Neonatal testosterone voids sexually differentiated microglia morphology and behavior.

The involvement of immunity in psychiatric disorders, such as anxiety, is typified by the morphologic adaptation of microglia, immune cells of the brain, to anxiogenic stimuli. We previously reported sexually differentiated microglia morphology in adult rodents, in brain locations implicated in anxiety, including the pre-frontal cortex. These physiologic differences likely drive sex-dependent patterns of microglia morphologic remodeling in response to varied stress conditions in different periods of life, that correlate with sex-dependent behavioral adaptation to anxiogenic stimuli. The time-window of appearance of sex differences in microglia, correlating with sex-specific behavioral performance in anxiogenic conditions are still unknown. In rodents, a postnatal peak of the sexual hormone testosterone is determinant for the so-called brain masculinization and sex-determined behavioral traits. In the present work we aim to clarify if differences in microglia morphology are present at birth or can be driven by postnatal testosterone and impacts on the ability to deal with an anxiogenic context. Differences in microglia morphology are not present at birth, but are observable at adolescence (increased complexity of male microglia, particularly in branches more proximal to the soma), when differences in behavior are also observed. Our data also show that adolescent females neonatally treated with testosterone exhibit masculinized microglia and behavior. Importantly, between adolescence and adulthood, a sex-determined shift in the pattern of complexity takes place and microglia from females become more complex. When testosterone is administered, this morphological effect is partially abolished, approximating microglia and behavior to the male phenotype.

Microglia sequelae: brain signature of innate immunity in schizophrenia.

Schizophrenia is a psychiatric disorder with significant impact on individuals and society. The current pharmacologic treatment, which principally alleviates psychosis, is focused on neurotransmitters modulation, relying on drugs with severe side effects and ineffectiveness in a significant percentage of cases. Therefore, and due to difficulties inherent to diagnosis and treatment, it is vital to reassess alternative cellular and molecular drug targets. Distinct risk factors - genetic, developmental, epigenetic, and environmental - have been associated with disease onset and progression, giving rise to the proposal of different pathophysiological mechanisms and putative pharmacological targets. Immunity is involved and, particularly microglia - innate immune cells of the central nervous system, critically involved in brain development - have captured attention as cellular players. Microglia undergo marked morphologic and functional alterations in the human disease, as well as in animal models of schizophrenia, as reported in several original papers. We cluster the main findings of clinical studies by groups of patients: (1) at ultra-high risk of psychosis, (2) with a first episode of psychosis or recent-onset schizophrenia, and (3) with chronic schizophrenia; in translational studies, we highlight the time window of appearance of particular microglia alterations in the most well studied animal model in the field (maternal immune activation). The organization of clinical and translational findings based on schizophrenia-associated microglia changes in different phases of the disease course may help defining a temporal pattern of microglia changes and may drive the design of novel therapeutic strategies.

The Duration of Stress Determines Sex Specificities in the Vulnerability to Depression and in the Morphologic Remodeling of Neurons and Microglia.

Stress exposure has been shown to induce a variety of molecular and functional alterations associated with anxiety and depression. Some studies suggest that microglia, the immune cells of the brain, play a significant role in determining neuronal and behavioral responses to chronic stress and also contribute to the development of stress-related psychopathologies. However, little is known about the impact of the duration of stress exposure upon microglia and neurons morphology, particularly considering sex differences. This issue deserves particular investigation, considering that the process of morphologic remodeling of neurons and microglia is usually accompanied by functional changes with behavioral expression. Here, we examine the effects of short and long unpredictable chronic mild stress (uCMS) protocols on behavior, evaluating in parallel microglia and neurons morphology in the dorsal hippocampus (dHIP) and in the nucleus accumbens (NAc), two brain regions involved in the etiology of depression. We report that long-term uCMS induced more behavioral alterations in males, which present anxiety and depression-like phenotypes (anhedonia and helplessness behavior), while females only display anxiety-like behavior. After short-term uCMS, both sexes presented anxiety-like behavior. Microglia cells undergo a process of morphologic adaptation to short-term uCMS, dependent on sex, in the NAc: we observed a hypertrophy in males and an atrophy in females, transient effects that do not persist after long-term uCMS. In the dHIP, the morphologic adaptation of microglia is only observed in females (hypertrophy) and after the protocol of long uCMS. Interestingly, males are more vulnerable to neuronal morphological alterations in a region-specific manner: dendritic atrophy in granule neurons of the dHIP and hypertrophy in the medium spiny neurons of the NAc, both after short- or long-term uCMS. The morphology of neurons in these brain regions were not affected in females. These findings raise the possibility that, by differentially affecting neurons and microglia in dHIP and NAc, chronic stress may contribute for differences in the clinical presentation of stress-related disorders under the control of sex-specific mechanisms.

