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 exposure during rat brain development causes memory impairment in a sex selective manner that is offset by caffeine consumption throughout life.

Caffeine is the psychostimulant most consumed worldwide. In moderate doses, it affords a beneficial effect in adults and upon aging, but has a deleterious effect during brain development. We now tested if caffeine consumption by rats (0.1, 0.3, 1.0 g/L in the drinking water, only during active cycle and weekdays) during adulthood could revert the potentially negative effects of caffeine during early life. Thus, we compared caffeine intake starting 15 days before mating and lasting either up to weaning (development) or up to adulthood, on behavior and synaptic proteins in male and female rats. Recognition memory was impaired only in female rats receiving caffeine (0.3 and 1.0 g/L) during development, coincident with increased proBDNF and unchanged BDNF levels in the hippocampus. Caffeine in both treatment regimens caused hyperlocomotion only in male rats, whereas anxiety-related behavior was attenuated in both sexes by caffeine (1.0 g/L) throughout life. Both caffeine treatment regimens decreased GFAP (as an astrocyte marker) and SNAP-25 (as a nerve terminals marker) in the hippocampus from male rats. TrkB receptor was decreased in the hippocampus from both sexes and treatment regimens. These findings revealed that caffeine intake during a specific time window of brain development promotes sex-dependent behavioral outcomes related to modification in BDNF signaling. Furthermore, caffeine throughout life can overcome the deleterious effects of caffeine on recognition memory during brain development in female rats.

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.

Treadmill running frequency on anxiety and hippocampal adenosine receptors density in adult and middle-aged rats.

Physical exercise protocols have varied widely across studies raising the question of whether there is an optimal intensity, duration and frequency that would produce maximal benefits in attenuating symptoms related to anxiety disorders. Although physical exercise causes modifications in neurotransmission systems, the involvement of neuromodulators such as adenosine has not been investigated after chronic exercise training. Anxiety-related behavior was assessed in the elevated plus-maze in adult and middle-aged rats submitted to 8 weeks of treadmill running 1, 3 or 7 days/week. The speed of running was weekly adjusted to maintain moderate intensity. The hippocampal adenosine A1 and A2A receptors densities were also assessed. Treadmill running protocol was efficient in increasing physical exercise capacity in adult and middle-aged rats. All frequencies of treadmill running equally decreased the time spent in the open arms in adult animals. Middle-aged treadmill control rats presented lower time spent in the open arms than adult treadmill control rats. However, treadmill running one day/week reversed this age effect. Adenosine A1 receptor was not changed between groups, but treadmill running counteracted the age-related increase in adenosine A2A receptors. Although treadmill running, independent from frequency, triggered anxiety in adult rats and treadmill running one day/week reversed the age-related anxiety, no consistent relationship was found with hippocampal adenosine receptors densities. Thus, our data suggest that as a complementary therapy in the management of mental disturbances, the frequency and intensity of physical exercise should be taken into account according to age. Besides, this is the first study reporting the modulation of adenosine receptors after chronic physical exercise, which could be important to prevent neurological disorders associated to increase in adenosine A2A receptors.

Enriched environment effects on behavior, memory and BDNF in low and high exploratory mice.

Environmental enrichment (EE) has been largely used to investigate behavioral modifications and neuroplasticity in the adult brain both in normal and pathological conditions. The interaction between individual behavioral traits with EE responsiveness has not been investigated within the same strain. By using two extremes of CF1 mice that differ by their exploratory behavior in the Open Field (OF) task (Kazlauckas V, 2005), denominated as Low (LE) and High (HE) Exploratory Mice, the present study evaluated if EE during adulthood could modify the putative differences between LE and HE mice on exploratory behavior, memory performance and hippocampal BDNF levels. To this end, we investigated the effect of adult LE and HE mice after 2 months of enriched or standard housing conditions on the open field, on novel object recognition, on the inhibitory avoidance task and on hippocampal BDNF immunocontent. LE showed low exploratory behavior, less retention in the inhibitory avoidance and lower hippocampal BDNF levels. EE enhanced exploratory behavior, memory performance and hippocampal BDNF levels both in LE and HE mice. Importantly, the general profile of LE mice submitted to EE was similar to HE mice housed in standard conditions. These results show that internalized behavior of LE mice can be significantly modified by exposure to an enriched environment even during adulthood. These observations may contribute to investigate biological mechanisms and therapeutical interventions for individuals with internalized psychiatric disorders.

Blockade of adenosine A(1) receptors prevents methylphenidate-induced impairment of object recognition task in adult mice.

