In 2015, Brazil faced a severe Zika virus (ZIKV) epidemic transmitted by Aedes aegypti mosquitoes, causing febrile illness. It prompted a study to detect ZIKV in the amniotic fluid of pregnant women with microcephalic babies, confirming its association with CNS malformations. During pregnancy, the maternal immune system adjusts to tolerate embryo cells carrying paternal genetic material, while maternal antibodies provide passive immunity. However, congenital Zika disrupts this immune balance, potentially leading to brain malformations. Our research investigates the origin of these malformations, exploring microchimerism, where cells from one individual enter another during pregnancy. We aim to shed light on the underlying mechanisms of Zika-related complications. Here, we performed an intrauterine surgery to expose the embryos and inoculate saline, inactivated ZIKV (inZIKV), and ZIKV into the amniotic fluid, analyzing after five days. We measured the levels of fetal cells in the maternal blood and three regions of the maternal brain: Olfactory bulb (OB), hippocampus (HC), and cerebellum (CRB). Furthermore, we evaluated microchimerism between siblings, comparing the embryo's brain and liver as the potential harboring sites for the microchimeric cells. Our results revealed no significant difference in the levels of exchanged fetal cells between the control groups (saline and inZIKV) and those inoculated with ZIKV, both in the maternal blood and brain regions. However, there was a notable difference in cell transfer between siblings, particularly in the brain, when comparing the Zika virus group with the saline and inZIKV groups. We conclude that the ZIKV increases the levels of microchimeric cells exchanged between siblings, indicating a disruption in the permeability of the placental and blood-brain barriers. This finding implies that a potential leakage of embryonic cells to the mother could enhance the likelihood of disrupting maternal tolerance mechanisms.

Phytocannabinoids, secondary metabolites produced by Cannabis plants, modulate cellular excitability and present promising effects in adjunctive treatment for refractory epilepsies. Tetrahydrocannabinol (THC) and cannabidiol (CBD), two phytocannabinoids with the highest concentration in Cannabis sp. inflorescences, have similar mass and molecular structure, but distinct pharmacological effects: THC has a clear psychotropic and pro-convulsant effect, while CBD does not affect behavior or perception, despite notable anti-seizure activity in clinical and preclinical studies. On the other hand, little is known about the pharmacological effects of other phytocannabinoids, such as cannabigerol (CBG, a precursor molecule of THC and CBD) and cannabinol (CBN, which results from the abiotic degradation of THC and CBD). CBG and CBN interact with different molecular targets, such as receptors of the endocannabinoid system (CB1r), but their pharmacological effects on neuronal hyperexcitability are unknown. In this study, we evaluated the antiseizure effect of CBG and CBN in a model of chemically-induced acute seizures by pentylenetetrazol (PTZ) in mice (CEUA/UFRN #043/2022). PTZ administration (60 mg/kg, s.c.) produced spasms, myoclonic jerks, and in some animals, generalized limbic seizures. Pretreatment (1 h before PTZ) with CBG or CBN at doses of 3, 30, or 300 mg/kg (i.p.) did not modify the latency, quantity, or severity of PTZ-induced ictal behaviors compared to the control group (CTR, animals receiving vehicle, i.e., corn oil; p>0.05, ANOVA). On the other hand, pretreatment with diazepam (5 mg/kg, a traditional anti-seizure drug, used here as positive control) increased the latency to the first spasm and the first myoclonus and significantly reduced the number of events compared to the control group (p<0.05, unpaired t-test). Surprisingly, the pretreatment with CBG or CBN increased the proportion of animals experiencing limbic seizures, regardless of the dose. The proportion of animals with seizures in the CTR group was 2/16 (13%), while in the CBG group, it was 8/17 (47%, CTR vs CBG, chi-square=4.66, p=0.031) and in the CBN group it was 5/16 (31%, CTR vs CBN, chi-square=1.65, p>0.05). Our results demonstrate that CBG and CBN do not have antiseizure activity in the PTZ-induced acute seizure model and suggest that these phytocannabinoids promote the expression of limbic seizures. We believe this study expands our understanding of the potential and limitations of the endocannabinoid system in controlling neuronal excitability.

Mental health issues among graduate students have sparked debate within the academic community. Previous studies identified anxiety and depression levels up to 6 times higher than the general population. Sleep disorders and substance abuse are also commonly reported. The phenomenon of academic suffering requires further research to identify etiological factors – intrinsic or extrinsic to the individual – for preventive actions and support. The present study aimed to investigate risk and protective factors for mental illness among a Brazilian sample. 476 graduate students (mean age 33.9, SD=8.62) from all Brazilian regions completed an online questionnaire, as approved by the local ethics committee (CAAE: 54127221.0.0000.5537). Questions covered sociodemographics aspects, lifestyle and health, COVID-19 impact, academic experiences, social context, and dream reports. Psychometric screening instruments for depression (PHQ-9: mean=13.8, SD=7.21), anxiety (GAD-7: mean=11.9, SD=6.23), sleep disorders (PSQI: mean=8.41, SD=3.85), and psychosocial safety climate (PSC-12: mean=35, SD=18) were also applied. Non-parametric t-test was performed with the PSC-12 classification for mental illness risk as a grouping variable. Participants in the high risk (HR) group had significantly higher scores than those in the low risk (LR) group, indicating a higher presence of symptoms for depression (HR: mean=14.4, SD=6.89; LR: mean=11, SD=7.92; U=12337, p<.001, r=0.271), anxiety (HR: mean=12.4, SD=6.13; LR: mean=9.62, SD=6.18; U=12661, p<.001, r=0.252), sleep disorders (HR: mean=8.68, SD=3.81; LR: mean=7.18, SD=3.85; U=13197, p=.001, r=0.220) and an increased frequency of nightmares (HR: mean=1.53, SD=0.33; LR: mean=1.44, SD=0.34; U=1438, p=0.022, r=0.150). Linear regression analysis with PSC-12 scores as a predictable variable were statistically significant for depression (R2=0.0570, F(1,474)=28.7, p<.001), anxiety (R2=0.0443, F(1;474)=22, p<.001), sleep disorders (R2=0.104, F(1;465)=53.9) and nightmare frequency (R2=0.0221, F(1;474)=10.7, p=0.001). The frequency of nightmares was also significantly associated with outcomes on depressive symptoms (PHQ-9: R2=0.102, F=53.9, p<.001; Rho(474)=0.326, t=7.34, p<.001), anxiety (GAD-7: R2=0.114, F=61.1, p<.001; Rho(474)=0.342, t=7.82, p<.001), and sleep disorders (PSQI: R2=0.104, F(1,465)=53.9, p<.001; Rho(465)= 0.323, t=6.61, p<.001), indicating that the occurrence of nightmares can be discussed as a clinical indicator of mental distress. Global mental health challenges have reached new proportions after the COVID-19 pandemic. Predictions about future pandemics and global warming-related crises are not optimistic, and the main hope for dealing with these issues should come from future scientific and technological advances. Taking into account that increasing diversity in science is a direct way to enhance the diversity of approaches and creativity in problem-solving, there’s a responsibility to protect scientists in their formative years in order to minimize mental health issues, career abandonment and suicide rates among academics. 

Breathing is a fundamental physiological process, yet its significance extends beyond cellular respiration. The rhythmic control of respiration has been associated with cognitive enhancement, increased self-awareness, and relaxation. However, how the breathing pattern relates to the brain and behavior remains unclear. Rodents synchronize sensorimotor rhythms to respiration during active sensing, such as head and whisker movements. It suggests that the breathing cycle could be relevant for environmental neural coding. Here, we characterize how 
the breathing cycle modulates frontal brain activity during animal engagement in active states. To do so, we look for episodes in which the brain and breathing synchronize in these contexts, called respiratory-coupled oscillations (RCOs). We conducted our experiments in an apparatus composed of two opposing platforms separated by a gap (gap paradigm), where they could interact spontaneously with an object or social stimulus at different times. During the experiments, we simultaneously recorded rats&#39; behavior, intranasal pressure, and frontal brain electrical activity. First, we analyzed the behavioral variables by tracking the head positions over time to extract occupancy and head speed during resting and active states. During the activity, animals were more encouraged to stay over the gap in social than object contexts. These events were characterized by rats guiding their heads toward the opposite platform at high respiratory rates (&gt; 5 Hz). We also observed active episodes away from the gap, mainly on the opposite wall. We performed coherence analysis to identify respiration-coupled oscillations in the local-field potentials of the olfactory bulb and, medial-orbital, pre-limbic, and anterior cingulate cortices. This revealed levels of coherence to breathing above chance in all brain regions, both in rest and active states, with values generally decreasing from the anterior-most (OB) to the posterior-most region (Cg). Our results show that these areas are distinctly modulated by the breathing cycle during classes of exploratory contexts. We conclude that the breathing cycle is relevant to rat exploratory behavior and that the RCOs could mediate the coordination between respiration, behavior, and cognition, representing an approach to study the mechanisms behind the behavioral modulation.

Anxiety is characterized as a state of perception of threats that can lead to danger. It is considered an evolutionarily adaptive behavior that results in different physiological changes. The emotional response evoked by anxiety causes activation of brain areas and temporal coordination of neuromodulation. Over the past 20 years, studies have shown that changes in the activity of the hippocampus, amygdala, and prefrontal cortex (PFC) would be related to fear and anxiety responses. The connections between these areas are thought to be coordinated by the theta oscillations (~ 6 - 12 Hz).  However, recent findings have shown that a larger network could be related to the modulation of anxiety, which would be orchestrated by the projections from the olfactory bulb (OB) to the PFC. Namely, both the OB and PFC exhibit neuronal oscillations that synchronize to the phase of the respiratory cycle and are named respiratory rhythms (RR) because of their co-variability in peak frequency with the breathing rate. These refer to actual LFP oscillations coupled with breathing and not the mechanical process of breathing itself. Lower LFP frequencies present themselves as a mechanism for long-range communication through the synchronization of cortical brain regions during different behavioral states, which could be the case of RR. In this work, we wanted to understand how the changes in the respiratory cycle could influence the appearance of these oscillations and how they could be related to the behavioral state during anxiety. For that, we recorded respiration through intranasal pressure and LFPs from the medial PFC (anterior cingulate, prelimbic and medial orbital), OB and parietal cortex of 7 male Wistar rats as they freely behaved in the elevated plus maze (EPM). We found that the respiratory rate changes depending on the behavioral state (anxious vs non-anxious) and locomotor activity. Furthermore, changes in the LFP power spectrum also depend on the behavioral state and reflect changes in respiratory activity. Finally, we observed RR in the same frequency range as theta during more anxious states. We hypothesize that the theta rhythm described in previous anxiety studies may have been confused with RR, and that this rhythm may coordinate long-range communication in the brain, being found both in the frontal brain and in more distant regions such as the parietal cortex.

General anesthetics are ubiquitous in medical practice, but the cellular mechanisms that promote amnesia, analgesia, immobility and unconsciousness by modulating voltage- and ligand-gated ion channels are still poorly understood. Recent studies are beginning to show the diversity of different anesthetics in regulating specific ion channels and that cortical pyramidal neurons are affected by lower concentrations than interneurons. This highlights that general anesthesia does not follow a "general" mechanism but depends on the choice of anesthetics with cell-type specific effects that need to be studied case by case. For example, the electrophysiological effects of volatile anesthetics such as sevoflurane are especially hard to study in in vitro preparations, yet, biophysical models predict stabilization of the open-state of low-threshold voltage-gated K+ channels (Kv1.2). Here we attempt a first step in identifying the electrophysiological profile of sevoflurane on layer 5 pyramidal neurons (L5 PNs) of the auditory cortex of adult mice. We applied sevoflurane by passing the carbogen mixture through a vaporizer for volatile anesthetic thereby both oxygenating and solubilizing sevoflurane in the artificial cerebrospinal fluid (aCSF) used for whole-cell patch-clamp recordings. The concentration of sevoflurane in the aCSF was quantified using gas chromatography-mass spectrometry. We further subdivided L5 PN into type A and B cells based on responses to small depolarizing and hyperpolarizing current steps, since type A and type B L5 PNs display subtle differences in membrane properties that might be modulated by sevoflurane. Quantification of passive and active membrane properties before and after application of low dose sevoflurane (approximately 22 µM) showed a decrease in the firing frequency for both cell types and a specific effect of depolarizing the action potential threshold of type A PNs, (APthreshold aCSF: -36.31 ± 0.94 mV; sevo: -21.75 ± 4.39 mV; p = 0.009; n = 8 cells) and increasing the action potential half-width of type B PNs (APHalf-width aCSF: 1.80 ± 0.13 ms; sevo: 2.29 ± 0.23 ms; p = 0.040; n = 10). Stable recordings after the washout of sevoflurane restored partially initial maximum firing frequency, AP threshold for type A and AP half-width for type B L5 PNs. Our preliminary results conclude that sevoflurane decreases the firing frequency of both types of L5 PNs, however, membrane properties are differentially affected depending on cell type. This work shows for the first time the direct effect of sevoflurane on L5 cortical pyramidal cells and adds to the cellular understanding of the mechanism of action of volatile anesthetics.

Motor behavior involves the adaptive control of movement and depends on mechanisms arising from perception and cognition that plan and execute different complex movement actions of an innate nature or acquired through learning. Steroid hormones, in turn, act as transcription factors participating in signaling pathways and modulation of neural functions, including motor function. Evidence shows that sex steroids influence neurotransmission in the central nervous system, affecting the quality of female performance in motor tasks, highlighting the importance of restriction and motor control in human behavior. The present study investigated whether cortical activity and variations related to skeletal muscle activity present changes in the phases of the menstrual cycle (menstrual, follicular and luteal) in 30 women during a Go/No-Go Protocol. This test included pairs of auditory stimuli (1st stimulus = S1; 2nd stimulus = S2) that were grouped into two general types: centered on S1 and centered on S2. In the S1-centered task, the decision is made after S1 and
motor execution/inhibition occurs after S2. In tasks centered on S2, decision making and execution occur only after S2. Cortical activity was recorded using electroencephalography (EEG) in the medial frontal regions (Fz), left center (C3) and center-right (C4) to study specifically the alpha (8-13Hz), beta (15-30Hz) and beta 1 (15-23Hz) rhythms. The relationships between these EEG measures and Go/No-Go behavioral outcomes and ERPs were assessed. Behavioral activity differed significantly in time of occurrence and accuracy between protocols S1 and S2, without any interaction with the phases of the menstrual cycle. A moderate positive correlation was found between reaction time and progesterone levels in protocol centered on S1. Changes in brain activity with the generation of beta-ERS during movement initiation in the medial frontal region indicated a likely increase in motor stimulus control associated with the decreased levels of HSFs found in the menstrual phase. Furthermore, beta and alpha oscillations in the left central region were significantly more pronounced in the follicular and luteal phases, suggesting a modulation resulting from the increase in sex hormones, estradiol and progesterone, in the processing and execution of motor tasks. In summary, this pioneering study highlights variations in motor cognition throughout the menstrual cycle in Go/No-Go tasks, which may have implications for optimizing motor performance and rehabilitation of women during the reproductive age, providing an important basis for future research and clinical approaches.

