Os efeitos das queimadas na Amazônia em nível celular e molecular

The Amazon holds over half of the planet's remaining tropical forests and comprises the largest biodiversity in the world, accounting for approximately 60 % of the Brazilian territory. However, deforestation fires in the region causes serious problems to exposed human. The aim of this study was to evaluate the chemical compounds as well as the cellular and molecular effects after exposure to organic material extracted from particulate matter less than 10 µm (PM10) in the Amazon region. As for the chemical composition, n-alkanes analysis showed a prevalence of anthropogenic influence during the fires in the region. In addition, there was a predominance of monosaccharides from biomass burning markers. Also, the Polycyclic Aromatic Hydrocarbons (PAH) and their derivatives have also been identified in samples collected in the Amazon. By using the PAH concentrations was possible to calculate the BaP-equivalent and it was found that the dibenz(a) anthracene contributes with 83% to potential carcinogenic risk. As for the potential mutagenic risk, the benzo (a) pyrene is the HPA that has a major contribution in this analysis. It may be noted that the retene was the most abundant PAH. This compound was genotoxic and cause death by necrosis in the human lung cells. In biological tests, the data showed that organic PM10 is capable of causing genetic damage in both plant cells and in human lung cells. This damage cause an arrest in the G1 phase of the cell cycle exposed, increasing the expression of p53 and p21. Additionally, the PM10 caused cell death by apoptosis, increasing the foci of histone - H2AX. Given these results, it is important to emphasize the reduction and better control of biomass burning in the Amazon region thus improving the quality of health of the population being exposed. As clearly stated recently by the World Health Organization, the reduction of air pollution could save millions of lives annually.

Avaliação de efeitos biológicos de antiinflamatórios derivados do ácido propiônico através de modelos experimentais em nível molecular e celular

Derivatives of propionic acid NSAIDs are irreversible inhibitors of cyclooxygenase enzyme widely used. The aim of this study was to evaluate, through different experimental models, biological effects of derivatives of propionic acid (fenoprofen, naproxen, ibuprofen and ketoprofen) in cellular and molecular level. The labeling of blood constituents with technetium-99m (99mTc) and morphological analysis of erythrocytes of blood of rats, as well as growth, survival of cultures of Escherichia coli (E. coli) and the assessment of bacterial plasmid electrophoretic profiles were models used for experimental evaluation of possible biological effects of antiinflammatory drugs. The results show that, in general, anti-inflammatory drugs evaluated were not able to alter the labeling of blood constituents with 99mTc, the morphology of red blood cells from blood of rats, as well as the growth of cultures of E. coli and the electrophoretic profile of plasmid DNA. However, naproxen appears to cytotoxic effect on bacterial cultures, plasmids and genotoxic effects in reducing the action of stannous chloride in cultures of E. coli. The use of experimental fast performance and low cost was important for assessment of biological effects, contributing to a better understanding of the properties of propionic acid derivatives studied. anti-inflammatory, blood constituents, technetium-99m, stannous chloride, Escherichia coli; DNA

Dificuldades de aprendizagem sobre conceitos de genética no ensino fundamental

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior

Caracterização dos acoplamentos fase-amplitude na região CA1 do hopocampo

Brain oscillation are not completely independent, but able to interact with each other through cross-frequency coupling (CFC) in at least four different ways: power-to-power, phase-to-phase, phase-to-frequency and phase-to-power. Recent evidence suggests that not only the rhythms per se, but also their interactions are involved in the execution of cognitive tasks, mainly those requiring selective attention, information flow and memory consolidation. It was recently proposed that fast gamma oscillations (60 150 Hz) convey spatial information from the medial entorhinal cortex to the CA1 region of the hippocampus by means of theta (4-12 Hz) phase coupling. Despite these findings, however, little is known about general characteristics of CFCs in several brain regions. In this work we recorded local field potentials using multielectrode arrays aimed at the CA1 region of the dorsal hippocampus for chronic recording. Cross-frequency coupling was evaluated by using comodulogram analysis, a CFC tool recently developted (Tort et al. 2008, Tort et al. 2010). All data analyses were performed using MATLAB (MathWorks Inc). Here we describe two functionally distinct oscillations within the fast gamma frequency range, both coupled to the theta rhythm during active exploration and REM sleep: an oscillation with peak activity at ~80 Hz, and a faster oscillation centered at ~140 Hz. The two oscillations are differentially modulated by the phase of theta depending on the CA1 layer; theta-80 Hz coupling is strongest at stratum lacunosum-moleculare, while theta-140 Hz coupling is strongest at stratum oriens-alveus. This laminar profile suggests that the ~80 Hz oscillation originates from entorhinal cortex inputs to deeper CA1 layers, while the ~140 Hz oscillation reflects CA1 activity in superficial layers. We further show that the ~140 Hz oscillation differs from sharp-wave associated ripple oscillations in several key characteristics. Our results demonstrate the existence of novel theta-associated high-frequency oscillations, and suggest a redefinition of fast gamma oscillations

