Avaliação dos efeitos do extrato de Passiflora cincinnata Masters em camundongos: efeito na ansiedade e potencial neuroprotetor

Anxiety disorders and Parkinson’s disease (PD) affect a large portion of the world population. Indeed, therapeutic alternatives available do not contribute to improve most clinical conditions and/or are linked with undesirable side effects. Thus, there is a great demand for the development of new drugs to treatment of these diseases. Passiflora cincinnata Mast. is a native species present in several Brazilian states, popularly known as “maracujá do mato”, “maracujá tubarão” or “maracujá mochila”. Additionally, species of Passiflora genus are traditionally known for their exotic flowers, edible fruits with pronounced flavor and for their sedative, tranquilizer and anxiolytic properties reported by folk medicine. These plants possess important organic compounds such as phenols, cyanogenic glycosides, flavonoids and alkaloids, which are responsible for the anxiolytic, antioxidant, anti-inflammatory, antihyperglycemic, among others activities when tested in mammals. Despite this fact, only a few studies have been conducted to investigate the possible in vivo biological effects of Passiflora cincinnata Mast extracts. Thereby, in this study we evaluated the effects of the alcoholic extract of this plant in anxiety and PD animal model. Mice acutely or chronically administered with ethanolic extract of P. cincinnata do not showed any anxiogenic- or anxyolitic-like effect in elevated plus maze (EPM). In order to reproduce PD symptom’s in mice, we administered repeated injections of reserpine which progressively induced motor impairments such as increase in catalepsy, oral movements, and reduction of the average speed of the animals in the open field, as well as depleted dopamine prodution in SNpc cells. Furthermore, this treatment resulted in the loss of aversive memory recall in mice when undergoing PMDAT. Yet, passiflora group also show this amnesic profile. However, animals treated concomitantly with the alcoholic extract of Passiflora cincinnata Mast. showed higher latency for the onset of motor impairment evaluated by catalepsy. Thus, our results shows that the alcoholic extract of the plant P. cincinnata was able to delay the onset of the catalepsy induced by reserpine administration, plus reverted the depletion of dopamine production in SNpc cells.

Caracterização citoarquitetônica e por imunoistoquímica para tirosina-hidroxilase da substância negra, área tegmentar ventral e zona retrorubral do Sagui (Callithrix jacchus)

It is known that the catecholamine group is constituted by dopamine, noradrenaline and adrenaline, in which the synthesis is regulated by an enzyme named tyrosine hydroxylase. Thus, 3-hydroxytyramine/dopamine (DA) is a precursor of the noradrenaline and adrenaline synthesis and acts as a neurotransmitter in the central nervous system. The three main nuclei, named the retrorubral field (A8 group), the substantia nigra pars compacta (A9 group) and the ventral tegmental area (A10 group), are arranged in the die-mesencephalic portion and are involved in three complexes circuitries - the mesostriatal, mesolimbic and mesocortical pathways. These pathways are related to behavioral manifestations, motricity, learning, reward and pathologies such as Parkinson’s Disease and Schizophrenia. Thus, the aim of this study was to perform de morphological analysis of the A8, A9 and A10 nuclei of the common marmoset (Callithrix jacchus). The marmoset is a neotropical primate, whose morphological and functional characteristics supports the suitability of use of this animal in biomedical research. Coronal sections of the marmoset brain were submitted to cytoarchitectonic characterization and TH-immunohistochemistry. Based on the morphology of the neurons, it was possible to subdivide the A10 group in seven regions: interfascicular nucleus, raphe rostral linear nucleus and raphe caudal linear nucleus, in the middle line; paranigral and parainterfascicular nucleus, in the middle zone; rostral portion of the ventral tegmental area nucleus and parabrachial pigmented nucleus, located in the dorsolateral portion of the mesencephalic tegmentum. A9 group was divided into four regions: substantia nigra compacta dorsal and ventral tiers; substantia nigra compacta lateral and medial clusters. No subdivisions were founded into A8 group. These results revealed that A8, A9 and A10 are phylogenetically conserved between species, but it’s necessary to expand the studies about this compartmentalization, investigating its occurrence in other primate species or investigating its functional relevance.