Breathing is an essential rhythm of life, and the main system that terrestrial mammals use to smell the world around them. Breathing cycles passively carry odorant molecules that bind to the olfactory receptors of the olfactory epithelium, and each cycle is sufficient as a unit of odor sensation. It is also known today that breathing modulates activity in the brain. In 1942, work by Edgar Adrian showed that the rhythmic flux of air in the olfactory epithelium of hedgehogs generates oscillations in the olfactory bulb. Recent work in rodents has shown that neuronal oscillations synchronized to respiratory activity occur not only at the olfactory bulb but also across distant brain areas, from prefrontal and visual cortices to subcortical areas, such as the hippocampus and the amygdala. Intriguingly, these widespread respiratory brain rhythms are not continuously present. Instead, the coupling between neuronal activity and respiration across brain regions changes dramatically with time. Previous work also showed that the power of these respiration entrained oscillations is higher in frontal areas. To understand these dynamics, we simultaneously recorded depth video along with respiration (intranasal pressure) and distributed brain signals (anterior cingulate, prelimbic, medial orbital  and parietal cortices and olfactory bulb) in 6 rats as they spontaneously behaved in an enriched open field arena (21 sessions of 30 minutes each). 

Animals displayed a rich set of behavior, ranging from immobility (putative sleep) to walking, grooming, rearing and object exploration. In this thesis, we will present and discuss results on the modulation by respiratory activity on low and high-frequency components of the local field potential (LFP) of the rats' frontal brain. We will show how the magnitude of this modulation is strongly dependent on the instantaneous respiratory rate of the rats and recording site. We found that all breathing frequencies modulate LFP activity in all areas, with higher modulation of low frequency LFP components in the olfactory bulb, and higher modulation of gamma LFP components in the medial orbitofrontal cortex.

To better dissect possible modulations of entrainment strength, we detected and selected the behavioral state of the animals and quantified the LFP-respiration entrainment for each one. We found significant modulation of respiratory LFP components for all behaviors. We saw that grooming behavior has the highest modulation of frontal brain LFP by respiratory activity. The results presented here may help to understand a few pieces of this puzzle, and will raise more questions about how and when breathing modulates frontal brain activity.

In the primary visual cortex, connections to the corpus callosum maintain selectivity for specific visual features in homologous regions of the hemispheres. Its central position allows comparing intra- and inter-hemispheric activities, representing movement trajectories that cross the vertical meridian of the visual field. Thus, the long-range horizontal connections allow the deduction of shapes and movements, following Gestalt principles that reflect functional architecture. To verify these interactions, neurons were recorded simultaneously with two arrays of electrodes, with sixteen channels each, implanted in homotopic parts of the visual cortex, that is, the transition zone between area 17 and 18, in cats (n = 4), selected from intrinsic signal images. Electrophysiological recordings were made before and during visual stimulation with gratings and natural scenes, designed to study this interhemispheric integration. Analysis was performed by lowgamma coherence (30-59 Hz) between pairs of neurons with overlapping receptive fields; by the Fano factor and by the identification of neuronal assemblies. Significant coherence 20% greater was found in intra-pairs than between inter-hemispheric pairs, increasing by 15% where unstimulated and evoked activity showed significant coherence, both with orientation and direction selectivity. Furthermore, the Fano factor was higher in intrahemispheric pairs and with opposite selectivity preferences, that is, with more variation than pairs with the same preference and interhemispheric. As for the neuronal assemblies, both intra- and inter-hemispheric were identified, both with directional selectivity. For gratings, there was a significant difference between the two types of assemblies in vertical conditions, with greater activity in intrahemispheric assemblies and for natural scenes, both types had similar activities, but greater activation in interhemispheric ones. In conclusion, coherences between pairs of neurons and interhemispheric assemblies were identified in V1, reflecting the selectivity of horizontal axons that link neurons with similar responses over long distances, anticipating clustering between neurons that represent the vertical meridian.

Synchronous excitatory discharges from the entorhinal cortex (EC) to the dentate gyrus (DG) generate fast and prominent patterns in the hilar local field potential (LFP), called dentate spikes (DSs). As sharp-wave ripples in CA1, DSs are more likely to occur in quiet behavioral states, when memory consolidation is thought to take place. However, their functions in mnemonic processes are yet to be elucidated. The classification of DSs into types 1 or 2 is determined by their origin in the lateral or medial EC, as revealed by current source density (CSD) analysis, which requires recordings from linear probes with multiple electrodes spanning the DG layers. To allow the investigation of the functional role of each DS type in recordings obtained from single electrodes and tetrodes, which are abundant in the field, we developed an unsupervised method using Gaussian mixture models to classify such events based on their waveforms. Our classification approach achieved high accuracies (> 80%) when validated in 8 mice with DG laminar profiles. The average CSDs, waveforms, rates, and widths of the DS types obtained through our method closely resembled those derived from CSD-based classification. As an example of application, we used the technique to analyze single-electrode LFPs from apolipoprotein (apo) E3 and apoE4 knock-in mice. We observed that the latter group, which serves as a model for Alzheimer’s disease, exhibited wider DSs of both types from a young age, with a larger effect size for DS type 2, likely reflecting early pathophysiological alterations in the EC-DG network, such as hyperactivity. In addition to the applicability of the method in expanding the study of DS types, our results show that their waveforms carry information about their origins, suggesting different underlying network dynamics and roles in memory processing.

Non-synonymous genetic variations in the PTK2B locus have been associated with an increased risk of Alzheimer’s disease and are thought to regulate PTK2B expression. However, both the physiological and pathophysiological roles of PTK2B are not fully understood. In the human brain, PTK2B expression is mainly observed in glutamatergic neurons and this expression declines during AD progression. This reduced PTK2B expression in the brain of patients with AD may contribute to neuronal dysfunctions observed in the disease, such as increased electrical excitability and synaptic alterations. Therefore, understanding the role of PTK2B in human neurons may contribute to reveal the mechanisms of neuronal dysfunctions in AD. In this work, we investigated the impact of altered PTK2B expression in human-induced neural cells and started to tackle the effect of one genetic variant identified in AD patients.

To that, we took advantage of two different isogenic human induced-pluripotent stem cells (hiPSCs) to generate neurons expressing different levels of PTK2B, as well as the introduction of AD patient specific SNPs. Next, we employed functional and molecular assays to probe the consequences of altered PTK2B expression both in a physiological and in an AD-like context.

PTK2B reduced expression alters neuronal activity and gene expression, mainly in glutamatergic neurons, with STAT3 as a key regulator of these genetic changes. Interestingly, reduced PTK2B expression in heterozygous cells, but not its complete deletion in knockout cells, led to a substantially higher TAU phosphorylation at several AD-associated sites. This result coincided with increased activity of p38 MAPK and GSK3β, and decreased activity of ERK1/2 and LKB1. Preliminary evidence indicates that a patient-specific SNP associated to increased risk of developing AD may lead to decreased expression of PTK2B and increased phosphorylation of TAU, thereby contributing to the development of Alzheimer's disease pathology.

Our findings also suggest a correlation between PTK2B and calcium transients’ dynamics, which are directly associated with neuronal electrical activity. Notably, exposure to Aβ1-42 amplified calcium transients in neurons with partial or endogenous PTK2B expression but did not induce any effect in neurons with complete PTK2B deletion. This suggests a role for PTK2B in mediating the effects of Aβ1-42 on neuronal electrical activity.

Overall, our study reveals the potential role of PTK2B in mediating important aspects of AD-related pathology. Furthermore, it indicates that PTK2B may be a critical intermediary linking Aβ, modifications in neuronal electrical activity and Tauopathy. This information could provide insights for a deeper understanding of AD mechanisms and potential therapeutic strategies.

During pregnancy, the mother&#39;s immune system undergoes several physiological changes that induce its tolerance to the cells of the embryo. As the embryo has half of its father&#39;s genome, any breach of tolerance can induce the production of antibodies by the mother&#39;s immune system against fetal antigens. Among the variables that can induce the breakdown of tolerance are infections by pathogens. Congenital Zika virus (ZIKV) infection leads in many cases to malformation of the central nervous system (CNS), which has been called congenital Zika virus syndrome (CZVS). In some cases, the patient is born without any clear sequelae and develops postnatal microcephaly, which suggests persistent inflammation of the CNS. Thus, in this project we tested the hypothesis that in addition to the direct damage caused by the viral infection, patients may experience a maternal autoimmune response. Here we show that mothers of children with CZVS have higher levels of antibodies against Engrailed1 (α-EN1), a protein expressed by the developing hindbrain. The level of α-EN1 correlates with the level of ZIKV antibody expression that each mother expresses. However, α-EN1 levels do not correlate with the occurrence of epilepsies or hindbrain malformations. We also tested whether serum from mothers of CZVS children reacts with slices of adult hippocampus or embryonic telencephalon from rodents, a method used to detect antibodies responsible for autoimmune encephalitis. Serum from these patients does not react against these tissues or even against organoid slices from human brains. Thus, this work suggests that ZIKV infection during pregnancy stimulates the production of maternal antibodies against a target expressed in the embryonic CNS. However, this production does not reflect on the severity of the case, which is probably more dependent on direct infection by the virus.

Anxiety is a worldwide prevalent circuitopathy, in other words, a circuit disorder that substantially affects people's quality of life. To treat it properly, it is necessary to reverse the processes that lead to the malfunctioning of neuronal circuits mainly implicated in anxiety behavior. There is still no completely effective treatment for such a disorder. Several studies have presented promising and safe results with serotonergic psychedelics, evidencing the therapeutic potential of these compounds. However, their mechanisms of action have not yet been fully elucidated. Many clinical studies have managed to durably mitigate the symptoms of depression and anxiety with just a single dose, which makes these compounds very interesting alternatives to the classical treatments available in psychiatry. These lasting effects can be explained by a possible induction of neuroplasticity, neuroprotection, and modulation of inflammation-related agents. Some studies have shown an increase in neuritogenesis, synaptogenesis, and neurogenesis from treatment with psychedelic compounds. In this study, we sought to identify how the serotonergic psychedelic 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT) durably affects neuronal activity and the expression of plasticity-related genes in brain structures classically related to anxiety, such as the basolateral amygdala, the ventral hippocampus, and the anterior cingulate cortex of adult mice one hour, five hours, and five days after treatment. Assessing the electrophysiological properties of mice after 5 days of 5-MeO-DMT treatment, we found differences in passive membrane properties, as well as changes in amplitude and frequency of neuronal firing. Evaluating gene expression one hour after treatment, we showed increased expression of ARC and ZIF268 immediate early genes in the anterior cingulate cortex and basolateral amygdala. After 5 hours of treatment, the NR2A gene was significantly decreased in ventral CA1. Finally, 5 days after the treatment with 5-MeO-DMT we found a significant increase of the TRIP8b gene in ventral CA1. We then intended to see if 5-MeO-DMT produces changes in the behavior of mice after 24hours and 5 days of treatment, with and without stress conditions. We also assessed mice basal corticosterone serum levels under and after acute restraint stress. We found that 5-MeO-DMT treated mice presented significantly less basal corticosterone levels after 5 days. They also presented less anxious behavior. These molecular, cellular, and behavioral findings suggest that 5-MeO-DMT produces immediate and long-lasting effects in mice, although further studies are necessary to consider the therapeutic possibility and to unravel which signaling pathways underlie the 5-MeO-DMT role on synaptic plasticity. 

Activity-dependent gene expression represents the link between neuronal  activity and memory formation. Among these genes, egr1 (a.k.a. zenk) was one of the  first examples of a behavioral-linked gene and has been associated with memory formation  in rodents. However, the role of zenk in vocal learning remains unknown. To  investigate the precise contribution of this gene for the acquisition and maintenance of  song memories in zebra finches (Taeniopigygia guttata), we developed a dominant  negative ZENK to manipulate the transcriptional activity of the endogenous protein.  Using this tool, we have found that normal ZENK activity is required for song development. Downregulation of ZENK transcriptional activity using an  alternative inhibitor, NAB1, confirms these results and provides support for the specificity  of our manipulation. Furthermore, our results shows that blocking ZENK consistently does not lead to alteration of innate components of singing, suggesting that this activity-dependent gene alters only learned behaviors. Our findings provide for the first time a  clear causal link between activity dependent genes and vocal learning.  More importantly, they represent one of the first steps towards understanding the molecular processes involved in the acquisition of vocal signals, a process that is also required for human speech.  

Placebo-controlled clinical trials show that cannabidiol (CBD) reduces seizure frequency, both convulsive
and non-convulsive, in patients with difficult-to-control epilepsy. Part of this effect depends on
pharmacokinetic interactions since in these studies, patients continue conventional antiseizure
medication. Furthermore, the CBD mechanism would reduce seizure frequency still unknown, especially
considering its affinity for endocannabinoid system receptors. Other phytocannabinoids, such as
tetrahydrocannabinol (THC), also modify the endocannabinoid system, altering synaptic transmission
and neuronal excitability. Thus, this thesis evaluated the antiseizure potential of Cannabis extracts with
different profiles of phytocannabinoids (phytocomplexes) on status epilepticus (SE), comparing their
responses with those observed after CBD. We worked with the hypothesis that phytocomplexes derived
from Cannabis, as they contain a variety of secondary metabolites, would be more effective in
controlling neuronal hyperexcitability than a pure phytocannabinoid (CBD). For this, we used
electrophysiological and behavioral recordings of mice submitted to the SE model induced by
pilocarpine intrahippocampal administration. Dose-response curves of two extracts, one with a
CBD/THC concentration ratio greater than 1 (XT-CBD) and the other with a CBD/THC ratio less than 1
(XT-THC), were compared with conventional treatments (e.g. , CBD and diazepam). The animals received
the experimental compound intraperitoneally (i.p.) thirty minutes before pilocarpine. We used the
ambulation between the two administrations as an indicator of possible sedative effects caused by the
extracts. Our results show that XT-CBD reduced the duration and severity of behavioral SE at all
evaluated doses (4, 36 and 360 mg/kg; n=6-9/group) compared to the control group (sterile corn oil
solution, 10 mL/kg; n=24, p&lt;0.05, Mann-Whitney test). Treatment with XT-THC showed a biphasic
effect, with high doses (450 and 1000 mg/kg; n=6-9/group) decreasing the severity of seizures, while the
10 mg/kg dose (n=7 ) increased the proportion of animals with severe ictal behaviors. However, the high
doses of XT-THC produced strong sedation. Comparatively, the decrease in SE severity at high doses of
XT-CBD and XT-THC was similar to that observed in groups treated with diazepam (5 mg/kg; n=6; p&gt;0.05;
Mann-Whitney test) and with CBD (200 mg/kg dose, n=11). Surprisingly, the treatment with CBD, XT-
THC and XT-CBD did not change macroscopic markers of electrographic paroxysms. Our results
demonstrate that phytocomplexes are effective in modulating the behavioral expression of
pharmacologically induced SE, and CBD rich extract are more effective than THC rich extracts. Future
experiments will be necessary to understand the dissociation between behavioral and electrographic
expression of phytocannabinoids on SE induced by intrahippocampal administration of pilocarpine.
Thereby, this study contributed to a better understanding of the use of phytocannabinoids in seizure
treatment.

