAergic) plasticity in many neuronal populations. The dorsomedial striatum (DMS), a brain region critically involved in addiction, contains medium spiny neurons (MSNs) expressing dopamine D1 or D2 receptors, which form direct and indirect pathways, respectively. It is unclear how alcohol-evoked plasticity in the DMS contributes to alcohol consumption in a cell type–specific manner. Full-Text PDF

Exposures to cocaine and morphine produce similar adaptations in nucleus accumbens (NAc)-based behaviors, yet produce very different adaptations at NAc excitatory synapses. In an effort to explain this paradox, we found that both drugs induced NMDA receptor-containing, AMPA receptor-silent excitatory synapses, albeit in distinct cell types through opposing cellular mechanisms. Cocaine selectively induced silent synapses in D1-type neurons, likely via a synaptogenesis process, whereas morphine induced silent synapses in D2-type neurons via internalization of AMPA receptors from pre-existing synapses. After drug withdrawal, cocaine-generated silent synapses became 'unsilenced' by recruiting AMPA receptors to strengthen excitatory inputs to D1-type neurons, whereas morphine-generated silent synapses were likely eliminated to weaken excitatory inputs to D2-type neurons. Thus, these cell type-specific, opposing mechanisms produced the same net shift of the balance between excitatory inputs to D1- and D2-type NAc neurons, which may underlie certain common alterations in NAc-based behaviors induced by both classes of drugs.

The nucleus accumbens (NAc) gates motivated behaviors through the functional output of principle medium spiny neurons (MSNs), whereas dysfunctional output of NAc MSNs contributes to a variety of psychiatric disorders. Fast-spiking interneurons (FSIs) are sparsely distributed throughout the NAc, forming local feedforward inhibitory circuits. It remains elusive how FSI-based feedforward circuits regulate the output of NAc MSNs. Here, we investigated a distinct subpopulation of NAc FSIs that express the cannabinoid receptor type-1 (CB1). Using a combination of paired electrophysiological recordings and pharmacological approaches, we characterized and compared feedforward inhibition of NAc MSNs from CB1+ FSIs and lateral inhibition from recurrent MSN collaterals. We observed that CB1+ FSIs exerted robust inhibitory control over a large percentage of nearby MSNs in contrast to local MSN collaterals that provided only sparse and weak inhibitory input to their neighboring MSNs. Furthermore, CB1+ FSI-mediated feedforward inhibition was preferentially suppressed by endocannabinoid (eCB) signaling, whereas MSN-mediated lateral inhibition was unaffected. Finally, we demonstrated that CB1+ FSI synapses onto MSNs are capable of undergoing experience-dependent long-term depression in a voltage- and eCB-dependent manner. These findings demonstrated that CB1+ FSIs are a major source of local inhibitory control of MSNs and a critical component of the feedforward inhibitory circuits regulating the output of the NAc.

The development of drug addiction involves diverse molecular and cellular adaptations in complex neural circuits. A critical node in these circuits is the nucleus accumbens (NAc). Excitatory inputs that innervate medium spiny projection neurons (MSNs) undergo extensive and dynamic adaptive changes after exposure to drugs of abuse that critically contribute to the expression of addictive behaviors ( 1 Wolf M.E. The Bermuda Triangle of cocaine-induced neuroadaptations. Trends Neurosci. 2010; 33: 391-398 Abstract Full Text Full Text PDF PubMed Scopus (154) Google Scholar ). An important adaptation at NAc excitatory synapses after withdrawal from cocaine is the progressive accumulation of synaptic alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid receptors (AMPARs), demonstrated by increased AMPAR surface expression and an increase in the relative ratio of AMPAR-mediated to N-methyl-D-aspartate receptor (NMDAR)-mediated synaptic responses ( 1 Wolf M.E. The Bermuda Triangle of cocaine-induced neuroadaptations. Trends Neurosci. 2010; 33: 391-398 Abstract Full Text Full Text PDF PubMed Scopus (154) Google Scholar ). Although involving different AMPAR subtypes, accumulation of AMPARs occurs after either noncontingent or contingent cocaine administration, and the strengthened responsiveness of the NAc MSNs to excitatory inputs contributes to a variety of withdrawal-associated–motivated behaviors, including intensified/incubated cocaine seeking ( 1 Wolf M.E. The Bermuda Triangle of cocaine-induced neuroadaptations. Trends Neurosci. 2010; 33: 391-398 Abstract Full Text Full Text PDF PubMed Scopus (154) Google Scholar ). Cocaine Experience Enhances Thalamo-Accumbens N-Methyl-D-Aspartate Receptor FunctionBiological PsychiatryVol. 80Issue 9PreviewExcitatory synaptic transmission in the nucleus accumbens (NAc) is a key biological substrate underlying behavioral responses to psychostimulants and susceptibility to relapse. Studies have demonstrated that cocaine induces changes in glutamatergic signaling at distinct inputs to the NAc. However, consequences of cocaine experience on synaptic transmission from the midline nuclei of the thalamus (mThal) to the NAc have yet to be reported. Full-Text PDF