Citoarquitetura e imunoistoquímica para tirosinahidroxilase dos núcleos dopaminérgicos mesencefálicos do morcego (Artibeus planirostris)

The 3-Hidroxytyramine/dopamine (DA) is a monoamine of catecholamine family and isthe precursor substance synthesis of noradrenaline and adrenaline, having the enzymeTyrosine Hydroxylase (TH) as this regulatory process. In addition, the DA has theability to act as a neurotransmitter in the Central Nervous System - SNC, being themain neurotransmitter of brain nuclei, namely of A8 to A16. The nuclei of the midbrainthat express DA are the Retrorubral Field (RRF, A8), the Substantia Nigra parsCompacta (SNc, A9) and the Ventral Tegmental Area (VTA, A10). Such nuclei areinvolved in complex three circuitry that are the mesostriatal, mesolimbic andmesocotical and are directly related with several behavioral manifestations as motricitycontrol, reward signaling in behavioral learning, motivation and pathologicalconditions, such as Parkinson's Disease and schizophrenia. Interestingly, many of themorphological bases of these neural disturbance remain unknown. Considering therelevance of mesencephalic dopaminergic nuclei, the aim of this research is tocharacterize morphologically the dopaminergic nuclei (clusters A8, A9 and A10) of themidbrain of the bat (Artibeus planirostris). The Artibeus planirostris is a common bat inRio Grande do Norte. Ten animals were used in this research. The animals wereanesthetized, perfused, and the brain was removed from the skull. After dehydration insucrose, the brain was underwent microtomy. Saggital and coronal sections wereobtained and collected in six separate series. The series were Nissl-stained to identifythe cytoarchitectonic boundaries and the other series were subjected toimmunohistochemistry for TH. After cytoarchitectonic analysis and TH+ cellsidentification was possible to establish the anatomical boundaries of the nuclei, as wellas the subdivisions of three of the midbrain dopaminergic nuclei. The SNc is the mostrostral nucleus observed in the midbrain and is identified throughout the rostrocaudalextension of the midbrain. The VTA neurons were seen immediately caudal to the SNcappearance. The RRF neurons were observed just in the caudal levels of the midbrain.The SNc in Artibeus planirostris shows a particular feature, the tail of the SNc. The tailhave been described just in two other studied species. The present work shows aparticular variation in the organizational morphology of the SNc in the artibeus andcontribute to understand the phylogenetic routes by which the dopaminergic system hasevolved.

Avaliação de duas variedades de Passiflora edulis sobre as respostas comportamentais de camundongos e um possível mecanismo de ação

The medicinal plants constitute a rich source of biologically active compounds used for the treatment of many psychiatric disorders, such as anxiety disorders and depression. Generalized anxiety disorder has increased significantly, being the second most prevalent disorder in care facilities to public health. Depression is considered a chronic and common psychiatric disorder that affects 350 million people of all ages around the world. In this context, the pharmacological intervention conduits have been employed, effective, although leave to be desired when observed adverse effects. The genus Passiflora is commonly commercially known by its fruit, but is also widely used in traditional Brazilian medicine. Passiflora edulis displays considerable morphological variability. This plant produces two types of fruit: Purple (Passiflora edulis Sims fo. edulis) and yellow (Passiflora edulis fo. flavicarpa Degener). This study investigated the central effects of aqueous extract of the leaves of the two varieties of the species Passiflora edulis in tests used to assess behavior related to anxiety and depression, as well as investigating the potential effect of the antidepressant-like fractions of edulis fo. edulis and neuropharmacological mechanisms responsible for this action. To conduct this study used male Swiss mice (2 months old, weighing 30-35 g). The animals received the aqueous extract of the leaves of the two species of Passiflora: edulis fo. edulis (100, 300, 1000 mg / kg) and fractions ethyl acetate, butanol and aqueous waste (25, 50, 75, 100 mg / kg) and edulis fo. flavicarpa (30, 100, 300, 1000 mg / kg) or saline by gavage 60 minutes prior to the maze tests at high cross the open field test, test forced swim test and sedation induced by thiopental. To investigate the mechanism of action of the activity of antidepressant type of fractions the following drugs were used: PCPA (inhibitor of 5-HT synthesis) AMPT (inhibitor of catecholamine synthesis), DSP-4 (noradrenergic neurotoxin) and Sulpiride (antagonist selective dopamine D2 receptor). They were used as a standard positive control, fluoxetine and nortriptyline. The results of the phytochemical profile show very different characteristics to the aqueous extract of the varieties of Passiflora edulis "flavicarpa" and "edulis". The aqueous extracts of both varieties of Passiflora edulis share anxiolytic activity type (edulis fo. edulis 300 mg/kg; edulis fo. flavicarpa 300 and 1000 mg/kg) and antidepressant (edulis fo. edulis 300 mg/kg; edulis fo flavicarpa 1000 mg/kg), while the effect hipolocomotor/sedative was only seen for edulis fo. edulis (1000 mg/kg). Both fractions ethyl acetate, butanol aqueous extract edulis fo. edulis showed activity type antidepressant at a dose of 50 mg/kg in the forced swim test. The data suggest that the effect of antidepressant-like fractions edulis fo. edulis involves catecholaminergic and serotonergic neurotransmission, particularly dopaminergic, there is seen that pre-treatment DSP-4 is not affected antidepressant action of fractions as was dependent activation of dopamine D2 receptors.

