Transcriptomic atlas of midbrain dopamine neurons uncovers differential vulnerability in a Parkinsonism lesion model.

Midbrain dopamine (mDA) neurons comprise diverse cells with unique innervation targets and functions. This is illustrated by the selective sensitivity of mDA neurons of the substantia nigra compacta (SNc) in patients with Parkinson's disease, while those in the ventral tegmental area (VTA) are relatively spared. Here, we used single nuclei RNA sequencing (snRNA-seq) of approximately 70,000 mouse midbrain cells to build a high-resolution atlas of mouse mDA neuron diversity at the molecular level. The results showed that differences between mDA neuron groups could best be understood as a continuum without sharp differences between subtypes. Thus, we assigned mDA neurons to several 'territories' and 'neighborhoods' within a shifting gene expression landscape where boundaries are gradual rather than discrete. Based on the enriched gene expression patterns of these territories and neighborhoods, we were able to localize them in the adult mouse midbrain. Moreover, because the underlying mechanisms for the variable sensitivities of diverse mDA neurons to pathological insults are not well understood, we analyzed surviving neurons after partial 6-hydroxydopamine (6-OHDA) lesions to unravel gene expression patterns that correlate with mDA neuron vulnerability and resilience. Together, this atlas provides a basis for further studies on the neurophysiological role of mDA neurons in health and disease.

Two Years after the Beginning of COVID-19: Comparing Families Who Had or Did not Have Patients with COVID-19 on Health Beliefs and Obsessive-Compulsive Symptoms.

Objective: This study aimed to compare health beliefs and obsessive-compulsive symptoms (OCS) in families with (FIM+) or without an infected member (FIM-) two years after the beginning of COVID-19. Additionally, this research intended to predict a decrease in OCS from baseline (T1) to 40 days later (T2) based on health beliefs. Method : In a longitudinal survey, 227 participants in two groups, including FIM+ (n = 98; M = 30.44; SD = 5.39) and FIM- (n = 129; M = 29.24; SD = 4.93), were selected through purposive sampling. They responded to measurements consisting of demographic characteristics, the Obsessive-Compulsive Inventory-Revised (OCI-R), Patient Health Questionnaire (PHQ-9), Impact of Event Scale-Revised (IES-R), and COVID-19 Health Belief Questionnaire (COVID-19-HBQ) at the final assessment phase (T2). To investigate differences between the two groups and predict OCS changes from T1 to T2, data were analyzed using Chi-squared, t-tests, U-Mann-Whitney, Kruskal-Wallis, Pearson correlations, and linear regression analyses. Results: At T1, FIM+ demonstrated significantly greater OCS, health beliefs, posttraumatic stress symptoms (PTS), and depressive symptoms than FIM-. Furthermore, FIM+ showed a decrease in OCS from T1 to T2 after its infected member recovered from COVID-19 (P < 0.001). A decrease in OCS was correlated with a decrease in perceived susceptibility, severity, and barriers. Lack of a vulnerable family member, lower educational attainment, and being a primary caregiver were associated with a greater decrease in OCS. Changes in perceived severity and self-efficacy accounted for 17% of variation in OCS. Conclusion: Even two years after the onset of the pandemic, COVID-19 not only impacts the life of patients with COVID-19 but family members who care for such patients respond to the disease by engaging in excessive health behaviors in the form of OCS.

PRC2-mediated repression is essential to maintain identity and function of differentiated dopaminergic and serotonergic neurons.

How neurons can maintain cellular identity over an entire life span remains largely unknown. Here, we show that maintenance of identity in differentiated dopaminergic and serotonergic neurons is critically reliant on the Polycomb repressive complex 2 (PRC2). Deletion of the obligate PRC2 component, Eed, in these neurons resulted in global loss of H3K27me3, followed by a gradual activation of genes harboring both H3K27me3 and H3K9me3 modifications. Notably, H3K9me3 was lost at these PRC2 targets before gene activation. Neuronal survival was not compromised; instead, there was a reduction in subtype-specific gene expression and a progressive impairment of dopaminergic and serotonergic neuronal function, leading to behavioral deficits characteristic of Parkinson's disease and anxiety. Single-cell analysis revealed subtype-specific vulnerability to loss of PRC2 repression in dopamine neurons of the substantia nigra. Our study reveals that a PRC2-dependent nonpermissive chromatin state is essential to maintain the subtype identity and function of dopaminergic and serotonergic neurons.

Anti-Disgust Cognitive Behavioral Therapy for Contamination-Based Obsessive Compulsive Disorder: A Randomized Controlled Clinical Trial.

