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Ageing is the main risk factor common to most primary neurodegenerative disorders. Indeed, age-related brain alterations have been long considered to predispose to neurodegeneration. Although protein misfolding and the accumulation of toxic protein aggregates have been contemplated as causative events in neurodegeneration, several biological pathways affected by brain ageing are also contributing to pathogenesis. Here, we discuss the evidence showing the involvement of the mechanisms controlling neuronal structure, gene expression, autophagy, cell metabolism, and neuroinflammation in the onset and progression of neurodegenerative disorders. Furthermore, we review the therapeutic strategies currently under development or as future approaches designed to normalize these pathways, which may then boost brain resilience to cope with toxic protein species. Therefore, in addition to therapies targeting the insoluble protein aggregates specifically associated with each neurodegenerative disorder, these novel pharmacological approaches may be part of combined therapies designed to rescue brain function.

Schizophrenia is a neuropsychiatric disorder, caused by a combination of genetic and environmental factors. Recently, metabolomic studies based on patients’ biofluids and post-mortem brain specimens have revealed altered levels of distinct metabolites between healthy individuals and patients with schizophrenia (SCZ). However, a putative link between dysregulated metabolites and distorted neurodevelopment has not been assessed and access to patients’ material is restricted. In this study, we aimed to investigate a presumed correlation between transcriptomics and metabolomics in a SCZ model using patient-derived induced pluripotent stem cells (iPSCs). iPSCs were differentiated towards cortical neurons and samples were collected longitudinally at defined developmental stages, such as neuroepithelium, radial glia, young and mature neurons. Samples were subsequently analyzed by bulk RNA-sequencing and targeted metabolomics. The transcriptomic analysis revealed dysregulations in several extracellular matrix-related genes in the SCZ samples observed in early neurogenesis, including members of the collagen superfamily. At the metabolic level, several lipid and amino acid discrepancies were correlated to the SCZ phenotype. By employing a novel in silico analysis, we correlated the transcriptome with the metabolome through the generation of integrative networks. The network comparison between SCZ and healthy controls revealed a number of consistently affected pathways in SCZ, related to early stages of cortical development, indicating abnormalities in membrane composition, lipid homeostasis and amino acid imbalances. Ultimately, our study suggests a novel approach of correlating in vitro metabolic and transcriptomic data obtained from a patient-derived iPSC model. This type of analysis will offer novel insights in cellular and genetic mechanisms underlying the pathogenesis of complex neuropsychiatric disorders, such as schizophrenia.

The concept of senescence as a phenomenon limited to proliferating cells has been challenged by growing evidence of senescence-like features in terminally differentiated cells, including neurons. The persistence of senescent cells late in life is associated with tissue dysfunction and increased risk of age-related disease. We found that Alzheimer's disease (AD) brains have significantly higher proportions of neurons that express senescence markers, and their distribution indicates bystander effects. AD patient-derived directly induced neurons (iNs) exhibit strong transcriptomic, epigenetic, and molecular biomarker signatures, indicating a specific human neuronal senescence-like state. AD iN single-cell transcriptomics revealed that senescent-like neurons face oncogenic challenges and metabolic dysfunction as well as display a pro-inflammatory signature. Integrative profiling of the inflammatory secretome of AD iNs and patient cerebral spinal fluid revealed a neuronal senescence-associated secretory phenotype that could trigger astrogliosis in human astrocytes. Finally, we show that targeting senescence-like neurons with senotherapeutics could be a strategy for preventing or treating AD.

The drivers of sporadic Alzheimer's disease (AD) remain incompletely understood. Utilizing directly converted induced neurons (iNs) from AD-patient-derived fibroblasts, we identified a metabolic switch to aerobic glycolysis in AD iNs. Pathological isoform switching of the glycolytic enzyme pyruvate kinase M (PKM) toward the cancer-associated PKM2 isoform conferred metabolic and transcriptional changes in AD iNs. These alterations occurred via PKM2's lack of metabolic activity and via nuclear translocation and association with STAT3 and HIF1α to promote neuronal fate loss and vulnerability. Chemical modulation of PKM2 prevented nuclear translocation, restored a mature neuronal metabolism, reversed AD-specific gene expression changes, and re-activated neuronal resilience against cell death.

