Péter Illés - Academia.edu (original) (raw)

Papers by Péter Illés

Trends in Pharmacological Sciences, 2014

The ATP-sensitive homomeric P2X7 receptor (P2X7R) has received particular attention as a potentia... more The ATP-sensitive homomeric P2X7 receptor (P2X7R) has received particular attention as a potential drug target because of its widespread involvement in inflammatory diseases as a key regulatory element of the inflammasome complex. However, it has only recently become evident that P2X7Rs also play a pivotal role in central nervous system (CNS) pathology. There is an explosion of data indicating that genetic deletion and pharmacological blockade of P2X7Rs alter responsiveness in animal models of neurological disorders, such as stroke, neurotrauma, epilepsy, neuropathic pain, multiple sclerosis (MS), amyotrophic lateral sclerosis (ALS), Alzheimer's disease, Parkinson's disease, and Huntington's disease. Moreover, recent studies suggest that P2X7Rs regulate the pathophysiology of psychiatric disorders, including mood disorders, implicating P2X7Rs as drug targets in a variety of CNS pathology.

Synapse, 2003

ATP and its metabolite adenosine activate membrane receptors, termed P2 and P1, respectively. In ... more ATP and its metabolite adenosine activate membrane receptors, termed P2 and P1, respectively. In the present study, the modulation of the mesolimbic neuronal circuit by ATPergic and adenosinergic mechanisms was investigated by microdialysis in the nucleus accumbens (NAc) and by telemetrically recorded EEG from both the NAc and the ventral tegmental area (VTA) of freely moving rats. The basal extracellular dopamine concentration was enhanced after accumbal perfusion with the ATP analog 2-methylthio ATP (2-MeSATP; 100 microM); by contrast, adenosine (100 microM) caused a reduction of extracellular dopamine. When given alone, the P2 receptor antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid (PPADS; 20 microM) decreased the concentration of dopamine, whereas the P1 receptor antagonist 8-(p-sulfophenyl)theophylline (8-SPT; 100 microM) increased it. In the same animals, P2 receptor stimulation by 2-MeSATP caused neuronal activation, indicated by an elevation of the absolute power in the EEG of the NAc mainly by enhancement of the relative power in the alpha band (8-13 Hz) of the EEG spectrum. By contrast, adenosine led to a depression of the absolute power in the VTA accompanied by an elevation of the delta-band power (0.4-6 Hz) in the NAc corresponding to a slowing of neuronal activity. When given alone, PPADS reduced the absolute EEG power in the NAc accompanied by a decrease in the high-frequency power, but had no effects on the VTA. 8-SPT on its own enhanced the total power in both the NAc and the VTA, reflected by an enhancement in the slow and the high-frequency bands. Whereas the 8-SPT-evoked changes of EEG pattern as well as of dopamine concentration in the NAc were abolished by the co-application of PPADS, the 8-SPT-induced EEG changes in the VTA persisted under these conditions. In conclusion, the accumbal neuronal output, reflected by both dopamine release and neuronal electrical activity, is modulated in a functionally antagonistic manner by P2 and P1 receptor stimulation. It is suggested that an inhibitory GABAergic feedback projection to the VTA is stimulated by adenosine, either directly or indirectly via glutamate release.

Purinergic Signalling, 2010

Locus coeruleus (LC) neurons in a rat brain slice preparation were superfused with a Mg 2+ -free ... more Locus coeruleus (LC) neurons in a rat brain slice preparation were superfused with a Mg 2+ -free and bicuculline-containing external medium. Under these conditions, glutamatergic spontaneous excitatory postsynaptic currents (sEPSCs) were recorded by means of the whole-cell patch-clamp method. ATP, as well as its structural analogue 2-methylthio ATP (2-MeSATP), both caused transient inward currents, which were outlasted by an increase in the frequency but not the amplitude of the sEPSCs. PPADS, but not suramin or reactive blue 2 counteracted both effects of 2-MeSATP. By contrast, α,β-methylene ATP (α,β-meATP), UTP and BzATP did not cause an inward current response. Of these latter agonists, only BzATP slightly facilitated the sEPSC amplitude and strongly potentiated its frequency. PPADS and Brilliant Blue G, as well as fluorocitric acid and aminoadipic acid prevented the activity of BzATP. Furthermore, BzATP caused a similar facilitation of the miniature (m)EPSC (recorded in the presence of tetrodotoxin) and sEPSC frequencies (recorded in its absence). Eventually, capsaicin augmented the frequency of the sEPSCs in a capsazepine-, but not PPADS-antagonizable, manner. In conclusion, the stimulation of astrocytic P2X7 receptors appears to lead to the outflow of a signalling molecule, which presynaptically increases the spontaneous release of glutamate onto LC neurons from their afferent fibre tracts. It is suggested, that the two algogenic compounds ATP and capsaicin utilise separate receptor systems to potentiate the release of glutamate and in consequence to increase the excitability of LC neurons.

Purinergic Signalling, 2012

Astrocytes are fundamental for central nervous system (CNS) physiology and are the fulcrum of neu... more 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.

