Diogo R. Lara

Caffeine and adenosine A2a receptor antagonists prevent β-amyloid (25–35)-induced cognitive deficits in mice

Consumption of caffeine, an adenosine receptor antagonist, was found to be inversely associated with the incidence of Alzheimers disease. Moreover, caffeine protects cultured neurons against beta-amyloid-induced toxicity, an effect mimicked by adenosine A(2A) but not A(1) receptor antagonists. We now tested if caffeine administration would prevent beta-amyloid-induced cognitive impairment in mice and if this was mimicked by A(2A) receptor blockade. One week after icv administration of the 25-35 fragment of beta-amyloid (Abeta, 3 nmol), mice displayed impaired performance in both inhibitory avoidance and spontaneous alternation tests. Prolonged treatment with caffeine (1 mg/ml) had no effect alone but prevented the Abeta-induced cognitive impairment in both tasks when associated with acute caffeine (30 mg/kg) 30 min treatment before Abeta administration. The same protective effect was observed after subchronic (4 days) treatment with daily injections of either caffeine (30 mg/kg) or the selective adenosine A(2A) receptor antagonist SCH58261 (0.5 mg/kg). This provides the first direct in vivo evidence that caffeine and A(2A) receptor antagonists afford a protection against Abeta-induced amnesia, which prompts their interest for managing Alzheimers disease.

Neuroprotection by caffeine and adenosine A2A receptor blockade of β‐amyloid neurotoxicity

Adenosine is a neuromodulator in the nervous system and it has recently been observed that pharmacological blockade or gene disruption of adenosine A2A receptors confers neuroprotection under different neurotoxic situations in the brain. We now observed that coapplication of either caffeine (1–25 μM) or the selective A2A receptor antagonist, 4‐(2‐[7‐amino‐2(2‐furyl)(1,2,4)triazolo (2,3‐a)(1,3,5)triazin‐5‐ylamino]ethyl)phenol (ZM 241385, 50 nM), but not the A receptor antagonist, 8‐cyclopentyltheophylline (200 nM), prevented the neuronal cell death caused by exposure of rat cultured cerebellar granule neurons to fragment 25–35 of β‐amyloid protein (25 μM for 48 h), that by itself caused a near three‐fold increase of propidium iodide‐labeled cells. This constitutes the first in vitro evidence to suggest that adenosine A2A receptors may be the molecular target responsible for the observed beneficial effects of caffeine consumption in the development of Alzheimers disease.

Guanosine and GMP prevent seizures induced by quinolinic acid in mice

In the mammalian CNS, glutamate and GABA are the principal neurotransmitters mediating excitatory and inhibitory synaptic events, respectively, and have been implicated in the neurobiology of seizures. Guanine-based purines, including the nucleoside guanosine and the nucleotide GMP, have been shown to antagonize glutamatergic activity at the receptor level and the other purine nucleoside adenosine is a well-known modulator of seizure threshold. In the present study we investigated the anticonvulsant effect of i. p. guanosine and GMP against seizures induced by the glutamate agonist quinolinic acid (QA) or the GABA(A) antagonist picrotoxin in mice. Animals were pretreated with an i.p. injection of saline, guanosine or GMP 30 min before either an i.c.v. injection of 4 microliter QA (36.8 nmol) or a subcutaneous injection of picrotoxin (3.2 mg/kg). All animals pretreated with vehicle followed by QA or picrotoxin presented seizures, which were completely prevented by the NMDA antagonist MK-801 and the GABA agonist phenobarbital, respectively. Guanosine and GMP dose-dependently protected against QA-induced seizures, up to 70 and 80% at 7.5 mg/kg, with ED(50)=2. 6+/-0.4 and 1.7+/-0.6 mg/kg, respectively. Conversely, neither guanosine, GMP nor MK-801 affected picrotoxin-induced seizures, indicating some degree of specificity towards the glutamatergic system. This study suggests anticonvulsant properties of i.p. guanosine and GMP, which may be related with antagonism of glutamate receptors.