Progressive aging is a physiological process that represents a central risk factor for the development of several human age-associated chronic diseases, including neurodegenerative diseases. A major focus in biomedical research is the pursuit for appropriate model systems to better model the biology of human aging and the interface between aging and disease mechanisms. Direct conversion of human fibroblasts into induced neurons (iNs) has emerged as a novel technology for the in vitro modeling of age-dependent neurological diseases. Similar to other cellular reprogramming techniques, e.g., iPSC-based cellular reprograming, direct conversion relies on the ectopic overexpression of transcription factors, typically including well-known pioneer factors. However, in contrast to alternative technologies to generate neurons, the entire process of direct conversion bypasses any proliferative or stem cell-like stage, which in fact renders it the unique aptitude of preserving age-associated hallmarks from the initial fibroblast source. In this chapter, we introduce direct conversion as a practical and easy-to-approach disease model for aging and neurodegenerative disease research. A focus here is to provide a stepwise protocol for the efficient and highly reproducible generation of iNs from adult dermal fibroblasts from human donors.

Progressive aging is a physiological process that represents a central risk factor for the development of several human age-associated chronic diseases, including neurodegenerative diseases. A major focus in biomedical research is the pursuit for appropriate model systems to better model the biology of human aging and the interface between aging and disease mechanisms. Direct conversion of human fibroblasts into induced neurons (iNs) has emerged as a novel technology for the in vitro modeling of age-dependent neurological diseases. Similar to other cellular reprogramming techniques, e.g., iPSC-based cellular reprograming, direct conversion relies on the ectopic overexpression of transcription factors, typically including well-known pioneer factors. However, in contrast to alternative technologies to generate neurons, the entire process of direct conversion bypasses any proliferative or stem cell-like stage, which in fact renders it the unique aptitude of preserving age-associated hallmarks from the initial fibroblast source. In this chapter, we introduce direct conversion as a practical and easy-to-approach disease model for aging and neurodegenerative disease research. A focus here is to provide a stepwise protocol for the efficient and highly reproducible generation of iNs from adult dermal fibroblasts from human donors.

Autism spectrum disorder (ASD) is thought to emerge during early cortical development. However, the exact developmental stages and associated molecular networks that prime disease propensity are elusive. To profile early neurodevelopmental alterations in ASD with macrocephaly, we monitored patient-derived induced pluripotent stem cells (iPSCs) throughout the recapitulation of cortical development. Our analysis revealed ASD-associated changes in the maturational sequence of early neuron development, involving temporal dysregulation of specific gene networks and morphological growth acceleration. The observed changes tracked back to a pathologically primed stage in neural stem cells (NSCs), reflected by altered chromatin accessibility. Concerted overrepresentation of network factors in control NSCs was sufficient to trigger ASD-like features, and circumventing the NSC stage by direct conversion of ASD iPSCs into induced neurons (iPSC-iNs) abolished ASD-associated phenotypes. Our findings identify heterochronic dynamics of a gene network that, while established earlier in development, contributes to subsequent neurodevelopmental aberrations in ASD.

With the advancing age of humans and with it, growing numbers of age-related diseases, aging has become a major focus in recent research. The lack of fitting aging models, especially in neurological diseases where access to human brain samples is limited, has highlighted direct conversion into induced neurons (iN) as an important method to overcome this challenge. Contrary to iPSC reprogramming and its corresponding cell rejuvenation, the generation of iNs enables us to retain aging signatures throughout the conversion process and beyond. In this review, we explore different cell reprogramming methods in light of age-associated neurodegenerative diseases and discuss different approaches, advances, and limitations.

Summary Sporadic Alzheimer’s disease (AD) exclusively affects elderly people. Using direct conversion of AD patient fibroblasts into induced neurons (iNs), we generated an age-equivalent neuronal model. AD patient-derived iNs exhibit strong neuronal transcriptome signatures characterized by downregulation of mature neuronal properties and upregulation of immature and progenitor-like signaling pathways. Mapping iNs to longitudinal neuronal differentiation trajectory data demonstrated that AD iNs reflect a hypo-mature neuronal identity characterized by markers of stress, cell cycle, and de-differentiation. Epigenetic landscape profiling revealed an underlying aberrant neuronal state that shares similarities with malignant transformation and age-dependent epigenetic erosion. To probe for the involvement of aging, we generated rejuvenated iPSC-derived neurons that showed no significant disease-related transcriptome signatures, a feature that is consistent with epigenetic clock and brain ontogenesis mapping, which indicate that fibroblast-derived iNs more closely reflect old adult brain stages. Our findings identify AD-related neuronal changes as age-dependent cellular programs that impair neuronal identity., Graphical abstract, Highlights • Alzheimer's patients’ induced neurons (iNs) show transcriptional and cellular defects • Alzheimer's iNs activate de-differentiation pathways and markers of neuronal fate loss • Epigenetic erosion underlies the pathogenic hypo-mature state of Alzheimer's iNs • Rejuvenated iPSC neurons show only very mild transcriptomic disease signatures, Mertens et al. generated directly induced neurons (iNs) from Alzheimer's patients’ fibroblasts that show neuronal defects and activate immature and de-differentiation signaling pathways. Aging- and cancer-associated epigenetic changes promote the underlying pathogenic hypo-mature neuronal state, and iPSC rejuvenation largely rescued the age-dependent disease phenotype.

