Neurons derived from radial glial cells establish radial units in neocortex

The neocortex of the adult brain consists of neurons and glia that are generated by precursor cells of the embryonic ventricular zone. In general, glia are generated after neurons during development, but radial glia are an exception to this rule. Radial glia are generated before neurogenesis and guide neuronal migration. Radial glia are mitotically active throughout neurogenesis, and disappear or become astrocytes when neuronal migration is complete. Although the lineage relationships of cortical neurons and glia have been explored, the clonal relationship of radial glia to other cortical cells remains unknown. It has been suggested that radial glia may be neuronal precursors, but this has not been demonstrated in vivo. We have used a retroviral vector encoding enhanced green fluorescent protein to label precursor cells in vivo and have examined clones 1-3 days later using morphological, immunohistochemical and electrophysiological techniques. Here we show that clones consist of mitotic radial glia and postmitotic neurons, and that neurons migrate along clonally related radial glia. Time-lapse images show that proliferative radial glia generate neurons. Our results support the concept that a lineage relationship between neurons and proliferative radial glia may underlie the radial organization of neocortex.     

Truncated TrkB-T1 mediates neurotrophin-evoked calcium signalling in glia cells

The neurotrophin receptor TrkB is essential for normal function of the mammalian brain. It is expressed in three splice variants. Full-length receptors (TrkB(FL)) possess an intracellular tyrosine kinase domain and are considered as those TrkB receptors that mediate the crucial effects of brain-derived neurotrophic factor (BDNF) or neurotrophin 4/5 (NT-4/5). By contrast, truncated receptors (TrkB-T1 and TrkB-T2) lack tyrosine kinase activity and have not been reported to elicit rapid intracellular signalling. Here we show that astrocytes predominately express TrkB-T1 and respond to brief application of BDNF by releasing calcium from intracellular stores. The calcium transients are insensitive to the tyrosine kinase blocker K-252a and persist in mutant mice lacking TrkB(FL). By contrast, neurons produce rapid BDNF-evoked signals through TrkB(FL) and the Na(v)1.9 channel. Expression of antisense TrkB messenger RNA strongly reduces BDNF-evoked calcium signals in glia. Thus, our results show that, unexpectedly, TrkB-T1 has a direct signalling role in mediating inositol-1,4,5-trisphosphate-dependent calcium release; in addition, they identify a previously unknown mechanism of neurotrophin action in the brain.     

MeCP2转基因食蟹猴所表征的类自闭症行为及种系传递

 自闭症是近年来公众关注度很高的一种神经系统疾病，甲基化CpG结合蛋白2（MeCP2）因其能够在转录水平调节基因表达和操控微小RNA（miRNA）的效应而在自闭症中扮演着重要的角色。当MeCP2因突变而功能缺失时会导致瑞特综合症（Rett syndrome），而当MeCP2拷贝数过多则会导致一种名为MeCP2重复综合症的自闭症。虽然目前科学家已经构建成功了MeCP2的转基因小鼠，但在这种小鼠模型中无法很好地观察到类似人类自闭症的表型。本研究组通过慢病毒侵染的方法构建了能在神经系统中特异表达人源MeCP2的转基因食蟹猴模型，并通过深度测序检测出了转基因插入位点以及通过免疫印迹（westernblot）确证了外源基因的表达。该转基因食蟹猴模型在行动、社交及情绪方面表现出明显的类似自闭症行为，并呈现转基因的种系传递现象。这些结果表明通过基因编辑技术构建非人灵长类模型在脑疾病研究中的重要性。     

Ciliary neurotrophic factor prevents degeneration of motor neurons in mouse mutant progressive motor neuronopathy.

Ciliary neurotrophic factor (CNTF) supports the survival of embryonic motor neurons in vitro and in vivo, and prevents lesion-mediated degeneration of rat motor neurons during early post-natal stages. Here we report that CNTF greatly reduces all the functional and morphological changes in pmn/pmn mice, an autosomal recessive mutant leading to progressive caudo-cranial motor neuron degeneration. The first manifestations of progressive motor neuronopathy in homozygous pmn/pmn mice become apparent in the hind limbs at the end of the third post-natal week, and all the mice die up to 6 or 7 weeks after birth from respiratory paralysis. Treatment with CNTF prolongs survival and greatly improves motor function of these mice. Moreover, morphological manifestations, such as loss of motor axons in the phrenic nerve and degeneration of facial motor neurons, were greatly reduced by CNTF, although the treatment did not start until the first symptoms of the disease had already become apparent and substantial degenerative changes were already present. The protective and restorative effects of CNTF in this mouse mutant give new perspectives for the treatment of human degenerative motor neuron diseases with CNTF.     