Songbirds are considered one of the best animal models to investigate the neurobiological basis of vocal learning. As in humans, these animals have distinct phases for vocal learning, that are influenced by social and motor experiences. More specifically, social deprivation in songbirds can generate deficits in the individual's vocal production, whereas singing prevention delays vocal maturation. However, it is not completely clear whether limited experience during development can affect the animal's auditory discrimination abilities, a well-known phenomenon in humans.To address this question, we manipulate social and motor practice experiences in zebra finches, the most widely used songbird species. We first confirmed that our manipulation altered vocal learning, without grossly affecting development. More precisely, socially deprived animals showed abnormal songs, typical of animals lacking a song model. In contrast, juvenile animals where singing practice was limited continued to produce immature vocalizations as adults. Notably, behavioral experiments demonstrated that control animals, i.e., animals with normal social experiences, showed a lower performance in auditory discrimination tasks compared to isolated and juvenile animals. Conversely, sing-limited did not differ from control birds. These results suggest that early development auditory experiences may influence the discriminatory capacity in adult zebra finches, while vocal practice appears to have little influence in the performance of adult birds. That is, birds that are exposed to normal social experiences during postnatal development may have diminished ability to discriminate conspecific songs. Such interpretation is in agreement with the increased specificity observed in auditory neurons, and would parallel the perceptual losses observed during speech acquisition in humans. 

High-frequency oscillations (HFOs) are spontaneous, transient, and fast oscillations (80- 500 Hz) observed in the cortical regions of mammals. Recorded mainly during slow-wave sleep and quiet waking, physiological HFOs (a.k.a. ripples) participate in sensory perception and memory consolidation. Conversely, pathological HFOs (a.k.a. fast ripples) occur at a higher frequency than ripples and are commonly recorded in seizure-prone structures of epileptic subjects. Despite the strong association between fast ripples and seizure onset zones, it is unclear whether they emerge from the degradation of the ripple generation network or if they develop de novo in the adult brain after cell death and synaptic reorganization triggered by a brain insult.

Here, we used electrophysiological recordings and an animal model of status epilepticus to study the expression of HFOs associated with hippocampal epileptogenesis in the mouse. Animals were chronically implanted with bilateral hippocampal and cortical electrodes and a guide cannula targeting the right dorsal hippocampus. After baseline recordings (for at least two days, minimum six h/day), animals received a single intrahippocampal administration of pilocarpine (700 µg/µl). As we have previously shown, a sustained status epilepticus (SE) evolves after that. 

Initially, our analysis focused on detecting, extracting, and classifying HFOs recorded in the hippocampal electrodes. First, features such as the central HFOs frequency, duration, power, and rate were obtained in a 5-minute slow-wave sleep episode using the RIPPLELAB, a Matlab-based toolbox for semi-automatic extraction of HFOs. Then, we merged the extracted features over five time-points: before SE (baseline period), one day after the SE (SE+1), two days after the SE (SE+2), the first week after the SE (SE+7), and between 10 and 30 days after SE (SE+30). 

We identified 1,334 true-positive ripples from 6 animals. Ripple rate monotonically decreased after the SE (F(4,24)= 2.98, p=0.039, factor time, one-way ANOVA), dropping from 16.2±7.2 events/min to 7.8±4.3 events/min in the baseline period and SE+1, respectively. We recorded the lowest ripple rate at the chronic period (SE+30) (6.9±6.2 events/min). Ripple central frequency also decreased after SE (F(4,1329)= 29.9, p<0.01, factor time, one-way ANOVA) although the most substantial reduction was observed in SE+2 (in comparison to baseline: unpaired mean difference= -9.9 Hz, 95%CI: [-11.8 and -8.0]; p<0.01, two-sided permutation t-test). We observed no statistical differences in ripple duration regardless of the time after SE. The average ripple duration was 39.8±12.8 msec. Finally, SE significantly reduced ripple power over time F(4,1329)= 10.8, p<0.01, factor time, one-way ANOVA) and reached the lowest power one day after SE (in comparison to baseline: unpaired power mean difference = -0.07 mV²/Hz, 95%CI: [-0.09 and -0.05]; p<0.01, two-sided permutation t-test). Spearman correlation analysis revealed a positive association between ripple frequency and power at SE+1 (r=0.30, p<0.01) and SE+7 (r=0.71, p<0.01) considering all detected events. In this line, looking at each animal at each time point, ripple rate and power showed a  positive correlation (r=0.49, p<0.01) as well as power and frequency (r=0.38, p<0.05). Surprisingly, latency to the first seizure, maximum Racine scale during SE, and length of SE did not correlate with any ripple feature. A positive trend was seen with body weight and rate of events between days, however, no significance was achieved (Spearman correlation, r=0.31, p=0.12).

We also analyzed the appearance of fast ripples in SE. In this case, we focused our analysis on transient events with frequencies above 200 Hz. Surprisingly, in all animals, fast ripples appeared in the first 10 minutes of the acute, pilocarpine-induced seizures (n=409; 68.1±34.0 events/animal within 30 min of SE). Fast ripples usually co-occurred with the ictal spike, and they were followed by slower oscillations (circa 60-100 Hz). We observed phase reversal between this electrode and adjacent ones. The fast ripple mean frequency was 283.8±28.6 Hz, the mean power was 0.5±0.1 mV²/Hz, and their mean duration was 31.7±13.7 msec. As SE evolved, the rate of those events decreased, and they were completely abolished by diazepam (10mg/kg, i.p.). All animals presenting fast ripples during acute SE also showed fast ripples in the following days. The rate of fast ripples during 5-min slow-wave sleep was higher on the first day (SE+1) and decreased in the following days (events per min: 14.0±6.8, 15.0±2.5, and 3.8±3.6 in SE+1, SE+2 and SE+30 periods, respectively; F(4,19)=5.4, p<0.01, one-way ANOVA). As expected, fast ripple mean frequency was higher than ripples (mean difference=+156 Hz, 95%CI: 153 and 158, p<0.05, unpaired t-test). Fast ripples also showed stronger power (mean difference=+0.119 mV²/Hz, 95%CI: 0.098 and 0.138, p<0.05, unpaired t-test), and shorter duration (mean difference=-8.12 msec, 95%CI: -9.55 and -9.58, p<0.05, unpaired t-test) than physiological ripples.

To our knowledge, this work is the first study to describe the temporal changes of ripples features during epileptogenesis extensively. Our results demonstrate that the ripple alterations are partially explained by SE severity. Also, we showed that pathological HFOs (fast ripples) occurred in the first minutes of the SE. This observation suggests that fast ripples do not need cell death or synaptic reorganization to be expressed but rather reflect abnormal balance between excitation and inhibition usually observed in acute and induced seizures. Future analysis will demonstrate if the reduction in ripple rate and power can explain the poor performance in memory and learning tasks observed in animals with spontaneous seizures.

Female sex steroids (FSS) have a wide influence on brain functions and pathologies. They can specifically affect the motor system, modulating the excitability of the motor cortex, as well as performance in tasks of dexterity and coordination. There is a lack of studies that investigate its effects on motor cognitive processing. Changes in the amplitude of the electroencephalography (EEG) alpha and beta frequency bands over the frontal, central (mu rhythm) and parietal regions during kinesthetic motor imaging (IM) and action observation (AO) are important measures to investigate the modulation of cortical areas involved in motor planning. Another tool used to investigate cognitive motor processing is the Manual Laterality Recognition Test (TRLM). It is able to modulate the P100 and P300 components of the Event-Related Evoked Potential (ERP) in the EEG and has behavioral measures that allow inferences about cognitive processes related to the spatial manipulation of parts of the body (in this specific case, the hands). This process is known as implicit motor imagery. In this study, we investigated whether
the cortical activity obtained through the EEG and the behavioral measures related to the aforementioned tasks (IM, AO, TRLM) are modulated according to the phases of the menstrual cycle. Thirty-two women of reproductive age participated in this study, which was carried out in a familiarization session and 3 experimental sessions. The FSS comprise estrogen and progesterone and their levels were quantified using the ELISA technique by chemiluminescence in the menstrual (low levels of both), follicular (high levels of estrogen) and luteal (high levels of progesterone) phases. Blood collections for hormonal dosage and experiments with the IM, AO and TRLM protocols were performed in the three phases of the volunteers&#39; menstrual cycle. EEG measurements (for obtaining alpha and beta power, P100-ERP and P300-ERP), reaction time and accuracy were collected during the practice of the tasks. In the statistical analysis, the Friedman test was used to compare the measurements between the phases of the menstrual cycle. Behavioral and electrophysiological measures were also correlated with hormone levels (estradiol and progesterone) through Spearman&#39;s correlation and the P value considered was &lt;0.05. The results showed that the desynchronization related to the beta event (beta-ERD) on the bilateral frontal region during the practice of MI was significantly greater in the follicular phase when compared to the menstrual and luteal phases, suggesting the effect of estrogens on the disinhibition of the control areas of motor cognitive processing. Beta-ERD in the medial parietal region and Alpha-ERD in the central medial region were significantly lower in the luteal phase when compared to the follicular and menstrual phases, areas that are related to the vividness of motor imagery. No difference between the phases of the menstrual cycle was observed on alpha-ERD and beta-ERD on the sensorimotor region or on any cortical region studied during the practice of AO, but a positive correlation was found between the amplitude of these frequency bands and estradiol levels in the follicular phase, suggesting the effect of estradiol on the activation of mirror neurons. The behavioral analysis in TRLM indicated better performance during the follicular and luteal phases when compared to the menstrual phase. According to what is observed in the literature, the spatial manipulation of objects is impaired by the ESF. According to the results presented, when the objects to be moved are hands, the FHS favor the task, suggesting its effect on the motor cognitive component present in the task. In the EEG analysis of the TRLM, it was possible to observe a difference between the medial and lateral orientations of the hands in the follicular phase, suggesting greater engagement in the intrinsic motor imagery in this phase of the menstrual cycle.

Neurons are electrically and chemically excitable, when excited enough, several ions quickly move across the membrane, causing a depolarization. Among these ions, Ca²⁺ is the only one that in addition to participating in action potential is also biologically active, participating in various cellular events, acting primarily as a cellular messenger. Studying the role of Ca²⁺ in the various events was possible through the development of techniques that allowed to measure its dynamics, such as the development of optical tools, being the first generation composed of dyes and later, with the advent of molecular engineering, the genetically encoded calcium indicators (GECIs), composed by hybridization of a fluorescent protein linked to a sensor protein. From early dyes to the most current GECIs, many advances have been made in increasing temporal and spatial resolution, Ca²⁺ sensitivity, photobleaching resistance, decreased phototoxicity, and high signal-to-noise ratio. However, in vivo mammalian preparations remained a challenge. Currently, GCaMP variants are one of the most commonly used GECIs, which are composed of calmodulin as a sensor protein, linked to a green fluorescent protein (GFP). In this work we will evaluate GCaMP expression with different viral promoters and calcium signals in vivo, in records made with adult mice walking in a treadmill at different speeds, using chronically implanted lenses and epifluorescence cameras.

Retinal gamma oscillations have been observed in a variety of vertebrates. Recent and former evidence points to their involvement in encoding stimulus properties such as size and continuity. To investigate these claims, we started by revisiting already established relationships between oscillation characteristics and different visual stimuli in the anesthetized animal. We analyzed single and multi-unit activity recorded from the retina and LGN of cats while they observed complex (short videos) and simple (light circles) stimuli. In follow-up experiments, the LGN of awake cats was also recorded. The animals were subsequently anesthetized with ketamine and later on, halothane. We observed that under natural stimuli and halothane anesthesia, the oscillatory response elicited by video stimuli was unreliable. It occurred in brief periods, which seemed to mainly reflect image contrast. Large circles presented to anesthetized cats, on the other hand, robustly generated gamma-band oscillations, with its strength modulated by luminance and stimulus size. Interestingly, in the awake or ketamine only anesthetized animals, no oscillations could be detected, although entrainment to the monitor refresh rate was observed up to 120 Hz. Furthermore, gamma oscillation in the cat's retina was dependent on halothane concentration. Gamma activity was drastically reduced as levels of halothane were decreased. Taken together, our results do not support a cognitive role for gamma oscillations in the retinogeniculate system.

Noise-induced tinnitus is a phantom sound, perceived without a physical source, caused by over-exposure to loud noise. The dorsal cochlear nucleus (DCN), a region known to integrate somatosensory and auditory pathways, has been identified as a potential key structure in the generation of phantom sound perception. Here, we first target the output layer of the DCN using viral vectors containing the CaMKIIa (CaMKIIa) promoter to express optogenetic proteins and investigate how exciting (Channelrhodopsin-2, ChR2) or inhibiting (enhanced Archaerhodopsin 3.0, eArch3.0) those DCN cells affects the DCN circuitry response to sound. We found that optogenetic stimulation of DCN cells expressing CaMKIIa-ChR2 or CaMKIIa-Arch can distort DCN units responses to sound. Next, we chemogenetically decrease activity of the same subgroup of DCN neurons using the Gi-coupled human M4 Designer Receptors Exclusively Activated by Designer Drugs (hM4Di DREADDs) to investigate their role in tinnitus perception. Mice were exposed to loud noise (90dBSPL, 1h, followed by 2h of silence) to induce tinnitus. Auditory brainstem responses (ABRs) and gap prepulse inhibition of acoustic startle (GPIAS) test were recorded two days before and two weeks after noise exposure to confirm tinnitus perception (significant decrease in GPIAS response) without permanent hearing loss. Activity of CaMKIIa-hM4Di+ neurons in the DCN was decreased by expressing and pharmacologically activating the hM4Di DREADDs. Perception of tinnitus was evaluated when inhibitory hM4Di DREADDs were activated by systemic clozapine-n-oxide (CNO, 0.5mg/kg) administration. We found CNO to decrease tinnitus-like responses (p = 0.038, n = 11 mice), compared to the control group that showed no improvement in GPIAS responses (control virus; CaMKIIa-YFP + CNO, p = 0.696, n = 7 mice). Unit recordings confirmed CNO to decrease DCN firing frequency, alter best frequency and broadness of response to sound. Next CaMKIIa-hM4Di-mCherry positive DCN neurons were instead chemogenetically inhibited during the noise exposure (n = 6 experimental and 6 control mice), but this did not prevent tinnitus induction, nor did CNO administration 2 weeks later improve gap-detection indicating tinnitus perception was not decreased. Our results suggest that CaMKIIa-hM4Di-mCherry positive cells in the DCN are not crucial for tinnitus induction but play a significant role in maintaining tinnitus perception in mice.

