Delayed Outgrowth in Response to the BDNF and Altered Synaptic Proteins in Neurons From SHR Rats.

Attention deficit hyperactivity disorder (ADHD) is a neurodevelopmental disorder characterized by inattention, hyperactivity, and impulsivity symptoms. Neuroimaging studies have revealed a delayed cortical and subcortical development pattern in children diagnosed with ADHD. This study followed up on the development in vitro of frontal cortical neurons from Spontaneously hypertensive rats (SHR), an ADHD rat model, and Wistar-Kyoto rats (WKY), control strain, over their time in culture, and in response to BDNF treatment at two different days in vitro (DIV). These neurons were also evaluated for synaptic proteins, brain-derived neurotrophic factor (BDNF), and related protein levels. Frontal cortical neurons from the ADHD rat model exhibited shorter dendrites and less dendritic branching over their time in culture. While pro- and mature BDNF levels were not altered, the cAMP-response element-binding (CREB) decreased at 1 DIV and SNAP-25 decreased at 5 DIV. Different from control cultures, exogenous BDNF promoted less dendritic branching in neurons from the ADHD model. Our data revealed that neurons from the ADHD model showed decreased levels of an important transcription factor at the beginning of their development, and their delayed outgrowth and maturation had consequences in the levels of SNAP-25 and may be associated with less response to BDNF. These findings provide an alternative tool for studies on synaptic dysfunctions in ADHD. They may also offer a valuable tool for investigating drug effects and new treatment opportunities.

Induced pluripotent stem cell-derived astrocytes from patients with schizophrenia exhibit an inflammatory phenotype that affects vascularization.

Molecular and functional abnormalities of astrocytes have been implicated in the etiology and pathogenesis of schizophrenia (SCZ). In this study, we examined the proteome, inflammatory responses, and secretome effects on vascularization of human induced pluripotent stem cell (hiPSC)-derived astrocytes from patients with SCZ. Proteomic analysis revealed alterations in proteins related to immune function and vascularization. Reduced expression of the nuclear factor kappa B (NF-κB) p65 subunit was observed in these astrocytes, with no incremental secretion of cytokines after tumor necrosis factor alpha (TNF-α) stimulation. Among inflammatory cytokines, secretion of interleukin (IL)-8 was particularly elevated in SCZ-patient-derived-astrocyte-conditioned medium (ASCZCM). In a chicken chorioallantoic membrane (CAM) assay, ASCZCM reduced the diameter of newly grown vessels. This effect could be mimicked with exogenous addition of IL-8. Taken together, our results suggest that SCZ astrocytes are immunologically dysfunctional and may consequently affect vascularization through secreted factors.

Sex differences in maternal odor preferences and brain levels of GAP-43 and sonic hedgehog proteins in infant SHR and Wistar Kyoto rats.

Attention Deficit Hyperactivity Disorder (ADHD) is a neurodevelopmental disorder that presents sex differences in the severity and presentation of symptoms, whose neurobiological basis is still unknown. Both Growth-associated Protein 43 (GAP-43) and Sonic hedgehog (Shh) are considered essential proteins for the appropriate brain development, but their participation in ADHD neurobiology have not been investigated yet. In this study, we hypothesized that alterations in these proteins could be related to behavioral traits to ADHD phenotype. Thus, both sexes of infant Spontaneously hypertensive rats (SHR, used as ADHD animal model) were evaluated for developmental milestones, locomotor activity, olfactory and recognition memory. Both GAP-43 and Shh were assessed in the olfactory bulb, frontal cortex and hippocampus in early and late infancy. During early infancy, SHR reached three developmental milestones later, and females showed olfactory memory impairment accompanied by increased levels of Shh in the olfactory bulb. In later infancy, hyperlocomotion, impaired recognition memory, and decreased Shh in the hippocampus were observed in SHR from both sexes. While in early infancy GAP-43 was not altered, it was decreased in the frontal cortex and hippocampus of female SHR in late infancy. Therefore, both Shh and GAP-43 are involved in the sex-dependent behavioral alterations showed by infant SHR. Despite the disorder's complexity and heterogeneity, our findings reveal important developmental parameters during SHR development and also emphasizes the relevance of studying sex differences in the ADHD context.

