Ionotropic ATP receptors in neuronal-glial communication

In the central nervous system ATP is released from both neurones and astroglial cells acting as a homo- and heterocellular neurotransmitter. Glial cells express numerous purinoceptors of both ionotropic (P2X) and metabotropic (P2Y) varieties. Astroglial P2X receptors can be activated by ongoing synaptic transmission and can mediate fast local signalling through elevation in cytoplasmic Ca(2+) and Na(+) concentrations. These ionic signals can be translated into various physiological messages by numerous pathways, including release of gliotransmitters, metabolic support of neurones and regulation of activity of postsynaptic glutamate and GABA receptors. Ionotropic purinoceptors represent a novel pathway of glia-driven modulation of synaptic signalling that involves the release of ATP from neurones and astrocytes followed by activation of P2X receptors which can regulate synaptic activity by variety of mechanisms expressed in both neuronal and glial compartments.     

ATP released by astrocytes mediates glutamatergic activity-dependent heterosynaptic suppression

Extracellular ATP released from axons is known to assist activity-dependent signaling between neurons and Schwann cells in the peripheral nervous system. Here we report that ATP released from astrocytes as a result of neuronal activity can also modulate central synaptic transmission. In cultures of hippocampal neurons, endogenously released ATP tonically suppresses glutamatergic synapses via presynaptic P2Y receptors, an effect that depends on the presence of cocultured astrocytes. Glutamate release accompanying neuronal activity also activates non-NMDA receptors of nearby astrocytes and triggers ATP release from these cells, which in turn causes homo- and heterosynaptic suppression. In CA1 pyramidal neurons of hippocampal slices, a similar synaptic suppression was also produced by adenosine, an immediate degradation product of ATP released by glial cells. Thus, neuron-glia crosstalk may participate in activity-dependent synaptic modulation.     

Neuroprotective roles of the P2Y2 receptor

Purinergic signaling plays a unique role in the brain by integrating neuronal and glial cellular circuits. The metabotropic P1 adenosine receptors and P2Y nucleotide receptors and ionotropic P2X receptors control numerous physiological functions of neuronal and glial cells and have been implicated in a wide variety of neuropathologies. Emerging research suggests that purinergic receptor interactions between cells of the central nervous system (CNS) have relevance in the prevention and attenuation of neurodegenerative diseases resulting from chronic inflammation. CNS responses to chronic inflammation are largely dependent on interactions between different cell types (i.e., neurons and glia) and activation of signaling molecules including P2X and P2Y receptors. Whereas numerous P2 receptors contribute to functions of the CNS, the P2Y(2) receptor is believed to play an important role in neuroprotection under inflammatory conditions. While acute inflammation is necessary for tissue repair due to injury, chronic inflammation contributes to neurodegeneration in Alzheimer's disease and occurs when glial cells undergo prolonged activation resulting in extended release of proinflammatory cytokines and nucleotides. This review describes cell-specific and tissue-integrated functions of P2 receptors in the CNS with an emphasis on P2Y(2) receptor signaling pathways in neurons, glia, and endothelium and their role in neuroprotection.     

Regulation of cell-to-cell communication mediated by astrocytic ATP in the CNS

It has become apparent that glial cells, especially astrocytes, not merely supportive but are integrative, being able to receive inputs, assimilate information and send instructive chemical signals to other neighboring cells including neurons. At first, the excitatory neurotransmitter glutamate was found to be a major extracellular messenger that mediates these communications because it can be released from astrocytes in a Ca(2+)-dependent manner, diffused, and can stimulate extra-synaptic glutamate receptors in adjacent neurons, leading to a dynamic modification of synaptic transmission. However, recently extracellular ATP has come into the limelight as an important extracellular messenger for these communications. Astrocytes express various neurotransmitter receptors including P2 receptors, release ATP in response to various stimuli and respond to extracellular ATP to cause various physiological responses. The intercellular communication "Ca(2+) wave" in astrocytes was found to be mainly mediated by the release of ATP and the activation of P2 receptors, suggesting that ATP is a dominant "gliotransmitter" between astrocytes. Because neurons also express various P2 receptors and synapses are surrounded by astrocytes, astrocytic ATP could affect neuronal activities and even dynamically regulate synaptic transmission in adjacent neurons as if forming a "tripartite synapse". In this review, we summarize the role of astrocytic ATP, as compared with glutamate, in gliotransmission and synaptic transmission in neighboring cells, mainly focusing on the hippocampus. Dynamic communication between astrocytes and neurons mediated by ATP would be a key event in the processing or integration of information in the CNS.     