Sex-specific changes in peripheral metabolism in a model of chronic anxiety induced by prenatal stress.

BACKGROUND: Prenatal stress is associated with increased susceptibility to psychiatric and metabolic disorders later in life. Prenatal exposure to stress mediators may have sex-dependent effects on offspring brain and metabolic function, promoting a sex-specific vulnerability to psychopathology and metabolic alterations at adulthood. In this work, the impact of prenatal stress on glucose homeostasis and peripheral metabolism of male and female offspring was investigated in a chronic anxiety animal model. METHODS: Pregnant Wistar rats were injected with saline or glucocorticoid (dexamethasone: 1 mg/kg, subcutaneous) at gestational days 18 and 19. Male and female offspring weight was monitored, and anxious-like behaviour and peripheral insulin-sensitive tissues were analysed at adulthood. RESULTS: At birth, females and males prenatally exposed to stress presented decreased body weight which remained low in females. At adulthood, a morphological disorganization of the Langerhans islets was observed in both sexes prenatally exposed to stress, yet not changes in insulin levels were detected. Also, prenatal stress increased glucose transporter 4 (GLUT-4) levels in female and male adipose tissues and decreased insulin receptor levels in the liver and skeleton muscle but only in females. CONCLUSIONS: Exposure to stress mediators in critical periods of development negatively affects behaviour and metabolism. Prenatal stress programmes offspring peripheral metabolism in a sex-specific manner, emphasizing that the response to stress in critical periods of development may be sex-specific having each sex different vulnerabilities to psychiatric and metabolic disorders. Considering sex-specificities may provide critical clues for the design of preventive strategies and for early therapeutic intervention.

Resilience to stress and sex-specific remodeling of microglia and neuronal morphology in a rat model of anxiety and anhedonia.

Prenatal exposure to stress or glucocorticoids (GC) is associated with the appearance of psychiatric diseases later in life. Microglia, the immune cells of the brain, are altered in stress-related disorders. Synthetic GC such as dexamethasone (DEX) are commonly prescribed in case of preterm risk labour in order to promote fetal lung maturation. Recently, we reported long-lasting differences in microglia morphology in a model of in utero exposure to DEX (iuDEX), that presents an anxious phenotype. However, it is still unclear if stress differentially affects iuDEX males and females. In this work, we evaluated how iuDEX animals of both sexes cope with chronic mild stress for 2 weeks. We evaluated emotional behavior and microglia and neuronal morphology in the dorsal hippocampus (dHIP) and nucleus accumbens (NAc), two brain regions involved in emotion-related disorders. We report that males and females prenatally exposed to DEX have better performance in anxiety- and depression-related behavioral tests after chronic stress exposure in adulthood than non-exposed animals. Interestingly, iuDEX animals present sex-dependent changes in microglia morphology in the dHIP (hypertrophy in females) and in the NAc (atrophy in females and hypertrophy in males). After chronic stress, these cells undergo sex-specific morphological remodeling. Paralleled to these alterations in cytoarchitecture of microglia, we report inter-regional differences in dendritic morphology in a sex-specific manner. iuDEX females present fewer complex neurons in the NAc, whereas iuDEX males presented less complex neuronal morphology in the dHIP. Interestingly, these alterations were modified by stress exposure. Our work shows that stressful events during pregnancy can exert a preserved sex-specific effect in adulthood. Although the role of the observed cellular remodeling is still unknown, sex-specific differences in microglia plasticity induced by long-term stress exposure may anticipate differences in drug efficacy in the context of stress-induced anxiety- or depression-related behaviors.

Sex differences in offspring neurodevelopment, cognitive performance and microglia morphology associated with maternal diabetes: Putative targets for insulin therapy.

