Tenascin-C (Tnc) is an astrocytic multifunctional extracellular matrix (ECM) glycoprotein that potentially promotes or inhibits neurite outgrowth. To investigate its possible functions for retinal development, explants from embryonic day 18 (E18) rat retinas were cultivated on culture substrates composed of poly-d-lysine (PDL), or PDL additionally coated with Tnc or laminin (LN)-1, which significantly increased fiber length. When combined with LN, Tnc induced axon fasciculation that reduced the apparent number of outgrowing fibers. In order to circumscribe the stimulatory region, Tnc-derived fibronectin type III (TNfn) domains fused to the human Ig-Fc-fragment TNfnD6-Fc, TNfnBD-Fc, TNFnA1A2-Fc and TNfnA1D-Fc were studied. The fusion proteins TNfnBD-Fc and to a lesser degree TNfnA1D-Fc were stimulatory when compared with the Ig-Fc-fragment protein without insert. In contrast, the combination TNfnA1A2-Fc reduced fiber outgrowth beneath the values obtained for the Ig-Fc domain, indicating potential inhibitory properties. The monoclonal J1/tn2 antibody (clone 578) that is specific for domain TNfnD blocked the stimulatory properties of the TNfn-Fc fusions. When postnatal day 7 retinal ganglion cells were used rather that explants, Tnc and Tnc-derived proteins proved permissive for neurite outgrowth. The present study highlights a strong retinal axon growth-promoting activity of the Tnc domain TNfnD, which is modulated by neighboring domains.

To analyze myelin structure and the composition of myelinated tissue in the African lungfish (Protopterus dolloi), we used a combination of ultrastructural and biochemical techniques. Electron microscopy showed typical multilamellar myelin: CNS sheaths abutted one another, and PNS sheaths were separated by endoneurial collagen. The radial component, prominent in CNS myelin of higher vertebrates, was suggested by the pattern of staining but was poorly organized. The lipid and myelin protein compositions of lungfish tissues more closely resembled those of teleost than those of higher vertebrates (frog, mouse). Of particular note, for example, lungfish glycolipids lacked hydroxy fatty acids. Native myelin periodicities from unfixed nerves were in the range of those for higher vertebrates rather than for teleost fish. Lungfish PNS myelin had wider inter-membrane spaces compared with other vertebrates, and lungfish CNS myelin had spaces that were closer in value to those in mammalian than to amphibian or teleost myelins. The membrane lipid bilayer was narrower in lungfish PNS myelin compared to other vertebrates, whereas in the CNS myelin the bilayer was in the typical range. Lungfish PNS myelin showed typical compaction and swelling responses to incubation in acidic or alkaline hypotonic saline. The CNS myelin, by contrast, did not compact in acidic saline but did swell in the alkaline solution. This lability was more similar to that for the higher vertebrates than for teleost.

Astrocytes were classically considered as simple supportive cells for neurons without a significant role in information processing by the nervous system. However, considerable amounts of evidence obtained by several groups during the past years demonstrated the existence of a bidirectional communication between astrocytes and neurons, which prompted a re-examination of the role of astrocytes in the physiology of the nervous system. While neurons base their excitability on electrical signals generated across the membrane, astrocytes base their cellular excitability on variations of the Ca2+ concentration in the cytosol. This article discusses our current knowledge of the properties of the synaptically evoked astrocyte Ca2+ signal, which reveals that astrocytes display integrative properties for synaptic information processing. Astrocytes respond selectively to different axon pathways, discriminate between the activity of different synapses and their Ca2+ signal is non-linearly modulated by the simultaneous activity of different synaptic inputs. Furthermore, this Ca2+ signal modulation depends on astrocyte cellular intrinsic properties and is bidirectionally regulated by the level of synaptic activity. Finally, astrocyte Ca2+ elevations can trigger the release of gliotransmitters, which modulate neuronal activity as well as synaptic transmission and plasticity, hence granting the bidirectional communication with neurons. Consequently, astrocytes can be considered as cellular elements involved in information processing by the nervous system.

Investigations examining the role of polysialic acid (PSA) on the neural cell adhesion molecule (NCAM) in synaptic plasticity have yielded inconsistent data. Here, we addressed this issue by determining whether homosynaptic long-term potentiation (LTP) and heterosynaptic long-term depression (LTD) induce changes in the distribution of PSA-NCAM in the dentate gyrus (DG) of rats in vivo. In addition, we also examined whether the observed modifications were initiated via the activation of N-methyl-d-aspartate (NMDA) receptors. Immunocytochemical analysis showed an increase in PSA-NCAM positive cells both at 2 and 24 h following high-frequency stimulation of either medial or lateral perforant paths, leading to homosynaptic LTP and heterosynaptic LTD, respectively, in the medial molecular layer of the DG. Analysis of sub-cellular distribution of PSA-NCAM by electron microscopy showed decreased PSA dendritic labelling in LTD rats and a sub-cellular relocation towards the spines in LTP rats. Importantly, these modifications were found to be independent of the activation of NMDA receptors. Our findings suggest that strong activation of the granule cells up-regulates PSA-NCAM synthesis which then incorporates into activated synapses, representing NMDA-independent plastic processes that act synergistically on LTP/LTD mechanisms without participating in their expression.