Even though several mammalian orders have orientation-selective neurons, these neurons can be organized in different functional cortical layouts. A prominent example is the columnar orientation preference map found in carnivores (cats, ferrets, tree shrews) and all primates (e.g. human, macaque, mouse lemur) studied so far, as opposed to the seemingly random arrangements commonly referred to as salt-and-pepper configurations, observed in rodents. Orientation selectivity is generally deduced from stimulus-driven spiking activity. However, it has also been observed in oscillations of the local field potential (LFP), especially in the gamma band. In several of the species expressing a columnar map, preference orientation of spiking activity and gamma power are similar. This is compatible with the interpretation that the LFP samples activity from contiguous cortical patches responding to the same orientation. Spikes can be strongly locked to certain phases of LFP oscillations, and the latter are thought to not only establish temporal windows coordinating spiking activity of local neuronal populations but to also enable communication between distant neuronal populations. We thus hypothesized that a columnar/non-columnar functional layout might also be reflected in spike-field interactions.

In the present thesis, we examined spikes and LFP oscillations in the low gamma (low-γ) band of +/-30-50 Hz obtained from multiple parallel recordings in visual cortex during grating stimulation in two representatives of different mammalian order and similar V1 size but different functional layouts – the domestic cat for carnivores, and the Amazonian agouti for rodents. Previously, we observed that although agoutis possess orientation selective neurons they do not express columns as cats do. We correlated the orientation tuning of spikes with that of the low-γ power of the same electrode. This analysis revealed that units exhibiting both orientation selectivity in spikes and LG power (OSI > 0.1) exhibit similar preference in cats (n = 95, circular correlation = 0.56, p = 1.44e-12), but much less in agouti (n = 39, circular correlation = 0.40, p = 0.0067). Further, we evaluated if single unit spiking activity is coordinated with the LFP in the Gamma range recorded from the electrode as reflected in phase-locked events. In cats,  phase-locked events frequently occurred in an orientation- selective manner and with similar orientation preference as the spiking activity (n = 33, circular correlation = 0.57, p = 0.0035). In contrast, agouti showed only very few phase-locked events and those were not correlated with spike orientation selectivity (n = 6, circular correlation coefficient = 0.11, p = 0.79).

Our results support the notion that the coordination of spiking and LFP gamma power in cats could be related to their orderly columnar layout. Along the same line of argumentation, we propose that the lower similarity of spike and gamma tuning and the lack of selective phase-locked events in agouti reflects a lesser or absent organization of orientation preference in a contiguous columnar layout in this species.

Over the past decades, Brazilian population over 60 years of age has increased, as well as chronic diseases related to aging. Among these, stroke, one of the main causes of hospitalizations and mortality in Brazil; causing some type of disability to perform activities of daily living to the majority of patients. Understanding how the rehabilitation process occurs, such as the remodeling of motor maps and motor pathways is of utmost importance. The study of this phenomenon can lead to optimization of rehabilitation and to a better understanding of how drugs can act to restore brain function following stroke. In addition, lesion location and severity are determinant for the level of motor impairment following the insult. Preliminary results obtained by PhD Numa Dancause and PhD Dale Corbett research teams indicated that when the lesion encompass the dorsal striatum, motor representations are extremely impaired. Therefore, it was hypothesized that the dorsal striatum has a determining role in the constitution and possible remodeling of the cortical maps during motor relearning. In addition, subcortical structures may increase activity and assume a more active role in the expression of motor behavior. In this thesis we develop a new methodology to quantify and better understand changes of upper limb movement following stroke in rodents (rats: Article 1; mice: Article 2). Next, we review the main pathways and mechanisms involved in motor relearning following stroke (Article 3). Finally, the main article of this thesis demonstrates how intensive rehabilitation promotes cortical motor map plasticity (Article 4). Intensive rehabilitation led to motor map expansion, increased kinematic movement of the paretic limb, and increased use-dependent activity of compensatory brain regions, such as the contralesional cortex and red nucleus. Here, we propose an intertwined mechanism for severe stroke recovery, which involves bilateral and bi-hemispheric interactions. In short, this thesis corroborates that rehabilitation promotes expansion of cortical motor representations; and advances the knowledge about compensatory recovery following severe stroke in rats.

Consolidated memories can become labile when reactivated and, to persist, must undergo a protein synthesis-dependent restabilization process called reconsolidation. The perirhinal cortex (PER) regulates the flow of information between different brain areas and although it has been implicated in object recognition memory (ORM) reconsolidation, the actual role of PER in this process remains controversial. Using the novel object recognition task we investigated the conditions that constrain PER involvement in ORM reconsolidation. We found that bilateral intra-PER infusions of the protein synthesis inhibitor anisomycin caused amnesia when administered immediately after ORM reactivation in the presence of a familiar object but not when memory was reactivated in the presence of a new object, two familiar objects, or a new object alongside a familiar one. Immunofluorescence analyses revealed a prominent increase in PER ZIF268 expression only after ORM reactivation in the presence of a single familiar object. We also found that PER NMDAr blockade with the selective NMDAr antagonist AP5 and ZIF268 expression knockdown with antisense oligonucleotides hindered ORM reconsolidation. Our results indicate that partial perception of recognition cues during memory reactivation is a boundary condition for the participation of PER in ORM reconsolidation, and suggest that this process involves NMDAr activation and ZIF268 expression in that cortex.

Organization of neocortical circuits is critical for sensory perception, learning and multisensory
integration. In the primary somatosensory area (S1) layer IV neurons receive thalamic inputs and
synapse onto layer II/III neurons. These superficial neurons may connect other neurons within S1 and
within other areas in the ipsi or contralateral hemisphere, thus cooperating to select an
interpretation consistent with their various cortical and subcortical inputs. Here we show that
expression of the transcription factor Zinc Finger And BTB Domain-Containing Protein 20 (Zbtb20) in
neocortical progenitors is necessary and sufficient to regulate the generation and hiring patterns of
upper layer neurons. Conditional deletion of this gene in progenitors leads to an increase in the
number and radial occupancy of RORb+/layer IV at the expense of Brn2+/layers II/III neurons. This
change in the laminar organization of the neocortex is accompanied by an expansion of thalamic
axonal arborization and barrel area in S1. Furthermore, upper layer neurons increase their intra-
hemispheric axonal projections, while reducing contralateral innervation in the absence of Zbtb20
expression. These alterations are also observed, albeit at a lesser extent, after Zbtb20 deletion in
post-mitotic neurons, indicating that Zbtb20 act at sequential stages of the lineage progression of
neocortical progenitors, fine-tuning neuronal fates in upper cortical layers and contributing to the
proper wiring of callosal projecting neurons (CPNs). Besides these effects in CPN fate specification,
we also show that ZBTB20 regulates astrogliogenesis in a time-specific fashion. ZBTB20
overexpression at E14, but not at E16, increases neocortical astrogliogenesis, whereas expression of
a dominant-negative (DN) ZBTB20 at E16, but not E14, reduces astrogliogenesis. Altogether, our
results indicate that ZBTB20 is an important regulator of cell type/subtype specification in the
developing neocortex.

Remembering facts and events requires object recognition memory (MRO). Reconsolidation integrates new information into MRO through bidirectional changes in hippocampal synaptic efficacy and BDNF signaling. In turn, BDNF enhances long term potentiation (LTP) through protein kinase Mζ (PKMζ), which might preserve memory by controlling AMPAR function. However, the possible involvement of PKMζ in ORM reconsolidation has not yet been studied. In rats, we found that hippocampal PKMζ inhibition with zeta-inhibitory peptide (ZIP) or antisense oligonucleotides, but not PKCι/λ inhibition with ICAP, hindered retention provided MRO was reactivated simultaneously with the introduction of a novel object. Similarly, ORM reactivation increased hippocampal PKMζ only when it happened in the presence of an unfamiliar object. BDNF co-infusion reversed the amnesia induced by post-reactivation hippocampal protein synthesis inhibition but not that triggered by ZIP, which did not affect spontaneous oscillatory activity. Moreover, reduction of hippocampal AMPAR surface expression after MRO reactivation hampered retention, whereas blockade of AMPAR endocytosis increased PSD GluA1/GluA2 and reversed the amnesic effect of ZIP. MRO consolidation, but not reconsolidation, requires protein synthesis in entorhinal cortex (CE). We found that animals rendered amnesic by intra-CA1 ZIP reacquired MRO upon retraining, but inhibition of CE protein synthesis impaired relearning as if MRO had to be consolidated anew. Our results show that hippocampal PKMζ acts downstream BDNF to regulate AMPAR recycling at the time of reconsolidation and indicate that PKMζ inhibition during this process deletes MRO.

Temporal Lobe Epilepsy (TLE) is the most common form of focal epilepsy. The human condition can be modeled in animals by the systemic administration of pilocarpine (PILO) or kainic acid (KA). The experimental approach involves an initial insult (status epilepticus - SE) resulting in wide-spread cell death, structural reorganization, chronic spontaneous seizures and, impaired performance in memory tasks and anhedonia. The identification of the anatomical substrates related to the cognitive impairments in those models is not entirely known, since systemic administration may lead to multiple epileptic foci. To minimize the impact of spatially distributed, numerous epileptic foci, on cognitive performance, we present a protocol in which the convulsant agent is administered directly in the target structure of interest (i.e., straight into the hippocampus). This approach has been used for KA in mice, but no systematic study has evaluated the effects of intrahippocampal administration of PILO. Here, we described the acute and chronic behavioral effects of the intrahippocampal administration of PILO (4 doses: 70, 245, 400 e 700 µg, in 1 µL) in mice during isoflurane anesthesia. KA (20 mM, 50 nL) and saline (0.9 %) were used as positive and negative controls, respectively. The results show a correlation between the severity of the SE and
the dose of PILO given (measured acutely by behavioral scores and indirectly through the evolution of weight). Interestingly, intrahippocampal PILO injection (N=62) did not elicit tonic seizures, as commonly observed after systemic administration which contributed to the low mortality rate of the model (4 out of 63 and 10 out of 11, respectively). In the chronic phase (1 month after SE), mice treated with high doses of PILO and KA presented spontaneous seizures. Behavioral tests revealed that epileptic animals (independent of the convulsive drug used) present higher stereotyped ambulation (in the open-field) than animals from the control group. In hippocampal-dependent memory tests, PILO treatment, especially at high doses, impaired performance in the object
recognition task, but not in the Barnes maze. Animals from the KA group presented impaired performance in all memory tests compared with the PILO group (high dose). Taken together, our results demonstrate that ihpc
administration of PILO in mice results in spontaneous and recurrent seizures, as well as moderate cognitive impairment (compatible with comorbidities observed in humans with TLE), and low mortality rate. We believe that the present model has face validity for human TLE, and may serve as an alternative to KA models (which are expensive) and to the route of administration (high mortality when PILO is administered systemically).

The reprogramming of different specialized cell types in others has been a field widely studied in recent years. More specifically, the generation of induced neurons (iNs) is used with the objective of applying these cells in the study of pathological models and in cellular therapy, focusing on the treatment of patients with neurodegenerative diseases or acute lesions in the central nervous system. Several methodologies have been used for this approach, including the reprogramming of cells from other lineages into neurons, either directly or indirectly. However, most of the cellular reprogramming protocols depend on the overexpression of ectopic genes, which may lead to other transient changes not desired in reprogrammed cells. In order to overcome these possible side effects, we investigated the possibility of reprogramming astrocytes from postnatal mice into neurons through the transient exposure of the cells to a cocktail of small molecules added to the cell culture medium. This cocktail includes molecules that act in different cellular pathways, among which the regulation of gene expression, the modulation of neurogenesis and the control of the cell cycle, previously been used in the reprogramming of fibroblasts into neurons. To demonstrate the phenotype of the cells after treatment with the cocktail, aspects such as the expression of typically neuronal and astrocytic proteins, morphology, survival, proliferation and gene expression were evaluated through the techniques of immunocytochemistry, time-lapse video-microscopy and RT-qPCR. Based on the data obtained it was observed that the drug cocktail used induced in the treated astrocytes an increase in the expression of genes related to the neuronal profile and a significant change in its morphology, although this is not typical of neurons. Thus indicating that the combination of small molecules used is not sufficient to effectively reprogram astrocyte cells into induced neurons.

Alzheimer’s disease (AD) is present in 25 million people in the world, and is characterized by the progressive decline of cognition, mainly episodic memory. AD is multifactorial, and the accumulation of amyloid peptides (Aβ) is the main proposed mechanism underlying neurodegeneration. Aβ are the main components of the amyloid plaques in the brain, that are physiopathological hallmarks of the disease. Intracerebral infusion of Aβ in rats is usually used as an animal model of DA and generates the accumulation of these peptides in the brain together with spatial memory impairment. However, most studies show moderate to severe deficits after chronic Aβ infusion, without evaluation of possible subtle initial deficits. The study of the initial stages of AD is relevant for mechanicistic and therapeutic investigations. The Barnes maze has been used for investigating deficits in the hippocampal function in rats. The animals are exposed to a circular apparatus with holes in its periphery. One of the holes is connected to a safe place. In this task, rats navigate guided by distal cues to find this safe compartment. The performance of the animals in the training phase (repeated exposures to the apparatus with the safe box placed in the same position) and test probe (reexposure after variable intervals without the safe box) is used to the evaluation of acquisition and retrieval of the spatial task, respectively. The evaluation is conduct by parameters of general performance (latency and distance to reach target, number of errors, among others) and by a specific analysis of search strategies. In this way, the animal use spatial information and move directly towards the target (direct strategy), visit sequential holes until reach the target (serial strategy) or visit holes in a non-systematic fashion until reach the target (aleatory search). The analysis of strategies allows the detection of alterations in the mode of solution of the task, even if a given experimental approach does not modify the general performance parameters. In the present study, we aimed to investigate possible initial cognitive signs of AD in Wistar rats submitted to intracerebral infusions of Aβ, as well as the effects of a potentially neuroprotective treatment, by the evaluation of spatial memory in the Barnes maze. We used the alkaloid extract of Erythrina velutina (“mulungu”), which was previously studied for anxiolytic, anti-inflammatory, antioxidant and cholinergic actions. First, we standardized the task to our experimental conditions, and verified a possible influence of repeated expositions to the apparatus (for future long-term protocols) or of the implantation of the cannulas in the brain. Rats went through (or not) to implantation of cannulas (bilaterally in the hippocampal CA1 and in the lateral ventricle) and were exposed to 5 sequences of exposition to the Barnes maze (4 trainning sessions with 4 trials each, a 24h test and a 10-day retest sessions). Results showed that rats learned the task, showing diminished latency and distance to target, decreased erros and increased target quadrant exploration across the trials of all sequences. Both groups showed task retrieval, and there was a slightly improved performance in the implanted group. We concluded that the task can be held repeatedly, and in implanted animals, without altering the performance. In the second phase, we investigated the use of different strategies in the Barnes maze by rats submitted to the presence of absence of distal cues. Rats were exposed to the training phase, and in the probe session (24 h later) half the animals were exposed to the maze in the presence of the same distal cues used in training (spatial group), while the other rats went through the probe test without those cues (a black curtain was placed around the maze – non-spatial group). Both groups learned the task, but the spatial group preferred the used of direct strategies, while the non-spatial group preferred other strategies. We concluded that the removal of distal cues does not hinder the execution of the task, and the animals use alternative search modes under this condition. In the third phase (two experiments) we verified the effects of the intracerebral infusion of Aβ on the acquisition and retrieval of Barnes task. In experiment 1, rats received 15 daily i.c.v. infusions of saline or Aβ (30, 100 ou 300 pmol) plus bilateral CA1 infusions in the first day, and were exposed to 3 sequences of Barnes task (training, 24h test and 10-day retest in each sequence). Sequence I was held before surgery (all the animals learned the task), II started at 11^th infusion and III started 10 days after II. The behavior of the Aβ-treated animals varied greatly at sequences II and III, and hence no differences were observed. There was high mortality due to treatment. At the end of the behavioral sessions, saline and Aβ 30pmol groups were euthanized for Aβ immunohistochemistry. The analysis by relative optical density showed increased Aβ staining in the hippocampus and neocortex. We concluded that, although there was Aβ deposition, the treatment did not induce performance deficits. In experiment 2 we investigated the effects of Aβ (30 pmol) infusion on the search strategies in the Barnes maze. Animals went through one sequence of Barnes task (4 trainings with 2 trials each and a 3-day test, in order to increase difficulty). In the probe test, although Aβ animals showed some retrieval, they showed preference for non-spatial strategies, opposed to saline-treated rats. We concluded that Aβ infusion induced subtle alterations in spatial memory, compatible with the initial stages of AD, which is relevant for investigations of potential neuroprotective approaches. In the last stage, animals submitted to the same infusion protocol described above were concomitantly treated orally with 200 mg/kg of mulungu extract and went through 2 Barnes task sequences. In general, no differences were observed among the groups in acquisition or retrieval. Thus, we concluded that the protocol used here was not able to detect a beneficial effect of mulungu extract in the Aβ infusion AD model. 

