Circuit-wide gene network analysis reveals sex-specific roles for phosphodiesterase 1b in cocaine addiction.

Cocaine use disorder is a significant public health issue without an effective pharmacological treatment. Successful treatments are hindered in part by an incomplete understanding of the molecular mechanisms that underlie long-lasting maladaptive plasticity and addiction-like behaviors. Here, we leverage a large RNA-sequencing dataset to generate gene co-expression networks across 6 interconnected regions of the brain's reward circuitry from mice that underwent saline or cocaine self-administration. We identify phosphodiesterase 1b (Pde1b), a Ca2+/calmodulin-dependent enzyme that increases cAMP and cGMP hydrolysis, as a central hub gene within a nucleus accumbens (NAc) gene module that was bioinformatically associated with addiction-like behavior. Chronic cocaine exposure increases Pde1b expression in NAc D2 medium spiny neurons (MSNs) in male but not female mice. Viral-mediated Pde1b overexpression in NAc reduces cocaine self-administration in female rats, but increases seeking in both sexes. In female mice, overexpressing Pde1b in D1 MSNs attenuates the locomotor response to cocaine, with the opposite effect in D2 MSNs. Overexpressing Pde1b in D1/D2 MSNs had no effect on the locomotor response to cocaine in male mice. At the electrophysiological level, Pde1b overexpression reduces sEPSC frequency in D1 MSNs, while increasing excitability of D2 MSNs. Lastly, Pde1b overexpression significantly reduced the number of differentially expressed genes (DEGs) in NAc following chronic cocaine, with discordant effects on gene transcription between sexes. Together, we identify novel gene modules across the brain's reward circuitry associated with addiction-like behavior and explore the role of Pde1b in regulating the molecular, cellular, and behavioral responses to cocaine.Significance Statement Cocaine use disorder is a major public health challenge without an effective pharmacological treatment. Here, we leverage a combination of genome-wide RNA sequencing, gene co-expression network analysis, and bioinformatic analyses of cocaine self-administration behavior to identify a role for phosphodiesterase 1b (Pde1b) in regulating maladaptive, addiction-like behavior. Our studies reveal cell-type- and sex-specific roles for Pde1b in regulating the molecular, cellular, and behavioral responses to cocaine, yielding insight into the molecular mechanisms by which cocaine induces maladaptive plasticity in the brain's reward circuity to drive addiction-like behavior. These discoveries guide directions for future research investigating the molecular basis of cocaine action and provide a pathway for therapeutic development for cocaine use disorder.

CREB Binding at the Zfp189 Promoter Within Medium Spiny Neuron Subtypes Differentially Regulates Behavioral and Physiological Adaptations Over the Course of Cocaine Use.

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 neuroadaptations 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 Drd2+ neurons as a molecular signature of chronic cocaine use that is causal in counteracting the reinforcing effects of cocaine.

Teenage drinking and adult neuropsychiatric disorders: An epigenetic connection.

Targeted epigenetic remodeling in the rat amygdala reverses the effects of adolescent alcohol consumption on excessive drinking and anxiety-like behavior in adulthood.

Key transcription factors mediating cocaine-induced plasticity in the nucleus accumbens.

Repeated cocaine use induces coordinated changes in gene expression that drive plasticity in the nucleus accumbens (NAc), an important component of the brain's reward circuitry, and promote the development of maladaptive, addiction-like behaviors. Studies on the molecular basis of cocaine action identify transcription factors, a class of proteins that bind to specific DNA sequences and regulate transcription, as critical mediators of this cocaine-induced plasticity. Early methods to identify and study transcription factors involved in addiction pathophysiology primarily relied on quantifying the expression of candidate genes in bulk brain tissue after chronic cocaine treatment, as well as conventional overexpression and knockdown techniques. More recently, advances in next generation sequencing, bioinformatics, cell-type-specific targeting, and locus-specific neuroepigenomic editing offer a more powerful, unbiased toolbox to identify the most important transcription factors that drive drug-induced plasticity and to causally define their downstream molecular mechanisms. Here, we synthesize the literature on transcription factors mediating cocaine action in the NAc, discuss the advancements and remaining limitations of current experimental approaches, and emphasize recent work leveraging bioinformatic tools and neuroepigenomic editing to study transcription factors involved in cocaine addiction.