Increased serum S100B protein in schizophrenia: a study in medication-free patients

S100B protein, a calcium binding protein produced and released by glial cells, has been used as a sensitive marker of brain damage. Previous studies have found alterations in peripheral S100B levels in schizophrenic patients on medication. We compared serum S100B levels of 20 medication-free DSM-IV schizophrenic patients and 20 age-gender matched healthy controls. Schizophrenic patients presented higher serum S100B levels (mean 0.120 ng/ml+/-S.D. 0.140) compared to controls (mean 0.066 ng/ml+/-S.D. 0.067; P=0.014) and there was a negative correlation with illness duration (r=-0.496, P=0.031). The results of this study indicate that serum S100B levels may be a state marker of a limited neurodegenerative process, particularly in the early course of schizophrenia or, at least, in a subgroup of schizophrenic patients.

Guanosine Enhances Glutamate Uptake in Brain Cortical Slices at Normal and Excitotoxic Conditions

Abstract1. The effect of guanosine on L-[2,3-3H]glutamate uptake was investigated in brain cortical slices under normal or oxygen–glucose deprivation (OGD) conditions.2. In slices exposed to physiological conditions, guanosine (1–100 μM) stimulated glutamate uptake (up to 100%) in a concentration-dependent manner when a high (100 μM) but not a low (1 μM) concentration of glutamate was used.3. In slices submitted to OGD, guanosine 1 and 100 μM also increased 100 μM glutamate uptake (38 and 70%, respectively).4. The increasing of glutamate and taurine released to the incubation medium in cortical slices submitted to OGD were significantly attenuated by the presence of guanosine in the incubation medium.5. Guanosine prevented the increase in propidium iodide incorporation into cortical slices induced by OGD, indicating a protective role against ischemic injury.6. These results support the hypothesis of a protective role for guanosine during brain ischemia, possibly by activating glutamate uptake into neural cells.

Involvement of adenosine in the neurobiology of schizophrenia and its therapeutic implications.

Based on the neuromodulatory and homeostatic actions of adenosine, adenosine dysfunction may contribute to the neurobiological and clinical features of schizophrenia. The present model of adenosine dysfunction in schizophrenia takes into consideration the dopamine and glutamate hypotheses, since adenosine exerts neuromodulatory roles on these systems, and proposes that adenosine plays a role in the inhibitory deficit found in schizophrenia. Given the role of adenosine activation of adenosine A1 receptor (A1R) in mediating neurotoxicity in early stages of brain development, pre- and peri-natal complications leading to excessive adenosine release could induce primary brain changes (i.e., first hit). These events would lead to an adenosine inhibitory deficit through a partial loss of A1R that may emerge as reduced control of dopamine activity and increased vulnerability to excitotoxic glutamate action in the mature brain (i.e., second hit). Adenosine dysfunction is reasonably compatible with symptoms, gray and white matter abnormalities, progressive brain loss, pre- and peri-natal risk factors, age of onset, response to current treatments, impaired sensory gating and increased smoking in schizophrenia. Pharmacological treatments enhancing adenosine activity could be effective for symptom control and for alleviating deterioration in the course of the illness. Accordingly, allopurinol, which may indirectly increase adenosine, has been effective and well tolerated in the treatment of schizophrenia. Since much of the evidence for the adenosine hypothesis is preliminary and theoretical, further investigation in the field is warranted.