Methylphenidate (MPH) is the preferred treatment used for attention-deficit/hyperactivity disorder (ADHD). Recently, misuse for MPH due to its apparent cognitive enhancer properties has been reported. Adenosine is a neuromodulator known to exert influence on the dopaminergic neurotransmission, which is the main pharmacological target of MPH. We have reported that an overdosage of MPH up-regulates adenosine A(1) receptors in the frontal cortex, but this receptor was not involved in its anxiolytic effects. In this study, the role of adenosine A(1) receptor was investigated on MPH-induced effects on aversive and recognition memory in adult mice. Adult mice received acute and chronic (15 days) administration of methylphenidate (5mg/kg, i.p.), or an acute overdosage (50mg/kg, i.p) in order to model misuse. Memory was assessed in the inhibitory avoidance and object recognition task. Acute administration 5mg/kg improved whereas 50mg/kg disrupted recognition memory and decreased performance in the inhibitory avoidance task. Chronic administration did not cause any effect on memory, but decreased adenosine A(1) receptors immunocontent in the frontal cortex. The selective adenosine A(1) receptor antagonist, (DPCPX 1mg/kg, i.p.), prevented methylphenidate-triggered recognition memory impairment. Our findings showed that recognition memory rather than aversive memory was differently affected by acute administration at both doses. Memory recognition was fully impaired by the overdosage, suggesting that misuse can be harmful for cognitive functions. The adenosinergic system via A(1) receptors may play a role in the methylphenidate actions probably by interfering with dopamine-enhancing properties of this drug.

Distinctive effects of unpredictable subchronic stress on memory, serum corticosterone and hippocampal BDNF levels in high and low exploratory mice.

Stress affects learning and memory processes and sensitivity to stress greatly varies between individuals. We studied behavioral and neurobiological effects of unpredictable subchronic stress (USCS) in two behavioral extremes of mice from the same strain (CF1) selected by their exploratory behavior of the central arena of an open field. The top and bottom 25% explorers were classified as low exploratory (LE) and high exploratory (HE) mice, respectively. The open field task, the novel object recognition task (NOR), sucrose intake and tail suspension task were evaluated in LE and HE groups exposed to USCS for two weeks or control conditions. Also serum corticosterone and hippocampal BDNF and S100B levels were analyzed. Both stressed groups exhibited less exploratory activity when submitted to USCS, but their difference in exploratory behavior remained. This short stress protocol did not induce changes in sucrose intake or immobility in the tail suspension task. Also, LE mice exhibited impaired NOR performance after USCS, whereas HE mice changed their pattern of exploration towards less exploration of the familiar object. HE had lower corticosterone levels than LE mice, but corticosterone levels increased after stress only in HE mice. Hippocampal BDNF in LE was lower than in HE but decreased after USCS only in HE mice, whereas S100B levels were not different between groups and did not change with USCS. In conclusion, our results suggest that individual differences in exploratory behavior in rodents from the same strain influence cognitive and biochemical response to stress.

Adenosine A1 receptors are modified by acute treatment with methylphenidate in adult mice.

In recent years misuse of methylphenidate (MPH) has been reported. The main pharmacological target of methylphenidate is the dopaminergic system. Adenosine is a neuromodulator that influences the dopaminergic neurotransmission, but studies on MPH and adenosine are still lacking. In this study, adult mice were acutely treated with MPH (5mg/kg, i.p.) and to model misuse, they received an acute overdosage (50mg/kg, i.p). The involvement of adenosine A(1) receptors in anxiety-related behavior and locomotor and exploratory activity was examined. The administration of methylphenidate (5 and 50mg/kg) 30 min before the exposure to open field arena did not modify locomotor activity. The anxiolytic-like behavior was observed with both doses of MPH as revealed by the increase on the number of entries and the time spent in the open arms in the elevated plus-maze. Pre treatment with selective adenosine A(1) receptor antagonist (DPCPX 1mg/kg, i.p.) did not prevent anxiolytic effect caused by MPH 50mg/kg. Immunoblotting of frontal cortex and hippocampal extracts revealed that MPH 50mg/kg increased 88% adenosine A(1) receptor density in the frontal cortex. Extracts from hippocampus did not reveal any differences in the adenosine A(1) receptor density. Our findings ruled out the participation of adenosine A(1) receptors on the MPH-triggered anxiolytic effects. However, the density of adenosine A(1) receptors increased in a brain area strictly involved in the MPH-mediated effects. Thus, the adenosinergic system may play a role in the methylphenidate actions in the central nervous system.

Caffeine prevents disruption of memory consolidation in the inhibitory avoidance and novel object recognition tasks by scopolamine in adult mice.

Caffeine is a psychostimulant with positive effects on cognition. Recent studies have suggested the participation of the cholinergic system in the effects of caffeine on wakefulness. However, there are few studies assessing the contribution of cholinergic system in the cognitive enhancer properties of caffeine. In the present study, the effects of a dose and schedule of administration of caffeine that improved memory recognition were investigated on scopolamine-induced impairment of memory in adult mice. Inhibitory avoidance and novel object recognition tasks were used to assess learning and memory. Caffeine (10mg/kg, i.p.) was administered during 4 consecutive days, and the treatment was interrupted 24h before scopolamine administration (2mg/kg, i.p.). Scopolamine was administered prior to or immediately after training. Short-term and long-term memory was evaluated in both tasks. In the novel object recognition task, pre treatment with caffeine prevented the disruption of short- and long-term memory by scopolamine. In the inhibitory avoidance task, caffeine prevented short- but not long-term memory disruption by pre training administration of scopolamine. Caffeine prevented short- and long-term memory disruption by post training administration of scopolamine. Both treatments did not affect locomotor activity of the animals. These findings suggest that acute treatment with caffeine followed by its withdrawal may be effective against cholinergic-induced disruption of memory assessed in an aversive and non-aversive task. Finally, our results revealed that the cholinergic system is involved in the positive effects of caffeine on cognitive functions.