Glucose sensing by POMC neurons regulates glucose homeostasis and is impaired in obesity

A subset of neurons in the brain, known as 'glucose-excited' neurons, depolarize and increase their firing rate in response to increases in extracellular glucose. Similar to insulin secretion by pancreatic beta-cells, glucose excitation of neurons is driven by ATP-mediated closure of ATP-sensitive potassium (K(ATP)) channels. Although beta-cell-like glucose sensing in neurons is well established, its physiological relevance and contribution to disease states such as type 2 diabetes remain unknown. To address these issues, we disrupted glucose sensing in glucose-excited pro-opiomelanocortin (POMC) neurons via transgenic expression of a mutant Kir6.2 subunit (encoded by the Kcnj11 gene) that prevents ATP-mediated closure of K(ATP) channels. Here we show that this genetic manipulation impaired the whole-body response to a systemic glucose load, demonstrating a role for glucose sensing by POMC neurons in the overall physiological control of blood glucose. We also found that glucose sensing by POMC neurons became defective in obese mice on a high-fat diet, suggesting that loss of glucose sensing by neurons has a role in the development of type 2 diabetes. The mechanism for obesity-induced loss of glucose sensing in POMC neurons involves uncoupling protein 2 (UCP2), a mitochondrial protein that impairs glucose-stimulated ATP production. UCP2 negatively regulates glucose sensing in POMC neurons. We found that genetic deletion of Ucp2 prevents obesity-induced loss of glucose sensing, and that acute pharmacological inhibition of UCP2 reverses loss of glucose sensing. We conclude that obesity-induced, UCP2-mediated loss of glucose sensing in glucose-excited neurons might have a pathogenic role in the development of type 2 diabetes.     

A new role for cryptochrome in a Drosophila circadian oscillator

Cryptochromes are flavin/pterin-containing proteins that are involved in circadian clock function in Drosophila and mice. In mice, the cryptochromes Cry1 and Cry2 are integral components of the circadian oscillator within the brain and contribute to circadian photoreception in the retina. In Drosophila, cryptochrome (CRY) acts as a photoreceptor that mediates light input to circadian oscillators in both brain and peripheral tissue. A Drosophila cry mutant, cryb, leaves circadian oscillator function intact in central circadian pacemaker neurons but renders peripheral circadian oscillators largely arrhythmic. Although this arrhythmicity could be caused by a loss of light entrainment, it is also consistent with a role for CRY in the oscillator. A peripheral oscillator drives circadian olfactory responses in Drosophila antennae. Here we show that CRY contributes to oscillator function and physiological output rhythms in the antenna during and after entrainment to light-dark cycles and after photic input is eliminated by entraining flies to temperature cycles. These results demonstrate a photoreceptor-independent role for CRY in the periphery and imply fundamental differences between central and peripheral oscillator mechanisms in Drosophila.     

Selective inhibition of neurite outgrowth on mature astrocytes by Thy-1 glycoprotein.

THY-1, the smallest member of the immunoglobulin superfamily, is a major cell-surface component expressed by several tissues. The protein, carbohydrate and gene structures of this molecule are known, yet its function is not. It is highly expressed in nervous tissue, where it appears on virtually all neurons after the cessation of axonal growth. Here we show that expression of Thy-1 by a neural cell line inhibits neurite outgrowth on mature astrocytes, but not on other cellular substrata which include Schwann cells and embryonic glia. This inhibition of neurite extension on astrocytes can be reversed by low concentrations (nanomolar) of soluble Thy-1. If a similar interaction between neuronal Thy-1 and astrocytes occurs in vivo, it could stabilize neuronal connections and suppress axonal regrowth after injury in the astrocyte-rich areas of adult central nervous system.     