Purinergic Signalling, 2004

The importance of purinergic signaling in the intact mesolimbic-mesocortical circuit of the brain... more The importance of purinergic signaling in the intact mesolimbic-mesocortical circuit of the brain of freely moving rats is reviewed. In the rat, an endogenous ADP/ATPergic tone reinforces the release of dopamine from the axon terminals in the nucleus accumbens as well as from the somatodendritic region of these neurons in the ventral tegmental area, as well as the release of glutamate, probably via P2Y 1 receptor stimulation. Similar mechanisms may regulate the release of glutamate in both areas of the brain. Dopamine and glutamate determine in concert the activity of the accumbal GABAergic, medium-size spiny neurons thought to act as an interface between the limbic cortex and the extrapyramidal motor system. These neurons project to the pallidal and mesencephalic areas, thereby mediating the behavioral reaction of the animal in response to a motivation-related stimulus. There is evidence that extracellular ADP/ATP promotes goaldirected behavior, e.g., intention and feeding, via dopamine, probably via P2Y 1 receptor stimulation. Accumbal P2 receptor-mediated glutamatergic mechanisms seem to counteract the dopaminergic effects on behavior. Furthermore, adaptive changes of motivation-related behavior, e.g., by chronic succession of starvation and feeding or by repeated amphetamine administration, are accompanied by changes in the expression of the P2Y 1 receptor, thought to modulate the sensitivity of the animal to respond to certain stimuli.

Purinergic Signalling, 2010

Psychopharmacology, 2000

Rationale: Previous experiments have shown that P 2 receptor activation increases the release of ... more Rationale: Previous experiments have shown that P 2 receptor activation increases the release of dopamine in the mesolimbic mesocortical system. Objective: In order to investigate the functional correlates of dopaminergic stimulation, EEG and behavioural responses to injection of the P 2 receptor agonist 2-methylthio ATP (2-MeSATP) into the nucleus accumbens (NAc) of rats were investigated. Methods: EEG electrodes were positioned into the NAc together with the guide cannula for intracerebral injection. Behavioural analysis was performed in an open field cage and was evaluated by a video activity measurement system. Rats were assigned to separate groups that were given artificial cerebrospinal fluid (aCSF) or drug treatment. Results: 2-MeSATP significantly extended the period of locomotor activity in the novel environment. The quantitative EEG was characterized by an elevation of the power in the alpha-1 range and a decrease in power in the delta range. The P 2 receptor antagonists reactive blue 2 but not pyridoxalphosphate-6-azophenyl-2'4'-disulphonic acid (PPADS) also enhanced locomotion when given alone, and elevated the alpha-1 and beta-2 bands. Both antagonists abolished the locomotor and EEG responses to 2-MeSATP. The dopamine D 1 receptor antagonist SCH 23390 and the D 2 /D 3 receptor antagonist sulpiride did not alter locomotor activity when given either alone or in combination. Only sulpiride and especially sulpiride in combination with SCH 23390 prevented the effect of 2-MeSATP. Sulpiride produced a selective increase in the alpha-1 band of the power spectrum whereas SCH 23390 elevated the power of the alpha-1, alpha-2 and beta-1 activities. Neither antagonist inhibited the effect of 2-MeSATP on the EEG when applied separately; however, the coadministration of SCH 23390 and sulpiride abolished the 2-MeSATP-induced alteration of power distribution. After a 6-hydroxydopamine (6-OHDA)-induced lesion of the accumbal dopaminergic terminals, 2-MeSATP failed to enhance the locomotor activity and to induce the characteristic EEG changes. Conclusions: The observed alterations in open field behaviour and quantitative EEG after injection of 2-MeSATP into the NAc may be mostly due to P 2 receptor-mediated dopamine release and subsequent receptor activation.

Pflügers Archiv - European Journal of Physiology, 2006

Extracellular adenosine 5′-triphosphate (ATP) was proposed to be an activity-dependent signaling ... more Extracellular adenosine 5′-triphosphate (ATP) was proposed to be an activity-dependent signaling molecule that regulates glia-glia and glia-neuron communications. ATP is a neurotransmitter of its own right and, in addition, a cotransmitter of other classical transmitters such as glutamate or GABA. The effects of ATP are mediated by two receptor families belonging either to the P2X (ligandgated cationic channels) or P2Y (G protein-coupled receptors) types. P2X receptors are responsible for rapid synaptic responses, whereas P2Y receptors mediate slow synaptic responses and other types of purinergic signaling involved in neuronal damage/regeneration. ATP may act at pre-and postsynaptic sites and therefore, it may participate in the phenomena of long-term potentiation and long-term depression of excitatory synaptic transmission. The release of ATP into the extracellular space, e.g., by exocytosis, membrane transporters, and connexin hemichannels, is a widespread physiological process. However, ATP may also leave cells through their plasma membrane damaged by inflammation, ischemia, and mechanical injury. Functional responses to the activation of multiple P2 receptors were found in neurons and glial cells under normal and pathophysiological conditions. P2 receptor-activation could either be a cause or a consequence of neuronal cell death/ glial activation and may be related to detrimental and/or beneficial effects. The present review aims at demonstrating that purinergic mechanisms correlate with the etiopathology of brain insults, especially because of the massive extracellular release of ATP, adenosine, and other neurotransmitters after brain injury. We will focus in this review on the most important P2 receptor-mediated neurodegenerative and neuroprotective processes and their beneficial modulation by possible therapeutic manipulations.