The aim of this study was to investigate the developmental expression of major histocompatibility complex class II (MHCII) by microglia and macrophages and their relationship to blood vessels in the retina, a representative tissue of the central nervous system. Such information is crucial to understanding the role of these cells in immune surveillance. Wholemount preparations of retinas from late embryonic, postnatal and adult rabbits were subjected to three-colour fluorescence microscopy using β2 integrin (CD18) and MHCII antibodies and biotinylated Griffonia simplicifolia B4 isolectin labelling of blood vessels. CD18+ cells consistently exhibited characteristics of macrophages or microglia in the vascularized and non-vascularized regions of the retina, respectively. At all ages, MHCII was expressed by a high proportion of cells in the vascularized region, which contained macrophage-like ‘parenchymal cells’ as well as typical perivascular macrophages. MHCII expression by ramified microglia, first detected on postnatal day 30, was lower in the peripheral retina and intermediate in the avascular region of the myelinated streak. The observed localization of MHCII+ cells in relation to blood vessels and location-dependent differences in MHCII expression point to the possibility that these cells may be distributed strategically within the retina to provide multiple lines of defence against immune challenge arriving via the retinal vasculature.

During development, multipotent neural precursors give rise to oligodendrocyte progenitor cells (OPCs), which migrate and divide to produce additional OPCs. Near the end of embryogenesis and during postnatal stages, many OPCs stop dividing and differentiate as myelinating oligodendrocytes, whereas others persist as nonmyelinating cells. Investigations of oligodendrocyte development in mice indicated that the Nkx2.2 transcription factor both limits the number of OPCs that are formed and subsequently promotes their differentiation, raising the possibility that Nkx2.2 plays a key role in determining myelinating versus nonmyelinating fate. We used in vivo time-lapse imaging and loss-of-function experiments in zebrafish to further explore formation and differentiation of oligodendrocyte lineage cells. Our data show that newly specified OPCs are heterogeneous with respect to gene expression and fate. Whereas some OPCs express the nkx2.2a gene and differentiate as oligodendrocytes, others that do not express nkx2.2a mostly remain as nonmyelinating OPCs. Similarly to mouse, loss of nkx2.2a function results in excess OPCs and delayed oligodendrocyte differentiation. Notably, excess OPCs are formed as a consequence of prolonged OPC production from neural precursor cells. We conclude that Nkx2.2 promotes timely specification and differentiation of myelinating oligodendrocyte lineage cells from species representing different vertebrate taxa.

Neuronal networks can adapt to global changes in activity levels through compensatory modifications in pre- and postsynaptic parameters of synaptic transmission. These forms of synaptic plasticity are known as synaptic homeostasis, and are thought to require specific cellular interactions and signaling across the entire neuronal network. However, the molecular mechanisms underlying synaptic homeostasis have so far been investigated mostly in primary cultures of dissociated neurons, a preparation that lacks the specificity of in vivo circuitry. Here, we show that there are critical differences in the properties of synaptic homeostasis between dissociated neuronal cultures and organotypic slices, a preparation that preserves more precisely in vivo connectivity. Moreover, the cell adhesion molecule β3 integrin, which regulates excitatory synaptic strength, is specifically required for a postsynaptic form of synaptic homeostasis called synaptic scaling in both dissociated cultures and organotypic slices. Conversely, another form of synaptic homeostasis that involves changes in presynaptic quantal content occurs independently of β3 integrin. Our findings define the differential involvement of β3 integrin in two forms of synaptic homeostasis.

Mounting evidence demonstrates that glial cells might have important roles in regulating the physiology and behavior of adult animals. We summarize some of this evidence here, with an emphasis on the roles of glia of the differentiated nervous system in controlling neuronal excitability, behavior and plasticity. In the review we highlight studies in Drosophila and discuss results from the analysis of mammalian astrocytes that demonstrate roles for glia in the adult nervous system.

