Oligodendrocytes enable rapid central nervous system signaling by myelinating axons. Here, to model key biomechanical cues regulating myelination, we developed a tunable hydrogel-based micropillar array system that mimics the three-dimensional architecture and softness of axons. This platform supports the long-term culture of oligodendrocytes and robust formation of multilayered compact myelin by rodent and human oligodendrocytes. Using confocal and transmission electron microscopy, we observed a strong linear correlation between immunostained myelin thickness and the number of myelin wraps, enabling high-content quantification of myelination. Systematic variation of pillar stiffness, diameter and surface chemistry within pathophysiological ranges revealed that both mechanical and geometric properties of axon-like substrates critically regulate oligodendrocyte differentiation and myelin wrapping. Importantly, we demonstrate that pharmacological agents exhibit stiffness-dependent effects on myelination, suggesting that overly rigid in vitro models may yield false-positive drug hits. This platform offers a physiologically relevant, high-throughput assay for dissecting oligodendrocyte biology and discovering remyelinating therapies for diseases such as multiple sclerosis.

Chromatin remodeling complexes (CRCs) participate in oligodendrocyte (OL) differentiation, survival, and maintenance. We asked whether CRCs also control the proliferation of OL precursors (OPs)-focusing on the INO80 complex, which is known to regulate the proliferation of a variety of other cell types during development and disease. CRISPR/Cas9-mediated inactivation of Ino80 in vitro, or Cre-mediated deletion in vivo, slowed the OP cell cycle substantially by prolonging G1. RNAseq analysis revealed that E2F target genes were dysregulated in OPs from INO80-deficient mice, but correlated RNAseq and ATAC-seq uncovered no general correlation between gene expression and altered nucleosome positioning at transcription start sites. Fluorescence photobleaching experiments in cultured OPs demonstrated that histone H2A.Z mobility increased following the loss of INO80, suggesting that INO80 regulates the cell cycle machinery in OPs through H2A.Z/H2A exchange. We also present evidence that INO80 associates with OLIG2, a master regulator of OL development.

This method paper details a protocol to test spatial working memory in mice using a semi-automated radial 8-arm maze (RAM). The RAM is a partially enclosed apparatus with 8 horizontal, equally spaced arms radiating from a central hub, from which access to each arm can be controlled individually by servo-controlled motorized doors. Animals start in the central hub and are allowed to explore the maze for a food reward at the end of each arm or selected arm. The RAM task was originally designed for rats, but we have adapted the protocol for mice, for example, by including more habituation steps. In our protocol, all arms are initially baited with sweetened condensed milk, and mice are admitted sequentially into four pseudo-randomly selected arms to collect the rewards ("forced run") before opening all doors together to allow the mice to run freely and find the remaining four rewards ("free run"). A 5 s delay is introduced between the forced and free runs to challenge working memory; an error is recorded if the mouse enters any previously visited arm during the free run. The task is complete when all rewards are recovered. After 6 days of habituation and 9 days of maze training, male C57BL/6 mice regularly achieve ≥ 80% daily success rate score, defined as 4/(4+E), where E is the number of errors. This semi-automated task could, in principle, be combined with in vivo monitoring methods such as electrophysiology, multiple-photon microscopy, or calcium imaging.

Oligodendrocytes continue to differentiate from their precursor cells even in adulthood, a process that can be modulated by neuronal activity and experience. Previous work has indicated that conditional ablation of oligodendrogenesis in adult mice leads to learning and memory deficits in a range of behavioral tasks. The current study replicated and re-evaluated evidence for a role of oligodendrogenesis in motor learning, using a complex running wheel task. Further, we found that ablating oligodendrogenesis alters brain microstructure (ex vivo MRI) and brain activity (in vivo EEG) independent of experience with the task. This suggests a role for adult oligodendrocyte formation in the maintenance of brain function and indicates that task-independent changes due to oligodendrogenesis ablation need to be considered when interpreting learning and memory deficits in this model.

In the mouse embryonic forebrain, developmentally distinct oligodendrocyte progenitor cell populations and their progeny, oligodendrocytes, emerge from three distinct regions in a spatiotemporal gradient from ventral to dorsal. However, the functional importance of this oligodendrocyte developmental heterogeneity is unknown. Using a genetic strategy to ablate dorsally derived oligodendrocyte lineage cells (OLCs), we show here that the areas in which dorsally derived OLCs normally reside in the adult central nervous system become populated and myelinated by OLCs of ventral origin. These ectopic oligodendrocytes (eOLs) have a distinctive gene expression profile as well as subtle myelination abnormalities. The failure of eOLs to fully assume the role of the original dorsally derived cells results in locomotor and cognitive deficits in the adult animal. This study reveals the importance of developmental heterogeneity within the oligodendrocyte lineage and its importance for homeostatic brain function. Here the authors show that ventrally derived oligodendrocytes (OLs) can myelinate areas usually populated by dorsally derived OLs but cannot functionally compensate, as animals populated only by ventrally derived OLs show locomotor and cognitive deficits.