Cell fate conversion generally requires reprogramming effectors to both introduce fate programs of the target cell type and erase the identity of starting cell population. Here, we reveal insights into the activity of microRNAs, miR-9/9* and miR-124 (miR-9/9*-124) as reprogramming agents that orchestrate direct conversion of human fibroblasts into motor neurons, by first eradicating fibroblast identity and promoting uniform transition to a neuronal state in sequence. We identify KLF-family transcription factors as direct target genes for miR-9/9*-124 and show their repression is critical for erasing fibroblast fate. Subsequent gain of neuronal identity requires upregulation of a small nuclear RNA, RN7SK, which induces accessibilities of chromatin regions and neuronal gene activation to push cells to a neuronal state. Our study defines deterministic components in the microRNA-mediated reprogramming cascade.

The human brain is heavily affected by cellular aging. Neurons are primarily generated during embryogenesis and early life with a limited capacity for renewal and replacement, making them some of the oldest cells in the human body. Our present understanding of neurodegenerative diseases points towards advanced neuronal age as a prerequisite for the development of these disorders. While significant progress has been made in understanding the relationship between aging and neurological disease, it will be essential to delve further into the molecular mechanisms of neuronal aging in order to develop therapeutic interventions targeting age-related brain dysfunction. In this mini review, we highlight recent findings on the relationship between the aging of nuclear structures and changes in the epigenetic landscape during neuronal aging and disease.

Direct conversion of human somatic fibroblasts into induced neurons (iNs) allows for the generation of functional neurons while bypassing any stem cell intermediary stages. Although iN technology has an enormous potential for modeling age-related diseases, as well as therapeutic approaches, the technology faces limitations due to variable conversion efficiencies and a lack of thorough understanding of the signaling pathways directing iN conversion. Here, we introduce a new all-in-one inducible lentiviral system that simplifies fibroblast transgenesis for the two pioneer transcription factors, Ngn2 and Ascl1, and markedly improves iN yields. Further, our timeline RNA-Seq data across the course of conversion has identified signaling pathways that become transcriptionally enriched during iN conversion. Small molecular modulators were identified for four signaling pathways that reliably increase the yield of iNs. Taken together, these advances provide an improved toolkit for iN technology and new insight into the mechanisms influencing direct iN conversion.

Technologies for deriving human neurons in vitro have transformed our ability to study cellular and molecular components of human neurotransmission. Three groups, including our own, have recently published methods for efficiently generating human serotonergic neurons in vitro. Remarkably, serotonergic neurons derived from each method robustly produce serotonin, express raphe genes, are electrically active, and respond to selective serotonin reuptake inhibitors in vitro. Two of the methods utilize transdifferentiation technology by overexpressing key serotonergic transcription factors. The third and most recent method involves differentiating induced pluripotent stem cells (iPSCs) to serotonergic neurons using developmental patterning cues. In this mini-review, we briefly describe the developmental programs governing serotonergic specification in vivo and how they have been harnessed to achieve serotonergic differentiation in vitro. We discuss the distinct and overlapping features of the recently published methodologies and their value in the context of in vitro disease modeling. Also see the video abstract here.

Age-associated neurological diseases represent a profound challenge in biomedical research as we are still struggling to understand the interface between the aging process and the manifestation of disease. Various pathologies in the elderly do not directly result from genetic mutations, toxins, or infectious agents but are primarily driven by the many manifestations of biological aging. Therefore, the generation of appropriate model systems to study human aging in the nervous system demands new concepts that lie beyond transgenic and drug-induced models. Although access to viable human brain specimens is limited and induced pluripotent stem cell models face limitations due to reprogramming-associated cellular rejuvenation, the direct conversion of somatic cells into induced neurons allows for the generation of human neurons that capture many aspects of aging. Here, we review advances in exploring age-associated neurodegenerative diseases using human cell reprogramming models, and we discuss general concepts, promises, and limitations of the field.

Summary Tau is the major constituent of neurofibrillary tangles in Alzheimer’s disease (AD), but the mechanism underlying tau-associated neural damage remains unclear. Here, we show that tau can directly interact with nucleoporins of the nuclear pore complex (NPC) and affect their structural and functional integrity. Pathological tau impairs nuclear import and export in tau-overexpressing transgenic mice and in human AD brain tissue. Furthermore, the nucleoporin Nup98 accumulates in the cell bodies of some tangle-bearing neurons and can facilitate tau aggregation in vitro. These data support the hypothesis that tau can directly interact with NPC components, leading to their mislocalization and consequent disruption of NPC function. This raises the possibility that NPC dysfunction contributes to tau-induced neurotoxicity in AD and tauopathies.