Neuroscience Letters, 1993

Membrane currents of cultured human monocytes and rat microglia were recorded with the whole-cell... more Membrane currents of cultured human monocytes and rat microglia were recorded with the whole-cell patch clamp technique. Freshly isolated Inonocytes or resting (proliferating) microglia express only inwardly rectifying K+ channels. However, incubation in teflon bags leads to the expression of additional, outwardly rectifying K+ channels. The outward K+ conductance of microglial cells was inhibited by intracellular Cs' and extracellular 4-aminopyridine or tetraethylammonium. Functional similarities with the microglial outwardly rectifying K+ channel were found in the K.-channel oflymphocytes which has recently been cloned (RGK5). The polymerase chain reaction (PCR) was used to demonstrate the presence of RGK5-like mRNA in microglia.

Neuroscience Letters, 2008

Mixed neuronal/astrocytic cortical cell cultures of the rat were incubated for 2 or 12 h under no... more Mixed neuronal/astrocytic cortical cell cultures of the rat were incubated for 2 or 12 h under normoxic or ischemic conditions. Subsequent flow cytometric analysis with an anti-P2X 7 receptor antibody directed against an extracellular epitope indicated the up-regulation of these receptors at the plasma membrane by 12 h of ischemia. Labelling of MAP-2 immunopositive neurons by an anti-P2X 7 antibody directed against a C-terminal epitope, documented the selectivity of the ischemia-induced increase in receptor-density for the neuronal population. By contrast, staining of GFAP immunopositive astrocytes by the same anti-P2X 7 antibody excluded any effect of ischemia on the astrocytic density of P2X 7 receptors. The ischemic upregulation of neuronal P2X 7 receptors is in perfect agreement with the previously reported facilitation of transmitter release from the GABAergic non-pyramidal cell type in such cultures [

Neuroscience Letters, 1992

Membrane currents of cultured rat microglia were recorded with the whole-cell patch clamp techniq... more Membrane currents of cultured rat microglia were recorded with the whole-cell patch clamp technique. Undifferentiated microglia express only inwardly rectifying K+ channels. However, treatment of the cells with bacterial lipopolysaccharide, interferon-gamma, or their incubation in hydrophobic teflon bags, procedures that promote microglial differentiation, induced the expression of an additional outward current. Cycloheximide prevented the development of this conductance indicating the synthesis of a new channel protein. The reversal potential of the outward current was near to the K+ equilibrium potential; the current was abolished by intracellular Cs+ or extracellular 4-aminopyridine, and was depressed by extracellular tetraethylammonium. Hence, the channels involved appear to be highly selective for K+; their possible function is a rapid termination of depolarizing shifts of the membrane potential.

NeuroReport, 2005

Whole-cell patch-clamp recordings were performed on HEK293 cells transiently transfected with the... more Whole-cell patch-clamp recordings were performed on HEK293 cells transiently transfected with the rat (r) wild-type transient receptor potential vanilloid 1 (TRPV1) (rTRPV1) receptor or with a mutant that lacks the potential N-glycosylation site at position N604 (rTRPV1-N604T). Replacement of Asn by Thr (N604T) depressed the maximum of the concentration-response curve for capsaicin and decreased the EC50 value of this agonist. Further, such a manipulation modified the sensitivity to the TRPV1 receptor-antagonist capsazepine and altered the dependence of the capsaicin effect on extracellular pH. Hence, glycosylation may affect the basic functional characteristics of the rTRPV1 receptor channel in accordance with the knowledge that N-glycosylation may regulate ligand binding or gating properties of ionotropic neurotransmitter receptors.

Neurochemistry International, 2013

Current responses from CA1 neurons and stratum oriens astrocytes were recorded from hippocampal b... more Current responses from CA1 neurons and stratum oriens astrocytes were recorded from hippocampal brain slices by means of the whole-cell patch-clamp technique. Anoxic depolarization (AD) was induced by an oxygen/glucose-deprived (OGD) medium also containing sodium iodoacetate and antimycin, in order to block glycolysis and oxidative phosphorylation, respectively. Anoxic depolarization has been reported to be due to the sudden increase of the extracellular K + concentration and the accompanying explosive rise in glutamate concentration. We asked ourselves whether the release of ATP activating P2X7 receptors is also involved in the AD. Although, the AD was evoked in absolute synchrony in neurons and astrocytes, and the NMDA receptor antagonistic AP-5 depressed these responses, neither the nonselective P2 receptor antagonist PPADS, nor the highly selective P2X7 receptor antagonist A438079 interfered with the AD or its delay time in neurons/astrocytes after inducing chemical hypoxia. However, A438079, but not PPADS increased in astrocytes the slow inward current observed in a hypoxic medium. It is concluded that ATP co-released with glutamate by hypoxic stimulation has only a minor function in the present brain slice system.