Warm-coding deficits and aberrant inflammatory pain in mice lacking P2X3 receptors

ATP activates damage-sensing neurons (nociceptors) and can evoke a sensation of pain. The ATP receptor P2X3 is selectively expressed by nociceptors and is one of seven ATP-gated, cation-selective ion channels. Here we demonstrate that ablation of the P2X3 gene results in the loss of rapidly desensitizing ATP-gated cation currents in dorsal root ganglion neurons, and that the responses of nodose ganglion neurons to ATP show altered kinetics and pharmacology resulting from the loss of expression of P2X(2/3) heteromultimers. Null mutants have normal sensorimotor function. Behavioural responses to noxious mechanical and thermal stimuli are also normal, although formalin-induced pain behaviour is reduced. In contrast, deletion of the P2X3 receptor causes enhanced thermal hyperalgesia in chronic inflammation. Notably, although dorsal-horn neuronal responses to mechanical and noxious heat application are normal, P2X3-null mice are unable to code the intensity of non-noxious 'warming' stimuli.     

Local generation of glia is a major astrocyte source in postnatal cortex

Glial cells constitute nearly 50% of the cells in the human brain. Astrocytes, which make up the largest glial population, are crucial to the regulation of synaptic connectivity during postnatal development. Because defects in astrocyte generation are associated with severe neurological disorders such as brain tumours, it is important to understand how astrocytes are produced. Astrocytes reportedly arise from two sources: radial glia in the ventricular zone and progenitors in the subventricular zone, with the contribution from each region shifting with time. During the first three weeks of postnatal development, the glial cell population, which contains predominantly astrocytes, expands 6-8-fold in the rodent brain. Little is known about the mechanisms underlying this expansion. Here we show that a major source of glia in the postnatal cortex in mice is the local proliferation of differentiated astrocytes. Unlike glial progenitors in the subventricular zone, differentiated astrocytes undergo symmetric division, and their progeny integrate functionally into the existing glial network as mature astrocytes that form endfeet with blood vessels, couple electrically to neighbouring astrocytes, and take up glutamate after neuronal activity.     

Suppression of basal autophagy in neural cells causes neurodegenerative disease in mice

Autophagy is an intracellular bulk degradation process through which a portion of the cytoplasm is delivered to lysosomes to be degraded. Although the primary role of autophagy in many organisms is in adaptation to starvation, autophagy is also thought to be important for normal turnover of cytoplasmic contents, particularly in quiescent cells such as neurons. Autophagy may have a protective role against the development of a number of neurodegenerative diseases. Here we report that loss of autophagy causes neurodegeneration even in the absence of any disease-associated mutant proteins. Mice deficient for Atg5 (autophagy-related 5) specifically in neural cells develop progressive deficits in motor function that are accompanied by the accumulation of cytoplasmic inclusion bodies in neurons. In Atg5-/- cells, diffuse, abnormal intracellular proteins accumulate, and then form aggregates and inclusions. These results suggest that the continuous clearance of diffuse cytosolic proteins through basal autophagy is important for preventing the accumulation of abnormal proteins, which can disrupt neural function and ultimately lead to neurodegeneration.     

Synaptic scaling mediated by glial TNF-alpha

Two general forms of synaptic plasticity that operate on different timescales are thought to contribute to the activity-dependent refinement of neural circuitry during development: (1) long-term potentiation (LTP) and long-term depression (LTD), which involve rapid adjustments in the strengths of individual synapses in response to specific patterns of correlated synaptic activity, and (2) homeostatic synaptic scaling, which entails uniform adjustments in the strength of all synapses on a cell in response to prolonged changes in the cell's electrical activity. Without homeostatic synaptic scaling, neural networks can become unstable and perform suboptimally. Although much is known about the mechanisms underlying LTP and LTD, little is known about the mechanisms responsible for synaptic scaling except that such scaling is due, at least in part, to alterations in receptor content at synapses. Here we show that synaptic scaling in response to prolonged blockade of activity is mediated by the pro-inflammatory cytokine tumour-necrosis factor-alpha (TNF-alpha). Using mixtures of wild-type and TNF-alpha-deficient neurons and glia, we also show that glia are the source of the TNF-alpha that is required for this form of synaptic scaling. We suggest that by modulating TNF-alpha levels, glia actively participate in the homeostatic activity-dependent regulation of synaptic connectivity.     