C9orf72 mutations are the most common cause of amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). Dipeptide repeat proteins (DPRs) produced by unconventional translation of the C9orf72 repeat expansions cause neurodegeneration in cell culture and in animal models. We performed two unbiased screens in Saccharomyces cerevisiae and identified potent modifiers of DPR toxicity, including karyopherins and effectors of Ran-mediated nucleocytoplasmic transport, providing insight into potential disease mechanisms and therapeutic targets.

Tau protein, which normally functions to stabilize microtubules, is the major constituent of neurofibrillary tangles in Alzheimer’s disease (AD). The mechanism underlying tau-associated neural damage remains unclear. We now show that tau can interact with nuclear pore complex (NPC) constituents and affect their structural and functional integrity. Pathological tau leads to dissociation of nuclear pore complex proteins, and impairs nuclear export and import in vitro, in tau overexpressing transgenic mouse models, and in human AD tissue. Moreover, a nuclear pore component, Nup98, surprisingly colocalizes with neurofibrillary tangles in neuronal soma, and both in vivo and in vitro directly interacts with tau to greatly facilitate its aggregation. These data support the hypothesis that tau directly interacts with nuclear pore complex constituents, leading to their mislocalization and to disruption of nuclear pore function, raising the possibility that nuclear pore dysfunction contributes to tau-induced neurotoxicity in Alzheimer’s disease.

Summary Mitochondria are a major target for aging and are instrumental in the age-dependent deterioration of the human brain, but studying mitochondria in aging human neurons has been challenging. Direct fibroblast-to-induced neuron (iN) conversion yields functional neurons that retain important signs of aging, in contrast to iPSC differentiation. Here, we analyzed mitochondrial features in iNs from individuals of different ages. iNs from old donors display decreased oxidative phosphorylation (OXPHOS)-related gene expression, impaired axonal mitochondrial morphologies, lower mitochondrial membrane potentials, reduced energy production, and increased oxidized proteins levels. In contrast, the fibroblasts from which iNs were generated show only mild age-dependent changes, consistent with a metabolic shift from glycolysis-dependent fibroblasts to OXPHOS-dependent iNs. Indeed, OXPHOS-induced old fibroblasts show increased mitochondrial aging features similar to iNs. Our data indicate that iNs are a valuable tool for studying mitochondrial aging and support a bioenergetic explanation for the high susceptibility of the brain to aging.

The functional connection of Alzheimer’s disease (AD) with adult neurogenesis remains a lingering question in the field. A deep understanding of the detrimental effects of AD pathology on neurogenesis and a potentially compensatory role for newborn neurons in dementia may lead to a more complete picture of AD pathology and new therapeutic avenues. The few human autopsy studies that have been conducted are as inconsistent as the results from mouse models, including diametrically opposed reports in some cases. The study by Ekonomou et al. (1) tries to resolve this issue by applying stringent exclusion criteria for the examination of a clearly defined population of postmortem AD brains at various stages. Using multiple neurogenic markers for different maturation levels along with glial markers, the aim of this study is to complement the pathologic picture. The main observation of the study by Ekonomou et al. is a decrease in newly generated neurons in the dentate gyrus (DG) with advancing Braak stages (more severe tau pathology). This decrease was found only at the level of immature

Cellular heterogeneity, for example, the intratumoral coexistence of cancer cells with and without stem cell characteristics, represents a potential root of therapeutic resistance and a significant challenge for modern drug development in glioblastoma (GBM). We propose here that activation of the innate immune system by stimulation of innate immune receptors involved in antiviral and antitumor responses can similarly target different malignant populations of glioma cells. We used short-term expanded patient-specific primary human GBM cells to study the stimulation of the cytosolic nucleic acid receptors melanoma differentiation-associated gene 5 (MDA5) and retinoic acid-inducible gene I (RIG-I). Specifically, we analyzed cells from the tumor core versus “residual GBM cells” derived from the tumor resection margin as well as stem cell-enriched primary cultures versus specimens without stem cell properties. A portfolio of human, nontumor neural cells was used as a control for these studies. The expression of RIG-I and MDA5 could be induced in all of these cells. Receptor stimulation with their respective ligands, p(I:C) and 3pRNA, led to in vitro evidence for an effective activation of the innate immune system. Most intriguingly, all investigated cancer cell populations additionally responded with a pronounced induction of apoptotic signaling cascades revealing a second, direct mechanism of antitumor activity. By contrast, p(I:C) and 3pRNA induced only little toxicity in human nonmalignant neural cells. Granted that the challenge of effective central nervous system (CNS) delivery can be overcome, targeting of RIG-I and MDA5 could thus become a quintessential strategy to encounter heterogeneous cancers in the sophisticated environments of the brain.