Naunyn-Schmiedeberg's Archives of Pharmacology, 2002

The aim of the present study was to investigate whether 5,7-dihydroxytryptamine (5,7-DHT), an aut... more The aim of the present study was to investigate whether 5,7-dihydroxytryptamine (5,7-DHT), an autofluorescent serotonin derivative, can be used as a specific marker for serotonergic or dopaminergic neurons in rat mesencephalic cultures. To this end, primary cultures were prepared from the ventral brain stem of 14-day-old Wistar rat foetuses and kept in culture for 10 days (DIV10). At DIV10, the cultures were characterized immunocytochemically with antibodies raised against tyrosine hydroxylase (TH; a marker for catecholaminergic/dopaminergic neurons) and serotonin (5-HT). 5,7-DHT labelling of the neurons was investigated after incubation with 25 µM of the serotonin derivative (plus 0.005% ascorbic acid) for 60 min at 37°C, followed by incubation with primary antibodies against TH or serotonin and a fluorescence (Cy3)-labelled secondary antibody. Using confocal laser scanning microscopy, this double immunofluorescence approach demonstrated that all cells which had accumulated 5,7-DHT additionally displayed anti-5-HT immunoreactivity, whereas no evidence was found for 5,7-DHT labelling of TH immunoreactive cells. Preincubation with the selective serotonin reuptake inhibitor fluvoxamine maleate (10 µM) prevented the loading of the 5-HT-positive cells with 5,7-DHT. In conclusion, the present data indicate that 5,7-DHT specifically labels serotonergic cells in rat midbrain cultures. Thus, 5,7-DHT can be used for the identification of living serotonergic neurons even in the presence of dopaminergic neurons.

Naunyn-Schmiedeberg's Archives of Pharmacology, 1992

Intracellular recordings were performed in a pontine slice preparation of the rat brain containin... more Intracellular recordings were performed in a pontine slice preparation of the rat brain containing the locus coeruleus (LC). The spontaneous firing of action potentials was prevented by passing continuous hyperpolarizing current via the recording electrode. Focal electrical stimulation evoked a synaptic depolarization (PSP) followed by a hyperpolarization (IPSP). Neuropeptide Y (NPY; 0.1 txmol/1) inhibited the IPSP only. Pressure ejection of noradrenaline produced hyperpolarization which was potentiated in the presence of NPY (0.1 Ixmol/1). Hence, NPY appears to inhibit the release of noradrenaline from dendrites or recurrent axon collaterals of LC neurones.

Naunyn-Schmiedeberg's Archives of Pharmacology, 1989

Naunyn-Schmiedeberg's Archives of Pharmacology, 1989

Naunyn-Schmiedeberg's Archives of Pharmacology, 1986

Slices of the rabbit hippocampus were preincubated with 3H-choline, rinsed and superfused continu... more Slices of the rabbit hippocampus were preincubated with 3H-choline, rinsed and superfused continuously. The release of 3H-acetylcholine in these slices, evoked by electrical field stimulation, was strongly reduced by the preferential kappa-agonists ethylketocyclazocine, dynorphin A (1-13) and dynorphin A (1-17). Dynorphin A (1-9) and (-)MR 2034 [(-)5,9-dimethyl-2'-OH-2-tetrahydrofurfuryl-6, 7-benzomorphan] were less potent, the (+)enantiomer of (-)MR 2034 was ineffective. Whereas the mu-agonist DAGO (D-Ala2-Gly-ol5-enkephalin) showed significant depressant effects, two other mu-agonists morphine and morphiceptine, as well as the delta-agonists DADLE (D-Ala2-D-Leu5-enkephalin) and Leu-enkephalin were much less inhibitory. The preferential mu-antagonist (-)naloxone as well as (-)MR 2266 [(-)N-(3-furylmethyl)-alpha-noretazocine], a preferential kappa-antagonist, did not increase acetylcholine release when given alone, but antagonized the effect of ethylketocyclazocine; (-)MR 2266 (Ke: 1.6 nmol/l) was about 4 times more potent than (-)naloxone (Ke: 6.3 nmol/l). The inhibitory effects of DAGO and DADLE were abolished by (-)MR 2266 (0.1 mumol/l) but not by the delta-antagonist ICI 174864 (N,N-diallyl-Tyr-Aib-Phe-Leu-OH, 0.3 mumol/l). It is concluded that the release of acetylcholine in the hippocampus of the rabbit is inhibited at the level of the axon terminals via kappa-receptors; in addition, mu-receptors may be present. An inhibitory tone of endogenous opioid peptides on hippocampal acetylcholine release could not be demonstrated. Experiments on rat hippocampal slices showed that in this species mu- rather than kappa-receptors may modulate acetylcholine release.

Naunyn-Schmiedeberg's Archives of Pharmacology, 1990

Naunyn-Schmiedeberg's Archives of Pharmacology, 1996

~2-Adrenoceptor agonists inhibit the firing of locus coeruleus (LC) neurons. It was recently obse... more ~2-Adrenoceptor agonists inhibit the firing of locus coeruleus (LC) neurons. It was recently observed that the ct-adrenoceptor agonists clonidine, rilmenidine and cirazoline, when injected intravenously in anaesthetized rats pretreated with the irreversible c~2-adrenoceptor antagonist N-ethoxycarbonyl-2-ethoxy-1,2dihydroquinoline (EEDQ), excite the LC. The effect was attributed to activation of 11 imidazoline receptors. The aim of the present experiments was to characterize the direct effect of c~2-adrenoceptor and I1 imidazoline receptor agonists on LC neurons.