Avaliação de duas variedades de Passiflora edulis sobre as respostas comportamentais de camundongos e um possível mecanismo de ação

The medicinal plants constitute a rich source of biologically active compounds used for the treatment of many psychiatric disorders, such as anxiety disorders and depression. Generalized anxiety disorder has increased significantly, being the second most prevalent disorder in care facilities to public health. Depression is considered a chronic and common psychiatric disorder that affects 350 million people of all ages around the world. In this context, the pharmacological intervention conduits have been employed, effective, although leave to be desired when observed adverse effects. The genus Passiflora is commonly commercially known by its fruit, but is also widely used in traditional Brazilian medicine. Passiflora edulis displays considerable morphological variability. This plant produces two types of fruit: Purple (Passiflora edulis Sims fo. edulis) and yellow (Passiflora edulis fo. flavicarpa Degener). This study investigated the central effects of aqueous extract of the leaves of the two varieties of the species Passiflora edulis in tests used to assess behavior related to anxiety and depression, as well as investigating the potential effect of the antidepressant-like fractions of edulis fo. edulis and neuropharmacological mechanisms responsible for this action. To conduct this study used male Swiss mice (2 months old, weighing 30-35 g). The animals received the aqueous extract of the leaves of the two species of Passiflora: edulis fo. edulis (100, 300, 1000 mg / kg) and fractions ethyl acetate, butanol and aqueous waste (25, 50, 75, 100 mg / kg) and edulis fo. flavicarpa (30, 100, 300, 1000 mg / kg) or saline by gavage 60 minutes prior to the maze tests at high cross the open field test, test forced swim test and sedation induced by thiopental. To investigate the mechanism of action of the activity of antidepressant type of fractions the following drugs were used: PCPA (inhibitor of 5-HT synthesis) AMPT (inhibitor of catecholamine synthesis), DSP-4 (noradrenergic neurotoxin) and Sulpiride (antagonist selective dopamine D2 receptor). They were used as a standard positive control, fluoxetine and nortriptyline. The results of the phytochemical profile show very different characteristics to the aqueous extract of the varieties of Passiflora edulis "flavicarpa" and "edulis". The aqueous extracts of both varieties of Passiflora edulis share anxiolytic activity type (edulis fo. edulis 300 mg/kg; edulis fo. flavicarpa 300 and 1000 mg/kg) and antidepressant (edulis fo. edulis 300 mg/kg; edulis fo flavicarpa 1000 mg/kg), while the effect hipolocomotor/sedative was only seen for edulis fo. edulis (1000 mg/kg). Both fractions ethyl acetate, butanol aqueous extract edulis fo. edulis showed activity type antidepressant at a dose of 50 mg/kg in the forced swim test. The data suggest that the effect of antidepressant-like fractions edulis fo. edulis involves catecholaminergic and serotonergic neurotransmission, particularly dopaminergic, there is seen that pre-treatment DSP-4 is not affected antidepressant action of fractions as was dependent activation of dopamine D2 receptors.

Parvalbumin-positive interneurons of the prefrontal cortex support working memory and cognitive flexibility

We were supported by the Biotechnology and Biological Sciences Research Council grant BB/H001123/1 (P.W.), the Medical Research Council grants G0601498 and G1100546/2 (P.W.), Tenovus Scotland Grant G09/17 (A.J.M.) and the University of Aberdeen (P.W.). We thank O. Tüscher for discussion, P. Teismann and the microscopy core facility at the University of Aberdeen for the use of microscopy equipment, L. Strachan, A. Plano, S. Deiana for help with behavioral testing.