Dietary omega-3 fatty acids prevent neonatal seizure-induced early alterations in the hippocampal glutamatergic system and memory deficits in adulthood.

OBJECTIVE: We investigated the influence of dietary omega-3 polyunsaturated fatty acids (n-3 PUFAs) on glutamatergic system modulation after a single episode of neonatal seizures and their possible effects on seizure-induced long-lasting behavioral deficits. METHODS: Male Wistar rats receiving an omega-3 diet (n-3) or an n-3 deficient diet (D) from the prenatal period were subjected to a kainate-induced seizure model at P7. Glutamate transporter activity and immunocontents (GLT-1 and GLAST) were assessed in the hippocampus at 12, 24, and 48 h after the seizure episode. Fluorescence intensity for glial cells (GFAP) and neurons (NeuN) was assessed 24 h after seizure in the hippocampus. Behavioral analysis (elevated-plus maze and inhibitory avoidance memory task) was performed at 60 days of age. RESULTS: The D group showed a decrease in glutamate uptake 24 h after seizure. In this group only, the GLT1 content increased at 12 h, followed by a decrease at 24 h. GLAST increased up to 24 h after seizure. GFAP fluorescence was higher, and NeuN fluorescence decreased, in the D group independent of seizures. In adulthood, the D group presented memory deficits independent of seizures, but short-term memory (1.5 h after a training session) was abolished in the D group treated with kainate. SIGNIFICANCE: N-3 PUFA positively influenced the glutamatergic system during seizure and prevented seizure-related memory deficits in adulthood.

The Age of Brain Organoids: Tailoring Cell Identity and Functionality for Normal Brain Development and Disease Modeling.

Over the past years, brain development has been investigated in rodent models, which were particularly relevant to establish the role of specific genes in this process. However, the cytoarchitectonic features, which determine neuronal network formation complexity, are unique to humans. This implies that the developmental program of the human brain and neurological disorders can only partly be reproduced in rodents. Advancement in the study of the human brain surged with cultures of human brain tissue in the lab, generated from induced pluripotent cells reprogrammed from human somatic tissue. These cultures, termed brain organoids, offer an invaluable model for the study of the human brain. Brain organoids reproduce the cytoarchitecture of the cortex and can develop multiple brain regions and cell types. Integration of functional activity of neural cells within brain organoids with genetic, cellular, and morphological data in a comprehensive model for human development and disease is key to advance in the field. Because the functional activity of neural cells within brain organoids relies on cell repertoire and time in culture, here, we review data supporting the gradual formation of complex neural networks in light of cell maturity within brain organoids. In this context, we discuss how the technology behind brain organoids brought advances in understanding neurodevelopmental, pathogen-induced, and neurodegenerative diseases.

Neuromodulation and neuroprotective effects of chlorogenic acids in excitatory synapses of mouse hippocampal slices.