Ionotropic NMDA and P2X1/5 receptors mediate synaptically induced Ca2+ signalling in cortical astrocytes

Local, global and propagating calcium (Ca(2+)) signals provide the substrate for glial excitability. Here we analyse Ca(2+) permeability of NMDA and P2X(1/5) receptors expressed in cortical astrocytes and provide evidence that activation of these receptors trigger astroglial Ca(2+) signals when stimulated by either endogenous agonists or by synaptic release of neurotransmitters. The Ca(2+) permeability of the ionotropic receptors was determined by reversal potential shift analysis; the permeability ratio P(Ca)/P(K) was 3.1 for NMDA receptors and 2.2 for P2X(1/5) receptors. Selective stimulation of ionotropic receptors (with NMDA and α,β-methyleneATP) in freshly isolated cortical astrocytes induced ion currents associated with transient increases in cytosolic Ca(2+) concentration ([Ca(2+)](i)). Stimulation of neuronal afferents in cortical slices triggered glial synaptic currents and [Ca(2+)](i) responses, which were partially blocked by selective antagonists of NMDA (D-AP5 and UBP141) and P2X(1/5) (NF449) receptors. We conclude that ionotropic receptors contribute to astroglial Ca(2+) signalling and may provide a specific mechanism for fast neuronal-glial signalling at the synaptic level.     

A dialogue between glia and neurons in the retina: modulation of neuronal excitability

Bidirectional signaling between neurons and glial cells has been demonstrated in brain slices and is believed to mediate glial modulation of synaptic transmission in the CNS. Our laboratory has characterized similar neuron-glia signaling in the mammalian retina. We find that light-evoked neuronal activity elicits Ca(2+) increases in Müller cells, which are specialized retinal glial cells. Neuron to glia signaling is likely mediated by the release of ATP from neurons and is potentiated by adenosine. Glia to neuron signaling has also been observed and is mediated by several mechanisms. Stimulation of glial cells can result in either facilitation or depression of synaptic transmission. Release of D-serine from Müller cells might also potentiate NMDA receptor transmission. Müller cells directly inhibit ganglion cells by releasing ATP, which, following hydrolysis to adenosine, activates neuronal A(1) receptors. The existence of bidirectional signaling mechanisms indicates that glial cells participate in information processing in the retina.     

Electroconvulsive therapy: a novel hypothesis for the involvement of purinergic signalling

It is proposed that ATP is released from both neurons and glia during electroconvulsive therapy (ECT) and that this leads to reduction of depressive behaviour via complex stimulation of neurons and glia directly via P2X and P2Y receptors and also via P1 receptors after extracellular breakdown of ATP to adenosine. In particular, A₁ adenosine receptors inhibit release of excitatory transmitters, and A_2A and P2Y receptors may modulate the release of dopamine. Sequential ECT may lead to changes in purinoceptor expression in mesolimbic and mesocortical regions of the brain implicated in depression and other mood disorders. In particular, increased expression of P2X7 receptors on glial cells would lead to increased release of cytokines, chemokines and neurotrophins. In summary, we suggest that ATP release following ECT involves neurons, glial cells and neuron–glial interactions acting via both P2 and after breakdown to adenosine via P1 receptors. We suggest that ecto-nucleotidase inhibitors (increasing available amounts of ATP) and purinoceptor agonists may enhance the anti-depressive effect of ECT.     

P2X and P2Y purinoreceptors mediate ATP-evoked calcium signalling in optic nerve glia in situ