Alterações da default mode network provocadas pela ingestão de Ayahuasca investigadas por Ressonância Magnética Funcional

Ayahuasca is psychotropic beverage that has been used for ages by indigenous populations in South America, notably in the Amazon region, for religious and medicinal purposes. The tea is obtained by the decoction of leaves from the Psychotria viridis with the bark and stalk of a shrub, the Banisteriopsis caapi. The first is rich in N-N-dimethyltryptamine (DMT), which has an important and well-known hallucinogenic effect due to its agonistic action in serotonin receptors, specifically 5-HT2A. On the other hand, β-carbolines present in B. caapi, particularly harmine and harmaline, are potent monoamine oxidase inhibitors (MAOi). In addition, the tetrahydroharmine (THH), also present in B. caapi, acts as mild selective serotonin reuptake inhibitor and a weak MAOi. This unique composition induces a number of affective, sensitive, perceptual and cognitive changes in individuals under the effect of Ayahuasca. On the other hand, there is growing interest in the Default Mode Network (DMN), which has been consistently observed in functional neuroimaging studies. The key components of this network include structures in the brain midline, as the anterior medial frontal cortex, ventral medial frontal cortex, posterior cingulate cortex, precuneus, and some regions within the inferior parietal lobe and middle temporal gyrus. It has been argued that DMN participate in tasks involving self-judgments, autobiographical memory retrieval, mental simulations, thinking in perspective, meditative states, and others. In general, these tasks require an internal focus of attention, hence the conclusion that the DMN is associated with introspective mental activity. Therefore, this study aimed to evaluate by functional magnetic resonance imaging (fMRI) changes in DMN caused via the ingestion of Ayahuasca by 10 healthy subjects while submitted to two fMRI protocols: a verbal fluency task and a resting state acquisition. In general, it was observed that Ayahuasca causes a reduction in the fMRI signal in central nodes of DMN, such as the anterior cingulate cortex, the medial prefrontal cortex, the posterior cingulate cortex, precuneus and inferior parietal lobe. Furthermore, changes in connectivity patterns of the DMN were observed, especially a decrease in the functional connectivity of the precuneus. Together, these findings indicate an association between the altered state of consciousness experienced by individuals under the effect of Ayahuasca, and changes in the stream of spontaneous thoughts leading to an increased introspective mental activity

Avaliação por ressonância magnética funcional e estimulação magnética transcraniana da intervenção única da terapia espelho em pacientes após acidente vascular cerebral isquêmico

