Résumés / Abstracts 2021

Développement de techniques d'analyse quantitatives des concentrations ioniques neuronales

INTRO : Des débalancements anormaux et néfastes de l’homéostasie ionique neuronale, au niveau du calcium et du chlore notamment, ont été rapportés dans des modèles cellulaires ou animaux de troubles psychiatriques. Chez l’adulte, le calcium joue un rôle d’indicateur de l’activité neuronale alors que le chlore en joue un rôle plutôt inhibiteur; le rôle de cet équilibre entre les deux concentrations ioniques est donc un sujet d’intérêt dans le développement de troubles psychiatriques.

MÉTHODE : L’utilisation d’indicateurs ioniques fluorescents tels que Twitch-2B (pour le calcium) et SuperClomeleon (pour le chlore) permet de suivre les concentrations ioniques correspondantes dans des cultures de cellules neuronales. La précision des techniques d’analyse à base d’intensité de fluorescence, de temps de vie de fluorescence (Fluorescence Lifetime Imaging Microscopy, FLIM) via la régression multiexponentielle et l’utilisation de plans phaseurs pour analyser des données FLIM seront comparées. Les paramètres de comparaison seront le nombre de photons détectés, la séparation des temps de vie de fluorescence dans le cas des techniques à base de FLIM, ainsi que de la précision de l’obtention de concentrations ioniques lorsque deux indicateurs ioniques sont présents dans le même échantillon.

RÉSULTATS : Les modèles mathématiques associés aux plans phaseurs ont été validés sur un simulateur conçu pour recréer des données FLIM, puis comparés à la technique de régression multiexponentielle (algorithme de Levenberg-Marquardt). Les modèles ont ensuite été utilisés sur des échantillons in-vitro de cultures de cellules HEK-293, puis de cellules neuronales d’hippocampe de souris et comparés à des techniques d’analyse ratiométriques d’intensité de fluorescence.

CONCLUSION : Les résultats obtenus sur simulateur ont montré que la précision d’obtention de temps de vie de fluoresence, qui peuvent ensuite être liés à des concentrations ioniques, est comparable entre les méthodes de plans phaseurs et les méthodes de régression multiexponentielles. Lorsque deux fluorophores sont présents dans le même échantillon, les résultats sont plus précis pour le modèle des plans phaseurs que pour les régressions multiexponentielles.

Propriétés des neurones glutamatergiques du noyau gigantocellulaire

Bien que les neurones glutamatergiques de la formation réticulée médullaire contribuent au contrôle moteur et locomoteur à travers la voie réticulospinale, leurs propriétés intrinsèques, et leur connectivité dans le circuit neuronal restent méconnues. Nous avons émis l’hypothèse que les neurones glutamatergique réticulospinaux sont organisés topographiquement dans le noyau gigantocellulaire réticulaire (Gi) et qu’ils possédent des propriétés électrophysiologiques spécifiques leur permettant d’activer le circuit locomoteur spinal.

OBJECTIF - OBJECTIVE

Évaluer l’organisation topographique et les propriétés électrophysiologiques des neurones glutamatergique réticulospinaux selon leur projection spinale.

Étudier la connectivité entre les neurones glutamatergiques réticulospinaux cervicaux et lombaires ainsi que leur connectivité dans la formation réticulée médullaire.

MÉTHODES - METHODS

Des souris VGluT2-CRE âgées entre P20-P30 ont été injectées dans le segment lombaire de la moelle épinière puis dans le segment cervical avec deux virus adéno-associé rétrograde munis de protéines fluorescentes différentes (AAV-DIO-Mcherry et AAVretro-DIO-GFP ou AAVretro-DIO-ChR2-YFP) afin d’identifier chaque population neuronale. Une à trois semaines après la dernière injection, les troncs cérébraux sont collectés et coupés en tranches pour nos études neuroanatomiques et électrophysiologiques lors d’enregistrements de type patch-clamp.

RÉSULTATS - RESULTS

Nos études neuroanatomiques ont révélé que les neurones glutamatergiques réticulospinaux cervicaux ou lombaires sont distribués à travers le Gi. Nos enregistrements électrophysiologiques ont aussi révélé l’existence de 4 types de patrons de décharges incluant ou non la présence d’un rebond post-inhibiteur avec ou sans potentiel d’action et une diminution du voltage (sag) en réponse à une hyperpolarization membranaire. Par ailleurs, les neurones présentant un rebond avec potentiel d’action et une sag sont localisés préférentiellement dans la portion alpha et ventrale du Gi et incluaient une grande proportion de neurones réticulospinaux projettant à la fois au niveau cervical et lombaire. À l’aide d’outils optogénétique, nous étudions présentement la connectivité locale de ces neurones.

