Vânia L. Batalha

A2A adenosine receptor deletion is protective in a mouse model of Tauopathy

Consumption of caffeine, a non-selective adenosine A2A receptor (A2AR) antagonist, reduces the risk of developing Alzheimer’s disease (AD) in humans and mitigates both amyloid and Tau burden in transgenic mouse models. However, the impact of selective A2AR blockade on the progressive development of AD-related lesions and associated memory impairments has not been investigated. In the present study, we removed the gene encoding A2AR from THY-Tau22 mice and analysed the subsequent effects on both pathological (Tau phosphorylation and aggregation, neuro-inflammation) and functional impairments (spatial learning and memory, hippocampal plasticity, neurotransmitter profile). We found that deleting A2ARs protect from Tau pathology-induced deficits in terms of spatial memory and hippocampal long-term depression. These effects were concomitant with a normalization of the hippocampal glutamate/gamma-amino butyric acid ratio, together with a global reduction in neuro-inflammatory markers and a decrease in Tau hyperphosphorylation. Additionally, oral therapy using a specific A2AR antagonist (MSX-3) significantly improved memory and reduced Tau hyperphosphorylation in THY-Tau22 mice. By showing that A2AR genetic or pharmacological blockade improves the pathological phenotype in a Tau transgenic mouse model, the present data highlight A2A receptors as important molecular targets to consider against AD and Tauopathies.

Adenosine A 2A receptor blockade reverts hippocampal stress-induced deficits and restores corticosterone circadian oscillation

Maternal separation (MS) is an early life stress model that induces permanent changes in the central nervous system, impairing hippocampal long-term potentiation (LTP) and spatial working memory. There are compelling evidences for a role of hippocampal adenosine A2A receptors in stress-induced modifications related to cognition, thus opening a potential window for therapeutic intervention. Here, we submitted rats to MS and evaluated the long-lasting molecular, electrophysiological and behavioral impairments in adulthood. We then assessed the therapeutic potential of KW6002, a blocker of A2A receptors, in stress-impaired animals. We report that the blockade of A2A receptors was efficient in reverting the behavior, electrophysiological and morphological impairments induced by MS. In addition, this effect is associated with restoration of the hypothalamic-pituitary-adrenal axis (HPA-axis) activity, as both the plasma corticosterone levels and hippocampal glucocorticoid receptor expression pattern returned to physiological-like status after the treatment. These results reveal the involvement of A2A receptors in the stress-associated impairments and directly in the stress response system by showing that the dysfunction of the HPA-axis as well as the long-lasting synaptic and behavioral effects of MS can be reverted by targeting adenosine A2A receptors. These findings provide a novel evidence for the use of adenosine A2A receptor antagonists as potential therapy against psychopathologies.

Maternal separation impairs long term-potentiation in CA1-CA3 synapses and hippocampal-dependent memory in old rats

Exposure to chronic stress during the neonatal period is known to induce permanent long-term changes in the central nervous system and hipothalamic-pituitary-adrenal axis reactivity that are associated with increased levels of depression, anxiety, and cognitive impairments. In rodents, a validated model of early life stress is the maternal separation (MS) paradigm, which has been shown to have long-term consequences for the pups that span to adulthood. We hypothesized that the early life stress-associated effects could be exacerbated with aging, because it is often accompanied by cognitive decline. Using a MS model in which rat pups were separated from their mothers for 3 hours daily, during postnatal days 2-14, we evaluated the long-term functional consequences to aged animals (70-week-old), by measuring synaptic plasticity and cognitive performance. The baseline behavioral deficits of aged control rats were further exacerbated in MS animals, indicating that early-life stress induces sustained changes in anxiety-like behavior and hippocampal-dependent memory that are maintained much later in life. We then investigated whether these differences are linked to impaired function of hippocampal neurons by recording hippocampal long-term potentiation from Schaffer collaterals/CA1 synapses. The magnitude of the hippocampal long-term potentiation induced by high-frequency stimulation was significantly lower in aged MS animals than in age-matched controls. These results substantiate the hypothesis that the neuronal and endocrine alterations induced by early-life stress are long lasting, and are able to exacerbate the mild age-associated deficits.