The scarcity of live human brain cells for experimental access has for a long time limited our ability to study complex human neurological disorders and elucidate basic neuroscientific mechanisms. A decade ago, the development of methods to reprogramme somatic human cells into induced pluripotent stem cells enabled the in vitro generation of a wide range of neural cells from virtually any human individual. The growth of methods to generate more robust and defined neural cell types through reprogramming and direct conversion into induced neurons has led to the establishment of various human reprogramming-based neural disease models.

Background Bipolar disorder is a serious and common psychiatric disorder characterized by manic and depressive mood switches and a relapsing and remitting course. The cornerstone of clinical management is stabilization and prophylaxis using mood-stabilizing medications to reduce both manic and depressive symptoms. Lithium remains the gold standard of treatment with the strongest data for both efficacy and suicide prevention. However, many patients do not respond to this medication, and clinically there is a great need for tools to aid the clinician in selecting the correct treatment. Large genome wide association studies (GWAS) investigating retrospectively the effect of lithium response are in the pipeline; however, few large prospective studies on genetic predictors to of lithium response have yet been conducted. The purpose of this project is to identify genes that are associated with lithium response in a large prospective cohort of bipolar patients and to better understand the mechanism of action of lithium and the variation in the genome that influences clinical response. Methods/Design This study is an 11-site prospective non-randomized open trial of lithium designed to ascertain a cohort of 700 subjects with bipolar I disorder who experience protocol-defined relapse prevention as a result of treatment with lithium monotherapy. All patients will be diagnosed using the Diagnostic Interview for Genetic Studies (DIGS) and will then enter a 2-year follow-up period on lithium monotherapy if and when they exhibit a score of 1 (normal, not ill), 2 (minimally ill) or 3 (mildly ill) on the Clinical Global Impressions of Severity Scale for Bipolar Disorder (CGI-S-BP Overall Bipolar Illness) for 4 of the 5 preceding weeks. Lithium will be titrated as clinically appropriate, not to exceed serum levels of 1.2 mEq/L. The sample will be evaluated longitudinally using a wide range of clinical scales, cognitive assessments and laboratory tests. On relapse, patients will be discontinued or crossed-over to treatment with valproic acid (VPA) or treatment as usual (TAU). Relapse is defined as a DSM-IV manic, major depressive or mixed episode or if the treating physician decides a change in medication is clinically necessary. The sample will be genotyped for GWAS. The outcome for lithium response will be analyzed as a time to event, where the event is defined as clinical relapse, using a Cox Proportional Hazards model. Positive single nucleotide polymorphisms (SNPs) from past genetic retrospective studies of lithium response, the Consortium on Lithium Genetics (ConLiGen), will be tested in this prospective study sample; a meta-analysis of these samples will then be performed. Finally, neurons will be derived from pluripotent stem cells from lithium responders and non-responders and tested in vivo for response to lithium by gene expression studies. SNPs in genes identified in these cellular studies will also be tested for association to response. Discussion Lithium is an extraordinarily important therapeutic drug in the clinical management of patients suffering from bipolar disorder. However, a significant proportion of patients, 30–40 %, fail to respond, and there is currently no method to identify the good lithium responders before initiation of treatment. Converging evidence suggests that genetic factors play a strong role in the variation of response to lithium, but only a few genes have been tested and the samples have largely been retrospective or quite small. The current study will collect an entirely unique sample of 700 patients with bipolar disorder to be stabilized on lithium monotherapy and followed for up to 2 years. This study will produce useful information to improve the understanding of the mechanism of action of lithium and will add to the development of a method to predict individual response to lithium, thereby accelerating recovery and reducing suffering and cost. © 2016 Oedegaard et al.Open AccessThis article is distributed under the terms of the Creative Commons Attribution 4.0International License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, distribution, andreproduction in any medium, provided you give appropriate credit to the original author(s) and the source, provide a link tothe Creative Commons license, and indicate if changes were made. The Creative Commons Public Domain Dedication waiver(http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated

Alzheimer's disease (AD) is the most frequent cause of dementia. There is compelling evidence that the proteolytic processing of the amyloid precursor protein (APP) and accumulation of amyloid-β (Aβ) peptides play critical roles in AD pathogenesis. Due to limited access to human neural tissue, pathogenetic studies have, so far, mostly focused on the heterologous overexpression of mutant human APP in non-human cells. In this study, we show that key steps in proteolytic APP processing are recapitulated in neurons generated from human embryonic and induced pluripotent stem cell-derived neural stem cells (NSC). These human NSC-derived neurons express the neuron-specific APP(695) splice variant, BACE1, and all members of the γ-secretase complex. The human NSC-derived neurons also exhibit a differentiation-dependent increase in Aβ secretion and respond to the pharmacotherapeutic modulation by anti-amyloidogenic compounds, such as γ-secretase inhibitors and nonsteroidal anti-inflammatory drugs. Being highly amenable to genetic modification, human NSCs enable the study of mechanisms caused by disease-associated mutations in human neurons. Interestingly, the AD-associated PS1(L166P) variant revealed a partial loss of γ-secretase function, resulting in the decreased production of endogenous Aβ40 and an increased Aβ42/40 ratio. The PS1(L166P) mutant is also resistant to γ-secretase modulation by nonsteroidal anti-inflammatory drugs. Pluripotent stem cell-derived neurons thus provide experimental access to key steps in AD pathogenesis and can be used to screen pharmaceutical compounds directly in a human neuronal system.