Caffeine improves adult mice performance in the object recognition task and increases BDNF and TrkB independent on phospho-CREB immunocontent in the hippocampus.

Caffeine is one of the most psychostimulants consumed all over the world that usually presents positive effects on cognition. In this study, effects of caffeine on mice performance in the object recognition task were tested in different intertrial intervals. In addition, it was analyzed the effects of caffeine on brain derived neurotrophic factor (BDNF) and its receptor, TrkB, immunocontent to try to establish a connection between the behavioral finding and BDNF, one of the neurotrophins strictly involved in memory and learning process. CF1 mice were treated during 4 consecutive days with saline (0.9g%, i.p.) or caffeine (10mg/kg, i.p., equivalent dose corresponding to 2-3 cups of coffee). Caffeine treatment was interrupted 24h before the object recognition task analysis. In the test session performed 15min after training session, caffeine-treated mice recognized more efficiently both the familiar and the novel object. In the test session performed 90min and 24h after training session, caffeine did not change the time spent in the familiar object but increased the object recognition index, when compared to control group. Western blotting analysis of hippocampus from caffeine-treated mice revealed an increase in BDNF and TrkB immunocontent, compared to their saline-matched controls. Phospho-CREB immunocontent did not change with caffeine treatment. Our results suggest that acute treatment with caffeine improves recognition memory, and this effect may be related to an increase of the BDNF and TrkB immunocontent in the hippocampus.

Selenium compounds counteract the stimulation of ecto-nucleotidase activities in rat cultured cerebellar granule cells: putative correlation with neuroprotective effects.

Glutamate is the main excitatory neurotransmitter in brain involved in pathophysiology of several brain injuries. In this context, glutamate showed to stimulate ecto-nucleotidase activities in cerebellar granule cells increasing extracellular adenosine levels, an important neuromodulator in the CNS able to prevent cell damage. The organoselenium compounds, such as ebselen and diphenyl diselenide [(PhSe)(2)], display neuroprotective activities mediated at least in part by their antioxidant and anti-inflammatory properties. Ebselen was described to prevent glutamate-induced lipid peroxidation and cell death in cerebellar granule cells and (PhSe)(2) modify glutamatergic synapse parameters in vitro and in vivo. In the present study, we investigated the effects of ebselen or (PhSe)(2) on glutamate-induced stimulation of ecto-nucleotidase activities in rat cultured cerebellar granule cells. Glutamate increased nucleotide hydrolysis at lower concentrations (10 and 100 microM) than described in the literature and this effect was counteracted by both organoselenium compounds tested. Based on these results, we investigated the association of organoselenium effects with their antioxidant properties searching for redox site modulation by using the alkylant agent N-ethylmaleimide (NEM). Our results suggest that selenium compounds, as well as the well-known antioxidant trolox, can avoid the increase on glutamate-induced stimulation of ecto-nucleotidase activities probably due to their antioxidant properties.

Diphenyl diselenide exerts anxiolytic-like effect in Wistar rats: putative roles of GABAA and 5HT receptors.

Diphenyl diselenide [(PhSe)2] is an organoselenium compound which presents pharmacological antioxidant, anti-inflammatory, antinociceptive and antidepressant properties. The present study was designed to investigate the anxiolytic effect of (PhSe)2 in rats, employing the elevated plus maze task. The involvement of 5HT and GABA receptors in the anxiolytic-like effect was also evaluated. (PhSe)2 (5, 25 and 50 micromol/kg, i.p.) did not affect locomotor activity as evaluated in the open open-field test, and learning and memory when assessed in the inhibitory foot-shock avoidance task. However, (PhSe)2 at the 50 micromol/kg dose produced signs of an anxiolytic action, namely a decreased number of fecal boli in the open-field arena and an increased time spent in as well as an increased number of entries to the open arms of the elevated plus maze test. To evaluate the role of GABA and 5HT receptors in the anxiolytic-like effect of (PhSe)2, a selective GABAA receptor antagonist bicuculline, (0.75 mg/kg, i.p.), a non-selective 5HT2A/2C receptor antagonist, ritanserin (2 mg/kg, i.p.), a selective 5HT2A receptor antagonist, ketanserin (1 mg/kg, i.p.), and a selective 5HT1A receptor antagonist, WAY100635 (0.1 mg/kg, i.p.) were used. All the antagonists used were able to abolish the anxiolytic effect of (PhSe)2 suggesting that GABAA and 5HT receptors may play a role in the pharmacological property of this selenocompound in the central nervous system.