Single nuclei transcriptomics in human and non-human primate striatum in opioid use disorder.

In brain, the striatum is a heterogenous region involved in reward and goal-directed behaviors. Striatal dysfunction is linked to psychiatric disorders, including opioid use disorder (OUD). Striatal subregions are divided based on neuroanatomy, each with unique roles in OUD. In OUD, the dorsal striatum is involved in altered reward processing, formation of habits, and development of negative affect during withdrawal. Using single nuclei RNA-sequencing, we identified both canonical (e.g., dopamine receptor subtype) and less abundant cell populations (e.g., interneurons) in human dorsal striatum. Pathways related to neurodegeneration, interferon response, and DNA damage were significantly enriched in striatal neurons of individuals with OUD. DNA damage markers were also elevated in striatal neurons of opioid-exposed rhesus macaques. Sex-specific molecular differences in glial cell subtypes associated with chronic stress were found in OUD, particularly female individuals. Together, we describe different cell types in human dorsal striatum and identify cell type-specific alterations in OUD.

Enhancing Fear Extinction: Pharmacological Approaches.

Extinction is the process by which the memory of a learned conditioned association decreases over time and with introduction of new associations. It is a vital part of fear learning, and it is critical to recovery in multiple fear-related disorders, including Specific and Social Phobias, Panic Disorder, Obsessive Compulsive Disorder (OCD), and Posttraumatic Stress Disorder (PTSD). The process of extinction is also the underlying mechanism for recovery in gold-standard therapies for PTSD, including prolonged exposure, cognitive processing therapy, eye movement desensitization and procession, as well as other empirically-based paradigms. Pharmacological modulators of extinction are thus promising targets for treatment of fear-related disorders. We focus here on emerging psychopharmacological treatments to facilitate extinction: D-cycloserine, scopolamine, losartan, ketamine, and 3,4-methylenedioxymethamphetamine. We also provide an overview of recent advances in molecular pathways that show promise as targets for extincion and inhibitory learning, including pathways related to cannabinoid, brain-derived neurotrophic factor, hypothalamic-pituitary-adrenal signaling, and promising work in neurosteroid compounds.

Prefrontal cortex, amygdala, and threat processing: implications for PTSD.

Posttraumatic stress disorder can be viewed as a disorder of fear dysregulation. An abundance of research suggests that the prefrontal cortex is central to fear processing-that is, how fears are acquired and strategies to regulate or diminish fear responses. The current review covers foundational research on threat or fear acquisition and extinction in nonhuman animals, healthy humans, and patients with posttraumatic stress disorder, through the lens of the involvement of the prefrontal cortex in these processes. Research harnessing advances in technology to further probe the role of the prefrontal cortex in these processes, such as the use of optogenetics in rodents and brain stimulation in humans, will be highlighted, as well other fear regulation approaches that are relevant to the treatment of posttraumatic stress disorder and involve the prefrontal cortex, namely cognitive regulation and avoidance/active coping. Despite the large body of translational research, many questions remain unanswered and posttraumatic stress disorder remains difficult to treat. We conclude by outlining future research directions related to the role of the prefrontal cortex in fear processing and implications for the treatment of posttraumatic stress disorder.

Genome-wide translational profiling of amygdala Crh-expressing neurons reveals role for CREB in fear extinction learning.

Fear and extinction learning are adaptive processes caused by molecular changes in specific neural circuits. Neurons expressing the corticotropin-releasing hormone gene (Crh) in central amygdala (CeA) are implicated in threat regulation, yet little is known of cell type-specific gene pathways mediating adaptive learning. We translationally profiled the transcriptome of CeA Crh-expressing cells (Crh neurons) after fear conditioning or extinction in mice using translating ribosome affinity purification (TRAP) and RNAseq. Differential gene expression and co-expression network analyses identified diverse networks activated or inhibited by fear vs extinction. Upstream regulator analysis demonstrated that extinction associates with reduced CREB expression, and viral vector-induced increased CREB expression in Crh neurons increased fear expression and inhibited extinction. These findings suggest that CREB, within CeA Crh neurons, may function as a molecular switch that regulates expression of fear and its extinction. Cell-type specific translational analyses may suggest targets useful for understanding and treating stress-related psychiatric illness.

Author Correction: Genome-wide translational profiling of amygdala Crh-expressing neurons reveals role for CREB in fear extinction learning.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Cell Type-Specific Transcriptomics Reveals that Mutant Huntingtin Leads to Mitochondrial RNA Release and Neuronal Innate Immune Activation.

The mechanisms by which mutant huntingtin (mHTT) leads to neuronal cell death in Huntington's disease (HD) are not fully understood. To gain new molecular insights, we used single nuclear RNA sequencing (snRNA-seq) and translating ribosome affinity purification (TRAP) to conduct transcriptomic analyses of caudate/putamen (striatal) cell type-specific gene expression changes in human HD and mouse models of HD. In striatal spiny projection neurons, the most vulnerable cell type in HD, we observe a release of mitochondrial RNA (mtRNA) (a potent mitochondrial-derived innate immunogen) and a concomitant upregulation of innate immune signaling in spiny projection neurons. Further, we observe that the released mtRNAs can directly bind to the innate immune sensor protein kinase R (PKR). We highlight the importance of studying cell type-specific gene expression dysregulation in HD pathogenesis and reveal that the activation of innate immune signaling in the most vulnerable HD neurons provides a novel framework to understand the basis of mHTT toxicity and raises new therapeutic opportunities.