Mirror therapy (MT) is being used as a rehabilitation tool in various diseases, including stroke. Although some studies have shown its effectiveness, little is known about neural mechanisms that underlie the rehabilitation process. Therefore, this study aimed at assessing cortical neuromodulation after a single MT intervention in ischemic stroke survivors, by means of by functional Magnetic Resonance Imaging (fMRI) and Transcranial Magnetic Stimulation (TMS). Fifteen patients participated in a single thirty minutes MT session. fMRI data was analyzed bilaterally in the following Regions of Interest (ROI): Supplementary Motor Area (SMA), Premotor cortex (PMC), Primary Motor cortex (M1), Primary Sensory cortex (S1) and Cerebellum. In each ROI, changes in the percentage of occupation and beta values were computed. Group fMRI data showed a significant decreased in the percentage of occupation in PMC and cerebellum, contralateral to the affected hand (p <0.05). Significant increase in beta values was observed in the following contralateral motor areas: SMA, Cerebellum, PMC and M1 (p<0,005). Moreover, a significant decrease was observed in the following ipsilateral motor areas: PMC and M1 (p <0,001). In S1 a bilateral significant decrease (p<0.0005) was observed.TMS consisted of the analysis of Motor Evoked Potential (MEP) of M1 hotspot. A significant increase in the amplitude of the MEP was observed after therapy in the group (p<0,0001) and individually in 4 patients (p <0.05). Altogether, our results imply that single MT intervention is already capable of promoting changes in neurobiological markers toward patterns observed in healthy subjects. Furthermore, the contralateral hemisphere motor areas changes are opposite to the ones in the ipsilateral side, suggesting an increase system homeostasis.

A cafeína exerce efeitos positivos sobre a memória tipo episódica em ratos adultos sem influenciar a sobrevivência neural no giro dentado

The caffeine is a mild psychostimulant that has positive cognitive effects at low doses, while promotes detrimental effects on these processes at higher doses. The episodic-like memory can be evaluated in rodents through hippocampus-dependent tasks. The dentate gyrus is a hippocampal subregion in which neurogenesis occurs in adults, and it is believed that this process is related to the function of patterns separation, such as the identification of spatial and temporal patterns when discriminating events. Furthermore, neurogenesis is influenced spatial and contextual learning tasks. Our goal was to evaluate the performance of male Wistar rats in episodic-like tasks after acute or chronic caffeine treatment (15mg/kg or 30mg/kg). Moreover, we assessed the chronic effect of the caffeine treatment, as well as the influence of the hippocampus-dependent learning tasks, on the survival of new-born neurons at the beginning of treatment. For this purpose, we used BrdU to label the new cells generated in the dentate gyrus. Regarding the acute treatment, we found that the saline group presented a tendency to have better spatial and temporal discrimination than caffeine groups. The chronic caffeine group 15 mg/kg (low dose) showed the best discrimination of the temporal aspect of episodic-like memory, whereas the chronic caffeine group 30mg/kg (high dose) was able to discriminate temporal order, only in a condition of greater difficulty. Assessment of neurogenesis using immunohistochemistry for evaluating survival of new-born neurons generated in the dentate gyrus revealed no difference among groups of chronic treatment. Thus, the positive mnemonic effects of the chronic caffeine treatment were not related to neuronal survival. However, another plastic mechanism could explain the positive mnemonic effect, given that there was no improvement in the acute caffeine groups

Efeitos da eliminação de neurônios infragranulares sobre a especificação de neurônios supragranulares do córtex cerebral

The cerebral cortex of mammals is histologically organized into different layers of excitatory neurons that have distinct patterns of connections with cortical or subcortical targets. During development, these cortical layers are established through an intricate combination of neuronal specification and migration in a radial pattern known as "insideout": deep-layer neurons are generated prior to upper-layer neurons. In the last few decades, several genes encoding transcription factors involved in the sequential specification of neurons destined to different cortical layers have been identified. However, the influence of early-generated neurons in the specification of subsequent neuronal cohorts remains unclear. To investigate this possible influence, we induced the selective death of cortical neurons from layer V and VI before the generation of layer II, III and IV neurons. Thus, we can evaluate the effects of ablation of early born neurons on the phenotype of late born neurons. Our data shows that one-day after ablation, layer VI neurons expressing the transcription factor TBR1 are newly generated while virtually no neuron expressing TBR1 was generated in the same age in control animals. This suggests that progenitors involved in the generation of neurons destined for superficial layers suffer interference from the selective death of neurons in deep layers, changing their specification. We also observed that while TBR1-positive neurons are located exclusively in deep cortical layers of control animals, many TBR1-positive neurons are misplaced in superficial layers of ablated animals, suggesting that the migration of cortical neurons could be controlled independently of neuronal phenotypes. Furthermore, we observed an increase in layer V neurons expressing CTIP2 and neurons expressing SATB2 and that these cells have changed their distributions. As a conclusion, our data indicate the existence of a mechanism of control exercised by the early-generated neurons in the cerebral cortex on the fate of the progenitors involved in the generation of the following cortical neurons. This mechanism could help to control the number of neurons in different layers and contribute to the establishment of different cortical areas

