Astrocytic Domains Research Articles

Proximity of excitatory synapses and astroglial gap junctions in layer IV of the mouse barrel cortex

Neurons and astrocytes, the two major cell populations in the adult brain, are characterized by their own mode of intercellular communication – the synapses and the gap junctions (GJ), respectively. In addition, there is increasing evidence for dynamic and metabolic neuroglial interactions resulting in the modulation of synaptic transmission at the so-called “tripartite synapse”. Based on this, we have investigated at the ultrastructural level how excitatory synapses (ES) and astroglial GJ are spatially distributed in layer IV of the barrel cortex of the adult mouse. We used specific antibodies for connexin (Cx) 30 and 43 to identify astroglial GJ, these two proteins are known to be present in the majority of astroglial GJ in the cerebral cortex. In electron-microscopic images, we measured the distance between two ES, between two GJ and between a GJ and its nearest ES. We found a ratio of two GJ per three ES in the hollow and septal areas. Taking into account the size of an astrocyte domain, the high density of GJ suggests the occurrence of reflexive type, i.e. GJ between processes of the same astrocyte. Interestingly, the distance between an ES and an astroglial GJ was found to be significantly lower than that between either two synapses or between two GJ. These observations indicate that the two modes of cell-to-cell communication are not randomly distributed in layer IV of the barrel cortex. Consequently, this feature may provide the morphological support for the recently reported functional interactions between neuronal circuits and astroglial networks.

Pathophysiology of Schizophrenia Based on Impaired Glial-Neuronal Interactions

The model of impaired glial-neuronal interactions in schizophrenia is based on the core hypothesis that non-functional astrocyte receptors may cause an unconstrained synaptic information flux such that glia lose their modulatory function in tripartite synapses. This may lead to a generalization of information processing in the neuronal networks responsible for delusions and hallucinations on the behavioral level. In this acute paranoid stage of schizophrenia, non-functional astrocytic receptors or their loss decompose the astrocyte domain organization with the effect that a gap between the neuronal and the glial networks arises. If the illness progresses the permanent synaptic neurotransmitter flux may additionally impair the oligodendrocyte-axonic interactions, accompanied by a “creeping” decay of oligodendroglia, axons and glial gap junctions responsible for severe cognitive impairment. Here we may deal with after-effects caused by the basic fault of information processing in tripartite synapses. The gaps between the neuronal and glial networks prohibit the neuronal reality testing of intentional programs presumably generated in the glial networks, called schizophrenic dysintentionality. In non-schizophrenic delusions glia may not be disturbed, but exhausted extrasynaptic information processing may cause an unconstrained synaptic flux responsible for delusions.

Spatial organization of NG2 glial cells and astrocytes in rat hippocampal CA1 region

Similar to astrocytes, NG2 glial cells are uniformly distributed in the central nervous system (CNS). However, little is known about the interspatial relationship, nor the functional interactions between these two star-shaped glial subtypes. Confocal morphometric analysis showed that NG2 immunostained cells are spatially organized as domains in rat hippocampal CA1 region and that each NG2 glial domain occupies a spatial volume of ∼178, 364 μm(3) . The processes of NG2 glia and astrocytes overlap extensively; each NG2 glial domain interlaces with the processes deriving from 5.8 ± 0.4 neighboring astrocytes, while each astrocytic domain accommodates processes stemming from 4.5 ± 0.3 abutting NG2 glia. In CA1 stratum radiatum, the cell bodies of morphologically identified glial cells often appear to make direct somatic-somata contact, termed as doublets. We used dual patch recording and postrecording NG2/GFAP double staining to determine the glial identities of these doublets. We show that among 44 doublets, 50% were NG2 glia-astrocyte pairs, while another 38.6% and 11.4% were astrocyte-astrocyte and NG2 glia-NG2 glia pairs, respectively. In dual patch recording, neither electrical coupling nor intercellular biocytin transfer was detected in astrocyte-NG2 glia or NG2 glia-NG2 glia doublets. Altogether, although NG2 glia and astrocytes are not gap junction coupled, their cell bodies and processes are interwoven extensively. The anatomical and physiological relationships revealed in this study should facilitate future studies to understand the metabolic coupling and functional communication between NG2 glia and astrocytes.