Unpredictable chronic stress model in zebrafish (Danio rerio): Behavioral and physiological responses

Zebrafish (Danio rerio) have emerged as a promising model organism to study development, toxicology, pharmacology, and neuroscience, among other areas. Despite the increasing number of studies using zebrafish, behavioral studies with this species are still elementary when compared to rodents. The aim of this study was to develop a model of unpredictable chronic stress (UCS) in zebrafish. We evaluated the effects of UCS protocol during 7 or 14 days on behavioral and physiological parameters. The effects of stress were evaluated in relation to anxiety and exploratory behavior, memory, expression of corticotrophin-releasing factor (CRF) and glucocorticoid receptor (GR), and cortisol levels. As expected, UCS protocol increased the anxiety levels, impaired cognitive function, and increased CRF while decreased GR expression. Moreover, zebrafish submitted to 7 or 14 days of UCS protocol presented increased cortisol levels. The protocol developed here is a complementary model for studying the neurobiology and the effects of chronic stress in behavioral and physiological parameters. In addition, this protocol is less time consuming than standard rodent models commonly used to study chronic stress. These results confirm UCS in zebrafish as an adequate model to preclinical studies of stress, although further studies are warranted to determine its predictive validity.

Effect of orally administered guanosine on seizures and death induced by glutamatergic agents.

Intraperitoneal guanosine has been shown to prevent quinolinic acid-induced seizures in mice. In this study, we investigated the effect of orally administered guanosine on seizures induced by the glutamate agonists quinolinic acid and kainate, and the endogenous glutamate releaser alpha-dendrotoxin. Guanosine (7.5 mg/kg, per os), administered 75 min in advance, prevented 70% of seizures induced by i.c.v. quinolinic acid, being as efficient as the NMDA channel blocker MK-801 administered intraperitoneally. Guanosine was ineffective against kainate-induced seizures, but significantly reversed the potentiation of seizures and death caused by the concomitant injection of MK-801. Guanosine also significantly prevented seizures and death induced by i.c.v. alpha-dendrotoxin, whereas MK-801 and phenobarbital only prevented death. Altogether, our findings underscore the therapeutic potential of oral administration of guanosine for treating diseases involving glutamatergic excitotoxicity, including epilepsy.

Activation of glutamate uptake by guanosine in primary astrocyte cultures.

Guanine-based purines have been shown to modulate the effects of glutamate, which is essential for brain function and mediates excitotoxicity. In the search for a mechanism involving the interaction between purine nucleoside guanosine and glutamate, we found that guanosine dose-dependently, significantly (63%) and potently (EC50 = 2.47 μM) enhanced glutamate uptake in cultured astrocytes. This effect was not inhibited by the blocker of nucleoside transporter dipyridamole nor by the adenosine antagonist theophylline, suggesting an extracellular site of action without the involvement of adenosine receptors. These results indicate a regulatory role of guanosine on extracellular levels of glutamate, possibly contributing for protecting neural cells against glutamate-induced excitotoxicity.

Caffeine, mental health, and psychiatric disorders.

Caffeine intake is so common that its pharmacological effects on the mind are undervalued. Since it is so readily available, individuals can adjust their own dose, time of administration and dose intervals of caffeine, according to the perceived benefits and side effects of each dose. This review focuses on human studies of caffeine in subjects with and without psychiatric disorders. Besides the possibility of mild drug dependence, caffeine may bring benefits that contribute to its widespread use. These benefits seem to be related to adaptation of mental energy to the context by increasing alertness, attention, and cognitive function (more evident in longer or more difficult tasks or situations of low arousal) and by elevating mood. Accordingly, moderate caffeine intake (< 6 cups/day) has been associated with less depressive symptoms, fewer cognitive failures, and lower risk of suicide. However, its putative therapeutic effects on depression and ADHD have been insufficiently studied. Conversely, in rare cases high doses of caffeine can induce psychotic and manic symptoms, and more commonly, anxiety. Patients with panic disorder and performance social anxiety disorder seem to be particularly sensitive to the anxiogenic effects of caffeine, whereas preliminary data suggests that it may be effective for some patients with obsessive compulsive disorder (OCD). The threshold for the anxiogenic effect of caffeine is influenced by a polymorphism of the A2A receptor. In summary, caffeine can be regarded as a pharmacological tool to increase energy and effortful behavior in daily activities. More populational (cross-sectional and prospective) and experimental studies are necessary to establish the role of caffeine intake in psychiatric disorders, especially its putative efficacy on depressive mood and cognitive/attentional disorders.