Brain glucose sensing is critical for survival during hypoglycaemia and tunes the level of defended blood glucose, which goes up in diabetes. Neuronal glucose sensing neurons and mechanisms have been identified, but how these neurons access blood concentrations of glucose to adjust their output and maintain glucose homeostasis is unclear. Here, we demonstrate that adult oligodendrogenesis in the median eminence (ME) is modulated by changes in circulating glucose levels and rapidly upregulated by hypoglycaemia. We show that genetic blockade of new OL production in adult mice impairs the regulation of glucose homeostasis, the integrity of the ME blood-hypothalamus barrier, and neuronal glucose sensing in the arcuate nucleus of the hypothalamus (ARH). Unexpectedly, functional integrity of adult-formed myelin is not required for the maintenance of glucose homeostasis. Instead, newly formed OLs exert their glucoregulatory actions via the synthesis of A disintegrin and metallopeptidase with thrombospondin motifs 4 (ADAMTS4), a metallopeptidase expressed exclusively by OLs and dependent on adult OL genesis to maintain its expression in the ME. Both lack of Adamts4 and ADAMTS4 gain-of-function are associated with impaired glucose homeostasis and remodelling of the blood-hypothalamus barrier, indicating that optimal ADAMTS4 expression is required for the integrity of vascular permeability and normal glycaemic control. Finally, we show that ME ADAMTS4 expression is regulated by changes in peripheral glycaemia and is dysregulated in diabetes, providing a mechanism by which ME OLs contribute to the regulation of glucose homeostasis. ### Competing Interest Statement The authors have declared no competing interest.

Abstract This methods paper details a protocol to test spatial working memory in mice using a semi-automated radial 8-arm maze (RAM). The RAM is a partially enclosed apparatus with 8 horizontal equally spaced arms radiating from a central hub, from which access to each arm can be controlled individually by servo-controlled motorized doors. Animals start in the central hub and are allowed to explore the maze for a food reward at the end of each arm or selected arms. The RAM task was originally designed for rats but we have adapted the protocol for mice – for example, by including more habituation steps. In our protocol, all arms are initially baited with sweetened condensed milk and mice are admitted sequentially into four pseudo-randomly selected arms to collect the rewards (“forced run”) before opening all doors together to allow the mice to run freely and find the remaining four rewards (“free run”). A 5-sec time delay is introduced between the forced and free runs to challenge working memory; an error is recorded if the mouse enters any previously-visited arm during the free run. The task is complete when all rewards are recovered. After 6 days of habituation and 9 days of maze training, male C57BL/6 mice regularly achieve ≥80% success rate, defined as 4/(4+E) where E is the number of errors. This semi-automated task could in principle be combined with in vivo monitoring methods such as electrophysiology or calcium imaging.

Motor skill learning stimulates and requires generation of myelinating oligodendrocytes (OLs) from their precursors (OLPs). We asked whether OL production is also required for non-motor learning and cognition, using T-maze and radial arm maze tasks that tax spatial working memory. Maze training stimulated OL production in the medial prefrontal cortex (mPFC), anterior corpus callosum (genu), dorsal thalamus and hippocampal formation; myelin sheath formation was also stimulated in the genu. Genetic blockade of OL differentiation and neo-myelination in Myrf conditional-knockout mice strongly impaired training-induced improvements in maze performance. Remarkably, there was a strong positive correlation between working memory performance of individual mice and the scale of OLP proliferation and OL generation during training, but not with the number or intensity of c-Fos+ neurons in the mPFC, underscoring the key role of OL lineage cells in cognitive performance.### Competing Interest StatementThe authors have declared no competing interest.

