Publications by authors named "Felicita Pedata"

62 Publications

Protective Effect of Adenosine A Receptor Agonist, BAY60-6583, Against Transient Focal Brain Ischemia in Rat.

Front Pharmacol 2020 11;11:588757. Epub 2021 Feb 11.

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), Division of Pharmacology and Toxicology, University of Florence, Florence, Italy.

Cerebral ischemia is a multifactorial pathology characterized first by an acute injury, due to excitotoxicity, followed by a secondary brain injury that develops hours to days after ischemia. During ischemia, adenosine acts as an endogenous neuroprotectant. Few studies have investigated the role of A receptor in brain ischemia because of the low potency of adenosine for it and the few selective ligands developed so far. A receptors are scarcely but widely distributed in the brain on neurons, glial and endothelial cells and on hematopoietic cells, lymphocytes and neutrophils, where they exert mainly anti-inflammatory effects, inhibiting vascular adhesion and inflammatory cells migration. Aim of this work was to verify whether chronic administration of the A agonist, BAY60-6583 (0.1 mg/kg i.p., twice/day), starting 4 h after focal ischemia induced by transient (1 h) Middle Cerebral Artery occlusion (tMCAo) in the rat, was protective after the ischemic insult. BAY60-6583 improved the neurological deficit up to 7 days after tMCAo. Seven days after ischemia BAY60-6583 reduced significantly the ischemic brain damage in cortex and striatum, counteracted ischemia-induced neuronal death, reduced microglia activation and astrocytes alteration. Moreover, it decreased the expression of TNF-α and increased that of IL-10 in peripheral plasma. Two days after ischemia BAY60-6583 reduced blood cell infiltration in the ischemic cortex. The present study indicates that A receptors stimulation can attenuate the neuroinflammation that develops after ischemia, suggesting that A receptors may represent a new interesting pharmacological target to protect from degeneration after brain ischemia.
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http://dx.doi.org/10.3389/fphar.2020.588757DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7905306PMC
February 2021

New Insight into the Role of Adenosine in Demyelination, Stroke and Neuropathic Pain.

Front Pharmacol 2020 29;11:625662. Epub 2021 Jan 29.

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), Section of Pharmacology and Toxicology, University of Florence, Florence, Italy.

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http://dx.doi.org/10.3389/fphar.2020.625662DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7878385PMC
January 2021

A Adenosine Receptors: When Outsiders May Become an Attractive Target to Treat Brain Ischemia or Demyelination.

Int J Mol Sci 2020 Dec 18;21(24). Epub 2020 Dec 18.

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), Section of Pharmacology and Toxicology, University of Florence, 50139 Florence, Italy.

Adenosine is a signaling molecule, which, by activating its receptors, acts as an important player after cerebral ischemia. Here, we review data in the literature describing AR-mediated effects in models of cerebral ischemia obtained in vivo by the occlusion of the middle cerebral artery (MCAo) or in vitro by oxygen-glucose deprivation (OGD) in hippocampal slices. Adenosine plays an apparently contradictory role in this receptor subtype depending on whether it is activated on neuro-glial cells or peripheral blood vessels and/or inflammatory cells after ischemia. Indeed, ARs participate in the early glutamate-mediated excitotoxicity responsible for neuronal and synaptic loss in the CA1 hippocampus. On the contrary, later after ischemia, the same receptors have a protective role in tissue damage and functional impairments, reducing inflammatory cell infiltration and neuroinflammation by central and/or peripheral mechanisms. Of note, demyelination following brain ischemia, or autoimmune neuroinflammatory reactions, are also profoundly affected by ARs since they are expressed by oligodendroglia where their activation inhibits cell maturation and expression of myelin-related proteins. In conclusion, data in the literature indicate the ARs as putative therapeutic targets for the still unmet treatment of stroke or demyelinating diseases.
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http://dx.doi.org/10.3390/ijms21249697DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7766015PMC
December 2020

Acetylcholine modulates K and Na currents in human basal forebrain cholinergic neuroblasts through an autocrine/paracrine mechanism.

J Neurochem 2020 Oct 8. Epub 2020 Oct 8.

Department of Experimental and Clinical Medicine, Section of Human Anatomy and Histology, University of Florence, Florence, Italy.

The Nucleus Basalis of Meynert (NBM) is the main source of cholinergic neurons in the basal forebrain to be crucially involved in cognitive functions and whose degeneration correlates with cognitive decline in major degenerative pathologies as Alzheimer's and Parkinson's diseases. However, knowledge concerning NBM neurons derived from human brain is very limited to date. We recently characterized a primary culture of proliferating neuroblasts isolated from the human fetal NBM (hfNBM) as immature cholinergic neurons expressing the machinery to synthetize and release acetylcholine. Here we studied in detail electrophysiological features and cholinergic effects in this cell culture by patch-clamp recordings. Our data demonstrate that atropine-blocked muscarinic receptor activation by acetylcholine or carbachol enhanced I and reduced I currents by stimulating G -coupled M2 or phospholipase C-coupled M3 receptors, respectively. Inhibition of acetylcholine esterase activity by neostigmine unveiled a spontaneous acetylcholine release from hfNBM neuroblasts that might account for an autocrine/paracrine signaling during human brain development. Present data provide the first description of cholinergic effects in human NBM neurons and point to a role of acetylcholine as an autocrine/paracrine modulator of voltage-dependent channels. Our research could be of relevance in understanding the mechanisms of cholinergic system development and functions in the human brain, either in health or disease.
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http://dx.doi.org/10.1111/jnc.15209DOI Listing
October 2020

Acute visceral pain relief mediated by A3AR agonists in rats: involvement of N-type voltage-gated calcium channels.

Pain 2020 09;161(9):2179-2190

Department of Neuroscience, Psychology, Drug Research and Child Health-Neurofarba-Division of Pharmacology and Toxicology, Florence, Italy.