Brain circuit dysfunction in post-traumatic stress disorder: from mouse to man.

Post-traumatic stress disorder (PTSD) is a prevalent, debilitating and sometimes deadly consequence of exposure to severe psychological trauma. Although effective treatments exist for some individuals, they are limited. New approaches to intervention, treatment and prevention are therefore much needed. In the past few years, the field has rapidly developed a greater understanding of the dysfunctional brain circuits underlying PTSD, a shift in understanding that has been made possible by technological revolutions that have allowed the observation and perturbation of the macrocircuits and microcircuits thought to underlie PTSD-related symptoms. These advances have allowed us to gain a more translational knowledge of PTSD, have provided further insights into the mechanisms of risk and resilience and offer promising avenues for therapeutic discovery.

Control of Huntington's Disease-Associated Phenotypes by the Striatum-Enriched Transcription Factor Foxp2.

Alteration of corticostriatal glutamatergic function is an early pathophysiological change associated with Huntington's disease (HD). The factors that regulate the maintenance of corticostriatal glutamatergic synapses post-developmentally are not well understood. Recently, the striatum-enriched transcription factor Foxp2 was implicated in the development of these synapses. Here, we show that, in mice, overexpression of Foxp2 in the adult striatum of two models of HD leads to rescue of HD-associated behaviors, while knockdown of Foxp2 in wild-type mice leads to development of HD-associated behaviors. We note that Foxp2 encodes the longest polyglutamine repeat protein in the human reference genome, and we show that it can be sequestered into aggregates with polyglutamine-expanded mutant Huntingtin protein (mHTT). Foxp2 overexpression in HD model mice leads to altered expression of several genes associated with synaptic function, genes that present additional targets for normalization of corticostriatal dysfunction in HD.

Identifying therapeutic targets by combining transcriptional data with ordinal clinical measurements.

The immense and growing repositories of transcriptional data may contain critical insights for developing new therapies. Current approaches to mining these data largely rely on binary classifications of disease vs. control, and are not able to incorporate measures of disease severity. We report an analytical approach to integrate ordinal clinical information with transcriptomics. We apply this method to public data for a large cohort of Huntington's disease patients and controls, identifying and prioritizing phenotype-associated genes. We verify the role of a high-ranked gene in dysregulation of sphingolipid metabolism in the disease and demonstrate that inhibiting the enzyme, sphingosine-1-phosphate lyase 1 (SPL), has neuroprotective effects in Huntington's disease models. Finally, we show that one consequence of inhibiting SPL is intracellular inhibition of histone deacetylases, thus linking our observations in sphingolipid metabolism to a well-characterized Huntington's disease pathway. Our approach is easily applied to any data with ordinal clinical measurements, and may deepen our understanding of disease processes.Identifying gene subsets affecting disease phenotypes from transcriptome data is challenge. Here, the authors develop a method that combines transcriptional data with disease ordinal clinical measurements to discover a sphingolipid metabolism regulator involving in Huntington's disease progression.

Cell type-specific mRNA purification by translating ribosome affinity purification (TRAP).

Cellular diversity and architectural complexity create barriers to understanding the function of the mammalian CNS at a molecular level. To address this problem, we have recently developed a methodology that provides the ability to profile the entire translated mRNA complement of any genetically defined cell population. This methodology, which we termed translating ribosome affinity purification, or TRAP, combines cell type-specific transgene expression with affinity purification of translating ribosomes. TRAP can be used to study the cell type-specific mRNA profiles of any genetically defined cell type, and it has been used in organisms ranging from Drosophila melanogaster to mice and human cultured cells. Unlike other methodologies that rely on microdissection, cell panning or cell sorting, the TRAP methodology bypasses the need for tissue fixation or single-cell suspensions (and the potential artifacts that these treatments introduce) and reports on mRNAs in the entire cell body. This protocol provides a step-by-step guide to implement the TRAP methodology, which takes 2 d to complete once all materials are in hand.

Molecular adaptations of striatal spiny projection neurons during levodopa-induced dyskinesia.

Levodopa treatment is the major pharmacotherapy for Parkinson's disease. However, almost all patients receiving levodopa eventually develop debilitating involuntary movements (dyskinesia). Although it is known that striatal spiny projection neurons (SPNs) are involved in the genesis of this movement disorder, the molecular basis of dyskinesia is not understood. In this study, we identify distinct cell-type-specific gene-expression changes that occur in subclasses of SPNs upon induction of a parkinsonian lesion followed by chronic levodopa treatment. We identify several hundred genes, the expression of which is correlated with levodopa dose, many of which are under the control of activator protein-1 and ERK signaling. Despite homeostatic adaptations involving several signaling modulators, activator protein-1-dependent gene expression remains highly dysregulated in direct pathway SPNs upon chronic levodopa treatment. We also discuss which molecular pathways are most likely to dampen abnormal dopaminoceptive signaling in spiny projection neurons, hence providing potential targets for antidyskinetic treatments in Parkinson's disease.