Ia-restricted encephalitogenic T lymphocytes mediating EAE lyse autoantigen-presenting astrocytes

T lymphocytes specific for myelin basic protein (MBP) are responsible for the cellular events leading to autoimmune disease within the central (CNS) and peripheral (PNS) nervous systems. Both in actively induced and T-cell transfer versions of experimental autoimmune encephalomyelitis (EAE) and neuritis (EAN), the autoaggressive T cells are activated outside the nervous system and reach their target tissue via the blood circulation. The target specificity of the autoaggressive T cells is impressive; T-cell lines specific for MBP predominantly home to and affect the white matter of the CNS whereas T cells specific for PNS myelin protein P2 exclusively infiltrate peripheral nerves. Having penetrated the tight blood tissue barriers, the lymphocytes seem to interact with local cells expressing the relevant autoantigen in an immunogenic form. Although the exact mechanism of target finding and destruction is unknown, studies from our laboratory have shown that astrocytes, a main component of the normal CNS glia, can actively present antigen to specific T cells. This observation suggests that astrocytes are involved in natural immune reactivity within the CNS, and that they may be involved in pathological aberrations, such as in the development of autoimmune lesions. Having studied astrocyte/T-cell interactions in more detail, we discovered that encephalitogenic T-cell lines recognizing MBP on astrocytes will subsequently proceed to kill the presenting cells. Here we report that astrocyte killing follows the rules governing 'classical' T-cell-mediated cytolysis; it is antigen-specific, restricted by antigens of the major histocompatibility complex (MHC) and apparently contact-dependent. Our data suggest that the nature of the recognized antigenic epitope determines whether or not antigen recognition is followed by killing; moreover, killing of antigen-presenting astrocytes seems to be correlated with the capacity to transfer encephalomyelitis to normal syngeneic rats.     

A specific amyloid-beta protein assembly in the brain impairs memory

Memory function often declines with age, and is believed to deteriorate initially because of changes in synaptic function rather than loss of neurons. Some individuals then go on to develop Alzheimer's disease with neurodegeneration. Here we use Tg2576 mice, which express a human amyloid-beta precursor protein (APP) variant linked to Alzheimer's disease, to investigate the cause of memory decline in the absence of neurodegeneration or amyloid-beta protein amyloidosis. Young Tg2576 mice (< 6 months old) have normal memory and lack neuropathology, middle-aged mice (6-14 months old) develop memory deficits without neuronal loss, and old mice (> 14 months old) form abundant neuritic plaques containing amyloid-beta (refs 3-6). We found that memory deficits in middle-aged Tg2576 mice are caused by the extracellular accumulation of a 56-kDa soluble amyloid-beta assembly, which we term Abeta*56 (Abeta star 56). Abeta*56 purified from the brains of impaired Tg2576 mice disrupts memory when administered to young rats. We propose that Abeta*56 impairs memory independently of plaques or neuronal loss, and may contribute to cognitive deficits associated with Alzheimer's disease.     

A stomatin-domain protein essential for touch sensation in the mouse

Touch and mechanical pain are first detected at our largest sensory surface, the skin. The cell bodies of sensory neurons that detect such stimuli are located in the dorsal root ganglia, and subtypes of these neurons are specialized to detect specific modalities of mechanical stimuli. Molecules have been identified that are necessary for mechanosensation in invertebrates but so far not in mammals. In Caenorhabditis elegans, mec-2 is one of several genes identified in a screen for touch insensitivity and encodes an integral membrane protein with a stomatin homology domain. Here we show that about 35% of skin mechanoreceptors do not respond to mechanical stimuli in mice with a mutation in stomatin-like protein 3 (SLP3, also called Stoml3), a mammalian mec-2 homologue that is expressed in sensory neurons. In addition, mechanosensitive ion channels found in many sensory neurons do not function without SLP3. Tactile-driven behaviours are also impaired in SLP3 mutant mice, including touch-evoked pain caused by neuropathic injury. SLP3 is therefore indispensable for the function of a subset of cutaneous mechanoreceptors, and our data support the idea that this protein is an essential subunit of a mammalian mechanotransducer.     

Presenilin is required for proper morphology and function of neurons in C. elegans

Mutations in the human presenilin genes cause the most frequent and aggressive forms of familial Alzheimer's disease (FAD). Here we show that in addition to its role in cell fate decisions in non-neuronal tissues, presenilin activity is required in terminally differentiated neurons in vivo. Mutations in the Caenorhabditis elegans presenilin genes sel-12 and hop-1 result in a defect in the temperature memory of the animals. This defect is caused by the loss of presenilin function in two cholinergic interneurons that display neurite morphology defects in presenilin mutants. The morphology and function of the affected neurons in sel-12 mutant animals can be restored by expressing sel-12 only in these cells. The wild-type human presenilin PS1, but not the FAD mutant PS1 A246E, can also rescue these morphological defects. As lin-12 mutant animals display similar morphological and functional defects to presenilin mutants, we suggest that presenilins mediate their activity in postmitotic neurons by facilitating Notch signalling. These data indicate cell-autonomous and evolutionarily conserved control of neural morphology and function by presenilins.     