α-synuclein interacts with PrP C to induce cognitive impairment through mGluR5 and NMDAR2B

Synucleinopathies, such as Parkinsons disease and dementia with Lewy bodies, are neurodegenerative disorders that are characterized by the accumulation of α-synuclein (aSyn) in intracellular inclusions known as Lewy bodies. Prefibrillar soluble aSyn oligomers, rather than larger inclusions, are currently considered to be crucial species underlying synaptic dysfunction. We identified the cellular prion protein (PrPC) as a key mediator in aSyn-induced synaptic impairment. The aSyn-associated impairment of long-term potentiation was blocked in Prnp null mice and rescued following PrPC blockade. We found that extracellular aSyn oligomers formed a complex with PrPC that induced the phosphorylation of Fyn kinase via metabotropic glutamate receptors 5 (mGluR5). aSyn engagement of PrPC and Fyn activated NMDA receptor (NMDAR) and altered calcium homeostasis. Blockade of mGluR5-evoked phosphorylation of NMDAR in aSyn transgenic mice rescued synaptic and cognitive deficits, supporting the hypothesis that a receptor-mediated mechanism, independent of pore formation and membrane leakage, is sufficient to trigger early synaptic damage induced by extracellular aSyn.

Adenosine A2A Receptors Modulate α-Synuclein Aggregation and Toxicity.

Abstract Abnormal accumulation of aggregated &agr;‐synuclein (aSyn) is a hallmark of sporadic and familial Parkinsons disease (PD) and related synucleinopathies. Recent studies suggest a neuroprotective role of adenosine A2A receptor (A2AR) antagonists in PD. Nevertheless, the precise molecular mechanisms underlying this neuroprotection remain unclear. We assessed the impact of A2AR blockade or genetic deletion (A2AR KO) on synaptic plasticity and neuronal cell death induced by aSyn oligomers. We found that impairment of LTP associated with aSyn exposure was rescued in A2AR KO mice or upon A2AR blockade, through an NMDA receptor‐dependent mechanism. The mechanisms underlying these effects were evaluated in SH‐SY5Y cells overexpressing aSyn and rat primary neuronal cultures exposed to aSyn. Cell death in both conditions was prevented by selective A2AR antagonists. Interestingly, blockade of these receptors did not interfere with aSyn oligomerization but, instead, reduced the percentage of cells displaying aSyn inclusions. Altogether, our data raise the possibility that the well‐documented effects of A2AR antagonists involve the control of the latter stages of aSyn aggregation, thereby preventing the associated neurotoxicity. These findings suggest that A2AR represent an important target for the development of effective drugs for the treatment of PD and related synucleinopathies.

Escitalopram improves memory deficits induced by maternal separation in the rat

Maternal separation (MS) induces depressive-like behavior and long-term changes in cognition in rats. Escitalopram is an antidepressant drug shown to reverse the depressive-like features caused by this stress model. However, it is not known if it can ameliorate the affected cognition. We now characterized the effect of escitalopram on hippocampal-dependent memory in rats submitted to the MS protocol. Male Wistar rats were assigned either to control (CTR) or maternal separated (MS) group. MS were separated from their dams between 2-14 postnatal days (PND) for 180min daily. Escitalopram was given in food pellets (0.34g/kg/day first 2 weeks and 0.41g/kg/day the subsequent period, average dose 25mg/kg) from PND 43 onwards, during 1 month. Depressive behavior was assessed in the forced swimming test (FST), and memory performance in the Morris water maze (MWM). Escitalopram significantly improved the FSTs latency to despair in the MS group (n=6), but did not change the immobility time. All groups showed a significant learning effect in the MWM over time, but no differences have been found upon treatment (n=6). However, escitalopram treatment significantly increased the time spent on the platform quadrant in the probe trial in the MS group. We report here that chronic treatment with escitalopram is able to improve hippocampal dependent memory in a chronic stress model, while not changing the learning ability. Moreover, this is accompanied by an amelioration of the depressive like behavior. These results support the use of escitalopram to tackle underlying cognitive deficits caused by stress in early-life.

Modulating Alzheimer's Disease Through Caffeine: A Putative Link to Epigenetics

Aging is the best-known risk factor for many disorders, including neurodegenerative diseases such as Alzheimers disease (AD). The effect of epigenetic modulation of gene expression on normal aging and in pathological conditions is still unclear, but it is likely it may explain some of the complexity that is characteristic of these processes. Caffeine is a widely consumed psychoactive drug, which is emerging as a protective agent against AD progression and in aging associated deficits. This occurs mainly through the blockade of adenosine A2A receptors, whose expression and function become aberrant throughout aging and in age-related pathologies. Here, we discuss the data supporting the effects of caffeine in AD, focusing on adenosine A2A receptors and epigenetic modulation of gene expression. In addition, we speculate on the potential of caffeine as an epigenetic modulator and the consequences it might have for preventive and therapeutic applications of caffeine in AD.

From epidemiology to pathophysiology: what about caffeine in Alzheimer's disease?