Trends in Pharmacological Sciences, 2014

The ATP-sensitive homomeric P2X7 receptor (P2X7R) has received particular attention as a potentia... more The ATP-sensitive homomeric P2X7 receptor (P2X7R) has received particular attention as a potential drug target because of its widespread involvement in inflammatory diseases as a key regulatory element of the inflammasome complex. However, it has only recently become evident that P2X7Rs also play a pivotal role in central nervous system (CNS) pathology. There is an explosion of data indicating that genetic deletion and pharmacological blockade of P2X7Rs alter responsiveness in animal models of neurological disorders, such as stroke, neurotrauma, epilepsy, neuropathic pain, multiple sclerosis (MS), amyotrophic lateral sclerosis (ALS), Alzheimer's disease, Parkinson's disease, and Huntington's disease. Moreover, recent studies suggest that P2X7Rs regulate the pathophysiology of psychiatric disorders, including mood disorders, implicating P2X7Rs as drug targets in a variety of CNS pathology.

Synapse, 2003

ATP and its metabolite adenosine activate membrane receptors, termed P2 and P1, respectively. In ... more ATP and its metabolite adenosine activate membrane receptors, termed P2 and P1, respectively. In the present study, the modulation of the mesolimbic neuronal circuit by ATPergic and adenosinergic mechanisms was investigated by microdialysis in the nucleus accumbens (NAc) and by telemetrically recorded EEG from both the NAc and the ventral tegmental area (VTA) of freely moving rats. The basal extracellular dopamine concentration was enhanced after accumbal perfusion with the ATP analog 2-methylthio ATP (2-MeSATP; 100 microM); by contrast, adenosine (100 microM) caused a reduction of extracellular dopamine. When given alone, the P2 receptor antagonist pyridoxalphosphate-6-azophenyl-2',4'-disulphonic acid (PPADS; 20 microM) decreased the concentration of dopamine, whereas the P1 receptor antagonist 8-(p-sulfophenyl)theophylline (8-SPT; 100 microM) increased it. In the same animals, P2 receptor stimulation by 2-MeSATP caused neuronal activation, indicated by an elevation of the absolute power in the EEG of the NAc mainly by enhancement of the relative power in the alpha band (8-13 Hz) of the EEG spectrum. By contrast, adenosine led to a depression of the absolute power in the VTA accompanied by an elevation of the delta-band power (0.4-6 Hz) in the NAc corresponding to a slowing of neuronal activity. When given alone, PPADS reduced the absolute EEG power in the NAc accompanied by a decrease in the high-frequency power, but had no effects on the VTA. 8-SPT on its own enhanced the total power in both the NAc and the VTA, reflected by an enhancement in the slow and the high-frequency bands. Whereas the 8-SPT-evoked changes of EEG pattern as well as of dopamine concentration in the NAc were abolished by the co-application of PPADS, the 8-SPT-induced EEG changes in the VTA persisted under these conditions. In conclusion, the accumbal neuronal output, reflected by both dopamine release and neuronal electrical activity, is modulated in a functionally antagonistic manner by P2 and P1 receptor stimulation. It is suggested that an inhibitory GABAergic feedback projection to the VTA is stimulated by adenosine, either directly or indirectly via glutamate release.

Purinergic Signalling, 2010

Locus coeruleus (LC) neurons in a rat brain slice preparation were superfused with a Mg 2+ -free ... more Locus coeruleus (LC) neurons in a rat brain slice preparation were superfused with a Mg 2+ -free and bicuculline-containing external medium. Under these conditions, glutamatergic spontaneous excitatory postsynaptic currents (sEPSCs) were recorded by means of the whole-cell patch-clamp method. ATP, as well as its structural analogue 2-methylthio ATP (2-MeSATP), both caused transient inward currents, which were outlasted by an increase in the frequency but not the amplitude of the sEPSCs. PPADS, but not suramin or reactive blue 2 counteracted both effects of 2-MeSATP. By contrast, α,β-methylene ATP (α,β-meATP), UTP and BzATP did not cause an inward current response. Of these latter agonists, only BzATP slightly facilitated the sEPSC amplitude and strongly potentiated its frequency. PPADS and Brilliant Blue G, as well as fluorocitric acid and aminoadipic acid prevented the activity of BzATP. Furthermore, BzATP caused a similar facilitation of the miniature (m)EPSC (recorded in the presence of tetrodotoxin) and sEPSC frequencies (recorded in its absence). Eventually, capsaicin augmented the frequency of the sEPSCs in a capsazepine-, but not PPADS-antagonizable, manner. In conclusion, the stimulation of astrocytic P2X7 receptors appears to lead to the outflow of a signalling molecule, which presynaptically increases the spontaneous release of glutamate onto LC neurons from their afferent fibre tracts. It is suggested, that the two algogenic compounds ATP and capsaicin utilise separate receptor systems to potentiate the release of glutamate and in consequence to increase the excitability of LC neurons.

Purinergic Signalling, 2012

Astrocytes are fundamental for central nervous system (CNS) physiology and are the fulcrum of neu... more 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.