Abstract: Pharmacological tools for chronic visceral pain management are still limited and inadequate. A3 adenosine receptor (A3AR) agonists are effective in different models of persistent pain. Recently, their activity has been related to the block of N-type voltage-gated Ca2+ channels (Cav2.2) in dorsal root ganglia (DRG) neurons. The present work aimed to evaluate the efficacy of A3AR agonists in reducing postinflammatory visceral hypersensitivity in both male and female rats. Colitis was induced by the intracolonic instillation of 2,4-dinitrobenzenesulfonic acid (DNBS; 30 mg in 0.25 mL 50% EtOH). Visceral hypersensitivity was assessed by measuring the visceromotor response and the abdominal withdrawal reflex to colorectal distension. The effects of A3AR agonists (MRS5980 and Cl-IB-MECA) were evaluated over time after DNBS injection and compared to that of the selective Cav2.2 blocker PD173212, and the clinically used drug linaclotide. A3AR agonists significantly reduced DNBS-evoked visceral pain both in the postinflammatory (14 and 21 days after DNBS injection) and persistence (28 and 35 days after DNBS) phases. Efficacy was comparable to effects induced by linaclotide. PD173212 fully reduced abdominal hypersensitivity to control values, highlighting the role of Cav2.2. The effects of MRS5980 and Cl-IB-MECA were completely abolished by the selective A3AR antagonist MRS1523. Furthermore, patch-clamp recordings showed that A3AR agonists inhibited Cav2.2 in dorsal root ganglia neurons isolated from either control or DNBS-treated rats. The effect on Ca2+ current was PD173212-sensitive and prevented by MRS1523. A3AR agonists are effective in relieving visceral hypersensitivity induced by DNBS, suggesting a potential therapeutic role against abdominal pain.
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http://dx.doi.org/10.1097/j.pain.0000000000001905DOI Listing
September 2020

Adenosine A receptors inhibit K currents and cell differentiation in cultured oligodendrocyte precursor cells and modulate sphingosine-1-phosphate signaling pathway.

Biochem Pharmacol 2020 07 3;177:113956. Epub 2020 Apr 3.

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), Section of Pharmacology and Toxicology, University of Florence, Italy.

Oligodendrocytes are the only myelinating cells in the brain and differentiate from their progenitors (OPCs) throughout adult life. However, this process fails in demyelinating pathologies. Adenosine is emerging as an important player in OPC differentiation and we recently demonstrated that adenosine A receptors inhibit cell maturation by reducing voltage-dependent K currents. No data are available to date about the A receptor (AR) subtype. The bioactive lipid mediator sphingosine-1-phosphate (S1P) and its receptors (S1P) are also crucial modulators of OPC development. An interaction between this pathway and the AR is reported in peripheral cells. We studied the role of ARs in modulating K currents and cell differentiation in OPC cultures and we investigated a possible interplay with S1P signaling. Our data indicate that the AR agonist BAY60-6583 and its new analogue P453 inhibit K currents in cultured OPC and the effect was prevented by the AR antagonist MRS1706, by K channel blockers and was differently modulated by the S1P analogue FTY720-P. An acute (10 min) exposure of OPCs to BAY60-6583 also increased the phosphorylated form of sphingosine kinase 1 (SphK1). A chronic (7 days) treatment with the same agonist decreased OPC differentiation whereas SphK1/2 inhibition exerted the opposite effect. Furthermore, AR was overexpressed during OPC differentiation, an effect prevented by the pan SphK1/2 inhibitor VPC69047. Finally, AR silenced cells showed increased cell maturation, decreased SphK1 expression and enhanced S1P lyase levels. We conclude that ARs inhibit K currents and cell differentiation and positively modulate S1P synthesis in cultured OPCs.
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http://dx.doi.org/10.1016/j.bcp.2020.113956DOI Listing
July 2020

Functional characterization of a novel adenosine A receptor agonist on short-term plasticity and synaptic inhibition during oxygen and glucose deprivation in the rat CA1 hippocampus.

Brain Res Bull 2019 09 24;151:174-180. Epub 2019 May 24.

Department of Neuroscience, Psychology, Drug Research and Child Health, NEUROFARBA, Section of Pharmacology and Toxicology, University of Florence, Florence, Italy. Electronic address:

Adenosine is an endogenous neuromodulator exerting its biological functions via four receptor subtypes, A, A, A, and A. A receptors (ARs) are expressed at hippocampal level where they are known to inhibit paired pulse facilitation (PPF), whose reduction reflects an increase in presynaptic glutamate release. The effect of ARs on PPF is known to be sensitive not only to AR blockade but also to the AR antagonist DPCPX, indicating that it involves AR activation. In this study we provide the first functional characterization of the newly synthesized non-nucleoside like AR agonist P453, belonging to the amino-3,5-dicyanopyridine series. By extracellular electrophysiological recordings, we demonstrated that P453 mimicked the effect of the prototypical AR agonist BAY60-6583 in decreasing PPF at Schaffer collateral-CA1 synapses in rat acute hippocampal slices. This effect was prevented by two different AR antagonists, PSB603 and MRS1754, and by the AR antagonist DPCPX. We also investigated the functional role of AR during a 2 min of oxygen and glucose deprivation (OGD) insult, known to produce a reversible fEPSP inhibition due to adenosine AR activation. We found that P453 and BAY60-6583 significantly delayed the onset of fEPSP reduction induced by OGD and the effect was blocked by PSB603. We conclude that P453 is a functional AR agonist whose activation decreases PPF by increasing glutamate release at presynaptic terminals and delays AR-mediated fEPSP inhibition during a 2-minute OGD insult.
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http://dx.doi.org/10.1016/j.brainresbull.2019.05.018DOI Listing
September 2019

Adenosine A3 receptor activation inhibits pronociceptive N-type Ca2+ currents and cell excitability in dorsal root ganglion neurons.

Pain 2019 05;160(5):1103-1118

Division of Pharmacology and Toxicology, Department of NEUROFARBA, University of Florence, Italy.

Recently, studies have focused on the antihyperalgesic activity of the A3 adenosine receptor (A3AR) in several chronic pain models, but the cellular and molecular basis of this effect is still unknown. Here, we investigated the expression and functional effects of A3AR on the excitability of small- to medium-sized, capsaicin-sensitive, dorsal root ganglion (DRG) neurons isolated from 3- to 4-week-old rats. Real-time quantitative polymerase chain reaction experiments and immunofluorescence analysis revealed A3AR expression in DRG neurons. Patch-clamp experiments demonstrated that 2 distinct A3AR agonists, Cl-IB-MECA and the highly selective MRS5980, inhibited Ca-activated K (KCa) currents evoked by a voltage-ramp protocol. This effect was dependent on a reduction in Ca influx via N-type voltage-dependent Ca channels, as Cl-IB-MECA-induced inhibition was sensitive to the N-type blocker PD173212 but not to the L-type blocker, lacidipine. The endogenous agonist adenosine also reduced N-type Ca currents, and its effect was inhibited by 56% in the presence of A3AR antagonist MRS1523, demonstrating that the majority of adenosine's effect is mediated by this receptor subtype. Current-clamp recordings demonstrated that neuronal firing of rat DRG neurons was also significantly reduced by A3AR activation in a MRS1523-sensitive but PD173212-insensitive manner. Intracellular Ca measurements confirmed the inhibitory role of A3AR on DRG neuronal firing. We conclude that pain-relieving effects observed on A3AR activation could be mediated through N-type Ca channel block and action potential inhibition as independent mechanisms in isolated rat DRG neurons. These findings support A3AR-based therapy as a viable approach to alleviate pain in different pathologies.
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http://dx.doi.org/10.1097/j.pain.0000000000001488DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6669900PMC
May 2019

A Selective Histamine H Receptor Antagonist, JNJ7777120, Is Protective in a Rat Model of Transient Cerebral Ischemia.