The increased healthspan afforded by coffee intake provides novel opportunities to identify new therapeutic strategies. Caffeine has been proposed to afford benefits through adenosine A2A receptors, which can control synaptic dysfunction underlying some brain disease. However, decaffeinated coffee and other main components of coffee such as chlorogenic acids, also attenuate brain dysfunction, although it is unknown if they control synaptic function. We now used electrophysiological recordings in mouse hippocampal slices to test if realistic concentrations of chlorogenic acids directly affect synaptic transmission and plasticity. 3-(3,4-dihydroxycinnamoyl)quinic acid (CA, 1-10 µM) and 5-O-(trans-3,4-dihydroxycinnamoyl)-D-quinic acid (NCA, 1-10 µM) were devoid of effect on synaptic transmission, paired-pulse facilitation or long-term potentiation (LTP) and long-term depression (LTD) in Schaffer collaterals-CA1 pyramidal synapses. However, CA and NCA increased the recovery of synaptic transmission upon re-oxygenation following 7 min of oxygen/glucose deprivation, an in vitro ischemia model. Also, CA and NCA attenuated the shift of LTD into LTP observed in hippocampal slices from animals with hippocampal-dependent memory deterioration after exposure to ß-amyloid 1-42 (2 nmol, icv), in the context of Alzheimer's disease. These findings show that chlorogenic acids do not directly affect synaptic transmission and plasticity but can indirectly affect other cellular targets to correct synaptic dysfunction. Unraveling the molecular mechanisms of action of chlorogenic acids will allow the design of hitherto unrecognized novel neuroprotective strategies.

Intermittent fasting promotes anxiolytic-like effects unrelated to synaptic mitochondrial function and BDNF support.

Anxiety disorders are linked to mitochondrial dysfunction and decreased neurotrophic support. Since anxiolytic drugs target mitochondria, non-pharmacological approaches to improve mitochondrial metabolism such as intermittent fasting (IF) may cause parallel behavioral benefits against anxiety disorders. Here, we investigated whether a chronic IF regimen could induce anxiolytic-like effects concomitantly to modulation in mitochondrial bioenergetics and trophic signaling in mice brain. A total of 44 Male C57BL/6 J mice (180 days old) were assigned to two dietary regimens: a normal, ad libitum diet (AL group) and an alternate-day fasting (IF group), where animals underwent 10 cycles of 24 h food restriction followed by 24 h ad libitum access. Animals underwent the open field test, dark/light box and elevated plus maze tasks. Isolated nerve terminals were obtained from mice brain and used for mitochondrial respirometry, hydrogen peroxide production and assessment of membrane potential dynamics, calcium handling and western blotting. We showed that IF significantly alters total daily food intake and food consumption patterns but not body weight. There were no differences in the exploratory and locomotory parameters. Remarkably, animals from IF showed decreased anxiety-like behavior. Mitochondrial metabolic responses in different coupling states and parameters linked with H2O2 production, Ca2+ buffering and electric gradient were not different between groups. Finally, no alterations in molecular indicators of apoptotic death (Bax/Bcl-2 ratio) and neuroplasticity (proBDNF/BDNF and synaptophysin were observed). In conclusion, IF exerts anxiolytic-like effect not associated with modulation in synaptic neuronergetics or expression of neurotrophic proteins. These results highlight a potential benefit of intermittent fasting as a nutritional intervention in anxiety-related disorders.

Caffeine and adenosine A2A receptors rescue neuronal development in vitro of frontal cortical neurons in a rat model of attention deficit and hyperactivity disorder.

Although some studies have supported the effects of caffeine for treatment of Attention deficit and hyperactivity disorder (ADHD), there were no evidences about its effects at the neuronal level. In this study, we sought to find morphological alterations during in vitro development of frontal cortical neurons from Spontaneoulsy hypertensive rats (SHR, an ADHD rat model) and Wistar-Kyoto rats (WKY, control strain). Further, we investigated the effects of caffeine and adenosine A1 and A2A receptors (A1R and A2AR) signaling. Cultured cortical neurons from WKY and SHR were analyzed by immunostaining of microtubule-associated protein 2 (MAP-2) and tau protein after treatment with either caffeine, or A1R and A2AR agonists or antagonists. Besides, the involvement of PI3K and not PKA signaling was also assessed. Neurons from ADHD model displayed less neurite branching, shorter maximal neurite length and decreased axonal outgrowth. While caffeine recovered neurite branching and elongation from ADHD neurons via both PKA and PI3K signaling, A2AR agonist (CGS 21680) promoted more neurite branching via PKA signaling. The selective A2AR antagonist (SCH 58261) was efficient in recovering axonal outgrowth from ADHD neurons through PI3K and not PKA signaling. For the first time, frontal cortical neurons were isolated from ADHD model and they presented disturbances in the differentiation and outgrowth. By showing that caffeine and A2AR may act at neuronal level rescuing ADHD neurons outgrowth, our findings strengthen the potential of caffeine and A2AR receptors as an adjuvant for ADHD treatment.