Forced expression of proneural transcription factors has been shown to direct neuronal conversion of fibroblasts. Because neurons are postmitotic, conversion efficiencies are an important parameter for this process. We present a minimalist approach combining two-factor neuronal programming with small molecule-based inhibition of glycogen synthase kinase-3β and SMAD signaling, which converts postnatal human fibroblasts into functional neuron-like cells with yields up to >200% and neuronal purities up to >80%.

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How metabolism is reprogrammed during neuronal differentiation is unknown. We found that the loss of hexokinase (HK2) and lactate dehydrogenase (LDHA) expression, together with a switch in pyruvate kinase gene splicing from PKM2 to PKM1, marks the transition from aerobic glycolysis in neural progenitor cells (NPC) to neuronal oxidative phosphorylation. The protein levels of c-MYC and N-MYC, transcriptional activators of the HK2 and LDHA genes, decrease dramatically. Constitutive expression of HK2 and LDHA during differentiation leads to neuronal cell death, indicating that the shut-off aerobic glycolysis is essential for neuronal survival. The metabolic regulators PGC-1α and ERRγ increase significantly upon neuronal differentiation to sustain the transcription of metabolic and mitochondrial genes, whose levels are unchanged compared to NPCs, revealing distinct transcriptional regulation of metabolic genes in the proliferation and post-mitotic differentiation states. Mitochondrial mass increases proportionally with neuronal mass growth, indicating an unknown mechanism linking mitochondrial biogenesis to cell size. DOI: http://dx.doi.org/10.7554/eLife.13374.001, eLife digest Structures called mitochondria act like the batteries of cells, and use several different metabolic processes to release energy. For example, neurons rely on a metabolic process called oxidative phosphorylation, while neural progenitor cells (which develop, or differentiate, into neurons) use a process called aerobic glycolysis instead. Little is known about why neurons prefer to use oxidative phosphorylation to provide them with energy, and it is also not clear why problems that affect this process are often seen in neurological disorders and neurodegenerative diseases. Zheng, Boyer et al. have now used human neural progenitor cells to explore the metabolic changes that occur as these cells develop into neurons. It appears that the loss of two metabolic enzymes, called hexokinase and lactate dehydrogenase, marks the transition from aerobic glycolysis to oxidative phosphorylation. In addition, the instructions to produce an enzyme called pyruvate kinase are altered or “alternatively spliced” when progenitor cells differentiate, which in turn changes the structure of the enzyme. The levels of the proteins that activate and regulate the production of these three metabolic enzymes also decrease dramatically during this transition. Further experiments showed that neurons that produce hexokinase and lactate dehydrogenase while they differentiate die, which means that neurons must shut off aerobic glycolysis in order to survive. The amounts of two proteins that regulate metabolism (called PGC-1α and ERRγ) increase significantly when a neuron differentiates. This sustains a constant level of activity for several metabolic and mitochondrial genes as neural progenitor cells differentiate to form neurons. Zheng, Boyer et al. also found that neurons build more mitochondria as they grow; this suggests that an unknown mechanism exists that links the creation of mitochondria to the size of the neuron. Zheng, Boyer et al. have mainly focused on how much of each metabolic enzyme is produced inside cells, but these levels may not completely reflect the actual level of enzyme activity. The next steps are therefore to investigate whether any other processes or modifications play a part in regulating the enzymes. Further investigation is also needed to determine the effects of changes in mitochondrial structure that occur as a neuron develops from a neural progenitor cell. DOI: http://dx.doi.org/10.7554/eLife.13374.002