Front Pharmacol 2018 29;9:1231. Epub 2018 Oct 29.

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), Division of Pharmacology and Toxicology, University of Florence, Florence, Italy.

Cerebral ischemia is a multifactorial pathology characterized by different events evolving in time. The acute injury, characterized by excitoxicity, is followed by a secondary brain injury that develops from hours to days after ischemia. Extracellular levels of histamine increase in the ischemic area after focal cerebral ischemia induced by occlusion of the middle cerebral artery (MCAo). The histamine H receptor (HR) is predominantly expressed in cell types of immune system where is involved in the regulation of immunological and inflammatory responses, and in numerous area of the Central Nervous System (CNS) including cortex and striatum. Our aim was to assess the putative neuroprotective effects of the potent and selective HR antagonist, JNJ7777120 (JNJ), chronically administered (1 mg/kg, i.p., twice/day for 7 days) on damage parameters in a rat model of focal ischemia induced by transient MCAo (tMCAo). Chronic treatment with the HR antagonist JNJ, significantly protected from the neurological deficit and from body weight loss after tMCAo. Seven days after the ischemic insult, JNJ reduced the volume of the ischemic cortical and striatal damage, the number of activated microglia and astrocytes in the ischemic cortex and striatum and decreased the plasma levels of IL-1β and TNF-α, while increased the levels of IL-10. Two days after ischemia, JNJ has reduced granulocyte infiltration in the ischemic area. Results demonstrate that the selective antagonist of HR, JNJ, systemically and chronically administered after ischemia, reduces the ischemic brain damage, improves the neurological deficit and decreases blood pro-inflammatory cytokines, suggesting that HR is a valuable pharmacological target after focal brain ischemia.
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http://dx.doi.org/10.3389/fphar.2018.01231DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6215858PMC
October 2018

The Selective Antagonism of Adenosine A Receptors Reduces the Synaptic Failure and Neuronal Death Induced by Oxygen and Glucose Deprivation in Rat CA1 Hippocampus .

Front Pharmacol 2018 24;9:399. Epub 2018 Apr 24.

Department of Neuroscience, Psychology, Drug Research and Child Health, NEUROFARBA, Section of Pharmacology and Toxicology, University of Florence, Florence, Italy.

Ischemia is a multifactorial pathology characterized by different events evolving in time. Immediately after the ischemic insult, primary brain damage is due to the massive increase of extracellular glutamate. Adenosine in the brain increases dramatically during ischemia in concentrations able to stimulate all its receptors, A, A, A, and A. Although adenosine exerts clear neuroprotective effects through A receptors during ischemia, the use of selective A receptor agonists is hampered by their undesirable peripheral side effects. So far, no evidence is available on the involvement of adenosine A receptors in cerebral ischemia. This study explored the role of adenosine A receptors on synaptic and cellular responses during oxygen and glucose deprivation (OGD) in the CA1 region of rat hippocampus . We conducted extracellular recordings of CA1 field excitatory post-synaptic potentials (fEPSPs); the extent of damage on neurons and glia was assessed by immunohistochemistry. Seven min OGD induced anoxic depolarization (AD) in all hippocampal slices tested and completely abolished fEPSPs that did not recover after return to normoxic condition. Seven minutes OGD was applied in the presence of the selective adenosine A receptor antagonists MRS1754 (500 nM) or PSB603 (50 nM), separately administered 15 min before, during and 5 min after OGD. Both antagonists were able to prevent or delay the appearance of AD and to modify synaptic responses after OGD, allowing significant recovery of neurotransmission. Adenosine A receptor antagonism also counteracted the reduction of neuronal density in CA1 stratum pyramidale, decreased apoptosis at least up to 3 h after the end of OGD, and maintained activated mTOR levels similar to those of controls, thus sparing neurons from the degenerative effects caused by the simil-ischemic conditions. Astrocytes significantly proliferated in CA1 stratum radiatum already 3 h after the end of OGD, possibly due to increased glutamate release. Areceptor antagonism significantly prevented astrocyte modifications. Both A receptor antagonists did not protect CA1 neurons from the neurodegeneration induced by glutamate application, indicating that the antagonistic effect is upstream of glutamate release. The selective antagonists of the adenosine A receptor subtype may thus represent a new class of neuroprotective drugs in ischemia.
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http://dx.doi.org/10.3389/fphar.2018.00399DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5928446PMC
April 2018

From the 7th Joint Italian-German Purine Club Meeting to European Purine Club Meetings.

Authors:
Felicita Pedata

Purinergic Signal 2017 12;13(4):683-685

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), University of Florence, Viale Pieraccini, 6, 50139, Florence, Italy.

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http://dx.doi.org/10.1007/s11302-017-9580-5DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5714843PMC
December 2017

The neuron-astrocyte-microglia triad in CA3 after chronic cerebral hypoperfusion in the rat: Protective effect of dipyridamole.

Exp Gerontol 2017 10 9;96:46-62. Epub 2017 Jun 9.

Department of Health Sciences, Section of Pharmacology and Clinical Oncology, University of Florence, Viale Pieraccini 6, 50139 Firenze, Italy. Electronic address:

We investigated the quantitative and morphofunctional alterations of neuron-astrocyte-microglia triads in CA3 hippocampus, in comparison to CA1, after 2 Vessel Occlusion (2VO) and the protective effect of dipyridamole. We evaluated 3 experimental groups: sham-operated rats (sham, n=15), 2VO-operated rats treated with vehicle (2VO-vehicle, n=15), and 2VO-operated rats treated with dipyridamole from day 0 to day 7 (2VO-dipyridamole, n=15), 90days after 2VO. We analyzed Stratum Pyramidalis (SP), Stratum Lucidum (SL) and Stratum Radiatum (SR) of CA3. 1) ectopic neurons increased in SL and SR of 2VO-vehicle, and 2VO-dipyridamole rats; 2) apoptotic neurons increased in SP of 2VO-vehicle rats and dipyridamole reverted this effect; 3) astrocytes increased in SP, SL and SR of 2VO-vehicle and 2VO-dipyridamole rats; 4) TNF-α expression increased in astrocytes, blocked by dipyridamole, and in dendrites in SR of 2VO-vehicle rats; 5) total microglia increased in SL and SR of 2VO-vehicle and 2VO-dipyridamole rats; 6) triads increased in SR of 2VO-vehicle rats and dipyridamole reverted this effect. Microglia cooperated with astrocytes to phagocytosis of apoptotic neurons and debris, and engulfed ectopic non-fragmented neurons in SL of 2VO-vehicle and 2VO-dipyridamole rats, through a new mechanism called phagoptosis. CA3 showed a better adaptive capacity than CA1 to the ischemic insult, possibly due to the different behaviour of astrocytes and microglial cells. Dipyridamole had neuroprotective effects.
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http://dx.doi.org/10.1016/j.exger.2017.06.006DOI Listing
October 2017

Equilibrative nucleoside transporter ENT1 as a biomarker of Huntington disease.