Caffeine prevents neurodegeneration and behavioral alterations in a mice model of agitated depression.

Longitudinal and some experimental studies have showed the potential of caffeine to counteract some depressive behaviors and synaptic dysfunctions. In this study, we investigated the potential of caffeine in preventing behavioral outcomes, neurodegeneration and synaptic proteins alterations in a mice model of agitated depression by bilateral olfactory bulbectomy (OB). For this purpose, bulbectomized mice received caffeine (0.3 g/L and 1.0 g/L, drinking water), during the active cycle, for seven weeks (two before the surgery and throughout five weeks after OB). Caffeine prevented OB-induced hyperactivity and recognition memory impairment and rescue self care and motivational behavior. In the frontal cortex, bulbectomized mice presented increase in the adenosine A1 receptors (A1R) and GFAP, while adenosine A2A receptors (A2AR) increased in the hippocampus and striatum and SNAP-25 was decreased in frontal cortex and striatum. Caffeine increased A1R in the striatum of bulbectomized mice and in SHAM-water group caffeine increased A2AR in the striatum and decreased SNAP-25 in the frontal cortex. Astrogliosis observed in the polymorphic layer of the dentate gyrus of OB mice was prevented by caffeine as well as the neurodegeneration in the striatum and piriform cortex. Based on these behavioral and neurochemical evidences, caffeine confirms its efficacy in preventing neurodegeneration associated with memory impairment and may be considered as a promising therapeutic tool in the prophylaxis and/or treatment of depression.

Caffeine and cannabinoid receptors modulate impulsive behavior in an animal model of attentional deficit and hyperactivity disorder.

Attention deficit and hyperactivity disorder (ADHD) is characterized by impaired levels of hyperactivity, impulsivity, and inattention. Adenosine and endocannabinoid systems tightly interact in the modulation of dopamine signaling, involved in the neurobiology of ADHD. In this study, we evaluated the modulating effects of the cannabinoid and adenosine systems in a tolerance to delay of reward task using the most widely used animal model of ADHD. Spontaneous Hypertensive Rats (SHR) and Wistar-Kyoto rats were treated chronically or acutely with caffeine, a non-selective adenosine receptor antagonist, or acutely with a cannabinoid agonist (WIN55212-2, WIN) or antagonist (AM251). Subsequently, animals were tested in the tolerance to delay of reward task, in which they had to choose between a small, but immediate, or a large, but delayed, reward. Treatment with WIN decreased, whereas treatment with AM251 increased the choices of the large reward, selectively in SHR rats, indicating a CB1 receptor-mediated increase in impulsive behavior. An acute pre-treatment with caffeine blocked WIN effects. Conversely, a chronic treatment with caffeine increased the impulsive phenotype and potentiated the WIN effects. The results indicate that both cannabinoid and adenosine receptors modulate impulsive behavior in SHR: the antagonism of cannabinoid receptors might be effective in reducing impulsive symptoms present in ADHD; in addition, caffeine showed the opposite effects on impulsive behavior depending on the length of treatment. These observations are of particular importance to consider when therapeutic manipulation of CB1 receptors is applied to ADHD patients who consume coffee.

Guanosine Attenuates Behavioral Deficits After Traumatic Brain Injury by Modulation of Adenosinergic Receptors.

