Implementation and validation of single-cell genomics experiments in neuroscience.

Single-cell or single-nucleus transcriptomics is a powerful tool for identifying cell types and cell states. However, hypotheses derived from these assays, including gene expression information, require validation, and their functional relevance needs to be established. The choice of validation depends on numerous factors.

Estrogen Receptor Chromatin Profiling by CUT&RUN.

Gonadal steroid hormones, namely, testosterone, progesterone, and estrogens, influence the physiological state of an organism through the regulation of gene transcription. Steroid hormones activate nuclear hormone receptor (HR), transcription factors (TFs), which bind DNA in a tissue- and cell type-specific manner to influence cellular function. Identifying the genomic binding sites of HRs is essential to understanding mechanisms of hormone signaling across tissues and disease contexts.

Area postrema neurons mediate interleukin-6 function in cancer cachexia.

Interleukin-6 (IL-6) has been long considered a key player in cancer cachexia. It is believed that sustained elevation of IL-6 production during cancer progression causes brain dysfunctions, which ultimately result in cachexia. However, how peripheral IL-6 influences the brain remains poorly understood.

The future of sex and gender in research.

Our understanding of sex and gender evolves. We asked scientists about their work and the future of sex and gender research. They discuss, among other things, interdisciplinary collaboration, moving beyond binary conceptualizations, accounting for intersecting factors, reproductive strategies, expanding research on sex-related differences, and sex's dynamic nature.

Chronic stress and its effects on behavior, RNA expression of the bed nucleus of the stria terminalis, and the M-current of NPY neurons.

Mood disorders, like major depressive disorder, can be precipitated by chronic stress and are more likely to be diagnosed in cisgender women than in cisgender men. This suggests that stress signaling in the brain is sexually dimorphic. We used a chronic variable mild stress paradigm to stress female and male mice for 6 weeks, followed by an assessment of avoidance behavior: the open field test, the elevated plus maze, the light/dark box emergence test, and the novelty suppressed feeding test.

Comparative analysis of gonadal hormone receptor expression in the postnatal house mouse, meadow vole, and prairie vole brain.

The socially monogamous prairie vole (Microtus ochrogaster) and promiscuous meadow vole (Microtus pennsylvanicus) are closely related, but only prairie voles display long-lasting pair bonds, biparental care, and selective aggression towards unfamiliar individuals after pair bonding. These social behaviors in mammals are largely mediated by steroid hormone signaling in the social behavior network (SBN) of the brain. Hormone receptors are reproducible markers of sex differences that can provide more information than anatomy alone and can even be at odds with anatomical dimorphisms.

Practical solutions for including sex as a biological variable (SABV) in preclinical neuropsychopharmacological research.

Recently, many funding agencies have released guidelines on the importance of considering sex as a biological variable (SABV) as an experimental factor, aiming to address sex differences and avoid possible sex biases to enhance the reproducibility and translational relevance of preclinical research. In neuroscience and pharmacology, the female sex is often omitted from experimental designs, with researchers generalizing male-driven outcomes to both sexes, risking a biased or limited understanding of disease mechanisms and thus potentially ineffective therapeutics. Herein, we describe key methodological aspects that should be considered when sex is factored into in vitro and in vivo experiments and provide practical knowledge for researchers to incorporate SABV into preclinical research.

Area postrema neurons mediate interleukin-6 function in cancer-associated cachexia.

Interleukin-6 (IL-6) has been long considered a key player in cancer-associated cachexia . It is believed that sustained elevation of IL-6 production during cancer progression causes brain dysfunctions, which ultimately result in cachexia . However, how peripheral IL-6 influences the brain remains poorly understood.

Epigenetic Mechanisms of Brain Sexual Differentiation.

Across vertebrate species, gonadal hormones coordinate physiology with behavior to facilitate social interactions essential for reproduction and survival. In adulthood, these hormones activate neural circuits that regulate behaviors presenting differently in females and males, such as parenting and territorial aggression. Yet long before sex-typical behaviors emerge at puberty, transient hormone production during sensitive periods of neurodevelopment establish the circuits upon which adult hormones act.

Oxytocin receptor behavioral effects and cell types in the bed nucleus of the stria terminalis.

Oxytocin is a neuropeptide that can produce anxiolytic effects and promote social approach. However, emerging evidence shows that under some conditions, oxytocin can instead induce anxiety-related behaviors. These diverse effects of oxytocin appear to be mediated by circuit-specific actions.

Oestrogen engages brain MC4R signalling to drive physical activity in female mice.

Oestrogen depletion in rodents and humans leads to inactivity, fat accumulation and diabetes, underscoring the conserved metabolic benefits of oestrogen that inevitably decrease with age. In rodents, the preovulatory surge in 17β-oestradiol (E2) temporarily increases energy expenditure to coordinate increased physical activity with peak sexual receptivity. Here we report that a subset of oestrogen-sensitive neurons in the ventrolateral ventromedial hypothalamic nucleus (VMHvl) projects to arousal centres in the hippocampus and hindbrain, and enables oestrogen to rebalance energy allocation in female mice.

Specificity in sociogenomics: Identifying causal relationships between genes and behavior.

There has been rapid growth in the use of transcriptomic analyses to study the interplay between gene expression and behavior. Experience can modify gene expression in the brain, leading to changes in internal state and behavioral displays, while gene expression variation between species is thought to specify many innate behavior differences. However, providing a causal association between a gene and a given behavior remains challenging as it is difficult to determine when and where a gene contributes to the function of a behaviorally-relevant neuronal population.