Neurobiol Dis 2016 Dec 24;96:47-53. Epub 2016 Aug 24.

Integrative Neurobiology Section, National Institute on Drug Abuse, Intramural Research Program, National Institutes of Health, Baltimore, MD 21224, United States. Electronic address:

The initial goal of this study was to investigate alterations in adenosine A receptor (AR) density or function in a rat model of Huntington disease (HD) with reported insensitivity to an AR antagonist. Unsuspected negative results led to the hypothesis of a low striatal adenosine tone and to the search for the mechanisms involved. Extracellular striatal concentrations of adenosine were measured with in vivo microdialysis in two rodent models of early neuropathological stages of HD disease, the Tg51 rat and the zQ175 knock-in mouse. In view of the crucial role of the equilibrative nucleoside transporter (ENT1) in determining extracellular content of adenosine, the binding properties of the ENT1 inhibitor [H]-S-(4-Nitrobenzyl)-6-thioinosine were evaluated in zQ175 mice and the differential expression and differential coexpression patterns of the ENT1 gene (SLC29A1) were analyzed in a large human cohort of HD disease and controls. Extracellular striatal levels of adenosine were significantly lower in both animal models as compared with control littermates and striatal ENT1 binding sites were significantly upregulated in zQ175 mice. ENT1 transcript was significantly upregulated in HD disease patients at an early neuropathological severity stage, but not those with a higher severity stage, relative to non-demented controls. ENT1 transcript was differentially coexpressed (gained correlations) with several other genes in HD disease subjects compared to the control group. The present study demonstrates that ENT1 and adenosine constitute biomarkers of the initial stages of neurodegeneration in HD disease and also predicts that ENT1 could constitute a new therapeutic target to delay the progression of the disease.
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http://dx.doi.org/10.1016/j.nbd.2016.08.013DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5102769PMC
December 2016

Imidazo[1,2-a]pyrazin-8-amine core for the design of new adenosine receptor antagonists: Structural exploration to target the A and A subtypes.

Eur J Med Chem 2017 Jan 26;125:611-628. Epub 2016 Sep 26.

Dipartimento di Neuroscienze, Psicologia, Area del Farmaco e Salute del Bambino, Sez. Farmaceutica e Nutraceutica, Universita' degli Studi di Firenze, Via Ugo Schiff 6, 50019, Sesto Fiorentino (FI), Italy.

The imidazo[1,2-a]pyrazine ring system has been chosen as a new decorable core skeleton for the design of novel adenosine receptor (AR) antagonists targeting either the human (h) A or the hA receptor subtype. The N-(hetero)arylcarboxyamido substituted compounds 4-14 and 21-30, bearing a 6-phenyl moiety or not, respectively, show good hA receptor affinity and selectivity versus the other ARs. In contrast, the 8-amino-6-(hetero)aryl substituted derivatives designed for targeting the hA receptor subtype (compounds 31-38) and also the 6-phenyl analogues 18-20 do not bind the hA AR, or show hA or balanced hA/hA AR affinity in the micromolar range. Molecular docking of the new hA antagonists was carried out to depict their hypothetical binding mode to our refined model of the hA receptor. Some derivatives were evaluated for their fluorescent potentiality and showed some fluorescent emission properties. One of the most active hA antagonists herein reported, i.e. the 2,6-diphenyl-8-(3-pyridoylamino)imidazo[1,2-a]pyrazine 29, tested in a rat model of cerebral ischemia, delayed the occurrence of anoxic depolarization caused by oxygen and glucose deprivation in the hippocampus and allowed disrupted synaptic activity to recover.
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http://dx.doi.org/10.1016/j.ejmech.2016.09.076DOI Listing
January 2017

Purinergic signalling in brain ischemia.

Neuropharmacology 2016 05 12;104:105-30. Epub 2015 Nov 12.

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), University of Florence, Viale Pieraccini, 6, 50139 Florence, Italy.

Ischemia is a multifactorial pathology characterized by different events evolving in the time. After ischemia a primary damage due to the early massive increase of extracellular glutamate is followed by activation of resident immune cells, i.e microglia, and production or activation of inflammation mediators. Protracted neuroinflammation is now recognized as the predominant mechanism of secondary brain injury progression. Extracellular concentrations of ATP and adenosine in the brain increase dramatically during ischemia in concentrations able to stimulate their respective specific P2 and P1 receptors. Both ATP P2 and adenosine P1 receptor subtypes exert important roles in ischemia. Although adenosine exerts a clear neuroprotective effect through A1 receptors during ischemia, the use of selective A1 agonists is hampered by undesirable peripheral effects. Evidence up to now in literature indicate that A2A receptor antagonists provide protection centrally by reducing excitotoxicity, while agonists at A2A (and possibly also A2B) and A3 receptors provide protection by controlling massive infiltration and neuroinflammation in the hours and days after brain ischemia. Among P2X receptors most evidence indicate that P2X7 receptor contribute to the damage induced by the ischemic insult due to intracellular Ca(2+) loading in central cells and facilitation of glutamate release. Antagonism of P2X7 receptors might represent a new treatment to attenuate brain damage and to promote proliferation and maturation of brain immature resident cells that can promote tissue repair following cerebral ischemia. Among P2Y receptors, antagonists of P2Y12 receptors are of value because of their antiplatelet activity and possibly because of additional anti-inflammatory effects. Moreover strategies that modify adenosine or ATP concentrations at injury sites might be of value to limit damage after ischemia. This article is part of the Special Issue entitled 'Purines in Neurodegeneration and Neuroregeneration'.
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http://dx.doi.org/10.1016/j.neuropharm.2015.11.007DOI Listing
May 2016

Role of adenosine in oligodendrocyte precursor maturation.

Front Cell Neurosci 2015 24;9:155. Epub 2015 Apr 24.

Department NEUROFARBA, Division of Pharmacology and Toxicology, University of Florence Florence, Italy.