So far, there is no evidence of a columnar orientation preference map in rodent primary visual cortex, such as commonly observed in carnivores and primates. At the same time, orientation selective neurons have been found in all rodent species investigated, though interspersed. This opens up the question whether the connectivity underlying the emergence of selective cortical response properties in animals with interspersed as compared to columnar maps follows a different blueprint. Rodent data are so far mainly available for species with nocturnal or crepuscular habits and small brain size, two factors that could also contribute to develop a different functional architecture.

Therefore, we set out to compare the functional architecture of the primary visual cortex of carnivores with that of a big rodent with diurnal habits, and a V1 size comparable to cats and small primates. To this end, we performed multi-site electrophysiological recordings using spatial arrays from both anesthetized cats’ (Felis catus) and agoutis’ (Dasyprocta aguti) visual cortex. Visual stimuli consisted of oriented gratings of several spatial and temporal frequencies.

Agoutis presented much smaller orientation selectivity indices (median OSI = 0.10) than cats (median OSI = 0.19). In order to describe the functional architecture based on the electrophysiological data, we quantified the orientation preference difference between neurons according to the cortical distance between them. As expected, this analysis revealed a characteristic slow decrease in neuronal orientation preference similarity for cats. No such “classical” modularity was found for agoutis, but a clustering of neurons with similar orientation preferences was observed for short ranges (< 250um).

Overall, our results are consistent with recent literature reporting ‘mini-columns’ of orientation preference in mice, and therefore further prove that the rodents’ interspersed maps are not random, as previously assumed. We cannot confirm, however, recent theoretical literature suggesting that agoutis might have “classical” columnar orientation preference maps. Future research should focus to understand the circuits, which lead to small selective receptive fields in agoutis and great visual performance while adopting a different functional architecture.

Extracellular electrophysiology is among the main tools used for studying neural activity. In addition to having a high temporal resolution, this technique is stable enough to allow for recording freely-moving animals. In the last decades, the electrophysiology of the hippocampal formation has received particular attention due to the discovery of its relationship with memory and spatial navigation. Specifically, many spatial correlates have been investigated to understand the basis of neural encoding, such as place cells, which increase their firing rate when the animal is at particular locations of the environment. In this thesis, we present three studies exploring hippocampal electrophysiology through different levels of analysis, which aim at better understanding the different coding mechanisms used by the brain. In the first work, we focus on the problem of spike sorting, which consists in the classification of the waveforms detected by the extracellular recording. We investigated the use of Gaussian mixture models to perform feature extraction and clusterization of the waveforms. After using real and simulated data to evaluate the best strategies of our algorithm, we compared our spike sorting to two other known methods. Our algorithm, which combines two main feature extraction techniques with a new method of dimensionality reduction, showed better, or similar, results compared to the two other spike sorters. In the second work, we analyzed the current metrics used to find neurons presenting some spatial correlate, or information, such as place cells, head-direction cells, and speed cells. Using real and simulated data, we evaluated how the spatial information estimated by each metric correlated to the empirical information obtained using a Naive-Bayes classifier. Our results showed that the two main current metrics fail to detect the information content of neurons with high basal firing rate, and thus bias the universe of spatial correlates to low-firing cells. Finally, in the third work, we explored the differences in the proposed mechanisms underlying place cell firing: rate coding and temporal coding. We analyzed the dynamics of coupling between place cells and theta oscillations, comparing it to the firing rate dynamics. Our results revealed an asymmetry between the two mechanisms, with the theta coupling place cells preceding its major changes in firing rate, which corroborate the hypothesis that temporal and rate coding are independent mechanisms. Beyond that, we found that temporal coding represents more extensively the future positions of the animal than the past ones, suggesting its involvement in trajectory planning. In summary, the results of this thesis contribute to both the understanding of neuronal encoding and the development of new techniques in the field of hippocampal electrophysiology

Mindfulness-based training have been used to facilitate more adaptative mechanisms aimed to reduce anxiety, depression, stress, and improve attention regulation, emotions and enhance quality of life. The present study integrates variables related to sustained attention, interoception, affect and stress response in young adults submitted to a three-day training using a mindfulness-based training. The aim of this study is to understand the mechanism by which the mindfulness meditation practice acts acutely regarding to the variables approached.  Electrophysiological, hormonal and behavioral data were collected in two moments, pre and post intervention, in an active control group (n= 20) and an experimental group (n= 20) Our results suggest a difference between groups related to interoception, mindfulness and sustained attention after training. These results are represented through psychological measures and analysis of event-related potentials in one well validated attention task. In addition, intragroup analyzes suggest reduction of anxiety, increased interoceptive sensitivity and state of mindfulness after intervention in the experimental group. Moreover, the data shows lower negative affect and perceived stress in both groups after intervention. In this sense, it is suggested that the brief mindfulness-based intervention impacts on the cognitive performance regarding to an attentional test, with decrease of latency and increase of amplitude of the P300 component. Besides that, increased state of mindfulness, interoception and decrease in the levels of negative affect and perceived stress are also important results consistent with previous findings related to more traditional interventions using mindfulness meditation in a variety of populations

The ayahuasca is a drink with psychedelic properties largely used by indigenous populations from the Amazon basin. It contains the psychedelic tryptamine N,N-dimethyltriptamine (DMT), and monoamine oxidase inhibitors (iMAO), such as harmine and harmaline. The ayahuasca is considered to be a serotonergic psychedelic, capable of inducing an altered state of consciousness with similarities to an oneiric experience, with intense alterations in perception, though, humor, emotion, and mystical-type experiences. The neural correlates of its acute effects have been investigated by different neuroimaging techniques, including electroencephalography (EEG). In this study, we explored EEG spectral changes in 50 healthy volunteers, using a randomized double-blind placebo-controlled experimental design. Half of the volunteers received ayahuasca, while the other half received placebo. After dosing, the volunteers were monitored by an EEG equipment for approximately 4-hours. Aiming to improve data quality, the volunteers were asked to perform 2 simple tasks in three specific moments: before intake, 2h and 4h after intake. For the first task, they should try to stay awake, and alternate moments of eyes open (20 seconds) and eyes closed (40 seconds) for 5 minutes. For the second task, they should keep their eyes closed, trying to stay awake, for another 5 minutes. The spectral analysis (2h) found the alpha power to be significantly lower for the ayahuasca group when compared to placebo in occipital and right temporoparietal regions. Moreover, we found a significant increase in theta power on the right temporoparietal region, also 2h after the intake. Correlation analysis revealed correspondences between the alpha power (2h) and individual scores on two scales used to measure psychedelic effects – the Hallucinogen Rating Scale (HRS) and the Mystical Experience Questionnaire (MEQ). Additionally, we also present curious events found during visual inspection of EEG tracings. Overall, our results suggest that the inhibition of alpha oscillations in posterior brain regions play an important role on the psychedelic experience, maybe sharing mechanisms present during the oneiric experience. 

In order to make sense out of a sequence of sounds in music, our brain must meaningfully fit and recombine acoustic events into a hierarchic online stream. Although these information units are auditively  delivered in sequences with local connections (one after the other), it is assumed that long term dependencies are established counting on memory traces to sustain recursiveness  in time. Despite theoretical and empirical consensus, there is yet no clear physiological evidence of the temporal dimension of syntactic relations in music.

We investigated whether there is quantifiable neural activity suggesting the existence of a mental representation for fundamental music syntax rules like tonic-dominant-tonic chords. For such, we compared brain electric activity in 24 subjects (12 musicians, 12 non-musicians) aroused by original and harmonically modified versions of J.S. Bach chorales, using electroencephalography (EEG). Chorales were built by two phrases: initiated by a tonic and arriving in a dominant chord two bars away (first phrase), and concluded in a tonic chord three bars after the dominant (second phrase). Modified versions were created either by elevating or lowering the first, therefore keeping the second phrase intact. We compared the brain electric response for the last chord in both versions.  

Cortical gamma oscillations (30 - 90 Hz) have been implicated in various cognitive processes, such as visual binding and attention. So far, most evidence in support of this hypothesis are based on studies that used artificial and simplified stimuli, such as moving gratings and bars. Recently, experimental work using natural images led to conflicting conclusions. In a paradigm that required human subjects to maintain fixation, electrocorticogram signals (ECoG) showed gamma for grating stimuli but not for static images or pink noise (Hermes et al., 2015). On the contrary, analysis of ECoG in the early visual cortex of macaque monkeys revealed strong gamma components for free viewing of natural scenes (Brunet et al., 2015). Here, we aim to clarify these discrepancies using a paradigm that allowed direct comparisons between fixation vs. free viewing conditions, for both simplified stimuli (moving and static gratings) and natural scenes (static and moving images). Recordings of spiking activity and local field potentials (LFPs) were obtained from the central and the peripheral representations of V1. Our results show that in capuchins (N= 3 monkeys), as previously described in macaques and in humans, gamma is characteristically strong when stimulus parameters, such as size, orientation and speed are set at to optimal values. Comparisons between fixation vs. free viewing conditions and gratings vs. natural stimuli revealed that gamma is always strong for optimal grating stimuli, regardless of viewing condition (N= 93 recording sites, 2 monkeys). However, gamma is surprisingly absent during free viewing of natural images and movies. Similar negative findings were also obtained when the monkeys were exposed to real-world scenes, such as objects and other animals in the laboratory. The present results suggest that strong, narrow-band, gamma responses in V1 is primarily associated with the selective activation of cell populations sharing similar response properties. Therefore, gamma may be seen as a resonance phenomenon of the underlying cortical connectivity. Overall, our results belittle the importance of gamma as a key cortical mechanism for vision.

In humans, the processing of sound information allowed the appearance of language and music. The pitch - which allows the melody of a song and prosody in spoken discourse - is one of these attributes. Pitch perception is a universal skill, however, just a few individuals are able to identify or produce a tone at a particular pitch without an external tonal reference, an ability known as the Absolute Pitch (AP). However, the neural mechanisms responsible for such ability are not yet fully understood. The present work has the objective to contribute to the understanding of the neural processes involved in the perception of pitch in AP possessors. In the first study, we evaluated AP prevalence in a local population of musicians (School of Music, UFRN). For this, we used psychophysical tools and a questionnaire. This first work showed that AP is not an "all-or-nothing" process, but rather that it has different levels of performance: from people that perform below chance, until it increases gradually to 100% of correct answers. While traditional thresholds (~85%) showed a prevalence of AP similar to that observed in musicians in Europe and the United States, the application of a statistical threshold resulted in prevalence similar to that described in populations of music conservatories. In addition, we showed that AP possessors hit more the natural notes (those with the white-keys of the piano) than those that are accidental (flat or sharp, i.e. the black keys of the piano). We also observed that AP possessors present an earlier onset of musical training. Finally, we showed that AP was more prevalent in a high proficiency group in comparison to the average proficiency group. In a second study, we used the auditory brainstem evoked potentials (ABR) to quantify the activation of brainstem nuclei in the processing of heights of individuals with OA. In this work, we showed that sustained responses (but not the transient ones) present a higher energy in individuals with AP than in control musicians. We also found that the amplitude of this sustained response correlates with the reaction time in a pitch naming test. These results suggest that individuals with AP have an increased refinement in the processing of acoustic information in the early stages of auditory processing, which would thus contribute to a greater automation of pitch identification. We believe that this thesis will contribute to the understanding of the relationship between nervous system development and musical learning, thus contributing to the development of new music teaching techniques and training programs.

The understanding of complex human behaviors such as language and its variations in different conditions and contexts has been an important research aim for many decades. Naturalistic and quantitative approaches to precisely measure language variations from the structural and semantic points of view have recently emerged, allowing the measurement of variations manifested in free speech that reflect atypical cognitive decline in pathological situations such as psychoses, or typical cognitive development in healthy children during alphabetization, and even the processing of memories in different states of consciousness, such as waking and dreaming. In this work we will start discussing 1) the construction of tools for the analysis of speech structure inspired by the psychopathological descriptions of mental illnesses, 2) their application to the differential diagnosis of psychosis and
dementias, and 3) the application of semantic tools to predict psychotic episodes. We will proceed by widening this view away from pathology, so as to determine 4) how graph-theoretical measures of language structure vary across healthy cognitive development, and 5) how they relate to indices of academic achievement. Next we will investigate 6) how speech structure varies within a large sample of healthy and psychotic subjects with large age and educational variation (N=200, ages 2-58, kindergarten to PhD), to 7) evaluate the impact of years of education and 8) compare with the development of literature across 5,000 years (N=448 texts; Syro-Mesopotamian (N=62), Egyptian (N=49), Hinduist (N=37), Persian (N=19), Judeo-Christian (N=76), Greek-Roman (N=134), Medieval (n=20), Modern (n=20) and Contemporary (n=31)). We will conclude by applying tools of semantic similarity as a strategy to 9) measure memory reverberation during dreams and their electrophysiological correlates in a sleep
transition experiment. The results indicate that the structural and semantic tools used in this work can greatly improve the precision of naturalistic measurements of the complex human behaviors expressed in speech.