Caractérisation de l’activité Nur77 et de ses complexes en essais BRET par complémentation

Dans le système nerveux central, la dopamine joue un rôle crucial dans de nombreuses fonctions physiologiques telles que : l’apprentissage, le mouvement volontaire, la motivation, la cognition et la production hormonale. Il a été aussi démontré que le système de signalisation dopaminergique est altéré dans plusieurs maladies neurologiques et psychiatriques comme la maladie de Parkinson et la schizophrénie. Des études, effectuées dans le laboratoire du Dr.Daniel Lévesque (laboratoire d’accueil), ont montré que les récepteurs nucléaires Nur77 (NR4A1, NGFI-B) et RXRγ (retinoid X receptors γ) sont impliqués dans la régulation des effets de la dopamine dans le système nerveux central. De plus, ces données suggèrent que le complexe Nur77 et RXR joueraient un rôle crucial dans l’effet des médicaments antipsychotiques et antiparkinsoniens. Toutefois, très peu de médicaments ciblant Nur77 ont été identifiés à ce jour et les médicaments agissant sur RXRγ restent mal caractérisés. En outre, les analyses actuellement disponibles ne peuvent pas résumer la complexité des activités des NRs et génèrent des mesures indirectes des activités des drogues. Afin de mieux comprendre comment est régulée l’interaction Nur77/RXRγ dans ces processus, mon projet a été de mettre au point un essai BRET (Bioluminescence Resonance Energy Transfer) et PCA-BRET (Protein Complementation Assay-BRET) basé sur le recrutement d'un motif mimant un co-activateur fusionné avec la YFP. Nos différents essais ont été validés par courbes dose-réponse en utilisant différents composés RXR . Les EC50 (concentration efficace médiane, qui permet de mesurer l'efficacité d'un composé) obtenues étaient très semblables aux valeurs précédemment rapportées dans la littérature. Nous avons aussi pu identifier un composé le SR11237 (BMS649) qui semble posséder une sélectivité pour le complexe Nur77/RXRγ par rapport aux complexes Nurr1/RXRγ et RXRγ /RXRγ. Nos résultats indiquent que ces essais de BRET peuvent être utilisés pour évaluer la sélectivité de nouveaux composés pour les complexes Nur77/RXRγ, Nurr1/RXRγ et RXRγ /RXRγ. Un autre aspect de mon projet de doctorat a été de mettre en évidence par BRET l’importance de la SUMOylation dans la régulation de l'activité de Nur77 dans sa forme monomèrique, homodimèrique et hétérodimèrique. Nous avons ainsi identifié que Nur77 recrute principalement SUMO2 sur sa lysine 577. Il est intéressant de noté que le recrutement de la SUMO2 à Nur77 est potentialisé en présence de la SUMO E3 Ligase PIASγ. Aussi, la perte de la SUMOylation sur la lysine 577 entraîne l'incapacité de Nur77 de recruter divers motifs de co-activation mais pas pour ses formes homo- et hétérodimèrique. Cependant, la présence de PIASγ ne potentialise pas le recrutement du co-activateur, suggérant que cette SUMO E3 Ligase est seulement impliqué dans le processus de recrutement de la SUMO mais pas dans celui du co-activateur. Nous avons ainsi déterminé une nouvelle modification post-traductionnelle sur Nur77 régulant spécifiquement son activité monomérique Ces projets pourraient donc apporter de nouvelles données cruciales pour l’amélioration du traitement de la maladie de Parkinson ou de la schizophrénie, ainsi que d'obtenir une meilleure compréhension sur les mécanismes permettant la régulation de la fonction de Nur77

The role of the homeobox transcription factor Pitx3 in the mesencephalic dopaminergic system

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Impact des facteurs génétiques et cliniques sur la réponse au traitement par méthylphénidate chez des enfants avec un trouble déficitaire de l'attention/hyperactivité

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Études des différences sexuelles dans les systèmes cérébraux régulateurs de la réponse de stress et des émotions chez le rat

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

L'effet de la quétiapine sur le phénomène de récompense

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Impact de l'expression du bFGF par les astrocytes du mésencéphale sur le développement et la fonction synaptique des neurones dopaminergiques

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

Régulation de la fonction du récepteur dopaminergique D2 dans les neurones dopaminergiques

Thèse numérisée par la Direction des bibliothèques de l'Université de Montréal.