It is known that ATP acts as an extracellular messenger mediating Ca2+ signalling in glial cells. Here, the mechanisms involved in the ATP-evoked increase in glial [Ca2+]i were studied in situ, in the acutely isolated rat optic nerve. ATP and agonists for P2X (a,b-metATP) and P2Y (2MeSATP) purinoreceptors triggered raised glial [Ca2+]i, and there was no significant difference between cells identified morphologically as astrocytes and oligodendrocytes. Dose-response curves indicated that P2Y receptors were activated at nanomolar concentrations, whereas P2X purinoreceptors were only activated above 10 microM. The rank order of potency for several agonists indicated optic nerve glia expressed heterogeneous purinoreceptors, with P2Y1< or = P2Y2/4< or = P2X. The ATP evoked increase in [Ca2+]i was reversibly blocked by the P2X/Y purinoreceptor antagonist suramin (100 microM) and markedly reduced by thapsigargin (10 microM), which blocks IP3-dependent release of Ca2+ from intracellular stores. Removal of extracellular Ca2+ reduced the ATP evoked increase in [Ca2+]i and completely blocked its recovery, indicating that refilling of intracellular stores was ultimately dependent on Ca2+ influx from the extracellular milieu. The results implicate ATP as an important signal in CNS white matter astrocytes and oligodendrocytes in situ, and indicate that metabotropic P2Y purinoreceptors mobilize intracellular Ca2+ at physiological concentrations of ATP, whereas ionotropic P2X purinoreceptors induce Ca2+ influx across the plasmalemma only at high concentrations of ATP, such as occur following CNS injury.     

Purinoceptors on Neuroglia

Purinergic transmission is one of the most ancient and widespread extracellular signalling systems. In the brain, purinergic signalling plays a unique role in integrating neuronal and glial cellular circuits, as virtually every type of glial cell possesses receptors to purines and pyrimidines. These receptors, represented by metabotropic P1 adenosine receptors, metabotropic P2Y purinoceptors and ionotropic P2X purinoceptors, control numerous physiological functions of glial cells and are intimately involved in virtually every form of neuropathology. In this essay, we provide an in depth overview of purinoceptor distribution in two types of CNS glia—in astrocytes and oligodendrocytes—and discuss their physiological and pathophysiological roles. An erratum to this article can be found at     

Purinergic signalling in neuron-glia interactions

Activity-dependent release of ATP from synapses, axons and glia activates purinergic membrane receptors that modulate intracellular calcium and cyclic AMP. This enables glia to detect neural activity and communicate among other glial cells by releasing ATP through membrane channels and vesicles. Through purinergic signalling, impulse activity regulates glial proliferation, motility, survival, differentiation and myelination, and facilitates interactions between neurons, and vascular and immune system cells. Interactions among purinergic, growth factor and cytokine signalling regulate synaptic strength, development and responses to injury. We review the involvement of ATP and adenosine receptors in neuron-glia signalling, including the release and hydrolysis of ATP, how the receptors signal, the pharmacological tools used to study them, and their functional significance.     

Exocytosis of ATP From Astrocytes Modulates Phasic and Tonic Inhibition in the Neocortex

Communication between neuronal and glial cells is important for many brain functions. Astrocytes can modulate synaptic strength via Ca²⁺-stimulated release of various gliotransmitters, including glutamate and ATP. A physiological role of ATP release from astrocytes was suggested by its contribution to glial Ca²⁺-waves and purinergic modulation of neuronal activity and sleep homeostasis. The mechanisms underlying release of gliotransmitters remain uncertain, and exocytosis is the most intriguing and debated pathway. We investigated release of ATP from acutely dissociated cortical astrocytes using “sniff-cell” approach and demonstrated that release is vesicular in nature and can be triggered by elevation of intracellular Ca²⁺ via metabotropic and ionotropic receptors or direct UV-uncaging. The exocytosis of ATP from neocortical astrocytes occurred in the millisecond time scale contrasting with much slower nonvesicular release of gliotransmitters via Best1 and TREK-1 channels, reported recently in hippocampus. Furthermore, we discovered that elevation of cytosolic Ca²⁺ in cortical astrocytes triggered the release of ATP that directly activated quantal purinergic currents in the pyramidal neurons. The glia-driven burst of purinergic currents in neurons was followed by significant attenuation of both synaptic and tonic inhibition. The Ca²⁺-entry through the neuronal P2X purinoreceptors led to phosphorylation-dependent down-regulation of GABAA receptors. The negative purinergic modulation of postsynaptic GABA receptors was accompanied by small presynaptic enhancement of GABA release. Glia-driven purinergic modulation of inhibitory transmission was not observed in neurons when astrocytes expressed dn-SNARE to impair exocytosis. The astrocyte-driven purinergic currents and glia-driven modulation of GABA receptors were significantly reduced in the P2X4 KO mice. Our data provide a key evidence to support the physiological importance of exocytosis of ATP from astrocytes in the neocortex.     