Interferindo com oscilações de alta frequência no hipocampo epiléptico: consequências para as crises espontâneas

Conselho Nacional de Desenvolvimento Científico e Tecnológico

Efeitos da sinalização via creb sobre a sobrevivência e diferenciação meuronal

The cortical development requires a precise process of proliferation, migration, survival and differentiation of newly formed neurons to finally achieve the development of a functional network. Different kinases, such as PKA, CaMKII, MAPK and PI3K, phosphorylate the transcription factors CREB, and thus activate it, inducing CREB-dependent gene expression. In order to identify the involvement of such signaling pathways mediated by CREB over neuronal differentiation and survival, in vitro experiments of cell culture were conducted using pharmacological kinase inhibitors and genetic techniques to express different forms of CREB (A-CREB and CREB-FY) in cortical neurons. Inhibition of PKA and CaMKII decreased the length of neuronal processes (neurites); whereas inhibition of MAPK did not affect the length, but increased the number of neurites. Blockade of PI3K do not appear to alter neuronal morphology, nor the soma size changed with the kinase blockades. CREB activation (CREB-FY) along with MAPK and PI3K blockades presented a negative side effect over neuritic growth and the expression of A-CREB leaded to a significant decrease in neuronal survival after 60h in vitro and mimicked some of the effects on neuronal morphology observed with PKA and CaMKII blockade. In summary the signaling through CREB influences the morphology of cortical neurons, particularly when phosphorylated by PKA, and CREB signaling is also important for survival of immature neurons prior to the establishment of fully functional synaptic contacts. Our data contribute to understanding the role of CREB signaling, activated by different routes, on survival and neuronal differentiation and may be valuable in the development of regenerative strategies in different neurological diseases

Indicadores de cálcio e de voltagem codificados geneticamente na detecção de potenciais de ação e inputs sinápticos em cultura de neurônios hipocampais

Recently, genetically encoded optical indicators have emerged as noninvasive tools of high spatial and temporal resolution utilized to monitor the activity of individual neurons and specific neuronal populations. The increasing number of new optogenetic indicators, together with the absence of comparisons under identical conditions, has generated difficulty in choosing the most appropriate protein, depending on the experimental design. Therefore, the purpose of our study was to compare three recently developed reporter proteins: the calcium indicators GCaMP3 and R-GECO1, and the voltage indicator VSFP butterfly1.2. These probes were expressed in hippocampal neurons in culture, which were subjected to patchclamp recordings and optical imaging. The three groups (each one expressing a protein) exhibited similar values of membrane potential (in mV, GCaMP3: -56 ±8.0, R-GECO1: -57 ±2.5; VSFP: -60 ±3.9, p = 0.86); however, the group of neurons expressing VSFP showed a lower average of input resistance than the other groups (in Mohms, GCaMP3: 161 ±18.3; GECO1-R: 128 ±15.3; VSFP: 94 ±14.0, p = 0.02). Each neuron was submitted to current injections at different frequencies (10 Hz, 5 Hz, 3 Hz, 1.5 Hz, and 0.7 Hz) and their fluorescence responses were recorded in time. In our study, only 26.7% (4/15) of the neurons expressing VSFP showed detectable fluorescence signal in response to action potentials (APs). The average signal-to-noise ratio (SNR) obtained in response to five spikes (at 10 Hz) was small (1.3 ± 0.21), however the rapid kinetics of the VSFP allowed discrimination of APs as individual peaks, with detection of 53% of the evoked APs. Frequencies below 5 Hz and subthreshold signals were undetectable due to high noise. On the other hand, calcium indicators showed the greatest change in fluorescence following the same protocol (five APs at 10 Hz). Among the GCaMP3 expressing neurons, 80% (8/10) exhibited signal, with an average SNR value of 21 ±6.69 (soma), while for the R-GECO1 neurons, 50% (2/4) of the neurons had signal, with a mean SNR value of 52 ±19.7 (soma). For protocols at 10 Hz, 54% of the evoked APs were detected with GCaMP3 and 85% with R-GECO1. APs were detectable in all the analyzed frequencies and fluorescence signals were detected from subthreshold depolarizations as well. Because GCaMP3 is the most likely to yield fluorescence signal and with high SNR, some experiments were performed only with this probe. We demonstrate that GCaMP3 is effective in detecting synaptic inputs (involving Ca2+ influx), with high spatial and temporal resolution. Differences were also observed between the SNR values resulting from evoked APs, compared to spontaneous APs. In recordings of groups of cells, GCaMP3 showed clear discrimination between activated and silent cells, and reveals itself as a potential tool in studies of neuronal synchronization. Thus, our results indicate that the presently available calcium indicators allow detailed studies on neuronal communication, ranging from individual dendritic spines to the investigation of events of synchrony in neuronal networks genetically defined. In contrast, studies employing VSFPs represent a promising technology for monitoring neural activity and, although still to be improved, they may become more appropriate than calcium indicators, since neurons work on a time scale faster than events of calcium may foresee