Expectation Modulates Episodic Memory Formation via Dopaminergic Circuitry

Episodic memory formation is shaped by expectation. Events that generate expectations have the capacity to influence memory. Additionally, whether subsequent events meet or violate expectations has consequences for memory. However, clarification is still required to illuminate the circumstances and direction of memory modulation. In the brain, the mechanisms by which expectation modulates memory formation also require consideration. The dopamine system has been implicated in signaling events associated with different states of expectancy; it has also been shown to modulate episodic memory formation in the hippocampus. Thus, the studies included in this dissertation utilized both functional magnetic resonance imaging (fMRI) and behavioral testing to examine when and how the dopaminergic system supports the modulation of memory by expectation. The work aimed to characterize the activation of dopaminergic circuitry in response to cues that generate expectancy, during periods of anticipation, and in response to outcomes that resolve expectancy. The studies also examined how each of these event types influenced episodic memory formation. The present findings demonstrated that novelty and expectancy violation both drive dopaminergic circuitry capable of contributing to memory formation. Consistent with elevated dopaminergic midbrain and hippocampus activation for each, expected versus expectancy violating novelty did not differentially affect memory success. We also showed that high curiosity expectancy states drive memory formation. This was supported by activation in dopaminergic circuitry that was greater for subsequently remembered information only in the high curiosity state. Finally, we showed that cues that generate high expected reward value versus high reward uncertainty differentially modulate memory formation during reward anticipation. This behavioral result was consistent with distinct temporal profiles of dopaminergic action having differential downstream effects on episodic memory formation. Integrating the present studies with previous research suggests that dopaminergic circuitry signals events that are unpredicted, whether cuing or resolving expectations. It also suggests that contextual differences change the contribution of the dopaminergic system during anticipation, depending on the nature of the expectation. And finally, this work is consistent with a model in which dopamine elevation in response to expectancy events positively modulates episodic memory formation.

Investigating the developmental and behavioral consequences of maternal immune activation on affected offspring

Maternal infection during pregnancy increases the risk of several neuropsychiatric disorders later in life, many of which have a component of dopaminergic (DA) dysfunction, including schizophrenia, autism spectrum disorders (ASD), and attention deficit hyperactivity disorder (ADHD). The majority of DA neurons are found in the adult midbrain; as such the midbrain is a key region of interest regarding these disorders. The literature is conflicting regarding the behavioral alterations following maternal immune activation (MIA) exposure, and the cellular and molecular consequences of MIA on the developing midbrain remain to be fully elucidated. Thus, this thesis aimed to establish the consequences of acute and mild MIA on offspring dopamine-related behaviors, as well as the associated cellular and molecular disturbances of MIA on offspring midbrains. We utilized a rat model of MIA using low dose (50μg/kg, I.P.) of LPS administered at different gestational ages. Our first study indicated that MIA at later gestational ages significantly increased pro-inflammatory IL-1β expression, and reduced HSD11B2 expression in the placenta, which is an important regulator of fetal development. In utero LPS exposure at later gestational ages also impaired the growth of neurons from affected offspring. This study identified key gestational stages during which MIA resulted in differential effects. We utilized these time points in subsequent studies, the next of which investigated neurobehavioral outcomes following MIA. Our results from that study showed that motor differences occurred in juvenile offspring following MIA at E16 only, and these differences were compensated for in adolescence. Then, there was a decline in motor behavior capabilities in adulthood, again only for animals exposed to MIA on E16 (and not E12). Furthermore, our results also demonstrated adolescent and adult offspring that were exposed to MIA at E12 had diminished responses to amphetamine in reward seeking behaviors. In our final study, we aimed to investigate the molecular and cellular changes following MIA which might explain these behavioral alterations. This final study showed a differential inflammatory response in fetal midbrains depending on gestational age of exposure as well as differential developmental alterations. For example, LPS exposure at E16 resulted in decreased VM neurosphere size after 7DIV and this was associated with an increased susceptibility to neurotoxic effects of pro-inflammatory cytokines for VM neurospheres and VM DA neurons treated in culture. In utero LPS exposure at E16 also reduced DA neuron count of fetal VM, measured by TH staining. However, there were no differences in DA neuron number in juvenile, adolescent, or adult offspring. Similarly, LPS exposure did not alter cell number or morphology of glial cells in the midbrains of affected offspring. In conclusion, this thesis indicated later rat pregnancy (E16) as vulnerable time for MIA to affect the development of the nigrostriatal pathway and subsequent behavioral outcomes, possibly implicating a role for MIA in increased risk for disorders associated with motor behavior, like PD. These effects may be mediated through alterations in the placenta and altered inflammatory mediators in the offspring brain. This thesis has also shown that MIA in earlier rat pregnancy (E12) results in altered mesocorticolimbic function, and in particular MIA on E12 resulted in a differential response to amphetamine in affected offspring, which may implicate a role for MIA in increasing the risk for disorders associated with this pathway, including drug tolerance and addiction.