Bipolar disorder is a complex neuropsychiatric disorder that is characterized by intermittent episodes of mania and depression; without treatment, 15% of patients commit suicide1. Hence, it has been ranked by the World Health Organization as a top disorder of morbidity and lost productivity2. Previous neuropathological studies have revealed a series of alterations in the brains of patients with bipolar disorder or animal models3, such as reduced glial cell number in the prefrontal cortex of patients4, upregulated activities of the protein kinase A and C pathways5–7 and changes in neurotransmission8–11. However, the roles and causation of these changes in bipolar disorder have been too complex to exactly determine the pathology of the disease. Furthermore, although some patients show remarkable improvement with lithium treatment for yet unknown reasons, others are refractory to lithium treatment. Therefore, developing an accurate and powerful biological model for bipolar disorder has been a challenge. The introduction of induced pluripotent stem-cell (iPSC) technology has provided a new approach. Here we have developed an iPSC model for human bipolar disorder and investigated the cellular phenotypes of hippocampal dentate gyrus-like neurons derived from iPSCs of patients with bipolar disorder. Guided by RNA sequencing expression profiling, we have detected mitochondrial abnormalities in young neurons from patients with bipolar disorder by using mitochondrial assays; in addition, using both patch-clamp recording and somatic Ca2+ imaging, we have observed hyperactive action-potential firing. This hyperexcitability phenotype of young neurons in bipolar disorder was selectively reversed by lithium treatment only in neurons derived from patients who also responded to lithium treatment. Therefore, hyperexcitability is one early endophenotype of bipolar disorder, and our model of iPSCs in this disease might be useful in developing new therapies and drugs aimed at its clinical treatment.

Regulatory networks for differentiation and pluripotency in embryonic stem (ES) cells have long been suggested to be mutually exclusive. However, with the identification of many new components of these networks ranging from epigenetic, transcriptional and translational to even post-translational mechanisms, the cellular states of pluripotency and early differentiation might not be strictly bi-modal, but differentiating stem cells appear to go through phases of simultaneous expression of stemness and differentiation genes. Translational regulators such as RNA binding proteins (RBPs) and micro RNAs (miRNAs) might be prime candidates for guiding a cell from pluripotency to differentiation. Using Trim71, one of two members of the Tripartite motif (Trim) protein family with RNA binding activity expressed in murine ES cells, we demonstrate that Trim71 is not involved in regulatory networks of pluripotency but regulates neural differentiation. Loss of Trim71 in mES cells leaves stemness and self-maintenance of these cells intact, but many genes required for neural development are up-regulated at the same time. Concordantly, Trim71−/− mES show increased neural marker expression following treatment with retinoic acid. Our findings strongly suggest that Trim71 keeps priming steps of differentiation in check, which do not pre-require a loss of the pluripotency network in ES cells.

Item does not contain fulltext The brain's serotonergic system centrally regulates several physiological processes and its dysfunction has been implicated in the pathophysiology of several neuropsychiatric disorders. While in the past our understanding of serotonergic neurotransmission has come mainly from mouse models, the development of pluripotent stem cell and induced fibroblast-to-neuron (iN) transdifferentiation technologies has revolutionized our ability to generate human neurons in vitro. Utilizing these techniques and a novel lentiviral reporter for serotonergic neurons, we identified and overexpressed key transcription factors to successfully generate human serotonergic neurons. We found that overexpressing the transcription factors NKX2.2, FEV, GATA2 and LMX1B in combination with ASCL1 and NGN2 directly and efficiently generated serotonergic neurons from human fibroblasts. Induced serotonergic neurons (iSNs) showed increased expression of specific serotonergic genes that are known to be expressed in raphe nuclei. iSNs displayed spontaneous action potentials, released serotonin in vitro and functionally responded to selective serotonin reuptake inhibitors (SSRIs). Here, we demonstrate the efficient generation of functional human serotonergic neurons from human fibroblasts as a novel tool for studying human serotonergic neurotransmission in health and disease.

Summary Neural progenitor cells (NeuPCs) possess a unique nuclear architecture that changes during differentiation. Nucleoporins are linked with cell-type-specific gene regulation, coupling physical changes in nuclear structure to transcriptional output; but, whether and how they coordinate with key fate-determining transcription factors is unclear. Here we show that the nucleoporin Nup153 interacts with Sox2 in adult NeuPCs, where it is indispensable for their maintenance and controls neuronal differentiation. Genome-wide analyses show that Nup153 and Sox2 bind and co-regulate hundreds of genes. Binding of Nup153 to gene promoters or transcriptional end sites correlates with increased or decreased gene expression, respectively, and inhibiting Nup153 expression alters open chromatin configurations at its target genes, disrupts genomic localization of Sox2, and promotes differentiation in vitro and a gliogenic fate switch in vivo. Together, these findings reveal that nuclear structural proteins may exert bimodal transcriptional effects to control cell fate.

