Vladimir Parpura

Tripartite synapses: glia, the unacknowledged partner

According to the classical view of the nervous system, the numerically superior glial cells have inferior roles in that they provide an ideal environment for neuronal-cell function. However, there is a wave of new information suggesting that glia are intimately involved in the active control of neuronal activity and synaptic neurotransmission. Recent evidence shows that glia respond to neuronal activity with an elevation of their internal Ca2+ concentration, which triggers the release of chemical transmitters from glia themselves and, in turn, causes feedback regulation of neuronal activity and synaptic strength. In view of these new insights, this article suggests that perisynaptic Schwann cells and synaptically associated astrocytes should be viewed as integral modulatory elements of tripartite synapses.

Glutamate‐dependent astrocyte modulation of synaptic transmission between cultured hippocampal neurons

The idea that astrocytes merely provide structural and trophic support for neurons has been challenged by the demonstration that astrocytes can regulate neuronal calcium levels. However, the physiological consequences of astrocyte–neuron signalling are unknown. Using mixed cultures of rat hippocampal astrocytes and neurons we have determined functional consequences of elevating astrocyte calcium levels on co‐cultured neurons. Electrical or mechanical stimulation of astrocytes to increase their calcium level caused a glutamate‐dependent slow inward current (SIC) in associated neurons. Microinjection of 1,2‐bis(2‐aminophenoxy)ethane‐N,N,N′,N′‐tetraacetic acid (BAPTA) into astrocytes to prevent the stimulus‐dependent increase in astrocyte calcium level, blocks the appearance of the neuronal SIC. Pharmacological manipulations indicate that this astrocyte‐dependent SIC is mediated by extracellular glutamate acting on N‐methyl‐d‐aspartate (NMDA) and non‐NMDA glutamate receptors. Additionally, stimulation of astrocytes reduced the magnitude of action potential‐evoked excitatory and inhibitory postsynaptic currents through the activation of metabotropic glutamate receptors. The demonstration that astrocytes modulate neuronal currents and synaptic transmission raises the possibility that astrocytes play a neuromodulatory role by controlling the extracellular level of glutamate.

Glial cells in (patho)physiology.

J. Neurochem. (2012) 121, 4–27.

Vesicular glutamate transporter-dependent glutamate release from astrocytes

Astrocytes exhibit excitability based on variations of their intracellular Ca2+ concentrations, which leads to glutamate release, that in turn can signal to adjacent neurons. This glutamate-mediated astrocyte–neuron signaling occurs at physiological intracellular Ca2+ levels in astrocytes and includes modulation of synaptic transmission. The mechanism underlying Ca2+-dependent glutamate release from astrocytes is most likely exocytosis, because astrocytes express the protein components of the soluble N-ethyl maleimide-sensitive fusion protein attachment protein receptors complex, including synaptobrevin 2, syntaxin, and synaptosome-associated protein of 23 kDa. Although these proteins mediate Ca2+-dependent glutamate release from astrocytes, it is not well understood whether astrocytes express functional vesicular glutamate transporters (VGLUTs) that are critical for vesicle refilling. Here, we find in cultured and freshly isolated astrocytes the presence of brain-specific Na+-dependent inorganic phosphate cotransporter and differentiation-associated Na+-dependent inorganic phosphate cotransporter that have recently been identified as VGLUTs 1 and 2. Indirect immunocytochemistry showed a punctate pattern of VGLUT immunoreactivity throughout the entire cell body and processes, whereas pharmacological inhibition of VGLUTs abolished mechanically and agonist-evoked Ca2+-dependent glutamate release from astrocytes. Taken together, these data indicate that VGLUTs play a functional role in exocytotic glutamate release from astrocytes.

Vesicular transmitter release from astrocytes

Astrocytes can release a variety of transmitters, including glutamate and ATP, in response to stimuli that induce increases in intracellular Ca2+ levels. This release occurs via a regulated, exocytotic pathway. As evidence of this, astrocytes express protein components of the vesicular secretory apparatus, including synaptobrevin 2, syntaxin, and SNAP‐23. Additionally, astrocytes possess vesicular organelles, the essential morphological elements required for regulated Ca2+‐dependent transmitter release. The location of specific exocytotic sites on these cells, however, remains to be unequivocally determined.

Expression of synaptobrevin II, cellubrevin and syntaxin but not SNAP-25 in cultured astrocytes

Astrocytes, a sub‐type of glial cell in the central nervous system, can release the excitatory transmitters glutamate and aspartate in response to elevated levels of internal calcium. To investigate potential release mechanisms that may be present in these cells we have determined whether protein components of the neuronal secretory apparatus are expressed in astrocytes. Western blots, immunocytochemistry and RT PCR demonstrate that astrocytes express cellubrevin, synaptobrevin II and syntaxin, proteins known to form a macromolecular fusion complex. However, SNAP‐25 which is another neuronal protein of the fusion complex, was not detected. Astrocyte cellubrevin and synaptobrevin II were also shown to be sensitive to the proteolytic activity of tetanus toxin. Together these data indicate that astrocytes express some proteins that are known to form a fusion complex indicating that regulated exocytosis might mediate calcium‐regulated transmitter release from these cells.