Traumatic brain injury (TBI) is a leading cause of disability worldwide, triggering chronic neurodegeneration underlying cognitive and mood disorder still without therapeutic prospects. Based on our previous observations that guanosine (GUO) attenuates short-term neurochemical alterations caused by TBI, this study investigated the effects of chronical GUO treatment in behavioral, molecular, and morphological disturbances 21 days after trauma. Rats subject to TBI displayed mood (anxiety-like) and memory dysfunction. This was accompanied by a decreased expression of both synaptic (synaptophysin) and plasticity proteins (BDNF and CREB), a loss of cresyl violet-stained neurons, and increased astrogliosis and microgliosis in the hippocampus. Notably, chronic GUO treatment (7.5 mg/kg i.p. daily starting 1 h after TBI) prevented all these TBI-induced long-term behavioral, neurochemical, and morphological modifications. This neuroprotective effect of GUO was abrogated in the presence of the adenosine A1 receptor antagonist DPCPX (1 mg/kg) but unaltered by the adenosine A2A receptor antagonist SCH58261 (0.05 mg/kg). These findings show that a chronic GUO treatment prevents the long-term mood and memory dysfunction triggered by TBI, which involves adenosinergic receptors.

Glutamate-induced and NMDA receptor-mediated neurodegeneration entails P2Y1 receptor activation.

Despite the characteristic etiologies and phenotypes, different brain disorders rely on common pathogenic events. Glutamate-induced neurotoxicity is a pathogenic event shared by different brain disorders. Another event occurring in different brain pathological conditions is the increase of the extracellular ATP levels, which is now recognized as a danger and harmful signal in the brain, as heralded by the ability of P2 receptors (P2Rs) to affect a wide range of brain disorders. Yet, how ATP and P2R contribute to neurodegeneration remains poorly defined. For that purpose, we now examined the contribution of extracellular ATP and P2Rs to glutamate-induced neurodegeneration. We found both in vitro and in vivo that ATP/ADP through the activation of P2Y1R contributes to glutamate-induced neuronal death in the rat hippocampus. We found in cultured rat hippocampal neurons that the exposure to glutamate (100 µM) for 30 min triggers a sustained increase of extracellular ATP levels, which contributes to NMDA receptor (NMDAR)-mediated hippocampal neuronal death through the activation of P2Y1R. We also determined that P2Y1R is involved in excitotoxicity in vivo as the blockade of P2Y1R significantly attenuated rat hippocampal neuronal death upon the systemic administration of kainic acid or upon the intrahippocampal injection of quinolinic acid. This contribution of P2Y1R fades with increasing intensity of excitotoxic conditions, which indicates that P2Y1R is not contributing directly to neurodegeneration, rather behaving as a catalyst decreasing the threshold from which glutamate becomes neurotoxic. Moreover, we unraveled that such excitotoxicity process began with an early synaptotoxicity that was also prevented/attenuated by the antagonism of P2Y1R, both in vitro and in vivo. This should rely on the observed glutamate-induced calpain-mediated axonal cytoskeleton damage, most likely favored by a P2Y1R-driven increase of NMDAR-mediated Ca2+ entry selectively in axons. This may constitute a degenerative mechanism shared by different brain diseases, particularly relevant at initial pathogenic stages.

Neuronal Adenosine A2A Receptors Are Critical Mediators of Neurodegeneration Triggered by Convulsions.

Neurodegeneration is a process transversal to neuropsychiatric diseases and the understanding of its mechanisms should allow devising strategies to prevent this irreversible step in brain diseases. Neurodegeneration caused by seizures is a critical step in the aggravation of temporal lobe epilepsy, but its mechanisms remain undetermined. Convulsions trigger an elevation of extracellular adenosine and upregulate adenosine A2A receptors (A2AR), which have been associated with the control of neurodegenerative diseases. Using the rat and mouse kainate model of temporal lobe epilepsy, we now tested whether A2AR control convulsions-induced hippocampal neurodegeneration. The pharmacological or genetic blockade of A2AR did not affect kainate-induced convulsions but dampened the subsequent neurotoxicity. This neurotoxicity began with a rapid A2AR upregulation within glutamatergic synapses (within 2 h), through local translation of synaptic A2AR mRNA. This bolstered A2AR-mediated facilitation of glutamate release and of long-term potentiation (LTP) in CA1 synapses (4 h), triggered a subsequent synaptotoxicity, heralded by decreased synaptic plasticity and loss of synaptic markers coupled to calpain activation (12 h), that predated overt neuronal loss (24 h). All modifications were prevented by the deletion of A2AR selectively in forebrain neurons. This shows that synaptic A2AR critically control synaptic excitotoxicity, which underlies the development of convulsions-induced neurodegeneration.