Astrocyte domains and the three-dimensional and seamless expression of consciousness and explicit memories

The expression of consciousness and the site of memory storage within the brain are unknown despite over a century of intense empirical scrutiny. Recent anatomical studies show that human protoplasmic astrocytes form innumerable uniform polyhedral tessellating domains that are arranged three-dimensionally. This complex geometric structure provides a matrix for seamless and three-dimensional expression of consciousness and explicit memories. These studies, in conjunction with physiological data, demonstrate how this may be achieved:1.Individual protoplasmic astrocytes occupy separate three-dimensional non overlapping (i.e., tessellating) territories known as domains. Thus, billions of contiguous and continuous domains tile mammalian cortical gray matter.2.Each domain subtends approximately 90,000 rodent and 2,000,000 human tripartite synapses that signal to perisynaptic astrocyte processes which encode and integrate synaptic information. Neuron to astrocyte signalling is as rapid as neuron to neuron signaling.3.Astrocytes are exquisitely sensitive to neural activity and distinguish synapse from numerous afferent pathways with different neurotransmitters or neuromodulators. Therefore, synaptic information is dynamically integrated within a global matrix of tessellating astrocyte domains.4.Astrocytes of the sensory cortex respond to peripheral stimulation in vivo, and some have the ability to distinguish sensory input in more refined detail than surrounding neurons (e.g., visual cortex). Additionally, astrocytes of the cortex and cerebellum react in concert with activity of awake and behaving animals.5.Domains are extensively interconnected by gap junctions that transmit molecules, many important for information processing and transcription, through complex syncytial networks. This adds an additional level of complexity to interactions between astrocyte domains that may extend over large areas including the entire neocortex.Hypothesis: Consciousness is seamlessly expressed in a three-dimensional matrix consisting of billions of tessellating cortical astrocyte domains that bind attended sensory information. The temporal sequence (stream of consciousness) depends on the global distribution of sequential neural activity. The matrix may also be utilized to encode and store explicit memories.

Astrocyte mega-domain hypothesis of the autistic savantism

Individuals with autism who show high abilities are called savants. Whereas in their brains a disconnection in and between neural networks has been identified, savantism is yet poorly understood. Focusing on astrocyte domain organization, it is hypothesized that local astrocyte mega-organizations may be responsible for exerting high capabilities in brains of autistic savants. Astrocytes, the dominant glial cell type, modulate synaptic information transmission. Each astrocyte is organized in non-overlapping domains. Formally, each astrocyte contacting n-neurons with m-synapses via its processes generates dynamic domains of synaptic interactions based on qualitative computation criteria, and hereby it structures neuronal information processing. If the number of processes is genetically significantly increased, these astrocytes operate in a mega-domain with a higher complexitiy of computation. From this model savant abilities are deduced.

Disruption of neuronal-glial-vascular units in the hippocampus of ovariectomized mice injected with d-galactose

Neuronal-glial-vascular units now have been regarded as the basic functional elements of the CNS; however, their pathophysiological changes in the neurodegenerative condition are poorly defined. Herein we addressed these issues in the hippocampus of an Alzheimer's disease mouse model which was established by ovariectomy and then followed with 8 weeks of d-galactose injection. Immunolocalization of glial fibrillary acidic protein showed that reactive astrogliosis destroyed astrocytic domain organization in the hippocampus of model mice. Electron microscopy further demonstrated microarchitectural damage of neuronal-glial units and gliovascular units in the CA1 stratum radiatum of model group. Moreover, model mice had decreased expressions of glutamate transporter 1, glutamate/aspartate transporter, glutamine synthetase and connexin 43, but an increased expression of water channel protein aquaporin-4 in the hippocampus as revealed by immunohistochemistry and immunoblotting analysis. These results highlight that disruptions in structural integrity of the neuronal-glial-vascular units and associated functions including glutamate clearance, water transport and astrocyte syncytium contribute to the neurodegeneration induced by long-term ovarian hormone deprivation and systemic d-galactose exposure.

