Neuroprotection by adenosine in the brain: From A1 receptor activation to A2A receptor blockade

Adenosine is a neuromodulator that operates via the most abundant inhibitory adenosine A1 receptors (A1Rs) and the less abundant, but widespread, facilitatory A2ARs. It is commonly assumed that A1Rs play a key role in neuroprotection since they decrease glutamate release and hyperpolarize neurons. In fact, A1R activation at the onset of neuronal injury attenuates brain damage, whereas its blockade exacerbates damage in adult animals. However, there is a down-regulation of central A1Rs in chronic noxious situations. In contrast, A2ARs are up-regulated in noxious brain conditions and their blockade confers robust brain neuroprotection in adult animals. The brain neuroprotective effect of A2AR antagonists is maintained in chronic noxious brain conditions without observable peripheral effects, thus justifying the interest of A2AR antagonists as novel protective agents in neurodegenerative diseases such as Parkinson’s and Alzheimer’s disease, ischemic brain damage and epilepsy. The greater interest of A2AR blockade compared to A1R activation does not mean that A1R activation is irrelevant for a neuroprotective strategy. In fact, it is proposed that coupling A2AR antagonists with strategies aimed at bursting the levels of extracellular adenosine (by inhibiting adenosine kinase) to activate A1Rs might constitute the more robust brain neuroprotective strategy based on the adenosine neuromodulatory system. This strategy should be useful in adult animals and especially in the elderly (where brain pathologies are prevalent) but is not valid for fetus or newborns where the impact of adenosine receptors on brain damage is different

Adenosine A2A receptor antagonism increases striatal glutamate outflow in dopamine-denervated rats

The objective of the work was to study, by in vivo microdialysis, the effect of the adenosine A2A receptor antagonist 7-(2-phenylethyl)-5-amino-2-(2-furyl)-pyrazolo-[4,3-e]-1,2,4-triazolo[1,5-c]pyrimidine (SCH 58261) on glutamate outflow in the striata of unilateral 6-hydroxydopamine-infused rats. Two vertical microdialysis probes were implanted bilaterally in both the denervated striatum and in the intact striatum. Glutamate concentrations in the dialysate were determined by high-performance liquid chromatography (HPLC). Infusion of the adenosine A2A receptor antagonist SCH 58261 (50 nM), through the microdialysis fiber, significantly increased glutamate outflow from the denervated striatum while it decreased glutamate outflow from the intact striatum. The opposite effects of SCH 58261 on glutamate outflow in the intact and 6-hydroxydopamine-lesioned striatum might be attributed to blockade of striatal adenosine A2A receptors located on either striatal indirect output pathways or glutamatergic terminals. These results may be relevant to our understanding of the mechanism of action of adenosine A2A receptor antagonists in Parkinson's diseas

The selective A(2A) receptor antagonist SCH 58261 protects from neurological deficit, brain damage and activation of p38 MAPK in rat focal cerebral ischemia

We investigated the protective effect of subchronic treatment of the A2A receptor antagonist, SCH 58261 (0.01 mg/kg, i.p.), administered 5 min, 6 h and 15 h after permanent right middle cerebral artery occlusion (MCAo). Twenty-four hours after ischemia, an extensive pallid area, evaluated by cresyl violet staining, is evident in the vascular territories supplied by the MCA, the striatum and the sensory motor cortex. The pallid area reflects the extent of necrotic neurons. Soon after waking, rats showed a definite contralateral turning behavior which was significantly reduced by SCH 58261 treatment. Twenty-four hours after MCAo, SCH 58261 significantly improved the neurol. deficit and reduced ischemic damage in the striatum and cortex. Phospho-p38 mitogen-activated protein kinase (MAPK), evaluated by Western Blot, increased by 500% in the ischemic striatum 24 h after MCAo. SCH 58261 treatment significantly reduced phospho-p38 MAPK by 70%. Microglia was immunostained using the OX-42 antibody. Phospho-p38 MAPK and OX-42-immunoreactive cells are localized in the ventral striatum and frontoparietal cortex. Furthermore, both OX-42 and phospho-p38 MAPK-immunoreactive cells have overlapping morphol. features, typical of reactive microglia. SCH 58261 reduced phospho-p38 MAPK immunoreactivity in the striatum and in the cortex without changing the microglial cell morphol. These results indicate that the protective effect of the adenosine antagonist SCH 58261 during ischemia is not due to reduced microglial activation but involves inhibition of phospho-p38 MAPK and suggest that treatment with the A2A antagonist from the first hour to several hours after ischemia may be a useful therapeutic approach in cerebral ischemia