Sex differences in the effects of pre- and postnatal caffeine exposure on behavior and synaptic proteins in pubescent rats.

Few studies have addressed the effects of caffeine in the puberty and/or adolescence in a sex dependent manner. Considering that caffeine intake has increased in this population, we investigated the behavioral and synaptic proteins changes in pubescent male and female rats after maternal consumption of caffeine. Adult female Wistar rats started to receive water or caffeine (0.1 and 0.3g/L in drinking water; low and moderate dose, respectively) during the active cycle at weekdays, two weeks before mating. The treatment lasted up to weaning and the offspring received caffeine until the onset of puberty (30-34days old). Behavioral tasks were performed to evaluate locomotor activity (open field task), anxious-like behavior (elevated plus maze task) and recognition memory (object recognition task) and synaptic proteins levels (proBDNF, BDNF, GFAP and SNAP-25) were verified in the hippocampus and cerebral cortex. While hyperlocomotion was observed in both sexes after caffeine treatment, anxiety-related behavior was attenuated by caffeine (0.3g/L) only in females. While moderate caffeine worsened recognition memory in females, an improvement in the long-term memory was observed in male rats for both doses. Coincident with memory improvement in males, caffeine increased pro- and BDNF in the hippocampus and cortex. Females presented increased proBDNF levels in both brain regions, with no effects of caffeine. While GFAP was not altered, moderate caffeine intake increased SNAP-25 in the cortex of female rats. Our findings revealed that caffeine promoted cognitive benefits in males associated with increased BDNF levels, while females showed less anxiety. Our findings revealed that caffeine promotes distinct behavioral outcomes and alterations in synaptic proteins during brain development in a sex dependent manner.

Caffeine Reverts Memory But Not Mood Impairment in a Depression-Prone Mouse Strain with Up-Regulated Adenosine A2A Receptor in Hippocampal Glutamate Synapses.

Caffeine prophylactically prevents mood and memory impairments through adenosine A2A receptor (A2AR) antagonism. A2AR antagonists also therapeutically revert mood and memory impairments, but it is not known if caffeine is also therapeutically or only prophylactically effective. Since depression is accompanied by mood and memory alterations, we now explored if chronic (4 weeks) caffeine consumption (0.3 g/L) reverts mood and memory impairment in helpless mice (HM, 12 weeks old), a bred-based model of depression. HM displayed higher immobility in the tail suspension and forced swimming tests, greater anxiety in the elevated plus maze, and poorer memory performance (modified Y-maze and object recognition). HM also had reduced density of synaptic (synaptophysin, SNAP-25), namely, glutamatergic (vGluT1; -22 ± 7 %) and GABAergic (vGAT; -23 ± 8 %) markers in the hippocampus. HM displayed higher A2AR density (72 ± 6 %) in hippocampal synapses, an enhanced facilitation of hippocampal glutamate release by the A2AR agonist, CGS21680 (30 nM), and a larger LTP amplitude (54 ± 8 % vs. 21 ± 5 % in controls) that was restored to control levels (30 ± 10 %) by the A2AR antagonist, SCH58261 (50 nM). Notably, caffeine intake reverted memory deficits and reverted the loss of hippocampal synaptic markers but did not affect helpless or anxiety behavior. These results reinforce the validity of HM as an animal model of depression by showing that they also display reference memory deficits. Furthermore, caffeine intake selectively reverted memory but not mood deficits displayed by HM, which are associated with an increased density and functional impact of hippocampal A2AR controlling synaptic glutamatergic function.