Differentiation and maturation of oligodendroglial cells are postnatal processes that involve specific morphological changes correlated with the expression of stage-specific surface antigens and functional voltage-gated ion channels. A small fraction of oligodendrocyte progenitor cells (OPCs) generated during development are maintained in an immature and slowly proliferative or quiescent state in the adult central nervous system (CNS) representing an endogenous reservoir of immature cells. Adenosine receptors are expressed by OPCs and a key role of adenosine in oligodendrocyte maturation has been recently recognized. As evaluated on OPC cultures, adenosine, by stimulating A1 receptors, promotes oligodendrocyte maturation and inhibits their proliferation; on the contrary, by stimulating A2A receptors, it inhibits oligodendrocyte maturation. A1 and A2A receptor-mediated effects are related to opposite modifications of outward delayed rectifying membrane K(+) currents (IK) that are involved in the regulation of oligodendrocyte differentiation. Brain A1 and A2A receptors might represent new molecular targets for drugs useful in demyelinating pathologies, such as multiple sclerosis (MS), stroke and brain trauma.
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http://dx.doi.org/10.3389/fncel.2015.00155DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4408841PMC
May 2015

Time-course of protection by the selective A2A receptor antagonist SCH58261 after transient focal cerebral ischemia.

Neurol Sci 2015 Aug 25;36(8):1441-8. Epub 2015 Mar 25.

Division of Pharmacology and Toxicology, Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), University of Florence, Viale Pieraccini, 6, 50139, Florence, Italy,

Evidence indicates that the adenosine A2A receptor subtype is of critical importance in stroke. In previous studies, in the model of permanent middle cerebral artery occlusion (pMCAo), the adenosine A2A receptor antagonist, SCH58261, administered soon after ischemia, proved protective against excessive glutamate outflow in the first 4 h after ischemia and against neurological deficit and tissue damage evaluated 24 h after pMCAo. In the present work, we investigated if neuroprotective effect of SCH58261 was maintained 7 days after transient MCAo (tMCAo). SCH58261 (0.01 mg/kg, i.p.), administered twice/day for 7 days, protected from neurological deficit 1 day after tMCAo, but no more after 5 and 7 days. Two days after tMCAo, SCH58261 did not reduce blood cell infiltration, evaluated as HIS-48 positive cells, into ischemic striatal and cortical tissue. Moreover, 7 days after tMCAo, SCH58261 has not protected ischemic areas from damage and has not ameliorated myelin organization into the ischemic striatum. Protection by the A2A receptor antagonist 24 h after ischemia is attributable to reduced excitotoxicity. Seven days after ischemia the early protective effect of the A2A receptor antagonist likely has been overwhelmed by a secondary damage due to blood cell infiltration and neuroinflammation.
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http://dx.doi.org/10.1007/s10072-015-2160-yDOI Listing
August 2015

The selective antagonism of P2X7 and P2Y1 receptors prevents synaptic failure and affects cell proliferation induced by oxygen and glucose deprivation in rat dentate gyrus.

PLoS One 2014 19;9(12):e115273. Epub 2014 Dec 19.

Dept. of Neuroscience, Psychology, Drug Research and Child Health, NEUROFARBA, Division of Pharmacology and Toxicology, University of Florence, Florence, Italy.

Purinergic P2X and P2Y receptors are broadly expressed on both neurons and glial cells in the central nervous system (CNS), including dentate gyrus (DG). The aim of this research was to determine the synaptic and proliferative response of the DG to severe oxygen and glucose deprivation (OGD) in acute rat hippocampal slices and to investigate the contribution of P2X7 and P2Y1 receptor antagonism to recovery of synaptic activity after OGD. Extracellular field excitatory post-synaptic potentials (fEPSPs) in granule cells of the DG were recorded from rat hippocampal slices. Nine-min OGD elicited an irreversible loss of fEPSP and was invariably followed by the appearance of anoxic depolarization (AD). Application of MRS2179 (selective antagonist of P2Y1 receptor) and BBG (selective antagonist of P2X7 receptor), before and during OGD, prevented AD appearance and allowed a significant recovery of neurotransmission after 9-min OGD. The effects of 9-min OGD on proliferation and maturation of cells localized in the subgranular zone (SGZ) of slices prepared from rats treated with 5-Bromo-2'-deoxyuridine (BrdU) were investigated. Slices were further incubated with an immature neuron marker, doublecortin (DCX). The number of BrdU+ cells in the SGZ was significantly decreased 6 hours after OGD. This effect was antagonized by BBG, but not by MRS2179. Twenty-four hours after 9-min OGD, the number of BrdU+ cells returned to control values and a significant increase of DCX immunofluorescence was observed. This phenomenon was still evident when BBG, but not MRS2179, was applied during OGD. Furthermore, the P2Y1 antagonist reduced the number of BrdU+ cells at this time. The data demonstrate that P2X7 and P2Y1 activation contributes to early damage induced by OGD in the DG. At later stages after the insult, P2Y1 receptors might play an additional and different role in promoting cell proliferation and maturation in the DG.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0115273PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4272279PMC
December 2015

The neuron-astrocyte-microglia triad in a rat model of chronic cerebral hypoperfusion: protective effect of dipyridamole.

Front Aging Neurosci 2014 27;6:322. Epub 2014 Nov 27.

Section of Pharmacology and Clinical Oncology, Department of Health Sciences, University of Florence Florence, Italy.

Chronic cerebral hypoperfusion during aging may cause progressive neurodegeneration as ischemic conditions persist. Proper functioning of the interplay between neurons and glia is fundamental for the functional organization of the brain. The aim of our research was to study the pathophysiological mechanisms, and particularly the derangement of the interplay between neurons and astrocytes-microglia with the formation of "triads," in a model of chronic cerebral hypoperfusion induced by the two-vessel occlusion (2VO) in adult Wistar rats (n = 15). The protective effect of dipyridamole given during the early phases after 2VO (4 mg/kg/day i.v., the first 7 days after 2VO) was verified (n = 15). Sham-operated rats (n = 15) were used as controls. Immunofluorescent triple staining of neurons (NeuN), astrocytes (GFAP), and microglia (IBA1) was performed 90 days after 2VO. We found significantly higher amount of "ectopic" neurons, neuronal debris and apoptotic neurons in CA1 Str. Radiatum and Str. Pyramidale of 2VO rats. In CA1 Str. Radiatum of 2VO rats the amount of astrocytes (cells/mm(2)) did not increase. In some instances several astrocytes surrounded ectopic neurons and formed a "micro scar" around them. Astrocyte branches could infiltrate the cell body of ectopic neurons, and, together with activated microglia cells formed the "triads." In the triad, significantly more numerous in CA1 Str. Radiatum of 2VO than in sham rats, astrocytes and microglia cooperated in the phagocytosis of ectopic neurons. These events might be common mechanisms underlying many neurodegenerative processes. The frequency to which they appear might depend upon, or might be the cause of, the burden and severity of neurodegeneration. Dypiridamole significantly reverted all the above described events. The protective effect of chronic administration of dipyridamole might be a consequence of its vasodilatory, antioxidant and anti-inflammatory role during the early phases after 2VO.
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http://dx.doi.org/10.3389/fnagi.2014.00322DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4245920PMC
December 2014

Adenosine receptors in cerebral ischemia.