Análise de grafos aplicada a relatos de sonhos: ferramenta diagnóstica objetiva e diferencial para psicose esquizofrênica e bipolar

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior

Encoding mechanisms based on fast oscillations in the retina of the cat and their dependencies on anesthesia

Processing in the visual system starts in the retina. Its complex network of cells with different properties enables for parallel encoding and transmission of visual information to the lateral geniculate nucleus (LGN) and to the cortex. In the retina, it has been shown that responses are often accompanied by fast synchronous oscillations (30 - 90 Hz) in a stimulus-dependent manner. Studies in the frog, rabbit, cat and monkey, have shown strong oscillatory responses to large stimuli which probably encode global stimulus properties, such as size and continuity (Neuenschwander and Singer, 1996; Ishikane et al., 2005). Moreover, simultaneous recordings from different levels in the visual system have demonstrated that the oscillatory patterning of retinal ganglion cell responses are transmitted to the cortex via the LGN (Castelo-Branco et al., 1998). Overall these results suggest that feedforward synchronous oscillations contribute to visual encoding. In the present study on the LGN of the anesthetized cat, we further investigate the role of retinal oscillations in visual processing by applying complex stimuli, such as natural visual scenes, light spots of varying size and contrast, and flickering checkerboards. This is a necessary step for understanding encoding mechanisms in more naturalistic conditions, as currently most data on retinal oscillations have been limited to simple, flashed and stationary stimuli. Correlation analysis of spiking responses confirmed previous results showing that oscillatory responses in the retina (observed here from the LGN responses) largely depend on the size and stationarity of the stimulus. For natural scenes (gray-level and binary movies) oscillations appeared only for brief moments probably when receptive fields were dominated by large continuous, flat-contrast surfaces. Moreover, oscillatory responses to a circle stimulus could be broken with an annular mask indicating that synchronization arises from relatively local interactions among populations of activated cells in the retina. A surprising finding in this study was that retinal oscillations are highly dependent on halothane anesthesia levels. In the absence of halothane, oscillatory activity vanished independent of the characteristics of the stimuli. The same results were obtained for isoflurane, which has similar pharmacological properties. These new and unexpected findings question whether feedfoward oscillations in the early visual system are simply due to an imbalance between excitation and inhibition in the retinal networks generated by the halogenated anesthetics. Further studies in awake behaving animals are necessary to extend these conclusions

Caracterização comportamental e distribuição de neurônios inibitórios em um modelo animal de autismo induzido por ácido valpróico