Aged mice receiving caffeine since adulthood show distinct patterns of anxiety-related behavior.

Caffeine is the psychostimulant most consumed worldwide. Anxiogenic effects of caffeine have been described in adult animals with controversial findings about its anxiogenic potential. Besides, the effects of caffeine on anxiety with aging are still poorly known. In this study, adult mice (6months old) started to receive caffeine (0.3 and 1.0mg/mL, drinking water) during 12-14months only in the light cycle and at weekdays. The open field (OF) and elevated plus maze (EPM) testing were used to determine the effects of caffeine on anxiety-related behavior in adult and aged mice (18-20months old). Because aging alters synaptic proteins, we also evaluated SNAP-25 (as a nerve terminals marker), GFAP (as an astrocyte marker) and adenosine A1 and A2A receptors levels in the cortex. According to the OF analysis, caffeine did not change both hypolocomotion and anxiety with aging. However, aged mice showed less anxiety behavior in the EPM, but after receiving caffeine (0.3mg/mL) during adulthood they were anxious as adult mice. While SNAP-25 and adenosine A2A receptors increased with aging, both GFAP and adenosine A1 receptors were not affected. Caffeine at moderate dose prevented the age-related increase of the SNAP-25, with no effect on adenosine A2A receptors. The absence of effect for the highest dose suggests that tolerance to caffeine may have developed over time. Aged mice showed high responsiveness to the OF, being difficult to achieve any effect of caffeine. On the other hand this substance sustained the adult anxious behavior over time in a less stressful paradigm, and this effect was coincident with changes in the SNAP-25, suggesting the involvement of this synaptic protein in the ability of caffeine to preserve changes related to emotionality with aging.

Mechanisms involved in the antinociception induced by spinal administration of inosine or guanine in mice.

It is well known that adenine-based purines exert multiple effects on pain transmission. Recently, we have demonstrated that guanine-based purines may produce some antinociceptive effects against chemical and thermal pain in mice. The present study was designed to investigate the antinociceptive effects of intrathecal (i.t.) administration of inosine or guanine in mice. Additionally, investigation into the mechanisms of action of these purines, their general toxicity and measurements of CSF purine levels were performed. Animals received an i.t. injection of vehicle (30mN NaOH), inosine or guanine (up to 600nmol) and submitted to several pain models and behavioural paradigms. Guanine and inosine produced dose-dependent antinociceptive effects in the tail-flick, hot-plate, intraplantar (i.pl.) glutamate, i.pl. capsaicin and acetic acid pain models. Additionally, i.t. inosine inhibited the biting behaviour induced by spinal injection of capsaicin and i.t. guanine reduced the biting behaviour induced by spinal injection of glutamate or AMPA. Intrathecal administration of inosine (200nmol) induced an approximately 115-fold increase on CSF inosine levels. This study provides new evidence on the mechanism of action of extracellular guanine and inosine presenting antinociceptive effects following spinal administration. These effects seem to be related, at least partially, to the modulation of A1 adenosine receptors.

Caffeine acts through neuronal adenosine A2A receptors to prevent mood and memory dysfunction triggered by chronic stress.