Background: Disgust is a strong and persistent emotion that frequently occurs during exposure-based treatments for contamination-based obsessive compulsive disorder (C-OCD). This study aimed to examine the efficacy of augmenting cognitive behavioral therapy (CBT) with a novel type of anti-disgust cognitive intervention in reducing the severity of OCD, disgust propensity/sensitivity, and refusal rate of exposure and response prevention, while simultaneously increasing acceptance of disgust. Materials and Methods: Fifty-five individuals with C-OCD (mean age 28.1 years, SD = 3.52; 77% female) were randomly assigned to 15 weekly sessions of anti-disgust plus CBT (AD-CBT) or CBT alone. They were evaluated for outcomes four times (pretreatment, prior to exposure and response prevention (ERP) sessions, posttreatment, and three-month follow-up), and mixed-design ANOVAs were used to analyze the data. Results: The findings indicated that when compared to CBT alone, AD-CBT significantly reduced OCD severity, disgust propensity/sensitivity, and concurrently increased disgust acceptance (p < 0.001). Additionally, engaging in an anti-disgust cognitive intervention was associated with lower ERP refusal rate (4% vs. 16%). The superiority of AD-CBT over CBT persisted through the three-month follow-up period. Conclusions: The current study suggests that supplementing CBT for C-OCD with an anti-disgust cognitive intervention significantly increased acceptance of disgust and decreased the refusal rate of ERP, OCD severity, and disgust-related factors.

Selective requirement for polycomb repressor complex 2 in the generation of specific hypothalamic neuronal subtypes.

The hypothalamus displays staggering cellular diversity, chiefly established during embryogenesis by the interplay of several signalling pathways and a battery of transcription factors. However, the contribution of epigenetic cues to hypothalamus development remains unclear. We mutated the polycomb repressor complex 2 gene Eed in the developing mouse hypothalamus, which resulted in the loss of H3K27me3, a fundamental epigenetic repressor mark. This triggered ectopic expression of posteriorly expressed regulators (e.g. Hox homeotic genes), upregulation of cell cycle inhibitors and reduced proliferation. Surprisingly, despite these effects, single cell transcriptomic analysis revealed that most neuronal subtypes were still generated in Eed mutants. However, we observed an increase in glutamatergic/GABAergic double-positive cells, as well as loss/reduction of dopamine, hypocretin and Tac2-Pax6 neurons. These findings indicate that many aspects of the hypothalamic gene regulatory flow can proceed without the key H3K27me3 epigenetic repressor mark, but points to a unique sensitivity of particular neuronal subtypes to a disrupted epigenomic landscape.

Variational autoencoding of gene landscapes during mouse CNS development uncovers layered roles of Polycomb Repressor Complex 2.

A prominent aspect of most, if not all, central nervous systems (CNSs) is that anterior regions (brain) are larger than posterior ones (spinal cord). Studies in Drosophila and mouse have revealed that Polycomb Repressor Complex 2 (PRC2), a protein complex responsible for applying key repressive histone modifications, acts by several mechanisms to promote anterior CNS expansion. However, it is unclear what the full spectrum of PRC2 action is during embryonic CNS development and how PRC2 intersects with the epigenetic landscape. We removed PRC2 function from the developing mouse CNS, by mutating the key gene Eed, and generated spatio-temporal transcriptomic data. To decode the role of PRC2, we developed a method that incorporates standard statistical analyses with probabilistic deep learning to integrate the transcriptomic response to PRC2 inactivation with epigenetic data. This multi-variate analysis corroborates the central involvement of PRC2 in anterior CNS expansion, and also identifies several unanticipated cohorts of genes, such as proliferation and immune response genes. Furthermore, the analysis reveals specific profiles of regulation via PRC2 upon these gene cohorts. These findings uncover a differential logic for the role of PRC2 upon functionally distinct gene cohorts that drive CNS anterior expansion. To support the analysis of emerging multi-modal datasets, we provide a novel bioinformatics package that integrates transcriptomic and epigenetic datasets to identify regulatory underpinnings of heterogeneous biological processes.

Genetic mechanisms controlling anterior expansion of the central nervous system.

In bilaterally-symmetric animals (Bilateria), condensation of neurons and ganglia into a centralized nervous system (CNS) constitutes a salient feature. In most, if not all, Bilateria another prominent aspect is that the anterior regions of the CNS are typically larger than the posterior ones. Detailed studies in Drosophila melanogaster (Drosophila) have revealed that anterior expansion in this species stems from three major developmental features: the generation of more progenitors anteriorly, an extended phase of proliferation of anterior progenitors, and more proliferative daughter cells in anterior regions. These brain-specific features combine to generate a larger average lineage size and higher cell numbers in the brain, when compared to more posterior regions. Genetic studies reveal that these anterior-posterior (A-P) differences are controlled by the modulation of temporal programs, common to all progenitors, as well as by Hox homeotic genes, expressed in the nerve cord, and brain-specific factors. All of these regulatory features are gated by the action of the PRC2 epigenetic complex. Studies in mammals indicate that most, if not all of these anterior expansion principles and the underlying genetic programs are evolutionarily conserved. These findings further lend support for the recently proposed idea that the brain and nerve cord may have originated from different parts of the nervous system present in the Bilaterian ancestor. This brain-nerve cord "fusion" concept may help explain a number of the well-known fundamental differences in the biology of the brain, when compared to the nerve cord.