Int Rev Neurobiol 2014 ;119:309-48

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), Division of Pharmacology and Toxicology, University of Florence, 50139 Florence, Italy.

Ischemic stroke is a complex pathology characterized by a sequence of events that evolve over time and space. It is the second leading cause of death and the main cause of adult long-term disability in developed countries. At the moment, there is no promising pharmacotherapy for acute ischemic stroke. Adenosine receptors (A1, A2A, A2B, A3) are important targets for therapeutic implementation in the treatment of stroke because extracellular adenosine concentrations increase dramatically soon after ischemia. Adenosine receptors located both on central nervous system cells and on immune blood cells exert important roles during ischemia. The neuroprotective role of adenosine through A1 receptor subtype during ischemia is accepted, but the use of selective A1 agonists is hampered by undesirable side effects such as sedation, bradycardia, and hypotension. Recently, the A2A receptor subtype emerged as a potential therapeutic attractive target in ischemia. Evidence suggests that A2A receptor has dual role: in a first phase of ischemia, it potentiates excitotoxicity, while hours and days after ischemia, A2A receptors on immune blood cells potentiate cell adhesion mechanisms and infiltration in the ischemic parenchyma. Consistently, the use of A2A receptor agonists/antagonists (administered at doses that do not modify blood pressure and heart rate) should be carefully evaluated in function of time after ischemia. Although much is still to be known about the role of A2B and A3 receptor subtypes in brain ischemia, most consistent information indicates their role in regulation of immunosuppression and inflammation.
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http://dx.doi.org/10.1016/B978-0-12-801022-8.00013-1DOI Listing
May 2015

Adenosine A2A receptors modulate acute injury and neuroinflammation in brain ischemia.

Mediators Inflamm 2014 5;2014:805198. Epub 2014 Aug 5.

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), University of Florence, Viale Pieraccini 6, 50139 Florence, Italy.

The extracellular concentration of adenosine in the brain increases dramatically during ischemia. Adenosine A(2A) receptor is expressed in neurons and glial cells and in inflammatory cells (lymphocytes and granulocytes). Recently, adenosine A(2A) receptor emerged as a potential therapeutic attractive target in ischemia. Ischemia is a multifactorial pathology characterized by different events evolving in the time. After ischemia the early massive increase of extracellular glutamate is followed by activation of resident immune cells, that is, microglia, and production or activation of inflammation mediators. Proinflammatory cytokines, which upregulate cell adhesion molecules, exert an important role in promoting recruitment of leukocytes that in turn promote expansion of the inflammatory response in ischemic tissue. Protracted neuroinflammation is now recognized as the predominant mechanism of secondary brain injury progression. A(2A) receptors present on central cells and on blood cells account for important effects depending on the time-related evolution of the pathological condition. Evidence suggests that A(2A) receptor antagonists provide early protection via centrally mediated control of excessive excitotoxicity, while A(2A) receptor agonists provide protracted protection by controlling massive blood cell infiltration in the hours and days after ischemia. Focus on inflammatory responses provides for adenosine A(2A) receptor agonists a wide therapeutic time-window of hours and even days after stroke.
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http://dx.doi.org/10.1155/2014/805198DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4138795PMC
May 2015

Low doses of the selective adenosine A2A receptor agonist CGS21680 are protective in a rat model of transient cerebral ischemia.

Brain Res 2014 Mar 20;1551:59-72. Epub 2014 Jan 20.

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), Division of Pharmacology and Toxicology, University of Florence, Viale Pieraccini 6, 50139 Florence, Italy. Electronic address:

Evidence indicate that adenosine A2A receptor subtype is of critical importance in stroke. An overexpression of A2A adenosine receptors occurs at central level on neurons and microglia of ischemic striatum and cortex after focal ischemia. Adenosine A2A receptor subtype is localized not only at central level but also peripherally on blood cells, where it is known to exert antiinflammatory effect. Purpose of the present work was to investigate the putative neuroprotective effect of the adenosine A2A receptor agonist CGS21680 in a rat model of transient medial cerebral artery occlusion (MCAo). Transient cerebral ischemia was induced by 1h occlusion of MCA. CGS21680 (0.01 and 0.1mg/kg, i.p.) was administered starting 4h after ischemia according to a chronic protocol (twice/day for 7 days). CGS21680, at the dose of 0.1mg/kg transiently increased heart frequency but did not modify blood pressure. At the dose of 0.01mg/kg the drug did not modify either heart frequency or blood pressure. Following transient MCAo, CGS21680 at both doses protected from neurological deficit from the first day up to 7 days thereafter. At this time, it has reduced microgliosis, astrogliosis and improved myelin organization in the striatum and cytoarchitecture of the ischemic cortex and striatum. Two days after transient MCAo, CGS21680 has reduced the number of infiltrated granulocytes into the ischemic tissue. Data indicate that CGS21680 systemically administered is protective by immunosuppressive effects.
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http://dx.doi.org/10.1016/j.brainres.2014.01.014DOI Listing
March 2014

Adenosine is present in rat brain synaptic vesicles.

Neuroreport 2013 Dec;24(17):982-7

Department of Neurosciences, Psychology, Drug Research and Child Health (NEUROFARBA), Division of Pharmacology and Toxicology, University of Florence, Florence, Italy.

Evidences in the central nervous system are in favor that adenosine under basal conditions is released by a direct excitation-secretion modality. However, till now, there is no direct evidence that adenosine is contained in synaptic vesicles. Eight synaptic vesicle fractions were recovered on a discontinuous sucrose gradient after ultracentrifugation of the crude synaptosomal fraction (pellet P2) of rat brain. The adenosine content in each fraction was measured by high-performance liquid chromatography coupled to a fluorescence detector (minimum sensitivity 10 femtomoles). The immunoblot analysis, to detect synaptophysin, a molecular marker for the vesicle membrane, showed that fractions from 3 to 8 were rich in synaptophysin. The sum of adenosine found in fractions 3-8 was (mean ± SEM, n = 4) 3325.6 ± 94.6 pmol/mg of tissue protein. We proved that adenosine measured in synaptic vesicle fractions was not contaminated by cytosolic adenosine, as adenosine exogenously added to the P2 preferentially distributed in fractions 1 and 2 that are synaptophysin-free and did not contaminate the vesicle pellet P3. Data provide direct demonstration that adenosine is present in rat brain synaptic vesicle fractions. This information is consistent with the notion that adenosine is stored in synaptic vesicles and is released under normoxic physiological conditions by an excitation-secretion mechanism typical of neuronal cells.
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http://dx.doi.org/10.1097/WNR.0000000000000033DOI Listing
December 2013

Adenosine A₂A receptors inhibit delayed rectifier potassium currents and cell differentiation in primary purified oligodendrocyte cultures.