Metabotropic Glutamate Receptors in Glial Cells

Glutamate is the major excitatory neurotransmitter in the mammalian central nervous system (CNS) and exerts its actions via a number of ionotropic glutamate receptors/channels and metabotropic glutamate (mGlu) receptors. In addition to being expressed in neurons, glutamate receptors are expressed in different types of glial cells including astrocytes, oligodendrocytes, and microglia. Astrocytes are now recognized as dynamic signaling elements actively integrating neuronal inputs. Synaptic activity can evoke calcium signals in astrocytes, resulting in the release of gliotransmitters, such as glutamate, ATP, and d-serine, which in turn modulate neuronal excitability and synaptic transmission. In addition, astrocytes, and microglia may play an important role in pathology such as brain trauma and neurodegeneration, limiting or amplifying the pathologic process leading to neuronal death. The present review will focus on recent advances on the role of mGlu receptors expressed in glial cells under physiologic and pathologic conditions. Special issue article in honor of Dr. Anna Maria Giuffrida-Stella.     

The role of nucleotides in the neuron--glia communication responsible for the brain functions

Accumulating findings indicate that nucleotides play an important role in cell-to-cell communication through P2 purinoceptors, even though ATP is recognized primarily to be a source of free energy and nucleotides are key molecules in cells. P2 purinoceptors are divided into two families, ionotropic receptors (P2X) and metabotropic receptors (P2Y). P2X receptors (7 types; P2X(1)-P2X(7)) contain intrinsic pores that open by binding with ATP. P2Y (8 types; P2Y(1, 2, 4, 6, 11, 12, 13,) and (14)) are activated by nucleotides and couple to intracellular second-messenger systems through heteromeric G-proteins. Nucleotides are released or leaked from non-excitable cells as well as neurons in physiological and pathophysiological conditions. One of the most exciting cells in non-excitable cells is the glia cells, which are classified into astrocytes, oligodendrocytes, and microglia. Astrocytes express many types of P2 purinoceptors and release the 'gliotransmitter' ATP to communicate with neurons, microglia and the vascular walls of capillaries. Microglia also express many types of P2 purinoceptors and are known as resident macrophages in the CNS. ATP and other nucleotides work as 'warning molecules' especially through activating microglia in pathophysiological conditions. Microglia play a key role in neuropathic pain and show phagocytosis through nucleotide-evoked activation of P2X(4) and P2Y(6) receptors, respectively. Such strong molecular, cellular and system-level evidence for extracellular nucleotide signaling places nucleotides in the central stage of cell communications in glia/CNS.     

Dual Electrophysiological Recordings of Synaptically-evoked Astroglial and Neuronal Responses in Acute Hippocampal Slices

Astrocytes form together with neurons tripartite synapses, where they integrate and modulate neuronal activity. Indeed, astrocytes sense neuronal inputs through activation of their ion channels and neurotransmitter receptors, and process information in part through activity-dependent release of gliotransmitters. Furthermore, astrocytes constitute the main uptake system for glutamate, contribute to potassium spatial buffering, as well as to GABA clearance. These cells therefore constantly monitor synaptic activity, and are thereby sensitive indicators for alterations in synaptically-released glutamate, GABA and extracellular potassium levels. Additionally, alterations in astroglial uptake activity or buffering capacity can have severe effects on neuronal functions, and might be overlooked when characterizing physiopathological situations or knockout mice. Dual recording of neuronal and astroglial activities is therefore an important method to study alterations in synaptic strength associated to concomitant changes in astroglial uptake and buffering capacities. Here we describe how to prepare hippocampal slices, how to identify stratum radiatum astrocytes, and how to record simultaneously neuronal and astroglial electrophysiological responses. Furthermore, we describe how to isolate pharmacologically the synaptically-evoked astroglial currents.     

Glial cell regulation of neuronal activity and blood flow in the retina by release of gliotransmitters

Astrocytes in the brain release transmitters that actively modulate neuronal excitability and synaptic efficacy. Astrocytes also release vasoactive agents that contribute to neurovascular coupling. As reviewed in this article, Müller cells, the principal retinal glial cells, modulate neuronal activity and blood flow in the retina. Stimulated Müller cells release ATP which, following its conversion to adenosine by ectoenzymes, hyperpolarizes retinal ganglion cells by activation of A1 adenosine receptors. This results in the opening of G protein-coupled inwardly rectifying potassium (GIRK) channels and small conductance Ca²⁺-activated K⁺ (SK) channels. Tonic release of ATP also contributes to the generation of tone in the retinal vasculature by activation of P2X receptors on vascular smooth muscle cells. Vascular tone is lost when glial cells are poisoned with the gliotoxin fluorocitrate. The glial release of vasoactive metabolites of arachidonic acid, including prostaglandin E₂ (PGE₂) and epoxyeicosatrienoic acids (EETs), contributes to neurovascular coupling in the retina. Neurovascular coupling is reduced when neuronal stimulation of glial cells is interrupted and when the synthesis of arachidonic acid metabolites is blocked. Neurovascular coupling is compromised in diabetic retinopathy owing to the loss of glial-mediated vasodilation. This loss can be reversed by inhibiting inducible nitric oxide synthase. It is likely that future research will reveal additional important functions of the release of transmitters from glial cells.     