Significance of the astrocyte domain organization for qualitative information structuring in the brain

Astrocytes, the dominant glial cell type, modulate synaptic information transmission. Each astrocyte is organized in non-overlapping domains. Here, a formally based model of the possible significance of astrocyte domain organization is proposed. It is hypothesized that each astrocyte contacting n neurons with m synapses via its processes generates dynamic domains of synaptic interactions based on qualitative criteria so that it exerts a structuring of neuronal information processing. The formalism (morpho-grammatics) describes the combinatorics of the various astrocytic receptor types for occupancy with cognate neurotransmitters. Astrocytic processes are able both to contact synapses and retract from them. Rhythmic oscillations of the astrocyte may program the domain organization, where clock genes may play a role in rhythm generation. For the interpretation of a domain organization a player of a string instrument is used as a paradigm. Since astrocytes form networks (syncytia), the interactions between astrocyte domains may be comparable to the improvisations in a jazz ensemble. Given the fact of a high combinational complexity of an astrocyte domain organization, which is formally demonstrable, and an uncomputable complexity of a network of astrocyte domains, the model proposed may not be testable in biological brains, but robotics could be a real alternative.

Special issue on neuron–glia interactions

Special issue on neuron–glia interactions

Astrocyte and oligodendrocyte connexins of the glial syncytium in relation to astrocyte anatomical domains and spatial buffering.

Astroctyes express a set of three connexins (Cx26, Cx30, and Cx43) that are contained in astrocyte-to-astrocyte (A/A) gap junctions; oligodendrocytes express a different set of three connexins (Cx29, Cx32, and Cx47) that are contained in the oligodendrocyte side of necessarily heterotypic astrocyte-to-oligodendrocyte (A/O) gap junctions, and there is little ultrastructural evidence for gap junction formation between individual oligodendrocytes. In addition, primarily Cx29 and Cx32 are contained deeper in myelin sheaths, where they form autologous gap junctions at sites of uncompacted myelin. The presence of six connexins in macroglial cell populations has revealed unprecedented complexity of potential connexin coupling partners, and with restricted deployment of gap junctional intercellular communication (GJIC) within the "pan-glial" syncytium. New implications for the organization and regulation of spatial buffering mediated by glial GJIC are derived from recent observations of the existence of separate astrocyte anatomical domains, with only narrow regions of overlap between astrocyte processes at domain borders. Thus, widespread spatial buffering in the CNS may occur not successively through a multitude of processes arising from different astrocytes, but rather in a more orderly fashion from one astrocyte domain to another via intercellular coupling that occurs only at restricted regions of overlap between astrocyte domains, augmented by autocellular coupling that occurs within each domain.

Astrocyte Networks and Epilepsy: When Stars Collide

Loss of Astrocytic Domain Organization in the Epileptic Brain. Oberheim NA, Tian GF, Han X, Peng W, Takano T, Ransom B, Nedergaard M. J Neurosci 2008;28(13):3264–3276. Gliosis is a pathological hallmark of posttraumatic epileptic foci, but little is known about these reactive astrocytes beyond their high glial fibrillary acidic protein (GFAP) expression. Using diolistic labeling, we show that cortical astrocytes lost their nonoverlapping domain organization in three mouse models of epilepsy: posttraumatic injury, genetic susceptibility, and systemic kainate exposure. Neighboring astrocytes in epileptic mice showed a 10-fold increase in overlap of processes. Concurrently, spine density was increased on dendrites of excitatory neurons. Suppression of seizures by the common antiepileptic, valproate, reduced the overlap of astrocytic processes. Astrocytic domain organization was also preserved in APP transgenic mice expressing a mutant variant of human amyloid precursor protein despite a marked upregulation of GFAP. Our data suggest that loss of astrocytic domains was not universally associated with gliosis, but restricted to seizure pathologies. Reorganization of astrocytes may, in concert with dendritic sprouting and new synapse formation, form the structural basis for recurrent excitation in the epileptic brain. Astroglial Metabolic Networks Sustain Hippocampal Synaptic Transmission. Rouach N, Koulakoff A, Abudara V, Willecke K, Giaume C. Science 2008;322(5907):1551–1555. Astrocytes provide metabolic substrates to neurons in an activity-dependent manner. However, the molecular mechanisms involved in this function, as well as its role in synaptic transmission, remain unclear. Here, we show that the gap-junction subunit proteins connexin 43 and 30 allow intercellular trafficking of glucose and its metabolites through astroglial networks. This trafficking is regulated by glutamatergic synaptic activity mediated by AMPA receptors. In the absence of extracellular glucose, the delivery of glucose or lactate to astrocytes sustains glutamatergic synaptic transmission and epileptiform activity only when they are connected by gap junctions. These results indicate that astroglial gap junctions provide an activity-dependent intercellular pathway for the delivery of energetic metabolites from blood vessels to distal neurons.