Sex-Specific Role for SLIT1 in Regulating Stress Susceptibility.

BACKGROUND: Major depressive disorder is a pervasive and debilitating syndrome characterized by mood disturbances, anhedonia, and alterations in cognition. While the prevalence of major depressive disorder is twice as high for women as men, little is known about the molecular mechanisms that drive sex differences in depression susceptibility. METHODS: We discovered that SLIT1, a secreted protein essential for axonal navigation and molecular guidance during development, is downregulated in the adult ventromedial prefrontal cortex (vmPFC) of women with depression compared with healthy control subjects, but not in men with depression. This sex-specific downregulation of Slit1 was also observed in the vmPFC of mice exposed to chronic variable stress. To identify a causal, sex-specific role for SLIT1 in depression-related behavioral abnormalities, we performed knockdown (KD) of Slit1 expression in the vmPFC of male and female mice. RESULTS: When combined with stress exposure, vmPFC Slit1 KD reflected the human condition by inducing a sex-specific increase in anxiety- and depression-related behaviors. Furthermore, we found that vmPFC Slit1 KD decreased the dendritic arborization of vmPFC pyramidal neurons and decreased the excitability of the neurons in female mice, effects not observed in males. RNA sequencing analysis of the vmPFC after Slit1 KD in female mice revealed an augmented transcriptional stress signature. CONCLUSIONS: Together, our findings establish a crucial role for SLIT1 in regulating neurophysiological and transcriptional responses to stress within the female vmPFC and provide mechanistic insight into novel signaling pathways and molecular factors influencing sex differences in depression susceptibility.

In vivo locus-specific editing of the neuroepigenome.

Studies over the past several decades have identified numerous epigenetic mechanisms associated with pathological states in psychiatric and neurological disease. Until recently, studies investigating chromatin-regulatory proteins, using overexpression or knockdown approaches, did not establish causal roles for epigenetic modifications at specific genes because these techniques typically affect hundreds or thousands of genomic loci. In this Review, we describe recent efforts in using locus-specific neuroepigenome editing in vivo to, for the first time, define causal relationships between a single chromatin modification at a specific gene in a defined cell population and downstream measures at the molecular, cellular, circuit and behavioural levels. We briefly introduce three epigenome-editing platforms: zinc-finger proteins, transcriptional activator-like effectors and clustered regularly interspaced short palindromic repeats (CRISPR). We then explore the development of in vivo neuroepigenome-editing tools and their applications to resolve epigenetic contributions to the pathophysiology of brain diseases. We also discuss technical considerations for in vivo neuroepigenome-editing experiments and ongoing innovations in the field, including new tools to investigate chromatin marks, manipulate chromatin topology and induce epigenetic modifications at multiple genes in the same cell. Lastly, we explore the potential clinical applications of in vivo neuroepigenome editing for treating brain pathology.

Intravenously Administered, Retinoid Activating Nanoparticles Increase Lifespan and Reduce Neurodegeneration in the SOD1G93A Mouse Model of ALS.

Dysregulation of the retinoic acid (RA) signaling pathway is observed in amyotrophic lateral sclerosis (ALS) and other neurodegenerative disorders. Here, we investigated the therapeutic potential of retinoid activation via the RA receptor ß (RARß) in the SOD1 G93A mouse model of ALS. Our approach utilized the RARß agonist adapalene, which we previously found to be neuroprotective in vitro. Adapalene, like most retinoids, is poorly water soluble, which has thus far prevented effective drug delivery in vivo. To address this challenge, we encapsulated adapalene within nanoparticles (Adap-NPs) composed of poly(lactic acid)-poly(ethylene glycol) (PLA-PEG). Our data demonstrate that intravenous administration of Adap-NPs robustly activates retinoid signaling in the CNS. Chronic administration of Adap-NPs resulted in improved motor performance, prolonged lifespan, and neuroprotection in SOD1 G93A mice. This study highlights retinoid signaling as a valuable therapeutic approach and presents a novel nanoparticle platform for the treatment of ALS.