P2X7 receptor modulation on microglial cells and reduction of brain infarct caused by middle cerebral artery occlusion in rat

Adenosine 5'-triphosphate outflow increases after an ischemic insult in the brain and may induce the expression of P2X7 receptors in resting microglia, determining its modification into an activated state. To assess the effects of P2X7 receptor blockade in preventing microglia activation and ameliorating brain damage and neurological impairment, we delivered the P2 unselective antagonist Reactive Blue 2 to rats after middle cerebral artery occlusion. In sham-operated animals, devoid of brain damage, double immunofluorescence verified the absence of P2X7 immunoreactivity on resting microglia, astrocytes, and neurons, identified, respectively, by OX-42, glial fibrillary acid protein, and neuronal nuclei (NeuN) immunoreactivity. After ischemia, vehicle-treated rats showed monolateral sensorimotor deficit and tissue damage in striatum and frontoparietal cortex. Moreover, P2X7 immunoreactivity was de novo expressed on activated microglia in infarcted and surrounding areas, as well as on a reactive form of microglia, resting in shape but P2X7 immunoreactive, present in ipsi- and contralateral cingulate and medial frontal cortex. Reactive Blue 2 improved sensorimotor deficit and restricted the volume of infarction, without preventing the expression of P2X7, but inducing it in the microglia of contralateral frontal and parietal cortex and striatum, which had lost reciprocal connections with the remote infarct area. De novo expression of P2X7 occurred in both activated and reactive microglia, suggesting their differentiated roles in the area of infarct and in remote regions. Reactive Blue 2 reduced ischemic brain damage, likely blocking the function of activated microglia in the infarct area, but in the remote brain regions promoted the expression of P2X7 on reactive microglia, developing defense and reparative processes

The selective A(2A) receptor antagonist SCH 58261 reduces striatal transmitter outflow, turning behavior and ischemic brain damage induced by permanent focal ischemia in the rat

Adenosine A(2A) receptor antagonists have been proved protective in different ischemia models. In this study we verified if the protective effect of the selective A(2A) antagonist, SCH 58261, could be attributed to the reduction of the excitatory amino acid outflow induced by cerebral focal ischemia. A vertical microdialysis probe was inserted into the striatum of male Wistar rats and, after 24 h, permanent right intraluminal middle cerebral artery occlusion (MCAo) was induced. Soon after waking, rats showed a definite contralateral turning behavior, which persisted up to 7 h after MCAo. During 4 h after MCAo, glutamate, aspartate, GABA, adenosine and taurine outflow increased. SCH 58261 (0.01 mg/kg, i.p.), administered 5 min after MCAo, suppressed turning behavior and significantly reduced the outflow of glutamate, aspartate, GABA and adenosine. At 24 h after MCAo, the rats showed severe sensorimotor deficit and damage in both the striatum and cortex. SCH 58261 significantly reduced cortical damage but did not protect against the sensorimotor deficit. The protective effect of SCH 58261 against turning behavior and increased outflow of excitatory amino acids in the first hours after MCAo suggests the potential utility of selective adenosine A(2A) antagonists when administered in the first hours after ischemia. Furthermore, this study, for the first time, proposes that turning behavior after permanent intraluminal MCAo, be used as a precocious index of neurological deficit and neuronal damage