Diabetes during pregnancy has been shown to affect the central nervous system (CNS) of the offspring, resulting in short- and long-term adverse effects. Children of diabetic mothers are more likely to develop cognitive impairment, also having increased susceptibility to psychiatric disorders. Microglia, the immune cells of the CNS, work as sensors of environmental changes, namely metabolic challenges, as early as the intrauterine period. During this period, microglia is actively involved in processes of neurogenesis, synaptic pruning and detection of any environmental alteration that may impact brain development. The remarkable sex dimorphism in neurodevelopment, as well as sex differences in the morphology and immune function of microglia during development, led us to clarify if maternal diabetes affects specific behavioral traits and microglia morphology during infancy in a sex-specific manner. Another important goal of this study was to clarify if insulin, the gold standard treatment of diabetes during gestation, could prevent maternal diabetes-induced behavioral changes, as well as microglia morphology, also considering sex specificities. Other molecular and cellular players potentially involved in the link between changes in metabolism and behavior were also analyzed in the hippocampus, a brain region implicated in cognition and other behavioral outcomes. Diabetes during pregnancy globally delayed female and male offspring development and was associated with impairments in recognition memory, but only in female offspring. In line with these results, at early and late infancy, some molecular and cellular markers were altered in offspring hippocampus in a sex-specific manner. The strict control of glycemia by insulin during pregnancy prevented most of the negative effects induced by uncontrolled hyperglycemia. Notably, insulin administration to diabetic dams may also modulate offspring development in a way that differs from what is observed in physiological conditions, since it promoted the expedited acquisition of developmental milestones and of discrimination ability at memory test, also inducing a hyper-ramification of male and female hippocampal microglia. Importantly, this study highlights the importance of analyzing the impact of maternal diabetes and insulin therapy, taking into account sex differences, since male and female present different vulnerabilities to hyperglycemia in this critical period of life.

Microglia cytoarchitecture in the brain of adenosine A2A receptor knockout mice: Brain region and sex specificities.

Microglia cells exert a critical role in brain development, mainly supported by their immune functions, which predicts an impact on the genesis of psychiatric disorders. In fact, microglia stress during gestation is, for instance, associated with chronic anxiety and cognitive deficits accompanied by long-lasting, region- and sex-specific changes in microglia morphology. We recently reported that the pattern of microglia morphologic plasticity, which is sex-determined, impacts on anxious-like behaviour and cognition. We also reported that the pharmacologic blockade of adenosine A2A receptors (A2A R) is able to reshape microglia morphology, in a sex-specific manner and with behavioural sequelae. In order to better understand the role of A2A R in the sex differentiation of microglia, we now compared their morphology in wild-type and A2A R knockout male and female C57BL/6 mice in two cardinal brain regions implicated in anxiety-like behaviour and cognition, the prefrontal cortex (PFC) and the dorsal hippocampus (dHIP). We report interregional differences between PFC and dHIP in a sex-specific manner: while males presented more complex microglia in the dHIP, microglia from females had a more complex morphology in the PFC. Surprisingly, the genetic deletion of A2A R did not alter these sex differences, but promoted the exclusive remodelling (increase in complexity) in PFC microglia from females. These findings further support the existence of a heterogeneous microglial network, distinct between sexes and brain regions, and help characterizing the role of A2A R in the sex- and brain region-specific morphologic differentiation of microglia.

Impairment of Axonal Transport in Diabetes: Focus on the Putative Mechanisms Underlying Peripheral and Central Neuropathies.

Diabetes mellitus is a chronic disease with numerous complications that severely impact on the quality of life of patients. Different neuropathies may arise as complications associated with the nervous system, both peripherally and at the central level. The mechanisms behind these neuronal complications are far from being clarified, but axonal transport impairment, a vital process for neuronal physiology, has been described in the context of experimental diabetes. Alterations in neuronal cytoskeleton and motor proteins, deficits in ATP supply or neuroinflammation, as processes that disturb the effective transport of cargoes along the axon, were reported as putative causes of axonal impairment, ultimately leading to axonal degeneration. The main goal of the present review is to reunite the main studies in the literature exploring diabetes-induced alterations likely involved in axonal transport deficits, and call the attention for the uttermost importance of further exploring the field. Understanding the mechanisms underlying neuronal deficits in diabetes is crucial for the development of new therapeutic strategies to prevent neuronal degeneration in diabetes and related neuropathies.

Region-specific control of microglia by adenosine A2A receptors: uncoupling anxiety and associated cognitive deficits in female rats.