Recent open label trials show that psychedelics hold promise as fast-onset antidepressants in treatment resistant depression. In Brazil, a psychedelic substance worth mentioning is the ayahuasca. This brew composed by the psychedelic N,N-dimethyltryptamine (DMT), a serotonin and sigma-1 receptors agonist, and reversible monoamine oxidase A inhibitors, such as harmine, harmaline, and tetrahydroharmine, causes changes in perception, emotion and cognition. Preliminary results from our group suggest its antidepressant potential, in addition to demonstrating its safety and tolerability. Although promising, none of these studies have controlled for the placebo effect, which is high in clinical trials for depression. To address this issue, we conducted a parallel-arm, double-blind randomized placebo-controlled trial in 35 patients with treatment-resistant depression. A healthy control group of 50 volunteers was also examined. Patients received a single dose of either ayahuasca or placebo. One day before dosing, and one day after dosing patients were enrolled in a series of tests and assessments, including psychometric scales and fMRI, conducted in the course of a four-day experimental session, to assess changes in different markers of depression. Two clinical scales for depression assessed changes in depression severity: the Montgomery–Åsberg Depression Rating Scale (MADRS) and the Hamilton Depression Rating scale (HAM-D). Assessments were made at baseline, and at 1, 2 and 7 days after dosing. We observed significant antidepressant effects of ayahuasca when compared to placebo at all endpoints. MADRS scores were significantly lower in the ayahuasca group, compared to placebo. Likewise, HAM-D scores were significantly different at D7. Between-group effect sizes increased from D1 to D7. Response rates were high for both groups at D1 and D2, and were significantly higher in the ayahuasca group at D7. Remission rate was marginally significant at D7. In addition, we examined the relationship between the antidepressant and acute psychedelic effects of ayahuasca. The psychedelic effects were assessed using the Hallucinogen Rating Scale (HRS) and the Mystical Experience Questionnaire (MEQ30) after the dosing session. We found that higher psychedelic effects measured by both HRS and MEQ30 correlated with improvements in clinical outcome at D7. Finally, to investigate the neural bases of the antidepressant effects of ayahuasca, we used fMRI data through an emotional and resting state protocols at baseline, and one day after dosing. We observed changes in key limbic regions, such as subgenual cingulate cortex, amygdala and insula in ayahuasca group when compared to placebo. To our knowledge this is the first controlled trial to test a psychedelic substance in treatment resistant depression. Overall, this study brings new evidence supporting the safety and therapeutic value of ayahuasca, dosed within an appropriate setting, to help treat depression.

Estudos recentes têm demonstrado que o reconhecimento da lateralidade de mãos em uma perspectiva de segunda pessoa engaja processos de imagética motora. No entanto, pouco se sabe sobre a atividade e contribuição das regiões sensório-motoras corticais nesse processo. O objetivo do presente estudo é mostrar como a rotação mental de estímulos visuais de partes do corpo influenciam o processamento neural dessas regiões através da modulação da atividade oscilatória de grupos neuronais nas faixas alfa e beta. Para tanto, realizamos a análise da perturbação espectral relacionada ao evento (ERSP) do registro eletroencefalográfico (EEG) de 20 sujeitos engajados no reconhecimento da lateralidade de mãos apresentadas em uma tela de computador. Os resultados mostram que a supressão da amplitude das frequências alfa e beta foi maior nas regiões sensório-motoras e que características do estímulo (como orientação da mão, visão, postura de dedos e lateralidade) são capazes de modular esses componentes oscilatórios em diferentes regiões, sugerindo etapas distintas para o processamento da tarefa cognitiva.

Temporal lobe epilepsy (TLE) is the most common form of epilepsy in adults. It is characterized by a progressive occurrence of epileptic seizures originating in the temporal lobe, particularly in the hippocampal formation (mesial TLE). Among the animal models used to investigate the physiopathological mechanisms of TLE, the most used are those that lead to spontaneous seizures after an initial insult, such as a prolonged convulsive state (status epilepticus, SE). This condition can be induced by the administration of the glutamatergic agonist, kainic acid (KA) or the cholinergic agonist, pilocarpine (PILO). However, the use of systemic injections and the lack of electrophysiological monitoring during SE lead to high mortality rates, widespread cell death and high behavioral variability during the chronic phase of epilepsy, which differs in several aspects from the human condition. These effects are mainly due to the lack of electrographic control of SE duration and the dynamics of the epileptogenesis process during the weeks following SE. Therefore, this project aimed to generate two animal models of TLE by intra-hippocampal injections of KA or PILO, and then, to analyze their behavioral and electrographic progression during SE. It is important to notice that no electrophysiological study has investigated the SE dynamics in animals infused with PILO into hippocampus so far. For that, we implanted two bundles of microelectrodes in the hippocampus bilaterally, one bundle in the medial prefrontal cortex and a cannula above the intermediate hippocampus for KA or PILO infusion. Following SE induction, we analyzed the behavioral and electrophysiological evolution of KA and PILO animals. SE was blocked after 2h by the injection of an anti-convulsant cocktail and the animals were continuously monitored by video-EEG for up to 72h. Seven days after SE, animals underwent euthanasia and had the brains removed for histological localization of cannula and electrodes. Video and EEG recordings were analyzed by visual inspection and spectral decomposition. Our results showed that PILO animals had shorter latency for first behavioral seizure than KA rats after drug injection. However, seizure severity showed higher variability among PILO rats (PILO: 50% animals had class 3 or higher vs.KA: 90% animals had class 1; Racine’s scale). PILO animals had a reduced number of wet-dog shake behaviors and shorter latency to SE onset as compared to KA rats. Electrophysiologically, we observed that high frequency oscillations (>150 Hz) occurred short after the injection of both drugs (15-40min before SE onset), as opposed to what is commonly reported to occur during the chronic phase of epilepsy in rodents. They were usually found within the first electrographic seizures. Finally, we have identified a distinct modular organization of paroxystic activity during the SE in each group, which consisted of blocks of nested rhythms. These findings thus suggest that PILO is more epileptogenic that KA and that these drugs produce distinct SE dynamics, which seem to be organized as periodically repeating modules of nested oscillations, modules of hypersynchrony with no nested oscillations and segments of asynchronous activity. Our data emphasizes the importance to conduct electrophysiological recordings during SE induction in order to better control individual brain responses. This can reduce variability during epileptogenesis and produce a more homogeneous model of chronic epilepsy.

Recent studies in computational psychiatry (Mota et al., 2012, Mota et al., 2014, Mota et al., in preparation) indicate significant structural differences between the speech of psychotic and healthy subjects. Graph analysis of these subjects’ autobiographic reports allow the quantification of psychotic speech symptoms, marked by poor, disconnected and repetitive speech. In the 1970’s, Julian Jaynes proposed that psychosis was socially prevalent until around 1,000 BC. This conjecture was first tested by Diuk et al. (2012), who used Latent Semantic Analysis to assess the semantic similarity to the introspection concept across Judeo-Christian and Greco-Roman texts from the first millenium BC. In this perspective, we seek to investigate this hypothesis proceeding graph analysis of 448 texts across history, from 3,000 BC to 2,010 AC. Graph measures of six different cultures of Afro-Eurasia (Syro-Mesopotamian, Egyptian, Hindu, Persian, Judeo-Christian, Greek-Roman) and three more recent literary categories (Medieval, Modern, and Contemporary) reveal a clear pattern: While lexical diversity and connectivity increase, recurrence decreases. These findings are compatible with the notion that psychosis is an early trait of our species.

A Doença de Parkinson (DP) é um distúrbio motor relacionado ao envelhecimento que atualmente acomete de 1-2% da população mundial acima dos 60 anos. No Brasil, estima-se que esta acometa aproximadamente 600 mil indivíduos, configurando-se como uma enfermidade de importância para países em processo de incremento da expectativa de vida. A epidemiologia da DP revela fatores de risco intrínsecos e extrínsecos ao paciente que definem a chance de desenvolvimento do distúrbio. Mutações pontuais e polimorfismos com significado funcional são tidos como fatores genéticos predisponentes, enquanto a exposição a pesticidas e toxinas destacam-se como fatores ambientais. No entanto, poucos estudos em modelos animais focam em investigar a interação entre estes fatores. Isto pode ser alcançado comparando-se os efeitos de substâncias indutoras de parkinsonismo em linhagens de ratos com diferentes contextos genéticos. Recentemente, o tratamento repetido com baixas doses de reserpina – um inibidor irreversível do transportador vesicular de monoaminas (VMAT2) – foi proposto como um modelo progressivo para a DP. Sob este regime de tratamento, roedores apresentam, de forma progressiva, comprometimento motor, cognitivo, e alterações neuroquímicas compatíveis com a fisiopatologia da DP. Em paralelo, comparados a ratos Wistar, animais da linhagem SHR (Spontaneously Hypertensive Rats) são resistentes à indução da discinesia oral pelo tratamento agudo com reserpina. Em vista destes achados, nós buscamos avaliar se ratos SHR seriam resistentes aos déficits motores e alterações neuroquímicas quando submetidos ao modelo progressivo para DP induzido por reserpina. Portanto, nós submetemos ratos Wistar e SHR ao tratamento agudo (1 mg/kg) ou repetido (15 injeções de 0,1 mg/kg, em dias alternados) com reserpina e investigamos a progressão dos déficits motores nas tarefas de catalepsia em barra, discinesia oral, e atividade espontânea em campo aberto. Observamos então que, para ambos os regimes de tratamento, animais SHR se mostram resilientes ao prejuízo motor em todas as dimensões motoras avaliadas. Ainda, estas diferenças se manifestaram tanto na latência para o surgimento do comprometimento motor como para a magnitude deste. Estas alterações foram ainda acompanhadas por decréscimo na expressão da tirosina hidroxilase (TH) e incremento na expressão de α-sinucleína na via nigro-estriatal de ambas linhagens submetidas ao tratamento com reserpina. Estas alterações neuroquímicas resultantes do tratamento com reserpina também se refletiram nos níveis de monoaminas – dopamina e serotonina – na via nigro-estriatal destes animais. De modo geral, como no comportamento motor, animais SHR apresentaram atraso para a depleção de monoaminas e menor magnitude deste efeito. Em conclusão, os resultados aqui apresentados claramente corroboram a resiliência de ratos SHR ao modelo progressivo da DP. Estes achados expõem novos alvos potenciais para as diferenças neuroquímicas, moleculares e genéticas na linhagem SHR relevantes para o estudo da susceptibilidade à DP.

Synchronization among neurons arises from the cooperative interaction of various cell types through excitation and inhibition. The mechanisms behind this type of neuronal coordination are as versatile as almost no other coordination task in the brain, making its comprehension heavily challenging. Among many others, the high number of involved cell types, the diversity of synaptic processes as well as the contribution of a multitude of ion channels and currents span the plurality of neuronal synchronization mechanisms in our brains. Focusing on two main brain areas, the hippocampus and the cortex, this thesis aims to understand the role of distal dendritetargeting interneurons in shaping pyramidal cell activity and the timing of their action potentials.

The distribution of ion channels and synaptic receptors in pyramidal cell dendrites is extremely anisotropic. Thus, interneurons innervating the proximal or distal areas of the dendrites cause different effects in the target cell when activated. For example, the distal portions of the pyramidal cell dendrites contain one of the most prominent pacemaker channels: the hyperpolarizationactivated cyclic nucleotide-gated channels. These channels produce a cationic depolarizing current (Ih) and play an important role in the regulation of neuronal excitability. Using computational modeling, this thesis shows how the amount of Ih in certain cell types changes their spike rate, synchrony as well as power and frequency of ongoing network oscillations. Moreover, since Ih differs between brain regions as well as cell types and location, this thesis electrophysiologically explores how Ih differs along the dorso-ventral axis of hippocampus in oriens-lacunosum moleculare (OLM) cells, the main distal dendrite-targeting interneurons of that region.

Utilizing the main distal dendrite-targeting interneuron of the cortex, the Martinotti cell, this thesis also shows how a defined population of interneurons can be manipulated in order to control and align pyramidal cell firing. By providing the right amount and frequency of inhibition, Martinotti cells are able to synchronize trains of subtype-specific pyramidal cells. Using optogenetic approaches to activate/inactivate populations of Martinotti cells, these dendrite-targeting interneurons are shown to trigger rebound action potentials in pyramidal cells when temporally
aligned. The rebound action potentials in turn are the result of strong Martinotti cell inhibition, giving these distal dendrite-targeting interneurons the potential to reset pyramidal cell firing.

Overall, Martinotti cells and OLM cells show quite striking similarities in morphological, neurochemical and electrophysiological properties. Especially, their long axonal projections to upper layers as well as their low-threshold, slow spiking fashion and the accommodating firing make these distal dendrite-targeting interneurons so special for neuronal synchronization

As principais vias aferentes ao hipocampo vêm do córtex entorrinal e fazem parte de um loop que retorna ao entorrinal após passar pelo giro denteado, e pelas subareas do hipocampo CA3 e CA1. Desde a descoberta nos anos 50 de que o hipocampo está envolvido na formação de memórias, esta região vem sendo extensivamente estudada. Além desta função mnemônica, o hipocampo também está associado a navegação espacial. Em camundongos e ratos, células de lugar exibem um aumento da taxa de disparo relacionado à posição do animal. O local do ambiente onde uma determinada célula de lugar se ativa é chamado de campo de lugar. A taxa de disparo das células de lugar é máxima quando o animal está no centro do campo de lugar, e diminui a medida que ele se afasta desse ponto, sugerindo a existência de uma codificação espacial baseada em taxa de disparos. Entretanto, pesquisas prévias vêm mostrando a existência de oscilações hipocampais em múltiplas frequências e ligadas a diferentes estados comportamentais, e muitos acreditam que estas oscilações são importantes para uma codificação temporal. Em particular, oscilações teta (5-12 Hz) possuem uma relação espaço-temporal com as células de lugar conhecida como precessão de fase. Na precessão, a fase de disparos da célula de lugar muda gradualmente do pico de teta para o fundo e, posteriormente, para a fase ascendente, a medida que o animal atravessa o campo de lugar. Além disso, as teorias vigentes sugerem que CA1, a porta de saída do hipocampo, intermediaria a comunicação com o córtex entorrinal e CA3 através de oscilações em diferentes frequências chamadas, respectivamente, de gama alto (60-100 Hz; HG) e gama baixo (30-60 Hz; LG). Essas oscilações se relacionam com teta, estando aninhadas dentro de cada ciclo desta frequência mais lenta. Nesta dissertação, utilizamos dados disponibilizados online para fazer análises computacionais visando reproduzir resultados clássicos e recentes acerca da atividade das células de lugar no hipocampo de ratos em livre movimento. Em particular, nós revisitamos o debate sobre a relação da precessão de fase com variações na taxa de disparos e na posição do animal no campo de lugar. Concluímos que este fenômeno não pode ser explicado por nenhuma dessas variáveis sozinha, e sim pela interação entre elas. Nós também realizamos novas análises investigando as propriedades das células de lugar em relação às oscilações. Nós mostramos que o nível de modulação dos disparos por teta afeta apenas levemente a informação espacial contida nas células de lugar, enquanto a fase de disparo média não tem nenhuma influência na informação espacial. Também encontramos que as células de lugar estão moduladas por teta quando disparam fora do campo de lugar. Além disso, nossos resultados mostram que o disparo das células de lugar dentro do ciclo de teta segue os padrões de modulação de HG e LG por teta presentes nos potenciais de campo local de CA1 e córtex entorrinal. Por último, achamos um acoplamento fase-amplitude em CA1 associado apenas aos disparos dentro do campo de lugar na faixa de 40-80 Hz. Concluímos que o disparo de células de lugar está ligado a estados de rede refletidos no potencial de campo local e sugerimos que a atividade dessas células sejam interpretadas como um estado dinâmico ao invés de uma propriedade fixa da célula.