Copolymerization of single-cell nucleic acids into balls of acrylamide gel.

We show the use of 5'-Acrydite oligonucleotides to copolymerize single-cell DNA or RNA into balls of acrylamide el (BAGs). Combining this step with split-and-pool techniques for creating barcodes yields a method with advantages in cost and scalability, depth of coverage, ease of operation, minimal cross-contamination, and efficient use of samples. We perform DNA copy number profiling on mixtures of cell lines, nuclei from frozen prostate tumors, and biopsy washes.

Sex Differences in the Epigenome: A Cause or Consequence of Sexual Differentiation of the Brain?

Females and males display differences in neural activity patterns, behavioral responses, and incidence of psychiatric and neurological diseases. Sex differences in the brain appear throughout the animal kingdom and are largely a consequence of the physiological requirements necessary for the distinct roles of the two sexes in reproduction. As with the rest of the body, gonadal steroid hormones act to specify and regulate many of these differences.

Signatures of sex: Sex differences in gene expression in the vertebrate brain.

Women and men differ in disease prevalence, symptoms, and progression rates for many psychiatric and neurological disorders. As more preclinical studies include both sexes in experimental design, an increasing number of sex differences in physiology and behavior have been reported. In the brain, sex-typical behaviors are thought to result from sex-specific patterns of neural activity in response to the same sensory stimulus or context.

A Central Extended Amygdala Circuit That Modulates Anxiety.

Both the amygdala and the bed nucleus of the stria terminalis (BNST) have been implicated in maladaptive anxiety characteristics of anxiety disorders. However, the underlying circuit and cellular mechanisms have remained elusive. Here we show that mice with gene deficiency in somatostatin-expressing (SOM) neurons exhibit heightened anxiety as measured in the elevated plus maze test and the open field test, two assays commonly used to assess anxiety-related behaviors in rodents.

Gene regulatory mechanisms underlying sex differences in brain development and psychiatric disease.

The sexual differentiation of the mammalian nervous system requires the precise coordination of the temporal and spatial regulation of gene expression in diverse cell types. Sex hormones act at multiple developmental time points to specify sex-typical differentiation during embryonic and early development and to coordinate subsequent responses to gonadal hormones later in life by establishing sex-typical patterns of epigenetic modifications across the genome. Thus, mutations associated with neuropsychiatric conditions may result in sexually dimorphic symptoms by acting on different neural substrates or chromatin landscapes in males and females.

Estrogen receptor alpha is required in GABAergic, but not glutamatergic, neurons to masculinize behavior.

Masculinization of the altricial rodent brain is driven by estrogen signaling during a perinatal critical period. Genetic deletion of estrogen receptor alpha (Esr1/ERα) results in altered hypothalamic-pituitary-gonadal (HPG) axis signaling and a dramatic reduction of male sexual and territorial behaviors. However, the role of ERα in masculinizing distinct classes of neurons remains unexplored.

Epithelial cell integrin β1 is required for developmental angiogenesis in the pituitary gland.

As a key component of the vertebrate neuroendocrine system, the pituitary gland relies on the progressive and coordinated development of distinct hormone-producing cell types and an invading vascular network. The molecular mechanisms that drive formation of the pituitary vasculature, which is necessary for regulated synthesis and secretion of hormones that maintain homeostasis, metabolism, and endocrine function, remain poorly understood. Here, we report that expression of integrin β1 in embryonic pituitary epithelial cells is required for angiogenesis in the developing mouse pituitary gland.

Notch-Dependent Pituitary SOX2(+) Stem Cells Exhibit a Timed Functional Extinction in Regulation of the Postnatal Gland.

Although SOX2(+) stem cells are present in the postnatal pituitary gland, how they are regulated molecularly and whether they are required for pituitary functions remain unresolved questions. Using a conditional knockout animal model, here we demonstrate that ablation of the canonical Notch signaling in the embryonic pituitary gland leads to progressive depletion of the SOX2(+) stem cells and hypoplastic gland. Furthermore, we show that the SOX2(+) stem cells initially play a significant role in contributing to postnatal pituitary gland expansion by self-renewal and differentiating into distinct lineages in the immediate postnatal period.

Required enhancer-matrin-3 network interactions for a homeodomain transcription program.

Homeodomain proteins, described 30 years ago, exert essential roles in development as regulators of target gene expression; however, the molecular mechanisms underlying transcriptional activity of homeodomain factors remain poorly understood. Here investigation of a developmentally required POU-homeodomain transcription factor, Pit1 (also known as Pou1f1), has revealed that, unexpectedly, binding of Pit1-occupied enhancers to a nuclear matrin-3-rich network/architecture is a key event in effective activation of the Pit1-regulated enhancer/coding gene transcriptional program. Pit1 association with Satb1 (ref.

A custody battle for the mind: evidence for extensive imprinting in the brain.

Relatively few genes (approximately 100) have previously been shown to be imprinted such that their expression in progeny derives from either the maternal or paternal copy. Two recent studies by Gregg et al. (2010a, 2010b) in Science expand this list by an order of magnitude, revealing complex patterns of parent-of-origin bias in gene expression in the brain that are developmentally and regionally restricted, and in many cases, sexually dimorphic.

The androgen receptor governs the execution, but not programming, of male sexual and territorial behaviors.

Testosterone and estrogen are essential for male behaviors in vertebrates. How these two signaling pathways interact to control masculinization of the brain and behavior remains to be established. Circulating testosterone activates the androgen receptor (AR) and also serves as the source of estrogen in the brain.