Epidemiologic studies have provided compelling evidence that prenatal stress, through excessive maternal glucocorticoids exposure, is associated with psychiatric disorders later in life. We have recently reported that anxiety associated with prenatal exposure to dexamethasone (DEX, a synthetic glucocorticoid) correlates with a gender-specific remodeling of microglia in the medial prefrontal cortex (mPFC), a core brain region in anxiety-related disorders. Gender differences in microglia morphology, the higher prevalence of anxiety in women and the negative impact of anxiety in cognition, led us to specifically evaluate cognitive behavior and associated circuits (namely mPFC-dorsal hippocampus, dHIP), as well as microglia morphology in female rats prenatally exposed to dexamethasone (in utero DEX, iuDEX). We report that iuDEX impaired recognition memory and deteriorated neuronal synchronization between mPFC and dHIP. These functional deficits are paralleled by microglia hyper-ramification in the dHIP and decreased ramification in the mPFC, showing a heterogeneous remodeling of microglia morphology, both postnatally and at adulthood in different brain regions, that differently affect mood and cognition. The chronic blockade of adenosine A2A receptors (A2A R), which are core regulators of microglia morphology and physiology, ameliorated the cognitive deficits, but not the anxiety-like behavior. Notably, A2A R blockade rectified both microglia morphology in the dHIP and the lack of mPFC-dHIP synchronization, further heralding their role in cognitive function.

Adenosine A2A Receptor Blockade Modulates Glucocorticoid-Induced Morphological Alterations in Axons, But Not in Dendrites, of Hippocampal Neurons.

The exposure to supra-physiological levels of glucocorticoids in prenatal life can lead to a long-term impact in brain cytoarchitecture, increasing the susceptibility to neuropsychiatric disorders. Dexamethasone, an exogenous glucocorticoid widely used in pregnant women in risk of preterm delivery, is associated with higher rates of neuropsychiatric conditions throughout life of the descendants. In animal models, prenatal dexamethasone exposure leads to anxious-like behavior and increased susceptibility to depressive-like behavior in adulthood, concomitant with alterations in neuronal morphology in brain regions implicated in the control of emotions and mood. The pharmacologic blockade of the purinergic adenosine A2A receptor, which was previously described as anxiolytic, is also able to modulate neuronal morphology, namely in the hippocampus. Additionally, recent observations point to an interaction between glucocorticoid receptors (GRs) and adenosine A2A receptors. In this work, we explored the impact of dexamethasone on neuronal morphology, and the putative implication of adenosine A2A receptor in the mediation of dexamethasone effects. We report that in vitro hippocampal neurons exposed to dexamethasone (250 nM), in the early phases of development, exhibit a polarized morphology alteration: dendritic atrophy and axonal hypertrophy. While the effect of dexamethasone in the axon is dependent on the activation of adenosine A2A receptor, the effect in the dendrites relies on the activation of GRs, regardless of the activation of adenosine A2A receptor. These results support the hypothesis of the interaction between GRs and adenosine A2A receptors and the potential therapeutic value of modulating adenosine A2A receptors activation in order to prevent glucocorticoid-induced alterations in developing neurons.

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.

Different danger signals differently impact on microglial proliferation through alterations of ATP release and extracellular metabolism.

Microglia rely on their ability to proliferate in the brain parenchyma to sustain brain innate immunity and participate in the reaction to brain damage. We now studied the influence of different danger signals activating microglia, both internal (typified by glutamate, associated with brain damage) and external (using a bacterial lipopolysaccharide, LPS), on the proliferation of microglia cells. We found that LPS (100 ng/mL) increased, whereas glutamate (0.5 mM) decreased proliferation. Notably, LPS decreased whereas glutamate increased the extracellular levels of ATP. In contrast, LPS increased whereas glutamate decreased the extracellular catabolism of ATP into adenosine through ecto-nucleotidases and ecto-5'-nucleotidase. Finally, apyrase (degrades extracellular ATP) abrogated glutamate-induced inhibition of microglia proliferation; conversely, inhibitors of ecto-nucleotidases (ARL67156 or α,ß-methylene ADP) and adenosine deaminase (degrades extracellular adenosine) abrogated the LPS-induced increase of microglia proliferation, which was blocked by a selective A2A receptor antagonist, SCH58261 (50 nM). Overall, these results highlight the importance of the extracellular purinergic metabolism to format microglia proliferation and influence the spatio-temporal profile of neuroinflammation in different conditions of brain damage.