Astrocytic adenosine: from synapses to psychiatric disorders

Although it is considered to be the most complex organ in the body, the brain can be broadly classified into two major types of cells, neuronal cells and glial cells. Glia is a general term that encompasses multiple types of non-neuronal cells that function to maintain homeostasis, form myelin, and provide support and protection for neurons. Astrocytes, a major class of glial cell, have historically been viewed as passive support cells, but recently it has been discovered that astrocytes participate in signalling activities both with the vasculature and with neurons at the synapse. These cells have been shown to release d-serine, TNF-α, glutamate, atrial natriuretic peptide (ANP) and ATP among other signalling molecules. ATP and its metabolites are well established as important signalling molecules, and astrocytes represent a major source of ATP release in the nervous system. Novel molecular and genetic tools have recently shown that astrocytic release of ATP and other signalling molecules has a major impact on synaptic transmission. Via actions at the synapse, astrocytes have now been shown to regulate complex network signalling in the whole organism with impacts on respiration and the sleep–wake cycle. In addition, new roles for astrocytes are being uncovered in psychiatric disorders, and astrocyte signalling mechanisms represents an attractive target for novel therapeutic agents.     

Ionotropic receptors in neuronal-astroglial signalling: What is the role of “excitable” molecules in non-excitable cells

Astroglial cells were long considered to serve merely as the structural and metabolic supporting cast and scenery against which the shining neurones perform their illustrious duties. Relatively recent evidence, however, indicates that astrocytes are intimately involved in many of the brain's functions. Astrocytes possess a diverse assortment of ionotropic transmitter receptors, which enable these glial cells to respond to many of the same signals that act on neurones. Ionotropic receptors mediate neurone-driven signals to astroglial cells in various brain areas including neocortex, hippocampus and cerebellum. Activation of ionotropic receptors trigger rapid signalling events in astroglia; these events, represented by local Ca²⁺ or Na⁺ signals provide the mechanism for fast neuronal-glial signalling at the synaptic level. Since astrocytes can detect chemical transmitters that are released from neurones and can release their own extracellular signals, gliotransmitters, they are intricately involved in homocellular and heterocellular signalling mechanisms in the nervous system. This article is part of a Special Issue entitled: 11th European Symposium on Calcium.     

Pathophysiology of astroglial purinergic signalling

Astrocytes are fundamental for central nervous system (CNS) physiology and are the fulcrum of neurological diseases. Astroglial cells control development of the nervous system, regulate synaptogenesis, maturation, maintenance and plasticity of synapses and are central for nervous system homeostasis. Astroglial reactions determine progression and outcome of many neuropathologies and are critical for regeneration and remodelling of neural circuits following trauma, stroke, ischaemia or neurodegenerative disorders. They secrete multiple neurotransmitters and neurohormones to communicate with neurones, microglia and the vascular walls of capillaries. Signalling through release of ATP is the most widespread mean of communication between astrocytes and other types of neural cells. ATP serves as a fast excitatory neurotransmitter and has pronounced long-term (trophic) roles in cell proliferation, growth, and development. During pathology, ATP is released from damaged cells and acts both as a cytotoxic factor and a proinflammatory mediator, being a universal "danger" signal. In this review, we summarise contemporary knowledge on the role of purinergic receptors (P2Rs) in a variety of diseases in relation to changes of astrocytic functions and nucleotide signalling. We have focussed on the role of the ionotropic P2X and metabotropic P2YRs working alone or in concert to modify the release of neurotransmitters, to activate signalling cascades and to change the expression levels of ion channels and protein kinases. All these effects are of great importance for the initiation, progression and maintenance of astrogliosis-the conserved and ubiquitous glial defensive reaction to CNS pathologies. We highlighted specific aspects of reactive astrogliosis, especially with respect to the involvement of the P2X(7) and P2Y(1)R subtypes. Reactive astrogliosis exerts both beneficial and detrimental effects in a context-specific manner determined by distinct molecular signalling cascades. Understanding the role of purinergic signalling in astrocytes is critical to identifying new therapeutic principles to treat acute and chronic neurological diseases.     