The consumption of caffeine (an adenosine receptor antagonist) correlates inversely with depression and memory deterioration, and adenosine A2A receptor (A2AR) antagonists emerge as candidate therapeutic targets because they control aberrant synaptic plasticity and afford neuroprotection. Therefore we tested the ability of A2AR to control the behavioral, electrophysiological, and neurochemical modifications caused by chronic unpredictable stress (CUS), which alters hippocampal circuits, dampens mood and memory performance, and enhances susceptibility to depression. CUS for 3 wk in adult mice induced anxiogenic and helpless-like behavior and decreased memory performance. These behavioral changes were accompanied by synaptic alterations, typified by a decrease in synaptic plasticity and a reduced density of synaptic proteins (synaptosomal-associated protein 25, syntaxin, and vesicular glutamate transporter type 1), together with an increased density of A2AR in glutamatergic terminals in the hippocampus. Except for anxiety, for which results were mixed, CUS-induced behavioral and synaptic alterations were prevented by (i) caffeine (1 g/L in the drinking water, starting 3 wk before and continued throughout CUS); (ii) the selective A2AR antagonist KW6002 (3 mg/kg, p.o.); (iii) global A2AR deletion; and (iv) selective A2AR deletion in forebrain neurons. Notably, A2AR blockade was not only prophylactic but also therapeutically efficacious, because a 3-wk treatment with the A2AR antagonist SCH58261 (0.1 mg/kg, i.p.) reversed the mood and synaptic dysfunction caused by CUS. These results herald a key role for synaptic A2AR in the control of chronic stress-induced modifications and suggest A2AR as candidate targets to alleviate the consequences of chronic stress on brain function.

Prenatal caffeine intake differently affects synaptic proteins during fetal brain development.

Caffeine is the psychostimulant most consumed worldwide. However, little is known about its effects during fetal brain development. In this study, adult female Wistar rats received caffeine in drinking water (0.1, 0.3 and 1.0 g/L) during the active cycle in weekdays, two weeks before mating and throughout pregnancy. Cerebral cortex and hippocampus from embryonic stages 18 or 20 (E18 or E20, respectively) were collected for immunodetection of the following synaptic proteins: brain-derived neurotrophic factor (BDNF), TrkB receptor, Sonic Hedgehog (Shh), Growth Associated Protein 43 (GAP-43) and Synaptosomal-associated Protein 25 (SNAP-25). Besides, the estimation of NeuN-stained nuclei (mature neurons) and non-neuronal nuclei was verified in both brain regions and embryonic periods. Caffeine (1.0 g/L) decreased the body weight of embryos at E20. Cortical BDNF at E18 was decreased by caffeine (1.0 g/L), while it increased at E20, with no major effects on TrkB receptors. In the hippocampus, caffeine decreased TrkB receptor only at E18, with no effects on BDNF. Moderate and high doses of caffeine promoted an increase in Shh in both brain regions at E18, and in the hippocampus at E20. Caffeine (0.3g/L) decreased GAP-43 only in the hippocampus at E18. The NeuN-stained nuclei increased in the cortex at E20 by lower dose and in the hippocampus at E18 by moderate dose. Our data revealed that caffeine transitorily affect synaptic proteins during fetal brain development. The increased number of NeuN-stained nuclei by prenatal caffeine suggests a possible acceleration of the telencephalon maturation. Although some modifications in the synaptic proteins were transient, our data suggest that caffeine even in lower doses may alter the fetal brain development.

The Janus face of caffeine.

Caffeine is certainly the psychostimulant substance most consumed worldwide. Over the past years, chronic consumption of caffeine has been associated with prevention of cognitive decline associated to aging and mnemonic deficits of brain disorders. While its preventive effects have been reported extensively, the cognitive enhancer properties of caffeine are relatively under debate. Surprisingly, there are scarce detailed ontogenetic studies focusing on neurochemical parameters related to the effects of caffeine during prenatal and earlier postnatal periods. Furthermore, despite the large number of epidemiological studies, it remains unclear how safe is caffeine consumption during pregnancy and brain development. Thus, the purpose of this article is to review what is currently known about the actions of caffeine intake on neurobehavioral and adenosinergic system during brain development. We also reviewed other neurochemical systems affected by caffeine, but not only during brain development. Besides, some recent epidemiological studies were also outlined with the control of "pregnancy signal" as confounding variable. The idea is to tease out how studies on the impact of caffeine consumption during brain development deserve more attention and further investigation.