Neuropharmacology 2013 Oct 13;73:301-10. Epub 2013 Jun 13.

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), Division of Pharmacology and Toxicology, University of Florence, Viale Pieraccini 6, 50139 Florence, Italy.

Oligodendrocyte progenitor cells (OPCs) are a population of cycling cells which persist in the adult central nervous system (CNS) where, under opportune stimuli, they differentiate into mature myelinating oligodendrocytes. Adenosine A(2A) receptors are Gs-coupled P1 purinergic receptors which are widely distributed throughout the CNS. It has been demonstrated that OPCs express A(2A) receptors, but their functional role in these cells remains elusive. Oligodendrocytes express distinct voltage-gated ion channels depending on their maturation. Here, by electrophysiological recordings coupled with immunocytochemical labeling, we studied the effects of adenosine A(2A) receptors on membrane currents and differentiation of purified primary OPCs isolated from the rat cortex. We found that the selective A(2A) agonist, CGS21680, inhibits sustained, delayed rectifier, K(+) currents (I(K)) without modifying transient (I(A)) conductances. The effect was observed in all cells tested, independently from time in culture. CGS21680 inhibition of I(K) current was concentration-dependent (10-200 nM) and blocked in the presence of the selective A(2A) antagonist SCH58261 (100 nM). It is known that I(K) currents play an important role during OPC development since their block decreases cell proliferation and differentiation. In light of these data, our further aim was to investigate whether A(2A) receptors modulate these processes. CGS21680, applied at 100 nM in the culture medium of oligodendrocyte cultures, inhibits OPC differentiation (an effect prevented by SCH58261) without affecting cell proliferation. Data demonstrate that cultured OPCs express functional A(2A) receptors whose activation negatively modulate I(K) currents. We propose that, by this mechanism, A(2A) adenosine receptors inhibit OPC differentiation.
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http://dx.doi.org/10.1016/j.neuropharm.2013.05.035DOI Listing
October 2013

UDP-glucose enhances outward K(+) currents necessary for cell differentiation and stimulates cell migration by activating the GPR17 receptor in oligodendrocyte precursors.

Glia 2013 Jul 2;61(7):1155-71. Epub 2013 May 2.

Divi Department of Neuroscience, Psychology, Drug Research and Child Health (NeuroFarBa), University of Florence, Viale Pieraccini, 6, 50139 Florence, Italy.

In the developing and mature central nervous system, NG2 expressing cells comprise a population of cycling oligodendrocyte progenitor cells (OPCs) that differentiate into mature, myelinating oligodendrocytes (OLGs). OPCs are also characterized by high motility and respond to injury by migrating into the lesioned area to support remyelination. K(+) currents in OPCs are developmentally regulated during differentiation. However, the mechanisms regulating these currents at different stages of oligodendrocyte lineage are poorly understood. Here we show that, in cultured primary OPCs, the purinergic G-protein coupled receptor GPR17, that has recently emerged as a key player in oligodendrogliogenesis, crucially regulates K(+) currents. Specifically, receptor stimulation by its agonist UDP-glucose enhances delayed rectifier K(+) currents without affecting transient K(+) conductances. This effect was observed in a subpopulation of OPCs and immature pre-OLGs whereas it was absent in mature OLGs, in line with GPR17 expression, that peaks at intermediate phases of oligodendrocyte differentiation and is thereafter downregulated to allow terminal maturation. The effect of UDP-glucose on K(+) currents is concentration-dependent, blocked by the GPR17 antagonists MRS2179 and cangrelor, and sensitive to the K(+) channel blocker tetraethyl-ammonium, which also inhibits oligodendrocyte maturation. We propose that stimulation of K(+) currents is responsible for GPR17-induced oligodendrocyte differentiation. Moreover, we demonstrate, for the first time, that GPR17 activation stimulates OPC migration, suggesting an important role for this receptor after brain injury. Our data indicate that modulation of GPR17 may represent a strategy to potentiate the post-traumatic response of OPCs under demyelinating conditions, such as multiple sclerosis, stroke, and brain trauma.
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http://dx.doi.org/10.1002/glia.22506DOI Listing
July 2013

Amyloid-β oligomer synaptotoxicity is mimicked by oligomers of the model protein HypF-N.

Neurobiol Aging 2013 Sep 18;34(9):2100-9. Epub 2013 Apr 18.

Department of Biomedical, Experimental and Clinical Sciences, University of Florence, Florence, Italy.

Protein misfolded oligomers are thought to be the primary pathogenic species in many protein deposition diseases. Oligomers by the amyloid-β peptide play a central role in Alzheimer's disease pathogenesis, being implicated in synaptic dysfunction. Here we show that the oligomers formed by a protein that has no link with human disease, namely the N-terminal domain of HypF from Escherichia coli (HypF-N), are also synaptotoxic. HypF-N oligomers were found to (i) colocalize with post-synaptic densities in primary rat hippocampal neurons; (ii) induce impairment of long-term potentiation in rat hippocampal slices; and (iii) impair spatial learning of rats in the Morris Water Maze test. By contrast, the native protein and control nontoxic oligomers had none of such effects. These results raise the importance of using HypF-N oligomers as a valid tool to investigate the pathogenesis of Alzheimer's disease, with advantages over other systems for their stability, reproducibility, and costs. The results also suggest that, in the context of a compromised protein homeostasis resulting from aggregation of the amyloid β peptide, a number of oligomeric species sharing common synaptotoxic activity can arise and cooperate in the pathogenesis of the disease.
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http://dx.doi.org/10.1016/j.neurobiolaging.2013.03.020DOI Listing
September 2013

Effects of oxygen and glucose deprivation on synaptic transmission in rat dentate gyrus: role of A2A adenosine receptors.

Neuropharmacology 2013 Apr 19;67:511-20. Epub 2012 Dec 19.

Department of Preclinical and Clinical Pharmacology, University of Florence, Viale Pieraccini 6, 50139 Florence, Italy.