Astrocytes: a central element in neurological diseases.

The neurone-centred view of the past disregarded or downplayed the role of astroglia as a primary component in the pathogenesis of neurological diseases. As this concept is changing, so is also the perceived role of astrocytes in the healthy and diseased brain and spinal cord. We have started to unravel the different signalling mechanisms that trigger specific molecular, morphological and functional changes in reactive astrocytes that are critical for repairing tissue and maintaining function in CNS pathologies, such as neurotrauma, stroke, or neurodegenerative diseases. An increasing body of evidence shows that the effects of astrogliosis on the neural tissue and its functions are not uniform or stereotypic, but vary in a context-specific manner from astrogliosis being an adaptive beneficial response under some circumstances to a maladaptive and deleterious process in another context. There is a growing support for the concept of astrocytopathies in which the disruption of normal astrocyte functions, astrodegeneration or dysfunctional/maladaptive astrogliosis are the primary cause or the main factor in neurological dysfunction and disease. This review describes the multiple roles of astrocytes in the healthy CNS, discusses the diversity of astroglial responses in neurological disorders and argues that targeting astrocytes may represent an effective therapeutic strategy for Alexander disease, neurotrauma, stroke, epilepsy and Alzheimer’s disease as well as other neurodegenerative diseases.

Astroglial excitability and gliotransmission: an appraisal of Ca2+ as a signalling route.

Astroglial cells, due to their passive electrical properties, were long considered subservient to neurons and to merely provide the framework and metabolic support of the brain. Although astrocytes do play such structural and housekeeping roles in the brain, these glial cells also contribute to the brains computational power and behavioural output. These more active functions are endowed by the Ca2+-based excitability displayed by astrocytes. An increase in cytosolic Ca2+ levels in astrocytes can lead to the release of signalling molecules, a process termed gliotransmission, via the process of regulated exocytosis. Dynamic components of astrocytic exocytosis include the vesicular-plasma membrane secretory machinery, as well as the vesicular traffic, which is governed not only by general cytoskeletal elements but also by astrocyte-specific IFs (intermediate filaments). Gliotransmitters released into the ECS (extracellular space) can exert their actions on neighbouring neurons, to modulate synaptic transmission and plasticity, and to affect behaviour by modulating the sleep homoeostat. Besides these novel physiological roles, astrocytic Ca2+ dynamics, Ca2+-dependent gliotransmission and astrocyte–neuron signalling have been also implicated in brain disorders, such as epilepsy. The aim of this review is to highlight the newer findings concerning Ca2+ signalling in astrocytes and exocytotic gliotransmission. For this we report on Ca2+ sources and sinks that are necessary and sufficient for regulating the exocytotic release of gliotransmitters and discuss secretory machinery, secretory vesicles and vesicle mobility regulation. Finally, we consider the exocytotic gliotransmission in the modulation of synaptic transmission and plasticity, as well as the astrocytic contribution to sleep behaviour and epilepsy.

Mechanisms of glutamate release from astrocytes: gap junction "hemichannels", purinergic receptors and exocytotic release

Neuronal exocytotic release of glutamate at synapses involves a highly specialized vesicular apparatus, consisting of a variety of proteins connected to the vesicles or required for vesicular fusion to the presynaptic membrane. Astrocytes also release glutamate, and recent evidence indicates that this release can modify neuronal function. Several mechanisms have been proposed for astrocytic release of glutamate under pathological conditions, such as reversal of glutamate transporters and opening of volume sensitive ion channels. In this review we limit our discussion to findings supporting the exocytotic release of glutamate, as well as two new pathways implicated in this release, the ionotropic (P2X) purinergic receptors and gap junction hemichannels.

Sodium dynamics: another key to astroglial excitability?

Astroglial excitability is largely mediated by fluctuations in intracellular ion concentrations. In addition to generally acknowledged Ca²⁺ excitability of astroglia, recent studies have demonstrated that neuronal activity triggers transient increases in the cytosolic Na⁺ concentration ([Na⁺](i)) in perisynaptic astrocytes. These [Na⁺](i) transients are controlled by multiple Na⁺-permeable channels and Na⁺-dependent transporters; spatiotemporally organized [Na⁺](i) dynamics in turn regulate diverse astroglial homeostatic responses such as metabolic/signaling utilization of lactate and glutamate, transmembrane transport of neurotransmitters and K⁺ buffering. In particular, near-membrane [Na⁺](i) transients determine the rate and the direction of the transmembrane transport of GABA and Ca²⁺. We discuss here the role of Na⁺ in the regulation of various systems that mediate fast bidirectional communication between neurones and glia at the single synapse level.