Integration of NG2-glia (synantocytes) into the neuroglial network.

NG2-glia are a distinct class of CNS glial cells that are generally classed as oligodendrocyte progenitor cells. However, in the adult CNS a large fraction of NG2 cells does not appear to divide or generate oligodendrocytes. The functions of these adult NG2-glia, which we have termed synantocytes, are unknown. NG2-glia (synantocytes) form interactive domains with astrocytes and neurons. Within their domains, NG2-glia and astrocytes contact the same neurons, form multiple heterologous contacts with each other, and contact pericytes which regulate cerebral blood flow. NG2-glia receive presynaptic input from neurons and respond to neurotransmitters released at synapses. In addition, NG2-glia are intimately associated with astroglia and respond to astroglial signals, a hitherto neglected aspect of NG2-glial cell physiology. The non-overlapping domain organisation of astrocytes is believed to be important in isolating and integrating activity at the synapses and blood vessels within their domains. The domains of NG2-glia overlap with astrocytes, suggesting they could play a role in integrating non-overlapping astrocyte domains.

Astroglia: Not Just Glue

Astroglia: Not Just Glue

Loss of Astrocytic Domain Organization in the Epileptic Brain

Gliosis is a pathological hallmark of posttraumatic epileptic foci, but little is known about these reactive astrocytes beyond their high glial fibrillary acidic protein (GFAP) expression. Using diolistic labeling, we show that cortical astrocytes lost their nonoverlapping domain organization in three mouse models of epilepsy: posttraumatic injury, genetic susceptibility, and systemic kainate exposure. Neighboring astrocytes in epileptic mice showed a 10-fold increase in overlap of processes. Concurrently, spine density was increased on dendrites of excitatory neurons. Suppression of seizures by the common antiepileptic, valproate, reduced the overlap of astrocytic processes. Astrocytic domain organization was also preserved in APP transgenic mice expressing a mutant variant of human amyloid precursor protein despite a marked upregulation of GFAP. Our data suggest that loss of astrocytic domains was not universally associated with gliosis, but restricted to seizure pathologies. Reorganization of astrocytes may, in concert with dendritic sprouting and new synapse formation, form the structural basis for recurrent excitation in the epileptic brain.

The Astrocyte as a Gatekeeper of Synaptic Information Transfer

We present a simple biophysical model for the coupling between synaptic transmission and the local calcium concentration on an enveloping astrocytic domain. This interaction enables the astrocyte to modulate the information flow from presynaptic to postsynaptic cells in a manner dependent on previous activity at this and other nearby synapses. Our model suggests a novel, testable hypothesis for the spike timing statistics measured for rapidly firing cells in culture experiments.

Growth-modulating molecules are associated with invading Schwann cells and not astrocytes in human traumatic spinal cord injury

Despite considerable progress in recent years, the underlying mechanisms responsible for the failure of axonal regeneration after spinal cord injury (SCI) remain only partially understood. Experimental data have demonstrated that a major impediment to the outgrowth of severed axons is the scar tissue that finally dominates the lesion site and, in severe injuries, is comprised of connective tissue and fluid-filled cysts, surrounded by a dense astroglial scar. Reactive astrocytes and infiltrating cells, such as fibroblasts, produce a dense extracellular matrix (ECM) that represents a physical and molecular barrier to axon regeneration. In the human situation, correlative data on the molecular composition of the scar tissue that forms following traumatic SCI is scarce. A detailed investigation on the expression of putative growth-inhibitory and growth-promoting molecules was therefore performed in samples of post-mortem human spinal cord, taken from patients who died following severe traumatic SCI. The lesion-induced scar could be subdivided into a Schwann cell dominated domain which contained large neuromas and a surrounding dense ECM, and a well delineated astroglial scar that isolated the Schwann cell/ECM rich territories from the intact spinal parenchyma. The axon growth-modulating molecules collagen IV, laminin and fibronectin were all present in the post-traumatic scar tissue. These molecules were almost exclusively found in the Schwann cell-rich domain which had an apparent growth-promoting effect on PNS axons. In the astrocytic domain, these molecules were restricted to blood vessel walls without a co-localization with the few regenerating CNS neurites located in this region. Taken together, these results favour the notion that it is the astroglial compartment that plays a dominant role in preventing CNS axon regeneration. The failure to demonstrate any collagen IV, laminin or fibronectin upregulation associated with the astroglial scar suggests that other molecules may play a more significant role in preventing axon regeneration following human SCI.