Depression as a Glial-Based Synaptic Dysfunction.

Recent studies combining pharmacological, behavioral, electrophysiological and molecular approaches indicate that depression results from maladaptive neuroplastic processes occurring in defined frontolimbic circuits responsible for emotional processing such as the prefrontal cortex, hippocampus, amygdala and ventral striatum. However, the exact mechanisms controlling synaptic plasticity that are disrupted to trigger depressive conditions have not been elucidated. Since glial cells (astrocytes and microglia) tightly and dynamically interact with synapses, engaging a bi-directional communication critical for the processing of synaptic information, we now revisit the role of glial cells in the etiology of depression focusing on a dysfunction of the "quad-partite" synapse. This interest is supported by the observations that depressive-like conditions are associated with a decreased density and hypofunction of astrocytes and with an increased microglia "activation" in frontolimbic regions, which is expected to contribute for the synaptic dysfunction present in depression. Furthermore, the traditional culprits of depression (glucocorticoids, biogenic amines, brain-derived neurotrophic factor, BDNF) affect glia functioning, whereas antidepressant treatments (serotonin-selective reuptake inhibitors, SSRIs, electroshocks, deep brain stimulation) recover glia functioning. In this context of a quad-partite synapse, systems modulating glia-synapse bidirectional communication-such as the purinergic neuromodulation system operated by adenosine 5'-triphosphate (ATP) and adenosine-emerge as promising candidates to "re-normalize" synaptic function by combining direct synaptic effects with an ability to also control astrocyte and microglia function. This proposed triple action of purines to control aberrant synaptic function illustrates the rationale to consider the interference with glia dysfunction as a mechanism of action driving the design of future pharmacological tools to manage depression.

Role of microglia adenosine A(2A) receptors in retinal and brain neurodegenerative diseases.

Neuroinflammation mediated by microglial cells in the brain has been commonly associated with neurodegenerative diseases. Whether this microglia-mediated neuroinflammation is cause or consequence of neurodegeneration is still a matter of controversy. However, it is unequivocal that chronic neuroinflammation plays a role in disease progression and halting that process represents a potential therapeutic strategy. The neuromodulator adenosine emerges as a promising targeting candidate based on its ability to regulate microglial proliferation, chemotaxis, and reactivity through the activation of its G protein coupled A2A receptor (A2AR). This is in striking agreement with the ability of A2AR blockade to control several brain diseases. Retinal degenerative diseases have been also associated with microglia-mediated neuroinflammation, but the role of A2AR has been scarcely explored. This review aims to compare inflammatory features of Parkinson's and Alzheimer's diseases with glaucoma and diabetic retinopathy, discussing the therapeutic potential of A2AR in these degenerative conditions.

Regulation of fear responses by striatal and extrastriatal adenosine A2A receptors in forebrain.

BACKGROUND: Adenosine A2A receptors (A2ARs) are enriched in the striatum but are also present at lower levels in the extrastriatal forebrain (i.e., hippocampus, cortex), integrating dopamine, glutamate, and brain-derived neurotrophic factor (BDNF) signaling, and are thus essential for striatal neuroplasticity and fear and anxiety behavior. METHODS: We tested two brain region-specific A2AR knockout lines with A2ARs selectively deleted either in the striatum (st-A2AR KO) or the entire forebrain (striatum, hippocampus, and cortex [fb-A2AR KO]) on fear and anxiety-related responses. We also examined the effect of hippocampus-specific A2AR deletion by local injection of adeno-associated virus type 5 (AAV5)-Cre into floxed-A2AR knockout mice. RESULTS: Selectively deleting A2ARs in the striatum increased Pavlovian fear conditioning (both context and tone) in st-A2AR KO mice, but extending the deletion to the rest of the forebrain apparently spared context fear conditioning and attenuated tone fear conditioning in fb-A2AR KO mice. Moreover, focal deletion of hippocampal A2ARs by AAV5-Cre injection selectively attenuated context (but not tone) fear conditioning. Deletion of A2ARs in the entire forebrain in fb-A2AR KO mice also produced an anxiolytic phenotype in both the elevated plus maze and open field tests, and increased the startle response. These extrastriatal forebrain A2AR behavioral effects were associated with reduced BDNF levels in the fb-A2AR KO hippocampus. CONCLUSIONS: This study provides evidence that inactivation of striatal A2ARs facilitates Pavlovian fear conditioning, while inactivation of extrastriatal A2ARs in the forebrain inhibits fear conditioning and also affects anxiety-related behavior.