OBJECTIVE:Oligodendrocyte progenitor cell differentiation is regulated by nutritional signals in the adult median eminence (ME), but the consequences on local myelination are unknown. The aim of this study was to characterize myelin plasticity in the ME of adult mice in health or in response to chronic nutritional challenge and determine its relevance to the regulation of energy balance.METHODS:We assessed new oligodendrocyte (OL) and myelin generation and stability in the ME of healthy adult male mice using bromodeoxyuridine labelling and genetic fate mapping tools. We evaluated the contribution of microglia to ME myelin plasticity in PLX5622-treated C57BL/6J mice and in Pdgfra-Cre/ERT2;R26R-eYFP;Myrffl/fl mice, where adult oligodendrogenesis is blunted. Next, we investigated how high-fat feeding or caloric restriction impact ME OL lineage progression and myelination. Finally, we characterized the functional relevance of adult oligodendrogenesis on energy balance regulation.RESULTS:We show that myelinating OLs are continuously and rapidly generated in the adult ME. Paradoxically, OL number and myelin amounts remain remarkably stable in the adult ME. In fact, the high rate of new OL and myelin generation in the ME is offset by continuous turnover of both. We show that microglia are required for continuous OL and myelin production, and that ME myelin plasticity regulates the recruitment of local immune cells. Finally, we provide evidence that ME myelination is regulated by the body's energetic status and demonstrate that ME OL and myelin plasticity are required for the regulation of energy balance and hypothalamic leptin sensitivity.CONCLUSIONS:This study identifies a new mechanism modulating leptin sensitivity and the central control of energy balance and uncovers a previously unappreciated form of structural plasticity in the ME.

The mediobasal hypothalamus (MBH; arcuate nucleus of the hypothalamus [ARH] and median eminence [ME]) is a key nutrient sensing site for the production of the complex homeostatic feedback responses required for the maintenance of energy balance. Here, we show that refeeding after an overnight fast rapidly triggers proliferation and differentiation of oligodendrocyte progenitors, leading to the production of new oligodendrocytes in the ME specifically. During this nutritional paradigm, ME perineuronal nets (PNNs), emerging regulators of ARH metabolic functions, are rapidly remodeled, and this process requires myelin regulatory factor (Myrf) in oligodendrocyte progenitors. In genetically obese ob/ob mice, nutritional regulations of ME oligodendrocyte differentiation and PNN remodeling are blunted, and enzymatic digestion of local PNN increases food intake and weight gain. We conclude that MBH PNNs are required for the maintenance of energy balance in lean mice and are remodeled in the adult ME by the nutritional control of oligodendrocyte differentiation.

Bronchopulmonary dysplasia (BPD) is a common complication of preterm birth characterized by arrested lung alveolarization, which generates lungs that are incompetent for effective gas exchange. We report here deregulated expression of miR‐34a in a hyperoxia‐based mouse model of BPD, where miR‐34a expression was markedly increased in platelet‐derived growth factor receptor (PDGFR)α‐expressing myofibroblasts, a cell type critical for proper lung alveolarization. Global deletion of miR‐34a; and inducible, conditional deletion of miR‐34a in PDGFRα+ cells afforded partial protection to the developing lung against hyperoxia‐induced perturbations to lung architecture. Pdgfra mRNA was identified as the relevant miR‐34a target, and using a target site blocker in vivo, the miR‐34a/Pdgfra interaction was validated as a causal actor in arrested lung development. An antimiR directed against miR‐34a partially restored PDGFRα+ myofibroblast abundance and improved lung alveolarization in newborn mice in an experimental BPD model. We present here the first identification of a pathology‐relevant microRNA/mRNA target interaction in aberrant lung alveolarization and highlight the translational potential of targeting the miR‐34a/Pdgfra interaction to manage arrested lung development associated with preterm birth.

Purpose To update evidence of diagnostic potential for identification of lumbar spinal stenosis (LSS) based on demographic and patient history, clinical findings, and physical tests, and report posttest probabilities associated with test findings. Methods An electronic search of PubMed, CINAHL and Embase was conducted combining terms related to low back pain, stenosis and diagnostic accuracy. Prospective or retrospective studies investigating diagnostic accuracy of LSS using patient history, clinical findings and/or physical tests were included. The risk of bias and applicability were assessed using the Quality Assessment of Diagnostic Accuracy Studies (QUADAS 2) tool. Diagnostic accuracy including sensitivities (SN), specificities (SP), likelihood ratios (+LR and −LR) and posttest probabilities (+PTP and −PTP) with 95% confidence intervals were summarized. Results Nine studies were included (pooled n = 36,228 participants) investigating 49 different index tests (30 demographic and patient history and 19 clinical findings/physical tests). Of the nine studies included, only two exhibited a low risk of bias and seven exhibited good applicability according to QUADAS 2. The demographic and patient history measures (self-reported history questionnaire, no pain when seated, numbness of perineal region) and the clinical findings/physical tests (two-stage treadmill test, symptoms after a March test and abnormal Romberg test) highly improved positive posttest probability by > 25% to diagnose LSS. Conclusion Outside of one study that was able to completely rule out LSS with no functional neurological changes none of the stand-alone findings were strong enough to rule in or rule out LSS. Graphic abstract These slides can be retrieved under Electronic Supplementary Material.