Shapes of astrocyte networks in the juvenile brain

The high level of intercellular communication mediated by gap junctions between astrocytes indicates that, besides individual astrocytic domains, a second level of organization might exist for these glial cells as they form communicating networks. Therefore,the contribution of astrocytes to brain function should also be considered to result from coordinated groups of cells. To evaluate the shape and extent of these networks we have studied the expression of connexin 43, a major gap junction protein in astrocytes, and the intercellular diffusion of gap junction tracers in two structures of the developing brain, the hippocampus and the cerebral cortex. We report that the shape of astrocytic networks depends on their location within neuronal compartments ina defined brain structure. Interestingly, not all astrocytes are coupled, which indicates that connections within these networks are restricted. As gap junctional communication in astrocytes is reported to contribute to several glial functions, differences in the shape of astrocytic networks might have consequences on neuronal activity and survival.

Maturation of astrocyte morphology and the establishment of astrocyte domains during postnatal hippocampal development

Mature protoplasmic astrocytes exhibit an extremely dense ramification of fine processes, yielding a ‘spongiform’ morphology. This complex morphology enables protoplasmic astrocytes to maintain intimate relationships with many elements of the brain parenchyma, most notably synapses. Recently, it has been demonstrated that astrocytes establish individual cellular-level domains within the neuropil, with limited overlap occurring between the extents of neighboring astrocytes. The highly ramified nature of protoplasmic astrocytes is closely associated with their ability to create such domains. This study was an attempt to characterize the development of spongiform processes and the establishment of astrocyte domains. A combination of immunolabeling for the astrocyte-specific markers glial fibrillary acidic protein and S100β with intracellular dye labeling in fixed tissue slices allowed for the identification of immature astrocytes and the elucidation of their complete, well-preserved morphologies. We find that during the first two postnatal weeks astrocytes extend stringy, filopodial processes. Fine, spongiform processes appear during the third week. Protoplasmic astrocytes are quite heterogeneous in morphology at 1-week postnatum, but there is a remarkable consistency in morphology by 2 weeks of age. Finally, protoplasmic astrocytes initially extend long, overlapping processes during the first two postnatal weeks. The subsequent elaboration of spongiform processes results in the development of boundaries between neighboring astrocyte domains. Stray processes that encroach on neighboring domains are eventually pruned by 1 month of age. These observations suggest that domain formation is largely the consequence of competition between astrocyte processes, similar to the well-studied competitive interactions between certain neuronal dendritic fields.

Micropatterned substrates: approach to probing intercellular communication pathways.

Intercellular signaling is critical for the normal development and physiology of the central nervous system (CNS). To study such signaling, it is vital to control where and when the cells make contact with one another. It is also important to determine whether the process used for cell localization has an impact on signaling. This paper describes a technique that controls the location for cell growth in vitro and demonstrates that the technique has minimal (if any) impact on intercellular signaling. By using photolithographic methods, poly(dimethylsiloxane) molds were fabricated to function as templates for micrometer-level patterning of a nonadhesive agar (agarose) onto glass coverslips coated with a cell adhesive film (poly(L-lysine)). This process yields a surface composed of well-defined adhesive and nonadhesive microdomains. When endothelia or astrocytes are plated onto these substrates, confluent domains of endothelia or astrocytes grow on the poly(L-lysine) domains. Cocultures of astrocytes and neurons can also successfully be used to form interwoven networks on the adhesive domains. Moreover, studies of calcium signaling revealed that astrocytes grown on such patterns retain their native physiological activity. This conclusion is based on the observed propagation rate for calcium waves within individual astrocyte domains and across neighboring, but spatially disconnected, astrocyte domains. The potential to apply these micropatterned substrates as platforms for interrogating communication pathways in key components of the CNS is discussed.