CONCLUSION

En conclusion, nos études révèlent que les neurones glutamatergique reticulospinaux projettant à la fois au niveau cervical et lombaire sont localisés préférentiellement dans la portion ventrale du Gi et présentent des propriétés électrophysiologiques uniques contribuant à la commande motrice descendante.  

Gaming disorders among psychotic patients: A scoping review

OBJECTIF - OBJECTIVE

There is a growing interest in understanding the impact of video games in the clinical field given that their excessive use is associated with health problems like depression, anxiety, and even suicide and can lead to severe addiction. To this day, little is known about the consequences of gaming disorder in vulnerable populations. Clinicians widely recognize the comorbidity of gaming and psychotic disorders, but it seems that the scientific community has yet to complete further research in their joint mechanisms.The aim of this scoping review is to understand the interactions between gaming and psychotic disorders and identify the knowledge gaps.  

MÉTHODES - METHODS

We used Levac’s six-stage methodology for scoping review. 212 articles from 7 databases were identified. Nine articles respected our inclusion and exclusion criteria. 

RÉSULTATS - RESULTS

No available study has assessed the prevalence or incidence of gaming disorder among psychotic patients. The cases reported highlight the possibility that excessive video game play or an abrupt gaming disruption could trigger psychosis in patients without any personal or family history of psychotic disorders.  

CONCLUSION

Our results show a significant lack of knowledge concerning psychotic disorders associated with gaming disorders and only a few reported cases and a single study exposed the direct association between those conditions.  

Rôle des protéines Nck dans le développement du circuit locomoteur

OBJECTIF - OBJECTIVE

Les protéines Nck1 et Nck2 sont des protéines adaptatrices impliquées dans le développement embryonnaire du circuit neuronal. Les souris mutées pour Nck1 et Nck2 ne marchent pas, mais sautent. L’objectif est de vérifier si les protéines Nck affectent le contrôle locomoteur de manière différente selon le degré de mutation de Nck1 et Nck2.

MÉTHODES - METHODS

Nous avons enregistré par vidéo la marche de souris contrôles et mutantes sur un tapis roulant à une vitesse constante de 15, 20 et 30 cm/s.  La technologie Cre-Lox a été utilisée afin de générer divers mutants présentant des délétions conditionnelles de Nck1 et/ou Nck2 sous le contrôle de différents promoteurs spécifiques : Nestin, Lhx3, En1 et Dbx1. Les vidéos ont été analysées afin d’étudier le patron locomoteur ainsi que le couplage bilatéral entre les pattes gauches et droites, le couplage homolatéral entre les pattes avant et arrière, et également le couplage diagonal entre la patte avant et la patte postérieure opposée.

RÉSULTATS - RESULTS

Comparativement aux souris sauvages qui ont une démarche exclusivement en alternance, les souris mutantes pour Nck dans les cellules neuronales et gliales (Nck1-/-;Nck2ff::Nes) ou dans les cellules commissurales V0 (Nck1-/-;Nck2ff::Dbx1) présentaient un déphasage du couplage bilatéral, homolatéral et diagonal. De plus, les souris mutantes pour Nck1 ou Nck2, ainsi que les souris mutantes pour Nck dans les populations exprimant V1 (Nck1-/-;Nck2ff::En1) et V2 (Nck1-/-;Nck2ff::Lhx3) présentaient un déphasage dans le couplage homolatéral et diagonal entre les pattes antérieur et postérieur.

CONCLUSION

Nos résultats supportent l’importance de Nck1 et Nck2 dans le développement du circuit locomoteur et dans la coordination bilatérale gauche-droite. De futures expériences in vitro utilisant des moelles épinières isolées de souris néonatales nous permettrons de déterminer si les changements observés sont supraspinaux ou spinaux.

Changements neuroanatomiques de la matière blanche et grise 24 mois après la chirurgie bariatrique

Notre étude récente a montré une récupération de la densité des matières blanche et grise 4 et 12 mois post-chirurgie bariatrique, suggérant une plasticité des structures du cerveau suivant une perte de poids et une amélioration des altérations métaboliques. Il est incertain si ces changements persistent à plus long terme.

OBJECTIF 

Caractériser les changements de matières blanche et grise observés 24 mois après une chirurgie bariatrique.

MÉTHODES

84 participants (âge = 49,8±6,5 ans, IMC = 44,0±4,1 kg/m2) ayant subi une gastrectomie pariétale, une diversion biliopancréatique avec commutation duodénale ou une dérivation gastrique Roux-en-Y ont été recrutés. Des images anatomiques pondérées en T1 ont été acquises avant, 4, 12 et 24 mois post-chirurgie. Les densités de matières blanche et grise ont été quantifiées par morphométrie cérébrale. Un modèle linéaire à effets mixtes a été utilisé pour comparer les changements neuroanatomiques pré- et post- chirurgie en contrôlant pour l’âge, le sexe, l’indice de masse corporelle initial, le type de chirurgie et le statut diabétique.