Os estudos revelam que nas ultimas décadas ocorreu uma diminuição na duração do sono.  Os compromissos sociais, como o trabalho e a escola, muitas vezes não estão alinhados ao “tempo biológico” dos indivíduos. Somado a isso, observa se uma menor força do zeitgeber causada pela menor exposição à luz de dia e maior à noite. Isso gera um débito de sono crônico que é compensado nos dias livres ocorrendo semanalmente uma restrição e extensão do sono denominada de Jet lag social. A privação de sono vem sendo associada à obesidade, risco cancerígeno e cardiovascular. Sugere-se que o sistema nervoso autonômico seja um caminho que relaciona os problemas do sono às doenças cardiovasculares. No entanto, além das evidências demonstradas por pesquisas com uso de modelos de privação de sono de forma aguda e controlada, são necessários estudos investigando efeitos da privação do sono de forma crônica como ocorre no Jet lag social. O objetivo deste estudo foi investigar a influência do Jet lag social em marcadores circadianos de atividade-repouso e cardíacos em estudantes de medicina. Trata-se de um estudo transversal e observacional realizado no departamento de fisiologia pelo Laboratório de Neurobiologia e Ritmicidade Biológica (LNRB) da UFRN. Participaram da pesquisa estudantes de medicina matriculados no 1º período do curso da UFRN. Foram utilizados os seguintes instrumentos: Questionário Cronotipo de Munique (MCTQ); Questionário para identificação de indivíduos matutinos e vespertinos (MEQ ou HO); Índice de qualidade do sono de Pittsburgh; Escala de Sonolência de Epworth; Actímetro; Cardiofrequencímetro. Foram analisadas variáveis de caracterização do sono, não paramétricas (IV60, IS60, L5 e M10) e índices cardíacos no domínio do tempo (intervalo RR, FC, SDNN, RMSSD, STD HR, NN50, pNN50), frequência (LF, HF, LF/HF) e não linear (SD1, SD2, SD1/SD2). Realizou-se análise estatística descritiva, comparativa e correlação com uso do programa SPSS versão 20. Participaram do estudo 41 estudantes, 48,8% (20) mulheres e 51,2% (21) homens, 19,63 ± 2,07 anos. O Jet lag social teve uma média de 02:39h ± 00:55h, 82,9% (34) com Jet lag social ≥ 1 hora e houve correlação negativa com escore cronotipo de Munique evidenciando maior privação do sono em indivíduos com tendência à vespertinidade. 90,2% (37) tiveram qualidade do sono ruim (X² = 26,56, p < 0,001) e 56,1% (23) sonolência diurna excessiva (X² = 0,61, p = 0,435). Observou-se diferença significativa dos valores de LFnu, HFnu e LF/HF entre os grupos de Jet lag social < 2h e ≥ 2h e houve correlação do Jet lag social com LFnu (rs = 0,354, p = 0,023), HFnu (rs = - 0,354, p = 0,023) e LF/HF (rs = 0,355, p = 0,023). Verificou-se ainda associação negativa entre IV60 e índices no domínio do tempo e não lineares. Sugere-se que a privação crônica de sono pode ter associação com maior atividade simpática promovendo maior risco cardiovascular.

Early physiologists were dazzled by the occurrence of high-amplitude, periodic oscillations, easily discernible in recording traces from the eye, optic tract and optic ganglia. Numerous studies thereafter pointed to retinal ganglion cell as the elements responsible for the generation of these fast rhythms, which were known to propagate to the lateral geniculate and to the cortex. Only recently, however, these early observations gained renewed interest, mainly in the light of recent theories linking neuronal oscillations to various cognitive processes, such as perceptual binding, attention and memory. In this context, fast retinal oscillations (FRO) have been associated to the binding of contiguous contours or surfaces, which in principle could support a fast feedforward segmentation process. In addition, a series of experiments in the cat have shown that FRO may convey global stimulus properties, such as size.

A limitation in these previous studies, however, was that most of them where were made in the anesthetized and paralyzed cat. Only a few early studies have been performed in the non-anesthetized but still paralyzed cat. Another concern was that, in these latter experiments, visual stimuli were often limited to ganzfeld flashes, far from natural vision conditions. Moreover, very recently we made the surprising discovery that FRO depend strongly on halothane (and isoflurane) anesthesia. It was therefore imperative to verify whether FRO are present or not in the awake cat, in naturalistic conditions, such as during free-viewing of a visual scene. This is the main goal of the present study.

Simultaneous multiple-electrode recordings were made from the LGN and the retina of anesthetized cats (N= 3) and from the LGN of an awake cat (N= 1). Comparisons were made for responses to natural movies and flashed stationary light stimuli. To test specifically the role of FRO in encoding stimulus size we designed a protocol made of a light circle of varying size along the trial. Spike sorting techniques allowed us to study separately the ON- and OFF-components of the responses. Analysis consisted in measuring synchronous oscillations for single cell spiking activity in the time (sliding correlation analysis) and spectral domains (multitaper spectral analysis, multitaper coherence). Our present results based on single-cells extend our previous findings in the anesthetized cat, which were restricted to an autocorrelation analysis of LGN mutiunitary responses. Both ON- and OFF-responses to varying size stimuli show that coherent oscillations appear only after the stimulus attained a minimum size of about 5°, suggesting that FRO is rather limited in encoding subtle changes in stimulus size. Recordings obtained directly from the retina showed that FRO are highly dependent on halothane anesthesia levels. Notably, in a series of sessions we were able to record LGN responses in an awake cat, which was subsequently anesthetized by halothane, keeping the same recording site. FRO were completely absent in the awake condition and appeared strong as usual during the halothane anesthesia.

   Os recentes avanços técnicos das duas últimas décadas para o registro de sinais neuroeletrofisiológicos foram essenciais para que se testassem hipóteses há muito propostas acerca de como células nervosas processam e armazenam informação. No entanto, ao permitir maior detalhamento dos dados coletados, as novas tecnologias levam inevitavelmente ao aumento de sua complexidade estatística e, consequentemente, à necessidade de novas ferramentas matemático-computacionais para sua análise.

Os objetivos da presente tese foram os de investigar padrões comportamentais e eletrofisiológicos associados à resiliência e suscetibilidade ao estresse social induzido em camundongos. Para isso, foi utilizado um protocolo de indução de estresse crônico contínuo a partir de derrotas sociais baseado no paradigma intruso-residente. Os resultados da tese são apresentados em dois estudos. No primeiro estudo, camundongos C57BL/6j submetidos a repetidos episódios de derrota social apresentaram motivação tardia para interagir com um camundongo desconhecido em sessões prolongadas (10 min) do teste de interação social. Utilizando uma abordagem etológica associada à análise computacional de vídeos foi possível rastrear precisamente a posição dos camundongos durante a realização de comportamentos de investigação social. Com isso, foi analisada a expressão detalhada de comportamentos defensivos, tais como investigação em postura estendida e fugas, associados ao comportamento de investigação social. A partir dessas análises, foi demonstrado que a realização do comportamento de investigação social em postura estendida foi significativamente maior para o grupo derrotado em comparação com o grupo controle. Ainda, um subgrupo de camundongos derrotados apresentou investigação social em postura estendida de forma persistente e sem habituação. Utilizando uma medida da distância de investigação durante as investigações sociais foi possível calcular um índice de aproximação (IA) para cada animal e separar um subgrupo apresentando fenótipo relacionado à ansiedade. A incidência de fugas também foi maior no grupo derrotado em comparação com os controles. A persistência na ocorrência desse comportamento foi observada em um subgrupo de camundongos submetidos às derrotas sociais. Calculamos então um índice de fugas (IF) que se correlacionou inversamente com a preferência por sacarose, sendo útil para identificar animais anedônicos. No segundo estudo, foram combinados análise etológica e registros eletrofisiológicos com tetrodos na área tegmentar ventral de camundongos submetidos à derrotas sociais. Utilizando critérios eletrofisiológicos e farmacológicos, foram classificadas as unidades registradas na área tegmentar ventral como supostos neurônios dopaminérgicos e não-dopaminérgicos. Foram analisadas a atividade desses neurônios durante o comportamento de investigação social e observado que a modulação da taxa de disparo dessas subpopulações neuronais distintas ocorreu de maneira oposta em animais suscetíveis e resilientes ao estresse social. Em suma, propomos que sessões prolongadas associadas à análise etológica detalhada durante os testes de interação social podem prover informação para classificação de camundongos em resilientes e susceptíveis após repetidas derrotas sociais. Ainda, a expressão do fenótipo suscetível parece estar associada ao comprometimento do sistema dopaminérgico mesolímbico na atribuição de valor de incentivo às interações sociais normalmente associadas ao aumento da atividade neuronal mesolímbica.

Hebb propôs que sinapses entre neurônios que disparam de forma síncrona são fortalecidas formando assembleias de células e sequências de fase. A primeira, numa escala menor, é um conjunto de células sincronizadas, que funcionam de forma transitória como um sistema fechado de processamento; a última, numa escala maior, corresponde à ativação sequencial de assembleias de células neuronais capazes de representar percepções e comportamentos. Atualmente, o registro de grandes populações neuronais permite a detecção simultânea de diversas assembleias neuronais. No âmbito da teoria de Hebb, o próximo passo lógico é a análise das sequências de fase. Neste trabalho investigamos seqüências de fase como padrões de ativações consecutivas de assembleias, analisando a relação entre comportamento animal e atributos de grafos de assembleias. Foram estudados trens de disparo neuronal registrados no hipocampo e neocórtex de 5 ratos adultos, antes, durante e depois da exploração de novos objetos (períodos experimentais). Para definir um grafo de assembleia, cada assembleia correspondeu a um nó, e cada aresta correspondeu à sequência temporal de ativação de nós consecutivos. A soma da ativação de todas as assembleias foi proporcional à taxa de disparo, mas a atividade de assembleias individuais não. O repertório de assembleias permaneceu estável ao longo dos períodos experimentais, indicando que a experiência com novos objetos não criou novas assembleias no rato adulto. Os atributos de grafos das assembleia, por outro lado, variaram significativamente entre os estados comportamentais e períodos experimentais e foram distintos o suficiente para permitir a classificação automática dos períodos experimentais (classificador Naive Bayes; AUROCs
máximas variaram entre 0,55 a 0,99) e estados comportamentais (vigília, sono de ondas lentas e sono de movimento rápido dos olhos; AUROCs máximas variaram entre 0,64 e 0,98). Nossos achados reforçam a teoria Hebbiana de que as assembleias neuronais correspondem a estruturas primitivas de representação, quase inalteradas na maturidade, enquanto as seqüências de fase são instáveis entre os estados comportamentais e mudam após novas experiências. Os resultados são compatíveis com um papel das sequências de fase no comportamento e cognição.

O ritmo teta consiste em uma oscilação eletrofisiológica hipocampal presente em várias espécies de mamíferos (4-12 Hz, com variações entre espécies). Essa oscilação está presente durante a vigília ativa de ratos e também é predominante no PCL desta espécie durante o sono de movimento rápido dos olhos (sono REM). Vários trabalhos demonstraram que o ritmo teta é importante em tarefas cognitivas. O septo medial é uma região importante na geração do ritmo teta hipocampal. Possui projeções colinérgicas, GABAérgicas e glutamatérgicas para o hipocampo, que por sua vez, possui projeções de feedback para o septo. Além do septo, outras regiões estão envolvidas na regulação do teta, formando uma rede complexa de interação e coordenação entre áreas que resultam no ritmo. A optogenética é uma ferramenta desenvolvida recentemente que tem sido amplamente utilizada em pesquisas de diversas áreas. Ela nos permite manipular a atividade elétrica de neurônios através de estimulação luminosa. A técnica consiste em, através de um vetor viral, induzir a expressão neuronal de canais iônicos associados a opsinas (ex.: ChR2), que uma vez infectados passam a ser sensíveis a luz de determinado comprimento de onda. O presente trabalho de pesquisa de mestrado teve como objetivo implantar a optogenética em animais em livre comportamento pioneiramente no Brasil, através de experimentos com implantes crônicos de eletrodos e fibras óptica em animais infectados com vetor viral para expressão de ChR2. Foram realizadas cirurgias de injeções de vírus no septo medial; resultados histológicos confirmaram a expressão de ChR2 através da marcação da
proteína repórter eYFP no septo e também em processos hipocampais. Além disso, foram realizados experimentos agudos com estimulação luminosa do septo medial e registro de potenciais de campo local (PCL) no próprio septo e hipocampo em animais anestesiados. Ainda nesses experimentos foi possível registrar potenciais de ação no septo. Nesses experimentos observamos aumento da taxa de disparo dos neurônios septais durante estimulação luminosa (n=300 estímulos). Além disso, encontramos uma resposta evocada no PCL do hipocampo no início do pulso luminoso. Também foram realizados experimentos crônicos com estimulação luminosa do septo medial e registro de PCL do hipocampo em animais em livre comportamento. Através de análise do PCL, verificamos se a estimulação luminosa do septo é capaz de induzir ritmo teta no hipocampo.

Since Hubel and Wiesel’s pioneer finding a vast body of literature has accumulated describing neuronal responses in the primary visual cortex (V1) to different visual stimuli. These stimuli mainly consisted of moving bars, dots or gratings which served to explore the responses to basic visual features such as orientation, direction of motion or contrast, among others, within a classical receptive field (CRF). However, in the last two decades it became increasingly evident that the activity of V1 neurons can be modulated by stimulation outside their CRF. Thus, early visual areas might be already involved in more complex visual tasks like, for example, the segmentation of an object or a figure from its (back)-ground. It is assumed that intrinsic long-range horizontal connections within V1 as well as feedback connections from higher visual areas are actively involved in the figure-ground segmentation process. Their possible role has been inferred from the analysis of the spike rate variations induced by stimuli placed outside the CRF of single neurons. Although it is very likely that those connections also have an impact on the joined activity of neurons involved in processing the figure and on their local field potentials (LFP), these issues remain understudied.

In order to examine the context-dependent modulation of those activities, we recorded spikes and LFPs in parallel from up to 48 electrodes in the primary visual cortex of anesthetized cats. We stimulated with composite grating and natural scene stimuli focusing on populations of neurons whose CRFs were situated on the foreground figure. In addition, in order to examine the influence of horizontal connections we removed the inter-hemispheric input of the isotopic contralateral visual areas by means of reversible cooling deactivation.