Anterior CNS expansion driven by brain transcription factors.

During CNS development, there is prominent expansion of the anterior region, the brain. In Drosophila, anterior CNS expansion emerges from three rostral features: (1) increased progenitor cell generation, (2) extended progenitor cell proliferation, (3) more proliferative daughters. We find that tailless (mouse Nr2E1/Tlx), otp/Rx/hbn (Otp/Arx/Rax) and Doc1/2/3 (Tbx2/3/6) are important for brain progenitor generation. These genes, and earmuff (FezF1/2), are also important for subsequent progenitor and/or daughter cell proliferation in the brain. Brain TF co-misexpression can drive brain-profile proliferation in the nerve cord, and can reprogram developing wing discs into brain neural progenitors. Brain TF expression is promoted by the PRC2 complex, acting to keep the brain free of anti-proliferative and repressive action of Hox homeotic genes. Hence, anterior expansion of the Drosophila CNS is mediated by brain TF driven 'super-generation' of progenitors, as well as 'hyper-proliferation' of progenitor and daughter cells, promoted by PRC2-mediated repression of Hox activity.

Evolutionarily conserved anterior expansion of the central nervous system promoted by a common PcG-Hox program.

A conserved feature of the central nervous system (CNS) is the prominent expansion of anterior regions (brain) compared with posterior (nerve cord). The cellular and regulatory processes driving anterior CNS expansion are not well understood in any bilaterian species. Here, we address this expansion in Drosophila and mouse. We find that, compared with the nerve cord, the brain displays extended progenitor proliferation, more elaborate daughter cell proliferation and more rapid cell cycle speed in both Drosophila and mouse. These features contribute to anterior CNS expansion in both species. With respect to genetic control, enhanced brain proliferation is severely reduced by ectopic Hox gene expression, by either Hox misexpression or by loss of Polycomb group (PcG) function. Strikingly, in PcG mutants, early CNS proliferation appears to be unaffected, whereas subsequent brain proliferation is severely reduced. Hence, a conserved PcG-Hox program promotes the anterior expansion of the CNS. The profound differences in proliferation and in the underlying genetic mechanisms between brain and nerve cord lend support to the emerging concept of separate evolutionary origins of these two CNS regions.

Anterior-Posterior Gradient in Neural Stem and Daughter Cell Proliferation Governed by Spatial and Temporal Hox Control.

A readily evident feature of animal central nervous systems (CNSs), apparent in all vertebrates and many invertebrates alike, is its "wedge-like" appearance, with more cells generated in anterior than posterior regions. This wedge could conceivably be established by an antero-posterior (A-P) gradient in the number of neural progenitor cells, their proliferation behaviors, and/or programmed cell death (PCD). However, the contribution of each of these mechanisms, and the underlying genetic programs, are not well understood. Building upon recent progress in the Drosophila melanogaster (Drosophila) ventral nerve cord (VNC), we address these issues in a comprehensive manner. We find that, although PCD plays a role in controlling cell numbers along the A-P axis, the main driver of the wedge is a gradient of daughter proliferation, with divisions directly generating neurons (type 0) being more prevalent posteriorly and dividing daughters (type I) more prevalent anteriorly. In addition, neural progenitor (NB) cell-cycle exit occurs earlier posteriorly. The gradient of type I > 0 daughter proliferation switch and NB exit combine to generate radically different average lineage sizes along the A-P axis, differing by more than 3-fold in cell number. We find that the Hox homeotic genes, expressed in overlapping A-P gradients and with a late temporal onset in NBs, trigger the type I > 0 daughter proliferation switch and NB exit. Given the highly evolutionarily conserved expression of overlapping Hox homeotic genes in the CNS, our results point to a common mechanism for generating the CNS wedge.

Global programmed switch in neural daughter cell proliferation mode triggered by a temporal gene cascade.

During central nervous system (CNS) development, progenitors typically divide asymmetrically, renewing themselves while budding off daughter cells with more limited proliferative potential. Variation in daughter cell proliferation has a profound impact on CNS development and evolution, but the underlying mechanisms remain poorly understood. We find that Drosophila embryonic neural progenitors (neuroblasts) undergo a programmed daughter proliferation mode switch, from generating daughters that divide once (type I) to generating neurons directly (type 0). This typeI>0 switch is triggered by activation of Dacapo (mammalian p21(CIP1)/p27(KIP1)/p57(Kip2)) expression in neuroblasts. In the thoracic region, Dacapo expression is activated by the temporal cascade (castor) and the Hox gene Antennapedia. In addition, castor, Antennapedia, and the late temporal gene grainyhead act combinatorially to control the precise timing of neuroblast cell-cycle exit by repressing Cyclin E and E2f. This reveals a logical principle underlying progenitor and daughter cell proliferation control in the Drosophila CNS.