Oxidant stress evoked by pacemaking in dopaminergic neurons is attenuated by DJ-1

Parkinson's disease is a pervasive, ageing-related neurodegenerative disease the cardinal motor symptoms of which reflect the loss of a small group of neurons, the dopaminergic neurons in the substantia nigra pars compacta (SNc). Mitochondrial oxidant stress is widely viewed as being responsible for this loss, but why these particular neurons should be stressed is a mystery. Here we show, using transgenic mice that expressed a redox-sensitive variant of green fluorescent protein targeted to the mitochondrial matrix, that the engagement of plasma membrane L-type calcium channels during normal autonomous pacemaking created an oxidant stress that was specific to vulnerable SNc dopaminergic neurons. The oxidant stress engaged defences that induced transient, mild mitochondrial depolarization or uncoupling. The mild uncoupling was not affected by deletion of cyclophilin D, which is a component of the permeability transition pore, but was attenuated by genipin and purine nucleotides, which are antagonists of cloned uncoupling proteins. Knocking out DJ-1 (also known as PARK7 in humans and Park7 in mice), which is a gene associated with an early-onset form of Parkinson's disease, downregulated the expression of two uncoupling proteins (UCP4 (SLC25A27) and UCP5 (SLC25A14)), compromised calcium-induced uncoupling and increased oxidation of matrix proteins specifically in SNc dopaminergic neurons. Because drugs approved for human use can antagonize calcium entry through L-type channels, these results point to a novel neuroprotective strategy for both idiopathic and familial forms of Parkinson's disease.     

Glial cells express N-CAM/D2-CAM-like polypeptides in vitro

The joining together of neurites to form fascicles and the growth of axons along glial surfaces during early development suggest that neurone-neurone and neurone-glial adhesion interactions are of considerable importance for defining nerve tracts. In vitro studies have indicated that adhesion between neurones involves a glycoprotein that has been independently studied under the names of N-CAM (for neural cell adhesion molecule), D2-CAM and BSP-2 (refs 10, 11). As N-CAM/D2-CAM appears to be a homophilic ligand that binds to N-CAM/D2-CAM polypeptide on adjacent cells, this glycoprotein is potentially important in adhesion interactions between any two N-CAM/D2-CAM-expressing cells. While it has been suggested that neurone-glial adhesion involves molecules other than N-CAM/D2-CAM, it is known that N-CAM/D2-CAM antigenic determinants are expressed by glial cells in vivo and that injection of anti-N-CAM antibodies into the eye-cup of chick embryos disrupts normal patterns of neuritic apposition to glial endfeet in the developing optic stalk. Do the molecules expressed by glia share restricted antigenic determinants, or binding domains, with N-CAM/D2-CAM, or are N-CAM/D2-CAM polypeptides expressed by glia? Here we present immunocytochemical evidence which suggests that all classes of macroglia express N-CAM/D2-CAM antigenic determinants on their surfaces and immunochemical analyses which indicate that the molecules expressed by purified astrocytes are closely similar, or identical, to at least some forms of N-CAM/D2-CAM obtained from whole brain or purified neurones. However, our results also suggest that different N-CAM/D2-CAM polypeptides may be separately expressed by neurones and astrocytes.     

Munc13-1 is essential for fusion competence of glutamatergic synaptic vesicles.

Neurotransmitter release at synapses between nerve cells is mediated by calcium-triggered exocytotic fusion of synaptic vesicles. Before fusion, vesicles dock at the presynaptic release site where they mature to a fusion-competent state. Here we identify Munc13-1, a brain-specific presynaptic phorbol ester receptor, as an essential protein for synaptic vesicle maturation. We show that glutamatergic hippocampal neurons from mice lacking Munc13-1 form ultrastructurally normal synapses whose synaptic-vesicle cycle is arrested at the maturation step. Transmitter release from mutant synapses cannot be triggered by action potentials, calcium-ionophores or hypertonic sucrose solution. In contrast, release evoked by alpha-latrotoxin is indistinguishable from wild-type controls, indicating that the toxin can bypass Munc13-1-mediated vesicle maturation. A small subpopulation of synapses of any given glutamatergic neuron as well as all synapses of GABA (gamma-aminobutyric acid)-containing neurons are unaffected by Munc13-1 loss, demonstrating the existence of multiple and transmitter-specific synaptic vesicle maturation processes in synapses.