We did so because i) the intrinsic horizontal connections cannot be easily manipulated without directly affecting the measured signals, ii) because inter-hemispheric connections share the major anatomical features with the intrinsic lateral network and can be seen as a functional continuation of the latter across the two hemispheres and iii) because cooling causally and reversibly deactivates input connections by temporarily silencing the sending neurons and thus enables direct conclusions on their contribution.

Our results demonstrate that the figure-ground segmentation mechanism is reflected in the spike rate of single neurons, as well as in their LFP power and its phase-relationship to the spike patterns produced by the population. In addition "lateral" inter-hemispheric connections modulate spike rates and LFP power depending on the stimulation of the neurons’ CRF surround. Further, we observe an influence of this lateral circuit on field-
field coherences between remote recording sites. In conclusion, our findings support the idea of complex figure-ground segmentation mechanism acting already in early visual areas on different time scales. This mechanism seems to involve groups of neurons firing synchronously and dependent on the LFP’s phase. Our results are also compatible with the hypothesis that long-range lateral connections contribute to that mechanism.

O autismo compreende um grupo heterogêneo de transtornos do desenvolvimento que afeta a maturação do encéfalo e produz alterações sensoriais, motoras, de linguagem e de interação social, de início na infância. Vários estudos indicam um importante envolvimento de fatores genéticos que levam à uma predisposição ao autismo, além de fatores ambientais que podem influenciar a vida embrionária e pós-natal. Estudos recentes, em modelos animais, indicam que alterações no controle epigenético durante o desenvolvimento possam gerar distúrbios de maturação neuronal e produzir circuitarias hiper-excitáveis, resultando em sintomas típicos do autismo. No modelo animal de autismo induzido por administração de ácido valpróico (VPA) durante a gestação de ratas, foram observadas alterações comportamentais, eletrofisiológicas e celulares semelhantes às observadas nos pacientes autistas. Entretanto, ainda são poucos os estudos que correlacionam as alterações comportamentais com a suposta hiper-excitabilidade neuronal desse modelo. Portanto, o objetivo deste trabalho foi inicialmente gerar o modelo animal de autismo por exposição pré-natal a ácido valpróico, e então avaliar o desenvolvimento e comportamento pós-natal em animais pré-púberes (P30). Além disso, pretendemos quantificar e analisar a distribuição de neurônios inibitórios no córtex pré-frontal e cerebelo dos animais VPA comparando-os com animais controle. Para isso, acompanhamos o desenvolvimento desses animais e realizamos testes de atividade exploratória e locomotora, auto-limpeza, reconhecimento de objetos e interação social. Quantificamos interneurônios imunorreativos para parvalbumina no córtex pré-frontal medial (CPFm) e células de Purkinje do cerebelo para avaliar a ocorrência de alterações celulares envolvidas em hiper-excitabilidade cortical. Nossos resultados mostram que o tratamento com VPA foi capaz de induzir alterações no desenvolvimento dos animais, refletindo-se em alterações comportamentais como hiperlocomoção, comportamento repetitivo e déficit de interação social. A quantificação celular revelou uma diminuição no número de interneurônios parvalbuminérgicos no córtex cingulado anterior e na região pré-límbica do CPFm, sugerindo um desbalanço excitatório-inibitório neste modelo de autismo. Observamos também que a redução neuronal ocorreu preferencialmente nas camadas II/III e V/VI do córtex. Esperamos que nossos resultados possam contribuir para o maior entendimento das alterações celulares neste modelo de autismo, assim como esclarecer suas implicações funcionais.

As epilepsias são desordens neurológicas caracterizadas por crises espontâneas e recorrentes resultado de uma atividade elétrica anormal de uma determinada rede neural. Dentre os diferentes tipos de epilepsia, a epilepsia mesial do lobo temporal (EMLT) é a mais comumente observada em seres humanos, sendo frequentemente associada a esclerose hipocampal. Infelizmente, nem todos os pacientes se beneficiam do tratamento farmacológico (pacientes fármaco-resistentes) e tornam-se, portanto, candidatos ao tratamento cirúrgico, um procedimento de alta complexidade e elevado custo. Hoje, a lobectomia temporal anterior com amigdalo-hipocampectomia seletiva é a cirurgia de epilepsia mais frequentemente realizada. Entretanto, uma parcela significativa de pacientes continua a apresentar crises debilitantes mesmo após o tratamento cirúrgico. Assim, compreender o encéfalo epiléptico humano é fundamental no refinamento do tratamento cirúrgico com o objetivo de aumentar a eficiência desse procedimento terapêutico.

O objetivo da presente dissertação foi identificar e quantificar a ocorrência de atividade epileptiforme (espículas interictais, EI e oscilações de alta frequência, OAF) em registros eletrocorticográficos (ECoG) realizados durante procedimento cirúrgico em pacientes com epilepsia mesial do lobo temporal refratária ao tratamento farmacológico (protocolo aprovado pelo CEP / Unifesp; #1038/03). 

Registros ECoG (Nihon-Koden, 32 canais @ 1kHz) foram realizados na superfície do lobo temporal e no giro para-hipocampal em 3 momentos cirúrgicos: no córtex intacto, após lobectomia temporal anterior e após amigdalo-hipocampectomia (duração média de cada um desses registros: 10 min; N=17 pacientes). A ocorrência de EI e OAF foi quantificada automaticamente, por meio de rotinas em Matlab, e validadas visualmente. Frequência dos eventos (número de eventos/canal) em cada um dos tempos cirúrgicos foram correlacionadas com resultado cirúrgico quanto ao controle das crises. 

De um total de 8 h e 40 min de registro, 36.858 EI e 2.326 EAF foram identificadas. Os pacientes com melhor prognóstico cirúrgico apresentaram maior quantidade de OAF antes da cirurgia, porém não diferiram quanto a frequência, morfologia e distribuição de EI. A ocorrência de OAF no registro basal apresentou melhor desempenho que as EI na previsão do controle total das crises no pós operatório (EI: AUC = 57%, S = 70% , E = 71% vs OAF: AUC = 77%, S = 70%, E=100%). O mesmo foi observado com o parâmetro a variação da área irritativa entre os momentos pré- e pós-ressecção (EI: AUC = 54%, S = 60%, E = 71% vs OAF: AUC = 84%, S = 80%, E = 100%). Nesse caso, o classificador foi capaz de identificar todos os pacientes de pior prognóstico, apresentando apenas dois falsos positivos. 

O presente trabalho demonstra que as OAF podem ser encontradas no registro ECoG intra-operatório, na presença de anestésicos, em uma curta sessão de registro, juntamente com as EI. A observação de que a ocorrência desses eventos no início da cirurgia permite classificar o paciente quanto ao prognóstico cirúrgico é surpreendente e abre caminho para aplicar o ECoG intra-operatório, por exemplo, na decisão sobre o uso de tratamento farmacológico adjuvante ou da conversão para ressecções individualizadas. O mecanismo responsável por esse efeito ainda é desconhecido.

A Ayahusca é uma bebida psicotrópica que tem sido utilizada há séculos por populações originais da América do Sul, notadamente da região Amazônica, com fins religiosos e medicinais. O chá é obtido pela decocção de folhas de Psychotria viridis com a casca e tronco de um arbusto, Banisteriopsis caapi. A primeira é rica em N,N-dimetiltriptamina (DMT), que tem importante e bem conhecido efeito alucinógeno devido a sua atuação agonista nos receptores de serotonina, especificamente 5-HT_2A. Por outro lado, as b-carbolinas presentes na B. caapi, particularmente a harmina e a harmalina, são potentes inibidores da monoamina oxidase (iMAO). Além disso, a tetrahidroharmina (THH), também presente na B. caapi, atua como leve inibidor seletivo da recaptação de serotonina e um fraco inibidor de MAO. A DMT, por si só não é ativa por via oral, uma vez que é degradada pela MAO. No entanto, a presença de iMAO na bebida permite que a DMT seja psicoativa quando ingerida. O acesso da DMT à circulação sistêmica e ao sistema nervoso central provoca uma série de alterações afetivas, perceptivas e cognitivas. Além disso, os seus efeitos têm sido associados ao aumento da atenção interoceptiva. Por outro lado, existe um interesse crescente na rede de modo padrão (DMN), que tem sido amplamente detectada em estudos de neuroimagem funcional e tem sido associada com atividade mental introspectiva. Assim, este estudo teve como objetivo avaliar, por meio de ressonância magnética funcional – fMRI, as possíveis mudanças da DMN causadas pela ingestão da Ayahuasca em 10 voluntários saudáveis enquanto executavam uma tarefa de fluência verbal e protocolo de resting state. De maneira geral, observa-se que a Ayahuasca provoca redução na amplitude do sinal BOLD nos nodos centrais da DMN, tais como o cíngulo anterior, o córtex pré-frontal medial, o cíngulo posterior, o pré-cuneus e o lobo parietal inferior. Além disso, também foram observadas alterações no padrão de conectividade funcional da DMN, em particular, diminuição na conectividade funcional no pré-cuneus. Juntos, esses achados indicam que, sob efeito da Ayahuasca, os indivíduos apresentam um estado alterado de consciência, no qual o fluxo de pensamentos espontâneos é alterado, e sugerem um potencial uso terapêutico da Ayahuasca em transtornos mentais nos quais a DMN se mostra alterada.

Terapia Espelho (TE) vem sendo usada como uma ferramenta de reabilitação para várias doenças, incluindo o Acidente Vascular Cerebral (AVC). Embora alguns estudos tenham mostrado a sua eficácia clínica, pouco se sabe sobre seus mecanismos neurais. Baseado nisso, este estudo teve como objetivo avaliar por meio da Ressonância Magnética funcional (fMRI) e da Estimulação Magnética Transcraniana (TMS) a neuromodulação cortical promovida pela intervenção única da Terapia Espelho em pacientes acometidos por Acidente Vascular Cerebral. Quinze pacientes participaram de sessão única de trinta minutos de TE. Os dados de fMRI foram analisados bilateralmente nas seguintes regiões de interesse (ROI): Área Motora Suplementar (AMS), córtex pré-motor (PM), córtex motor primário (M1), córtex sensorial primário (S1) e Cerebelo. Em cada ROI, as mudanças na porcentagem de ocupação e os valores de beta foram avaliados. No TMS foi analisado o Potencial Evocado Motor (PEM) sobre o hot spot M1. Um aumento significativo na amplitude do PEM foi observado após a terapia no grupo (p<0,0001) e em 4 pacientes (p <0,05). Nos resultados da fMRI houve uma redução significativano percentual deocupação noPMe cerebelocontralateralàmão afetada(p <0,05). Além disso, foi observado aumento significativonos valores debetanas seguintes áreasmotoras contralaterais: AMS, Cerebelo,PM eM1(p <0,005) e diminuiçãosignificativa nas áreasmotoras ipsilaterais: PM e M1 (p < 0,001).Nas áreas sensoriais foi observada redução em S1bilateralmente(p <0,0005). Assim, nossos resultados indicam que intervençãoúnica de TE mudamarcadoresneurobiológicosem direção aopadrão observadoem indivíduos saudáveis.Além disso, as alterações nas áreas motoras do hemisfério contralateralsãoopostas asdo ladoipsilateral, sugerindo um aumento na homeostase do sistema.

Crises epilépticas são desordens paroxísticas do sistema nervoso central (SNC) caracterizadas por uma descarga elétrica neuronal anormal, com ou sem perda de consciência e com sintomas clínicos variados. Nas epilepsias do lobo temporal as crises tem início focal, em estruturas do sistema límbico. Dados clínicos e experimentais mostram que essas regiões apresentam morte neuronal (esclerose hipocampal), reorganização sináptica (brotamento aberrante das fibras musgosas) e gliose reativa, sendo esses marcadores biológicos da zona epileptogênica. Registros extracelulares mostram que além das alterações anatômicas mencionadas acima, a zona epileptogênica também apresenta oscilações de alta frequência patológicas (pOAF). As pOAF são oscilações transientes (50 – 100 ms de duração), de baixa amplitude (200 µV - 1.5 mV) e de frequências variáveis (80 – 800 Hz). A relação entre essas oscilações e a gênese das crises espontâneas ainda é desconhecida. O objetivo do presente trabalho foi avaliar os efeitos da estimulação elétrica intracerebral (EIC) nas pOAF e frequência de crises espontâneas de animais cronicamente epilépticos (modelo da epilepsia do lobo temporal). Atualmente, a EIC é utilizada no tratamento de distúrbios do movimento (e.g., mal de Parkinson) e em alguns casos de dor crônica, e experimentalmente, no tratamento das epilepsias de difícil controle. A hipótese de trabalho dessa dissertação é de que a indução de depressão de longa duração por EIC, ao reduzir a excitabilidade neuronal local, modulará as pOAF, bem como a frequência de crises espontâneas. Para isso, comparamos as características espectrais das pOAF e a frequência de crises espontâneas antes e depois de um protocolo de 12 horas de estimulação elétrica de baixa frequência (0,2 Hz) aplicado na via perforante. De fato, esse protocolo reduziu a amplitude do potencial de ação coletivo registrado no giro denteado (GD) do hipocampo dorsal em 45% (amplitude média da primeira e da última hora de estimulação: 7,3 ± 3,0 mV e 4,1 ± 1,5 mV, respectivamente; p<0,05; teste T). O monitoramento contínuo do potencial de campo local, realizado no GD e em CA3 simultaneamente, mostrou que o protocolo de estimulação empregado foi eficaz em (i) aumentar a duração (64,6 ± 9,3 ms vs. 70,5 ± 11,5 ms) e reduzir (ii) a entropia (3,72 ± 0,28 vs. 3,58 ± 0,30), (iii) o índice pOAF (0,20 ± 0,08 vs. 0,15 ± 0,07) e (iv) o modo espectral (237,5 ± 15,8 Hz vs. 228,7 ± 15,2 Hz) das pOAF (valores do GD, expressos como média ± desvio-padrão, para os períodos “pré” e “pós” estimulação respectivamente; p<0,05; teste T). Ainda, este protocolo reduziu significativamente a frequência de crises espontâneas (1,8 ± 0,4 vs. 1,0 ± 0,3 crises/hora; “pré” e “pós” estimulação, respectivamente; p<0,05; teste T). Curiosamente, observamos um aumento na duração média das crises espontâneas após o término do protocolo (39,7 ± 6,0 vs. 51,6 ± 12,5 s; “pré” e “pós” estimulação respectivamente; p<0,05; teste T). Estes resultados sugerem que a redução da excitabilidade neuronal, por meio de protocolos de estimulação elétrica, reorganiza o perfil espectral das pOAF. Esse efeito foi acompanhado de redução na frequência de crises espontâneas. Apesar de preliminar, o presente trabalho contribui para o desenvolvimento de terapias baseadas em EIC para as epilepsias.