Trigeminal nerve activation in response to inflammatory stimuli has been shown to increase neuron–glia communication via gap junctions in trigeminal ganglion. The goal of this study was to identify changes in the expression of gap junction proteins, connexins (Cxs), in trigeminal ganglia in response to acute or chronic joint inflammation. Although mRNA for Cxs 26, 36, 40 and 43 was detected under basal conditions, protein expression of only Cxs 26, 36 and 40 increased following capsaicin or complete Freund's adjuvant (CFA) injection into the temporomandibular joint (TMJ). While Cx26 plaque formation between neurons and satellite glia was transiently increased following capsaicin injections, Cx26 plaque formation between neurons and satellite glia was sustained in response to CFA. Interestingly, levels of Cx36 and Cx40 were only elevated in neurons following capsaicin or CFA injections, but the temporal response was similar to that observed for Cx26. In contrast, Cx43 expression was not increased in neurons or satellite glial cells in response to CFA or capsaicin. Thus, trigeminal ganglion neurons and satellite glia can differentially regulate Cx expression in response to the type and duration of inflammatory stimuli, which likely facilitates increased neuron–glia communication during acute and chronic inflammation and pain in the TMJ.

The insulative properties of myelin sheaths in the central and peripheral nervous systems (CNS and PNS) are widely thought to derive from the high resistance and low capacitance of the constituent membranes. Although this view adequately accounts for myelin function in large diameter fibers, it poorly reflects the behavior of small fibers that are prominent in many regions of the CNS. Herein, we develop a computational model to more accurately represent conduction in small fibers. By incorporating structural features that, hitherto, have not been simulated, we demonstrate that myelin tight junctions (TJs) improve saltatory conduction by reducing current flow through the myelin, limiting axonal membrane depolarization and restraining the activation of ion channels beneath the myelin sheath. Accordingly, our simulations provide a novel view of myelin by which TJs minimize charging of the membrane capacitance and lower the membrane time constant to improve the speed and accuracy of transmission in small diameter fibers. This study establishes possible mechanisms whereby TJs affect conduction in the absence of overt perturbations to myelin architecture and may in part explain the tremor and gait abnormalities observed in Claudin 11-null mice.

NG2 cells represent a unique glial cell population that is distributed widely throughout the developing and adult CNS and is distinct from astrocytes, mature oligodendrocytes and microglia. The ability of NG2 cells to differentiate into myelinating oligodendrocytes has been documented in vivo and in vitro. We reported recently that NG2 cells in the forebrain differentiate into myelinating oligodendrocytes but into a subpopulation of protoplasmic astrocytes (Zhu et al., 2008). However, the in vivo fate of NG2 cells in the spinal cord and cerebellum has remained unknown. To investigate the fate of NG2 cells in caudal central nervous system (CNS) regions in vivo, we examined the phenotype of cells that express EGFP in mice that are double transgenic for NG2CreBAC and the Cre reporter Z/EG. The fate of NG2 cells can be studied in these mice by permanent expression of EGFP in cells that have undergone Cre-mediated recombination in NG2 cells. We find that NG2 cells give rise to oligodendrocytes in both gray and white matter of the spinal cord and cerebellum, and to protoplasmic astrocytes in the gray matter of the spinal cord. However, NG2 cells do not give rise to astrocytes in the white matter of the spinal cord and cerebellum. These observations indicate that NG2 cells serve as precursor cells for oligodendrocytes and a subpopulation of protoplasmic astrocytes throughout the rostrocaudal axis of the CNS.

In the postnatal forebrain, the extracellular matrix protein reelin is expressed and secreted by subsets of GABAergic neurons, whereas in the cerebellum reelin is detected in glutamatergic cells of the granule cell layer. Thus, various regions of the postnatal brain present different patterns of reelin expression, whose significance remains unknown. We combined immunocytochemical and pharmacological approaches to characterize the phenotypic and temporal profiles of reelin expression in dissociated cultures of cerebellar granule neurons. A single type of reelin immunoreactivity, identified by a punctate labelling, was present in the somata of the majority of neurons. This immunoreactivity was observed throughout maturation and was exclusively present in glutamatergic neurons expressing the vesicular glutamate transporter 1. Neurons containing the reelin receptors apolipoprotein E receptor 2 (Apoer2) and very low-density lipoprotein receptor (Vldlr) represented about 80% of cerebellar neurons. The vast majority of reelin-positive neurons coexpressed Apoer2, suggesting that reelin immunoreactivity resulted in part from receptor-bound reelin. Inhibition of protein synthesis with cycloheximide completely abolished reelin immunoreactivity. In contrast, blocking protein secretion with brefeldin A did not affect the proportion of punctate neurons but revealed a subpopulation of neurons characterized by a solid reelin staining. These data show for the first time that a homogeneous population of glutamatergic neurons can synthesize and secrete reelin in cerebellar granule cells in vitro.