Autism comprises a heterogeneous group of neurodevelopmental disorders that affects the brain maturation and produces sensorial, motor, language and social interaction deficits in early childhood. Several studies have shown a major involvement of genetic factors leading to a predisposition to autism, which are possibly affected by environmental modulators during embryonic and post-natal life. Recent studies in animal models indicate that alterations in epigenetic control during development can generate neuronal maturation disturbances and produce a hyper-excitable circuit, resulting in typical symptoms of autism. In the animal model of autism induced by valproic acid (VPA) during rat pregnancy, behavioral, electrophysiological and cellular alterations have been reported which can also be observed in patients with autism. However, only a few studies have correlated behavioral alterations with the supposed neuronal hyper-excitability in this model. The aim of this project was to generate an animal model of autism by pre-natal exposure to VPA and evaluate the early post-natal development and pre-puberal (PND30) behavior in the offspring. Furthermore, we quantified the parvalbumin-positive neuronal distribution in the medial prefrontal cortex and Purkinje cells in the cerebellum of VPA animals. Our results show that VPA treatment induced developmental alterations, which were observed in behavioral changes as compared to vehicle-treated controls. VPA animals showed clear behavioral abnormalities such as hyperlocomotion, prolonged stereotipies and reduced social interaction with an unfamiliar mate. Cellular quantification revealed a decrease in the number of parvalbumin-positive interneurons in the anterior cingulate cortex and in the prelimbic cortex of the mPFC, suggesting an excitatory/inhibitory unbalance in this animal model of autism. Moreover, we also observed that the neuronal reduction occurred mainly in the cortical layers II/III and V/VI. We did not detect any change in the density of Purkinje neurons in the Crus I region of the cerebellar cortex. Together, our results strengthens the face validity of the VPA model in rats and shed light on specific changes in the inhibitory circuitry of the prefrontal cortex in this autism model. Further studies should address the challenges to clarify particular electrophysiological correlates of the cellular alterations in order to better understand the behavioral dysfunctions

Revisitando o eletrocorticograma intra-operatório na epilepsia mesial do lobo temporal: relevância das oscilações de alta frequência

Epilepsies are neurological disorders characterized by recurrent and spontaneous seizures due to an abnormal electric activity in a brain network. The mesial temporal lobe epilepsy (MTLE) is the most prevalent type of epilepsy in adulthood, and it occurs frequently in association with hippocampal sclerosis. Unfortunately, not all patients benefit from pharmacological treatment (drug-resistant patients), and therefore become candidates for surgery, a procedure of high complexity and cost. Nowadays, the most common surgery is the anterior temporal lobectomy with selective amygdalohippocampectomy, a procedure standardized by anatomical markers. However, part of patients still present seizure after the procedure. Then, to increase the efficiency of this kind of procedure, it is fundamental to know the epileptic human brain in order to create new tools for auxiliary an individualized surgery procedure. The aim of this work was to identify and quantify the occurrence of epilepticform activity -such as interictal spikes (IS) and high frequency oscillations (HFO) - in electrocorticographic (ECoG) signals acutely recorded during the surgery procedure in drug-resistant patients with MTLE. The ECoG recording (32 channels at sample rate of 1 kHz) was performed in the surface of temporal lobe in three moments: without any cortical resection, after anterior temporal lobectomy and after amygdalohippocampectomy (mean duration of each record: 10 min; N = 17 patients; ethic approval #1038/03 in Research Ethic Committee of Federal University of São Paulo). The occurrence of IS and HFO was quantified automatically by MATLAB routines and validated manually. The events rate (number of events/channels) in each recording time was correlated with seizure control outcome. In 8 hours and 40 minutes of record, we identified 36,858 IS and 1.756 HFO. We observed that seizure-free outcome patients had more HFO rate before the resection than non-seizure free, however do not differentiate in relation of frequency, morphology and distribution of IS. The HFO rate in the first record was better than IS rate on prediction of seizure-free patients (IS: AUC = 57%, Sens = 70%, Spec = 71% vs HFO: AUC = 77%, Sens = 100%, Spec = 70%). We observed the same for the difference of the rate of pre and post-resection (IS: AUC = 54%, Sens = 60%, Spec = 71%; vs HFO: AUC = 84%, Sens = 100%, Spec = 80%). In this case, the algorithm identifies all seizure-free patients (N = 7) with two false positives. To conclude, we observed that the IS and HFO can be found in intra-operative ECoG record, despite the anesthesia and the short time of record. The possibility to classify the patients before any cortical resection suggest that ECoG can be important to decide the use of adjuvant pharmacological treatment or to change for tailored resection procedure. The mechanism responsible for this effect is still unknown, thus more studies are necessary to clarify the processes related to it