AD (Alzheimers disease) is the most prevalent form of dementia in the aged population. Definitive diagnosis of AD is based on the presence of senile plaques and neurofibrillary tangles that are identified in post-mortem brain specimens. A third pathological component is inflammation. AD results from multiple genetic and environmental risk factors. Among other factors, epidemiological studies report beneficial effects of caffeine, a non-selective antagonist of adenosine receptors. In the present review, we discuss the impact of caffeine and the adenosinergic system in AD pathology as well as consequences in terms of pathology and therapeutics.

Neuroprotection afforded by adenosine A2A receptor blockade is modulated by corticotrophin-releasing factor (CRF) in glutamate injured cortical neurons

In situations of hypoxia, glutamate excitotoxicity induces neuronal death. The release of extracellular adenosine is also triggered and is accompanied by an increase of the stress mediator, corticotrophin‐releasing factor (CRF). Adenosine A2A receptors contribute to glutamate excitoxicity and their blockade is effective in stress‐induced neuronal deficits, but the involvement of CRF on this effect was never explored. We now evaluated the interaction between A2A and CRF receptors (CRFR) function, upon glutamate insult. Primary rat cortical neuronal cultures (9 days in vitro) expressing both CRF1R and CRF2R were challenged with glutamate (20–1000 μM, 24 h). CRF1R was found to co‐localize with neuronal markers and CRF2R to be present in both neuronal and glial cells. The effects of the CRF and A2A receptors ligands on cell viability were measured using propidium iodide and Syto‐13 fluorescence staining. Glutamate decreased cell viability in a concentration‐dependent manner. Urocortin (10 pM), an agonist of CRF receptors, increased cell survival in the presence of glutamate. This neuroprotective effect was abolished by blocking either CRF1R or CRF2R with antalarmin (10 nM) or anti‐Sauvagine‐30 (10 nM), respectively. The blockade of A2A receptors with a selective antagonist SCH 58261 (50 nM) improved cell viability against the glutamate insult. This effect was dependent on CRF2R, but not on CRF1R activation. Overall, these data show a protective role of CRF in cortical neurons, against glutamate‐induced death. The neuroprotection achieved by A2A receptors blockade requires CRF2R activation. This interaction between the adenosine and CRF receptors can explain the beneficial effects of using A2A receptor antagonists against stress‐induced noxious effects.

Adenosine Kinase Deficiency in the Brain Results in Maladaptive Synaptic Plasticity.

Adenosine kinase (ADK) deficiency in human patients (OMIM:614300) disrupts the methionine cycle and triggers hypermethioninemia, hepatic encephalopathy, cognitive impairment, and seizures. To identify whether this neurological phenotype is intrinsically based on ADK deficiency in the brain or if it is secondary to liver dysfunction, we generated a mouse model with a brain-wide deletion of ADK by introducing a Nestin-Cre transgene into a line of conditional ADK deficient Adkfl/fl mice. These AdkΔbrain mice developed a progressive stress-induced seizure phenotype associated with spontaneous convulsive seizures and profound deficits in hippocampus-dependent learning and memory. Pharmacological, biochemical, and electrophysiological studies suggest enhanced adenosine levels around synapses resulting in an enhanced adenosine A1 receptor (A1R)-dependent protective tone despite lower expression levels of the receptor. Theta-burst-induced LTP was enhanced in the mutants and this was dependent on adenosine A2A receptor (A2AR) and tropomyosin-related kinase B signaling, suggesting increased activation of these receptors in synaptic plasticity phenomena. Accordingly, reducing adenosine A2A receptor activity in AdkΔbrain mice restored normal associative learning and contextual memory and attenuated seizure risk. We conclude that ADK deficiency in the brain triggers neuronal adaptation processes that lead to dysregulated synaptic plasticity, cognitive deficits, and increased seizure risk. Therefore, ADK mutations have an intrinsic effect on brain physiology and may present a genetic risk factor for the development of seizures and learning impairments. Furthermore, our data show that blocking A2AR activity therapeutically can attenuate neurological symptoms in ADK deficiency. SIGNIFICANCE STATEMENT A novel human genetic condition (OMIM #614300) that is based on mutations in the adenosine kinase (Adk) gene has been discovered recently. Affected patients develop hepatic encephalopathy, seizures, and severe cognitive impairment. To model and understand the neurological phenotype of the human mutation, we generated a new conditional knock-out mouse with a brain-specific deletion of Adk (AdkΔbrain). Similar to ADK-deficient patients, AdkΔbrain mice develop seizures and cognitive deficits. We identified increased basal synaptic transmission and enhanced adenosine A2A receptor (A2AR)-dependent synaptic plasticity as the underlying mechanisms that govern these phenotypes. Our data show that neurological phenotypes in ADK-deficient patients are intrinsic to ADK deficiency in the brain and that blocking A2AR activity therapeutically can attenuate neurological symptoms in ADK deficiency.