GDNF control of the glutamatergic cortico-striatal pathway requires tonic activation of adenosine A receptors.

Glial cell line-derived neurotrophic factor (GDNF) affords neuroprotection in Parkinson's disease in accordance with its ability to bolster nigrostriatal innervation. We previously found that GDNF facilitates dopamine release in a manner dependent on adenosine A(2A) receptor activation. As motor dysfunction also involves modifications of striatal glutamatergic innervation, we now tested if GDNF and its receptor system, Ret (rearranged during transfection) and GDNF family receptor alpha1 controlled the cortico-striatal glutamatergic pathway in an A(2A) receptor-dependent manner. GDNF (10 ng/mL) enhanced (by approximately 13%) glutamate release from rat striatal nerve endings, an effect potentiated (up to approximately 30%) by the A(2A) receptor agonist CGS 21680 (10 nM) and prevented by the A(2A) receptor antagonist, SCH 58261 (50 nM). Triple immunocytochemical studies revealed that Ret and GDNF family receptor alpha1 were located in 50% of rat striatal glutamatergic terminals (immunopositive for vesicular glutamate transporters-1/2), where they were found to be co-located with A(2A) receptors. Activation of the glutamatergic system upon in vivo electrical stimulation of the rat cortico-striatal input induced striatal Ret phosphorylation that was prevented by pre-treatment with the A(2A) receptor antagonist, MSX-3 (3 mg/kg). The results provide the first functional and morphological evidence that GDNF controls cortico-striatal glutamatergic pathways in a manner largely dependent on the co-activation of adenosine A(2A) receptors.

Glial cell line-derived neurotrophic factor (GDNF) enhances dopamine release from striatal nerve endings in an adenosine A2A receptor-dependent manner.

Both glial cell line-derived neurotrophic factor (GDNF) and adenosine influence dopaminergic function in the striatum. We now evaluated the GDNF effect on dopamine release from rat striatal nerve endings and if this effect of GDNF is modulated by adenosine A(2A) receptors. Dopamine release was evoked twice (S(1) and S(2)); GDNF was added before S(2) and drugs used to modify GDNF actions were present during both stimulation periods. The effect of GDNF was taken as the change in the S(2)/S(1) ratio in the absence and in the presence of GDNF in the same experimental conditions. GDNF (3-30 ng/ml) increased dopamine release from K(+) (20 mM, 2 min) stimulated synaptosomes and electrically (2 Hz, 2 min) stimulated striatal slices, an effect dependent upon tonic adenosine A(2A) receptor activation, since it was blocked by the A(2A) receptor antagonist, SCH 58261 (50 nM). Activation of A(2A) receptors with CGS 21680 (10 nM) potentiated the effect of GDNF in synaptosomes. CGS 21680 also potentiated the effect of GDNF in striatal slices, providing that GABAergic transmission was inhibited; if not, the action of GDNF was attenuated by CGS 21680. Blockade of GABAergic transmission per se increased dopamine release, but attenuated the effect of GDNF upon dopamine release in slices. The results suggest that GDNF enhances dopamine release by acting presynaptically at the striatum, an action that requires adenosine A(2A) receptor activity. Furthermore, in striatal slices, the action of GDNF as well as its modulation by adenosine A(2A) receptor activation appears to be also under control of GABAergic transmission.

Anticancer activity of phenolic acids of natural or synthetic origin: a structure-activity study.

Several phenolic acids-caffeic and gallic acid derivatives-were synthesized and screened for their potential antiproliferative and cytotoxic properties, in different human cancer cell lines: mammary gland and cervix adenocarcinomas and lymphoblastic leukemia. The selected phenols were structurally related, which allowed us to gather important information regarding the structure-activity relationships underlying the biological activity of such compounds. This is proposed to be due to a balance between the antioxidant and pro-oxidant properties of this kind of agent. Distinct effects were found for different cell lines, which points to a significant specificity of action of the drugs tested. It was verified, for the types of cancer investigated, that the trihydroxylated derivatives yielded better results than the dihydroxylated ones. Tests in noncancerous cells, human lung fibroblasts, were also undertaken, in view of determining the toxic side effects of the compounds studied.