VSTM2L is a novel secreted antagonist of the neuroprotective peptide Humanin

Humanin (HN) is a 24-residue peptide displaying a protective activity in vitro against a range of cytotoxic and neurotoxic insults, as well as mediating in vivo amelioration of Alzheimer disease (AD)-related memory impairment in experimental models. Published evidence suggests that the mechanisms through which HN exerts its cyto- and neuroprotective activity may include its secretion and binding to membraneassociated receptors. Here, we describe the identification of a new modulator of HN neuroprotective activity, V-set and transmembrane domain containing 2 like (VSTM2L), previously known as C20orf102. VSTM2L interacts with HN in both yeast and mammalian cells, is secreted in cultured cells, is present in serum, and is selectively expressed in the central nervous system. VSTM2L colocalizes with HN in distinct brain areas as well as in primary cultured neurons, where it plays a role in the modulation of neuronal viability. When tested in HN neuroprotection bioassays, VSTM2L acts as a strong antagonist of HN neuroprotective activity. In summary, VSTM2L is the first example of a secreted antagonist of HN and may play a role in the modulation of HN biological function

Procognitive and neuroprotective activity of a novel alpha7 nicotinic acetylcholine receptor agonist for treatment of neurodegenerative and cognitive disorders

ABSTRACT The ␣7 nicotinic acetylcholine receptor (nAChR) is a promising target for treatment of cognitive dysfunction associated with Alzheimer's disease and schizophrenia. Here, we report the pharmacological properties of 5-morpholin-4-yl-pentanoic acid (4-pyridin-3-yl-phenyl)-amide [SEN12333 (WAY-317538)], a novel selective agonist of ␣7 nAChR. SEN12333 shows high affinity for the rat ␣7 receptor expressed in GH4C1 cells (K i ϭ 260 nM) and acts as full agonist in functional Ca 2ϩ flux studies (EC 50 ϭ 1.6 M). In whole-cell patch-clamp recordings, SEN12333 activated peak currents and maximal total charges similar to acetylcholine (EC 50 ϭ 12 M). The compound did not show agonist activity at other nicotinic receptors tested and acted as a weak antagonist at ␣3-containing receptors. SEN12333 treatment (3 mg/kg i.p.) improved episodic memory in a novel object recognition task in rats in conditions of spontaneous forgetting as well as cognitive disruptions induced via glutamatergic [5H-dibenzo[a,d]cyclohepten-5,10-imine (dizocilpine maleate); MK-801] or cholinergic (scopolamine) mechanisms. This improvement was blocked by the ␣7-selective antagonist methyllycaconitine, indicating that it is mediated by ␣7 activation. SEN12333 also prevented a scopolamine-induced deficit in a passive avoidance task. In models targeting other cognitive domains, including attention and perceptual processing, SEN12333 normalized the apomorphine-induced deficit of prepulse inhibition. Neuroprotection of SEN12333 was demonstrated in quisqualate-lesioned animals in which treatment with SEN12333 (3 mg/kg/day i.p.) resulted in a significant protection of choline acetyltransferase-positive neurons in the lesioned hemisphere. Cumulatively, our results demonstrate that the novel ␣7 nAChR agonist SEN12333 has procognitive and neuroprotective properties, further demonstrating utility of ␣7 agonists for treatment of neurodegenerative and cognitive disorders. The family of nicotinic acetylcholine receptors, which comprises 16 different subunits in human (␣1-7, ␣9 -10, ␤1-4, ␦, ε, and ␥) that can form many functional homo-and heteropentameric receptor ion channel combinations, contributes to cholinergic neurotransmission in the nervous system and at the neuromuscular junction. The ␣7 nicotinic acetylcholine receptors (nAChRs) are rapidly desensitizing ligand-gated ion channels that are abundantly expressed in the cerebral cortex and the hippocampus, a limbic structure intimately linked to attention processing and memory formatio