Sophisticated protocols for the derivation of defined somatic cell cultures from human pluripotent stem cells have opened fascinating prospects for modeling human diseases in vitro. This is particularly relevant for the study of nervous system disorders, where so far the lack of primary tissue has precluded the development of standardized, cell-based in vitro models. We have recently described the derivation of long-term, self-renewing neuroepithelial stem cells (lt-NES cells) from human pluripotent stem cells. Here we report on how this stable somatic stem cell population can be used to study pathogenic mechanisms underlying Alzheimer’s disease (AD) and polyglutamine disorders. Specifically, we demonstrate that human neurons derived from lt-NES cells exhibit the entire machinery required for proteolytic processing of the amyloid precursor protein (APP) and are suitable for studying mutants associated with familial variants of AD as well as pharmaceutical compounds modulating the formation of amyloid beta (Aβ). Using induced pluripotent stem cell-derived lt-NES cells from patients with Machado-Joseph disease as an example, we further show that this cellular model provides experimental access to the molecular events initiating pathological protein aggregation – one of the most common denominators of human neurodegenerative disease.

Alterations in the microtubule (MT)–associated protein, tau, have emerged as a pivotal phenomenon in several neurodegenerative disorders, including frontotemporal dementia and Alzheimer's disease. Although compelling lines of evidence from various experimental models suggest that hyperphosphorylation and conformational changes of tau can cause its aggregation into filaments, the actual tau species and effective mechanisms that conspire to trigger the degeneration of human neurons remain obscure. Herein, we explored whether human embryonic stem cell–derived neural stem cells can be exploited to study consequences of an overexpression of 2N4R tau (two normal N-terminal and four MT-binding domains; n-tau) versus pseudohyperphosphorylated tau (p-tau) directly in human neurons. Given the involvement of tau in MT integrity and cellular homeostasis, we focused on the effects of both tau variants on subcellular transport and neuronal survival. By using inducible lentiviral overexpression, we show that p-tau, but not n-tau, readily leads to an MC-1–positive protein conformation and impaired mitochondrial transport. Although these alterations do not induce cell death under standard culture conditions, p-tau–expressing neurons cultured under non–redox-protected conditions undergo degeneration with formation of axonal varicosities sequestering transported proteins and progressive neuronal cell death. Our data support a causative link between the phosphorylation and conformational state of tau, microtubuli-based transport, and the vulnerability of human neurons to oxidative stress. They further depict human embryonic stem cell–derived neurons as a useful experimental model for studying tau-associated cellular alterations in an authentic human system.

Summary Increasing evidence suggests that elevated Aβ42 fractions in the brain cause Alzheimer’s disease (AD). Although γ-secretase modulators (GSMs), including a set of nonsteroidal anti-inflammatory drugs (NSAIDs), were found to lower Aβ42 in various model systems, NSAID-based GSMs proved to be surprisingly inefficient in human clinical trials. Reasoning that the nonhuman and nonneuronal cells typically used in pharmaceutical compound validation might not adequately reflect the drug responses of human neurons, we used human pluripotent stem cell-derived neurons from AD patients and unaffected donors to explore the efficacy of NSAID-based γ-secretase modulation. We found that pharmaceutically relevant concentrations of these GSMs that are clearly efficacious in conventional nonneuronal cell models fail to elicit any effect on Aβ42/Aß40 ratios in human neurons. Our work reveals resistance of human neurons to NSAID-based γ-secretase modulation, highlighting the need to validate compound efficacy directly in the human cell type affected by the respective disease., Graphical Abstract, Highlights • iPSC-derived neurons from Alzheimer patients exhibit elevated Aβ42/Aß40 ratios • Human neurons are resistant to NSAID-based γ-secretase modulation, Koch, Brüstle, and colleagues employed induced pluripotent stem cell-derived neurons (iPSC-Ns) to explore the clinical failure of NSAID-based γ-secretase modulators in the treatment of Alzheimer’s disease (AD). In contrast to the nonneuronal cells typically used in pharmaceutical screening, iPSC-Ns from healthy donors and AD patients were found to exhibit a remarkable resistance to this compound family, thereby supporting the value of iPSCs for predicting drug responsiveness.

Item does not contain fulltext Human cell reprogramming technologies offer access to live human neurons from patients and provide a new alternative for modeling neurological disorders in vitro. Neural electrical activity is the essence of nervous system function in vivo. Therefore, we examined neuronal activity in media widely used to culture neurons. We found that classic basal media, as well as serum, impair action potential generation and synaptic communication. To overcome this problem, we designed a new neuronal medium (BrainPhys basal + serum-free supplements) in which we adjusted the concentrations of inorganic salts, neuroactive amino acids, and energetic substrates. We then tested that this medium adequately supports neuronal activity and survival of human neurons in culture. Long-term exposure to this physiological medium also improved the proportion of neurons that were synaptically active. The medium was designed to culture human neurons but also proved adequate for rodent neurons. The improvement in BrainPhys basal medium to support neurophysiological activity is an important step toward reducing the gap between brain physiological conditions in vivo and neuronal models in vitro.