Nogo-A is possibly the best characterized myelin-derived inhibitor of nerve growth in the adult central nervous system (CNS). It is a member of the ancient reticulon family of mainly endoplasmic reticulum resident proteins with representatives found throughout the eukaryotic domain. Orthologs of the nogo gene were identified in tetrapods and teleost fish but none have been detected in invertebrates. Evolution of the nogo gene has been non-homogeneous. The exon–intron arrangement is conserved from amphibians (Xenopus) to mammals, but partly deviates from that found in several teleost fish species, indicating that the recruitment of nogo exons proceeded along at least two independent lines during early vertebrate evolution. This might have far-reaching consequences. Tetrapod nogo orthologs encode two neurite growth inhibitory domains whereas in fish nogo only one of the inhibitory domains is present. These distinct paths in nogo evolution have potentially contributed to the regeneration permissive CNS in fish as opposed to the non-regenerating CNS in higher vertebrates.

The astrocytic enzyme adenosine kinase (ADK) is a key negative regulator of the brain’s endogenous anticonvulsant adenosine. Astrogliosis with concomitant upregulation of ADK is part of the epileptogenic cascade and contributes to seizure generation. To molecularly dissect the respective roles of astrogliosis and ADK-expression for seizure generation, we used a transgenic approach to uncouple ADK-expression from astrogliosis: in Adk-tg mice the endogenous Adk-gene was deleted and replaced by a ubiquitously expressed Adk-transgene with novel ectopic expression in pyramidal neurons, resulting in spontaneous seizures. Here, we followed a unique approach to selectively injure the CA3 of these Adk-tg mice. Using this strategy, we had the opportunity to study astrogliosis and epileptogenesis in the absence of the endogenous astrocytic Adk-gene. After triggering epileptogenesis we demonstrate astrogliosis without upregulation of ADK, but lack of seizures, whereas matching wild-type animals developed astrogliosis with upregulation of ADK and spontaneous recurrent seizures. By uncoupling ADK-expression from astrogliosis, we demonstrate that global expression levels of ADK rather than astrogliosis per se contribute to seizure generation.

Glia are an indispensable structural and functional component of the synapse. They modulate synaptic transmission and also play important roles in synapse formation and maintenance. The vertebrate neuromuscular junction (NMJ) is a classic model synapse. Due to its large size, simplicity and accessibility, the NMJ has contributed greatly to our understanding of synapse development and organization. In the past decade, the NMJ has also emerged as an effective model for studying glia–synapse interactions, in part due to the development of various labeling techniques that permit NMJs and associated Schwann cells (the glia at NMJs) to be visualized in vitro and in vivo. These approaches have demonstrated that Schwann cells are actively involved in synapse remodeling both during early development and in post-injury reinnervation. In vivo imaging has also recently been combined with serial section transmission electron microscopic (ssTEM) reconstruction to directly examine the ultrastructural organization of remodeling NMJs. In this review, we focus on the anatomical studies of Schwann cell dynamics and their roles in formation, maturation and remodeling of vertebrate NMJs using the highest temporal and spatial resolution methods currently available.

Adhesive and repellent molecular cues guide migrating cells and growing neurites during development. They also contribute to synaptic function, learning and memory in adulthood. Here, we review the roles of cell adhesion molecules of the immunoglobulin superfamily (Ig-CAMs) and semaphorins (some of which also contain Ig-like domains) in regulation of synaptic transmission and plasticity. Interestingly, among the seven studied Ig-CAMs, the neuronal cell adhesion molecule proved to be important for all tested forms of hippocampal plasticity, while its associated unusual glycan polysialic acid is necessary and sufficient part for synaptic plasticity only at CA3-CA1 synapses. In contrast, Thy-1 and L1 specifically regulate long-term potentiation (LTP) at synapses formed by entorhinal axons in the dentate gyrus and cornu ammonis, respectively. Contactin-1 is important for long-term depression but not for LTP at CA3-CA1 synapses. Analysis of CHL1-deficient mice illustrates that at intermediate stages of development a deficit in a cell adhesion molecule is compensated but appears as impaired LTP during early and late postnatal development. The emerging mechanisms by which adhesive Ig-CAMs contribute to synaptic plasticity involve regulation of activities of NMDA receptors and L-type Ca2+ channels, signaling via mitogen-activated protein kinase p38, changes in GABAergic inhibition and motility of synaptic elements. Regarding repellent molecules, available data for semaphorins demonstrate their activity-dependent regulation in normal and pathological conditions, synaptic localization of their receptors and their potential to elevate or inhibit synaptic transmission either directly or indirectly.

Cell contact-dependent signaling is a major regulatory mechanism in the organization of developing tissues and in the reorganization (post-injury responses) of specialized tissues in multicellular organisms. In this review we contribute to the further understanding of post-injury recovery processes in adult nervous tissue. We emphasize evidence that supports the involvement of cell contact-inhibition signaling in the cell proliferation, growth and differentiation that occurs during healing and neural reorganization after brain damage.