Extracellular ATP-prinoceptor signaling and AMP-activated protein kinase regulate astrocytic glucose transporter 3 in an in vitro ischemia

Astrocytes become hypertrophic reactive in response to the ischemic stress, and they contribute to either protect or exacerbate neuronal damage, depending on the depth or duration of the stress. Astrocytes have more resistance to the ischemic stress than neurons, which is apparently due to active anerobic metabolic pathway in the emergency situation. We have been focused on the functional role of astrocytic glucose transporters in the ischemic condition. Under the physiological conditions, cultured astrocytes primarily express glucose transporter1 (GLUT1), and GLUT3 is only detected at extremely low levels. But astrocytes enhance GLUT3 expression through the signaling of nuclear factor-κ-light-chain-enhancer of activated B cells (NF-κB) under mild ischemic condition. It is reasonable since GLUT3 transports extracellular glucose about seven times faster than GLUT1, so astrocytes enhance the storage of intracellular glucose during the ischemia. However, other signaling cascades that regulate GLUT3 production remain unknown. Here we demonstrate that extracellular adenosine 5′-triphosphate (ATP)-P2Y receptor signaling also regulates GLUT3 expression. Under mild ischemic condition, astrocytes positively released existing intracellular or newly synthesized ATP by AMP-activated protein kinase (AMPK) signaling. The released extracellular ATP from pore channels activated ATP-sensitive P2Y receptor signaling, resulting in an increase in c-Fos and c-Jun proteins. Newly synthesized GLUT3 was regulated by those signaling since the inhibition of P2Y receptors or c-Fos/c-Jun signaling significantly reduced GLUT3 expression. Furthermore, the inhibition of P2Y receptors during the ischemic condition sustained intracellular ATP concentration, leading to a decrease in AMPK proteins. These results suggest AMPK-regulated ATP production triggers the release of ATP to activate P2Y receptor signaling, which is another candidate that regulates GLUT3 expression under the ischemic condition.     

Bi-functional effects of ATP/P2 receptor activation on tumor necrosis factor-alpha release in lipopolysaccharide-stimulated astrocytes

Neuroinflammation is associated with a variety of CNS pathologies. Levels of tumor necrosis factor-alpha (TNF-alpha), a major proinflammatory cytokine, as well as extracellular ATP, are increased following various CNS insults. Here we report on the relationship between ATP/P2 purinergic receptor activation and lipopolysaccharide (LPS)-induced TNF-alpha release from primary cultures of rat cortical astrocytes. Using ELISA, we confirmed that treatment with LPS stimulated the release of TNF-alpha in a concentration and time dependent manner. ATP treatment alone had no effect on TNF-alpha release. LPS-induced TNF-alpha release was attenuated by 1 mm ATP, a concentration known to activate P2X7 receptors. Consistent with this, 3'-O-(4-Benzoyl)benzoyl-ATP (BzATP), a P2X7 receptor agonist, also attenuated LPS-induced TNF-alpha release. This reduction in TNF-alpha release was not due to loss of cell viability. Adenosine and 2-chloroadenosine were ineffective, suggesting that attenuation of LPS-induced TNF-alpha release by ATP was not due to ATP breakdown and subsequent activation of adenosine/P1 receptors. Interestingly, treatment of astrocyte cultures with 10 microm or 100 microm ATP potentiated TNF-alpha release induced by a submaximal concentration of LPS. UTP and 2methylthioADP (2-MeSADP), P2Y receptor agonists, also enhanced this LPS-induced TNF-alpha release. Our observations demonstrate opposing effects of ATP/P2 receptor activation on TNF-alpha release, i.e. P2X receptor activation attenuates, whereas P2Y receptor activation potentiates TNF-alpha release in LPS-stimulated astrocytes. These observations suggest a mechanism whereby astrocytes can sense the severity of damage in the CNS via ATP release from damaged cells and can modulate the TNF-alpha mediated inflammatory response depending on the extracellular ATP concentration and corresponding type of astrocyte ATP/P2 receptor activated.