Purinergic Signalling, 2004

The importance of purinergic signaling in the intact mesolimbic-mesocortical circuit of the brain... more The importance of purinergic signaling in the intact mesolimbic-mesocortical circuit of the brain of freely moving rats is reviewed. In the rat, an endogenous ADP/ATPergic tone reinforces the release of dopamine from the axon terminals in the nucleus accumbens as well as from the somatodendritic region of these neurons in the ventral tegmental area, as well as the release of glutamate, probably via P2Y 1 receptor stimulation. Similar mechanisms may regulate the release of glutamate in both areas of the brain. Dopamine and glutamate determine in concert the activity of the accumbal GABAergic, medium-size spiny neurons thought to act as an interface between the limbic cortex and the extrapyramidal motor system. These neurons project to the pallidal and mesencephalic areas, thereby mediating the behavioral reaction of the animal in response to a motivation-related stimulus. There is evidence that extracellular ADP/ATP promotes goaldirected behavior, e.g., intention and feeding, via dopamine, probably via P2Y 1 receptor stimulation. Accumbal P2 receptor-mediated glutamatergic mechanisms seem to counteract the dopaminergic effects on behavior. Furthermore, adaptive changes of motivation-related behavior, e.g., by chronic succession of starvation and feeding or by repeated amphetamine administration, are accompanied by changes in the expression of the P2Y 1 receptor, thought to modulate the sensitivity of the animal to respond to certain stimuli.

Purinergic Signalling, 2010

Psychopharmacology, 2000

Rationale: Previous experiments have shown that P 2 receptor activation increases the release of ... more Rationale: Previous experiments have shown that P 2 receptor activation increases the release of dopamine in the mesolimbic mesocortical system. Objective: In order to investigate the functional correlates of dopaminergic stimulation, EEG and behavioural responses to injection of the P 2 receptor agonist 2-methylthio ATP (2-MeSATP) into the nucleus accumbens (NAc) of rats were investigated. Methods: EEG electrodes were positioned into the NAc together with the guide cannula for intracerebral injection. Behavioural analysis was performed in an open field cage and was evaluated by a video activity measurement system. Rats were assigned to separate groups that were given artificial cerebrospinal fluid (aCSF) or drug treatment. Results: 2-MeSATP significantly extended the period of locomotor activity in the novel environment. The quantitative EEG was characterized by an elevation of the power in the alpha-1 range and a decrease in power in the delta range. The P 2 receptor antagonists reactive blue 2 but not pyridoxalphosphate-6-azophenyl-2'4'-disulphonic acid (PPADS) also enhanced locomotion when given alone, and elevated the alpha-1 and beta-2 bands. Both antagonists abolished the locomotor and EEG responses to 2-MeSATP. The dopamine D 1 receptor antagonist SCH 23390 and the D 2 /D 3 receptor antagonist sulpiride did not alter locomotor activity when given either alone or in combination. Only sulpiride and especially sulpiride in combination with SCH 23390 prevented the effect of 2-MeSATP. Sulpiride produced a selective increase in the alpha-1 band of the power spectrum whereas SCH 23390 elevated the power of the alpha-1, alpha-2 and beta-1 activities. Neither antagonist inhibited the effect of 2-MeSATP on the EEG when applied separately; however, the coadministration of SCH 23390 and sulpiride abolished the 2-MeSATP-induced alteration of power distribution. After a 6-hydroxydopamine (6-OHDA)-induced lesion of the accumbal dopaminergic terminals, 2-MeSATP failed to enhance the locomotor activity and to induce the characteristic EEG changes. Conclusions: The observed alterations in open field behaviour and quantitative EEG after injection of 2-MeSATP into the NAc may be mostly due to P 2 receptor-mediated dopamine release and subsequent receptor activation.

Pflügers Archiv - European Journal of Physiology, 2006

Extracellular adenosine 5′-triphosphate (ATP) was proposed to be an activity-dependent signaling ... more Extracellular adenosine 5′-triphosphate (ATP) was proposed to be an activity-dependent signaling molecule that regulates glia-glia and glia-neuron communications. ATP is a neurotransmitter of its own right and, in addition, a cotransmitter of other classical transmitters such as glutamate or GABA. The effects of ATP are mediated by two receptor families belonging either to the P2X (ligandgated cationic channels) or P2Y (G protein-coupled receptors) types. P2X receptors are responsible for rapid synaptic responses, whereas P2Y receptors mediate slow synaptic responses and other types of purinergic signaling involved in neuronal damage/regeneration. ATP may act at pre-and postsynaptic sites and therefore, it may participate in the phenomena of long-term potentiation and long-term depression of excitatory synaptic transmission. The release of ATP into the extracellular space, e.g., by exocytosis, membrane transporters, and connexin hemichannels, is a widespread physiological process. However, ATP may also leave cells through their plasma membrane damaged by inflammation, ischemia, and mechanical injury. Functional responses to the activation of multiple P2 receptors were found in neurons and glial cells under normal and pathophysiological conditions. P2 receptor-activation could either be a cause or a consequence of neuronal cell death/ glial activation and may be related to detrimental and/or beneficial effects. The present review aims at demonstrating that purinergic mechanisms correlate with the etiopathology of brain insults, especially because of the massive extracellular release of ATP, adenosine, and other neurotransmitters after brain injury. We will focus in this review on the most important P2 receptor-mediated neurodegenerative and neuroprotective processes and their beneficial modulation by possible therapeutic manipulations.