The hippocampus is comprised of two distinct subfields that show different responses to hypoxic-ischemic brain injury: the CA1 region is particularly susceptible whereas the dentate gyrus (DG) is quite resistant. Our aim was to determine the synaptic and proliferative response of the DG to severe oxygen and glucose deprivation (OGD) in acute rat hippocampal slices and to investigate the contribution of A(2A) adenosine receptor antagonism to recovery of synaptic activity after OGD. Extracellular recordings of field excitatory post-synaptic potentials (fEPSPs) in granule cells of the DG in brain slices prepared from male Wistar rats were used. A 9-min OGD is needed in the DG to always induce the appearance of anoxic depolarization (AD) and the irreversible block of synaptic activity, as recorded up to 24 h from the end of the insult, whereas only 7-min OGD is required in the CA1 region. Selective antagonism of A(2A) adenosine receptors by ZM241385 significantly prevents or delays the appearance of AD and protects from the irreversible block of neurotransmission induced by 9-min OGD in the DG. The effects of 9-min OGD on proliferation and maturation of cells localized in the subgranular zone of DG in slices prepared from 5-bromo-2'-deoxyuridine (BrdU) treated rats was investigated. Slices were further incubated with an immature neuronal marker, doublecortin (DCX). The number of BrdU(+) cells was significantly decreased 6 h after 9-min OGD and this effect was antagonized by ZM241385. After 24 h from the end of 9-min OGD, the number of BrdU(+) cells returned to that found before OGD and increased arborization of tertiary dendrites of DCX(+) cells was observed. The adenosine A(2A) antagonist ZM241385 protects from synaptic failure and from decreased proliferation of immature neuronal cells at a precocious time after OGD.
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http://dx.doi.org/10.1016/j.neuropharm.2012.12.002DOI Listing
April 2013

3-Hydroxy-1H-quinazoline-2,4-dione derivatives as new antagonists at ionotropic glutamate receptors: molecular modeling and pharmacological studies.

Eur J Med Chem 2012 Aug 4;54:470-82. Epub 2012 Jun 4.

Dipartimento di Scienze Farmaceutiche, Laboratorio di Progettazione, Sintesi e Studio di Eterocicli Biologicamente Attivi, Universita' di Firenze, Polo Scientifico, Via Ugo Schiff 6, 50019 Sesto Fiorentino, FI, Italy.

Based on our 3-hydroxy-7-chloroquinazoline-2,4-dione derivatives, previously reported as antagonists at ionotropic glutamate receptors, we synthesized new 3-hydroxyquinazoline-2,4-diones bearing a trifluoromethyl group at the 7-position and different groups at position 6. Glycine/NMDA, AMPA and kainate receptor binding data showed that the 7-trifluoromethyl residue increased AMPA and kainate receptor affinity and selectivity, with respect to the 7-chlorine atom. Among the probed 6-substituents, the 6-(1,2,4-triazol-4-yl) group (compound 8) was the most advantageous for AMPA receptor affinity and selectivity. Derivative 8 demonstrated to be effective in decreasing neuronal damage produced by oxygen and glucose deprivation in organotypic rat hippocampal slices and also showed anticonvulsant effects in pentylenetetrazole-induced convulsions. The previously reported kainate receptor antagonist 6-(2-carboxybenzoyl)-amino-7-chloro-3-hydroxyquinazoline-2,4-dione 3 prevented the failure of neurotransmission induced by oxygen and glucose deprivation in the CA1 region of rat hippocampal slices.
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http://dx.doi.org/10.1016/j.ejmech.2012.05.036DOI Listing
August 2012

Cognitive impairment with vascular impairment and degeneration.

Curr Neurovasc Res 2011 Nov;8(4):342-50

Laboratory of Ischemic and Neurodegenerative Brain Research, Department of Neurodegenerative Disorders, Mossakowski Medical Research Centre, Polish Academy of Sciences, Warsaw, Poland.

Ischemic stroke is a leading cause of death and cognitive impairment worldwide. However, the mechanisms of progressive cognitive decline following brain ischemia are not yet certain. Ongoing interest in cerebrovascular diseases research has provided data showing that Alzheimer's proteins and other factors may be involved in the pathogenesis of gradual ischemic brain injury. Thus, both focal and global brain ischemia in rodents produces a stereotyped pattern of selective neuronal degeneration, which is just the same as in Alzheimer's type dementia. Data from animal models and clinical studies of ischemic stroke have demonstrated an increase in expression and processing of amyloid precursor protein (APP) to a neurotoxic form of oligomeric β-amyloid peptide (Aβ) and hyperphosphorylation of tau protein. The authors of this review are using advances in methods and technologies to study cerebrovascular diseases and this review examines the hypothesis that pathological mechanisms common to both brain ischemia and Alzheimer's dementia are contributing to cognitive impairment and brain ischemia-related dementia.
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http://dx.doi.org/10.2174/156720211798120981DOI Listing
November 2011

Ecto-ATPase inhibition: ATP and adenosine release under physiological and ischemic in vivo conditions in the rat striatum.

Exp Neurol 2012 Jan 6;233(1):193-204. Epub 2011 Oct 6.

Department of Preclinical and Clinical Pharmacology, University of Florence, Viale Pieraccini 6, 50139 Florence, Italy.

In the central nervous system (CNS) ATP and adenosine act as transmitters and neuromodulators on their own receptors but it is still unknown which part of extracellular adenosine derives per se from cells and which part is formed from the hydrolysis of released ATP. In this study extracellular concentrations of adenosine and ATP from the rat striatum were estimated by the microdialysis technique under in vivo physiological conditions and after focal ischemia induced by medial cerebral artery occlusion. Under physiological conditions, adenosine and ATP concentrations were in the range of 130 nmol/L and 40 nmol/L, respectively. In the presence of the novel ecto-ATPase inhibitor, PV4 (100 nmol/L), the extracellular concentration of ATP increased 12-fold to ~360 nmol/L but the adenosine concentration was not altered. This demonstrates that, under physiological conditions, adenosine is not a product of extracellular ATP. In the first 4h after ischemia, adenosine increased to ~690 nmol/L and ATP to ~50 nmol/L. In the presence of PV4 the extracellular concentration of ATP was in the range of 450 nmol/L and a significant decrease in extracellular adenosine (to ~270 nmol/L) was measured. The contribution of extracellular ATP to extracellular adenosine was maximal in the first 20 min after ischemia onset. Furthermore we demonstrated, by immunoelectron microscopy, the presence of the concentrative nucleoside transporter CNT2 on plasma and vesicle membranes isolated from the rat striatum. These results are in favor that adenosine is transported in vesicles and is released in an excitation-secretion manner under in vivo physiological conditions. Early after ischemia, extracellular ATP is hydrolyzed by ecto-nucleotidases which significantly contribute to the increase in extracellular adenosine. To establish the contribution of extracellular ATP to adenosine might constitute the basis for devising a correct putative purinergic strategy aimed at protection from ischemic damage.
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http://dx.doi.org/10.1016/j.expneurol.2011.09.036DOI Listing
January 2012