Neuroscience Letters, 1993

Membrane currents of cultured human monocytes and rat microglia were recorded with the whole-cell... more Membrane currents of cultured human monocytes and rat microglia were recorded with the whole-cell patch clamp technique. Freshly isolated Inonocytes or resting (proliferating) microglia express only inwardly rectifying K+ channels. However, incubation in teflon bags leads to the expression of additional, outwardly rectifying K+ channels. The outward K+ conductance of microglial cells was inhibited by intracellular Cs' and extracellular 4-aminopyridine or tetraethylammonium. Functional similarities with the microglial outwardly rectifying K+ channel were found in the K.-channel oflymphocytes which has recently been cloned (RGK5). The polymerase chain reaction (PCR) was used to demonstrate the presence of RGK5-like mRNA in microglia.

Neuroscience Letters, 2008

Mixed neuronal/astrocytic cortical cell cultures of the rat were incubated for 2 or 12 h under no... more Mixed neuronal/astrocytic cortical cell cultures of the rat were incubated for 2 or 12 h under normoxic or ischemic conditions. Subsequent flow cytometric analysis with an anti-P2X 7 receptor antibody directed against an extracellular epitope indicated the up-regulation of these receptors at the plasma membrane by 12 h of ischemia. Labelling of MAP-2 immunopositive neurons by an anti-P2X 7 antibody directed against a C-terminal epitope, documented the selectivity of the ischemia-induced increase in receptor-density for the neuronal population. By contrast, staining of GFAP immunopositive astrocytes by the same anti-P2X 7 antibody excluded any effect of ischemia on the astrocytic density of P2X 7 receptors. The ischemic upregulation of neuronal P2X 7 receptors is in perfect agreement with the previously reported facilitation of transmitter release from the GABAergic non-pyramidal cell type in such cultures [

Neuroscience Letters, 1992

Membrane currents of cultured rat microglia were recorded with the whole-cell patch clamp techniq... more Membrane currents of cultured rat microglia were recorded with the whole-cell patch clamp technique. Undifferentiated microglia express only inwardly rectifying K+ channels. However, treatment of the cells with bacterial lipopolysaccharide, interferon-gamma, or their incubation in hydrophobic teflon bags, procedures that promote microglial differentiation, induced the expression of an additional outward current. Cycloheximide prevented the development of this conductance indicating the synthesis of a new channel protein. The reversal potential of the outward current was near to the K+ equilibrium potential; the current was abolished by intracellular Cs+ or extracellular 4-aminopyridine, and was depressed by extracellular tetraethylammonium. Hence, the channels involved appear to be highly selective for K+; their possible function is a rapid termination of depolarizing shifts of the membrane potential.

NeuroReport, 2005

Whole-cell patch-clamp recordings were performed on HEK293 cells transiently transfected with the... more Whole-cell patch-clamp recordings were performed on HEK293 cells transiently transfected with the rat (r) wild-type transient receptor potential vanilloid 1 (TRPV1) (rTRPV1) receptor or with a mutant that lacks the potential N-glycosylation site at position N604 (rTRPV1-N604T). Replacement of Asn by Thr (N604T) depressed the maximum of the concentration-response curve for capsaicin and decreased the EC50 value of this agonist. Further, such a manipulation modified the sensitivity to the TRPV1 receptor-antagonist capsazepine and altered the dependence of the capsaicin effect on extracellular pH. Hence, glycosylation may affect the basic functional characteristics of the rTRPV1 receptor channel in accordance with the knowledge that N-glycosylation may regulate ligand binding or gating properties of ionotropic neurotransmitter receptors.

Neurochemistry International, 2013

Current responses from CA1 neurons and stratum oriens astrocytes were recorded from hippocampal b... more Current responses from CA1 neurons and stratum oriens astrocytes were recorded from hippocampal brain slices by means of the whole-cell patch-clamp technique. Anoxic depolarization (AD) was induced by an oxygen/glucose-deprived (OGD) medium also containing sodium iodoacetate and antimycin, in order to block glycolysis and oxidative phosphorylation, respectively. Anoxic depolarization has been reported to be due to the sudden increase of the extracellular K + concentration and the accompanying explosive rise in glutamate concentration. We asked ourselves whether the release of ATP activating P2X7 receptors is also involved in the AD. Although, the AD was evoked in absolute synchrony in neurons and astrocytes, and the NMDA receptor antagonistic AP-5 depressed these responses, neither the nonselective P2 receptor antagonist PPADS, nor the highly selective P2X7 receptor antagonist A438079 interfered with the AD or its delay time in neurons/astrocytes after inducing chemical hypoxia. However, A438079, but not PPADS increased in astrocytes the slow inward current observed in a hypoxic medium. It is concluded that ATP co-released with glutamate by hypoxic stimulation has only a minor function in the present brain slice system.

Naunyn-Schmiedeberg's Archives of Pharmacology, 2002

The aim of the present study was to investigate whether 5,7-dihydroxytryptamine (5,7-DHT), an aut... more The aim of the present study was to investigate whether 5,7-dihydroxytryptamine (5,7-DHT), an autofluorescent serotonin derivative, can be used as a specific marker for serotonergic or dopaminergic neurons in rat mesencephalic cultures. To this end, primary cultures were prepared from the ventral brain stem of 14-day-old Wistar rat foetuses and kept in culture for 10 days (DIV10). At DIV10, the cultures were characterized immunocytochemically with antibodies raised against tyrosine hydroxylase (TH; a marker for catecholaminergic/dopaminergic neurons) and serotonin (5-HT). 5,7-DHT labelling of the neurons was investigated after incubation with 25 µM of the serotonin derivative (plus 0.005% ascorbic acid) for 60 min at 37°C, followed by incubation with primary antibodies against TH or serotonin and a fluorescence (Cy3)-labelled secondary antibody. Using confocal laser scanning microscopy, this double immunofluorescence approach demonstrated that all cells which had accumulated 5,7-DHT additionally displayed anti-5-HT immunoreactivity, whereas no evidence was found for 5,7-DHT labelling of TH immunoreactive cells. Preincubation with the selective serotonin reuptake inhibitor fluvoxamine maleate (10 µM) prevented the loading of the 5-HT-positive cells with 5,7-DHT. In conclusion, the present data indicate that 5,7-DHT specifically labels serotonergic cells in rat midbrain cultures. Thus, 5,7-DHT can be used for the identification of living serotonergic neurons even in the presence of dopaminergic neurons.

Naunyn-Schmiedeberg's Archives of Pharmacology, 1992

Intracellular recordings were performed in a pontine slice preparation of the rat brain containin... more Intracellular recordings were performed in a pontine slice preparation of the rat brain containing the locus coeruleus (LC). The spontaneous firing of action potentials was prevented by passing continuous hyperpolarizing current via the recording electrode. Focal electrical stimulation evoked a synaptic depolarization (PSP) followed by a hyperpolarization (IPSP). Neuropeptide Y (NPY; 0.1 txmol/1) inhibited the IPSP only. Pressure ejection of noradrenaline produced hyperpolarization which was potentiated in the presence of NPY (0.1 Ixmol/1). Hence, NPY appears to inhibit the release of noradrenaline from dendrites or recurrent axon collaterals of LC neurones.

Naunyn-Schmiedeberg's Archives of Pharmacology, 1989

Naunyn-Schmiedeberg's Archives of Pharmacology, 1989

Naunyn-Schmiedeberg's Archives of Pharmacology, 1986

Slices of the rabbit hippocampus were preincubated with 3H-choline, rinsed and superfused continu... more Slices of the rabbit hippocampus were preincubated with 3H-choline, rinsed and superfused continuously. The release of 3H-acetylcholine in these slices, evoked by electrical field stimulation, was strongly reduced by the preferential kappa-agonists ethylketocyclazocine, dynorphin A (1-13) and dynorphin A (1-17). Dynorphin A (1-9) and (-)MR 2034 [(-)5,9-dimethyl-2'-OH-2-tetrahydrofurfuryl-6, 7-benzomorphan] were less potent, the (+)enantiomer of (-)MR 2034 was ineffective. Whereas the mu-agonist DAGO (D-Ala2-Gly-ol5-enkephalin) showed significant depressant effects, two other mu-agonists morphine and morphiceptine, as well as the delta-agonists DADLE (D-Ala2-D-Leu5-enkephalin) and Leu-enkephalin were much less inhibitory. The preferential mu-antagonist (-)naloxone as well as (-)MR 2266 [(-)N-(3-furylmethyl)-alpha-noretazocine], a preferential kappa-antagonist, did not increase acetylcholine release when given alone, but antagonized the effect of ethylketocyclazocine; (-)MR 2266 (Ke: 1.6 nmol/l) was about 4 times more potent than (-)naloxone (Ke: 6.3 nmol/l). The inhibitory effects of DAGO and DADLE were abolished by (-)MR 2266 (0.1 mumol/l) but not by the delta-antagonist ICI 174864 (N,N-diallyl-Tyr-Aib-Phe-Leu-OH, 0.3 mumol/l). It is concluded that the release of acetylcholine in the hippocampus of the rabbit is inhibited at the level of the axon terminals via kappa-receptors; in addition, mu-receptors may be present. An inhibitory tone of endogenous opioid peptides on hippocampal acetylcholine release could not be demonstrated. Experiments on rat hippocampal slices showed that in this species mu- rather than kappa-receptors may modulate acetylcholine release.

Naunyn-Schmiedeberg's Archives of Pharmacology, 1990

Naunyn-Schmiedeberg's Archives of Pharmacology, 1996

~2-Adrenoceptor agonists inhibit the firing of locus coeruleus (LC) neurons. It was recently obse... more ~2-Adrenoceptor agonists inhibit the firing of locus coeruleus (LC) neurons. It was recently observed that the ct-adrenoceptor agonists clonidine, rilmenidine and cirazoline, when injected intravenously in anaesthetized rats pretreated with the irreversible c~2-adrenoceptor antagonist N-ethoxycarbonyl-2-ethoxy-1,2dihydroquinoline (EEDQ), excite the LC. The effect was attributed to activation of 11 imidazoline receptors. The aim of the present experiments was to characterize the direct effect of c~2-adrenoceptor and I1 imidazoline receptor agonists on LC neurons.