RÉSULTATS

La perte de poids totale moyenne à 24 mois était de 33,5±7,2%. Une augmentation étendue de la matière blanche a été observée dans le cervelet, le tronc cérébral et le corps calleux, et une augmentation étendue de la matière grise a été observée dans le cortex occipital, temporal, le gyrus post central, le cervelet, l’hippocampe, l’insula, le gyrus fusiforme et l’amygdale (p<0,05, FDR). Ces augmentations étaient similaires à celles observées 12 mois post-chirurgie.

CONCLUSION

Les augmentations étendues de la densité des matières blanche et grise observées 4 et 12 mois post-chirurgie sont encore présentes après 24 mois. Les prochaines étapes sont d’examiner les prédicteurs neuronaux de la perte de poids à 24 mois et les mécanismes potentiels expliquant ces augmentations de densité post-chirurgie.

Étude du rôle des systèmes angiotensinergiques périphérique et central dans la vulnérabilité au trauma psychologique chez la souris

PROBLÉMATIQUE: Le trouble de stress post-traumatique (TSPT) est associé à une augmentation du risque de développer de l’hypertension. Ces deux maladies sont aussi associées à un niveau élevé du statut inflammatoire chez les individus. Cependant, la relation entre les deux maladies et les mécanismes sous-jacents au rôle de l’inflammation demeurent incompris. L’hypothèse était qu’un trauma psychologique activerait les récepteurs angiotensinergiques de type 1 (AT1aR) endothéliaux, perturbant ainsi la perméabilité de la barrière hémato-encéphalique (BHE) et facilitant le passage des cytokines inflammatoires au cerveau. OBJECTIFS: L’objectif est de déterminer l’effet de l’activation de l’AT1aR endothélial sur la pression sanguine et le phénotype de type post-traumatique dans un modèle murin de TSPT. MÉTHODOLOGIE: Une lignée de souris surexprimant le récepteur AT1aR (ehM3Dq) sous traitement à la clozapine N-oxide (CNO; 0.05 mg/kg p.o.) a été exposée à un stress de prédation (chat; 10 min). La pression sanguine a été mesurée 24h après le stress. Le phénotype de type post-traumatique a été évalué via l’open field (OF), le light-dark box (LD) et le trauma reminder (TR), mesurant l’anxiété généralisée et l’évitement d’un stimulus rappelant le trauma. RÉSULTATS: Les souris ehM3Dq ont une hausse significative de leur pression sanguine moyenne relativement aux souris contrôles 24h après le stress (p < 0.05). Dans l’OF, une diminution du phénotype anxieux a été observée chez les souris ehM3Dq stressées comparativement aux souris contrôles stressées (p < 0.01). Aucune différence n’a été observée entre les souris ehM3Dq et contrôles en réponse au stress dans le LD et le TR. CONCLUSION: Ces résultats montrent que la surexpression du récepteur AT1aR est associée à une hypertension en réponse au stress, mais à une diminution de certains phénotypes anxieux. Des cohortes supplémentaires nous permettront de déterminer les effets sexe-dépendants, l’inflammation et l’intégrité de la BHE.

Sex-specific retinal anomalies induced by chronic social defeat stress in mice

OBJECTIVE: Major depressive disorder (MDD) is one of the most common consequences of chronic stress. Still, there is currently no reliable biomarker to detect individuals at risk to develop the disease. Recently, the retina emerged as an effective way to investigate psychiatric disorders using the electroretinogram (ERG). In this study, we assessed the suitability of the ERG as a biomarker of MDD and its ability to predict susceptibility or resilience to stress in mice.

METHODS: Rod ERGs were performed in male and female C57BL/6 mice before and after chronic social defeat stress (CSDS). Mice were then divided as susceptible or resilient to stress.

RESULTS: Our results suggest that CSDS reduces the amplitude of both oscillatory potentials and a-waves of the rods in resilient, but not susceptible male mice. In females, rod ERGs mainly reveals age-related changes. Finally, our analysis suggests that baseline ERG can predict with up to 71% of efficacy the expression of susceptibility and resilience before stress exposition in males and females.

CONCLUSION: Overall, our findings suggest that the retinal activity is a valid biomarker of stress response that could potentially serve as a tool to predict whether males and females will become susceptible or resilient when facing CSDS.

Clozapine rechallenge or continuation despite neutropenia, an extended follow-up of a consecutive Quebec case series

INTRODUCTION - INTRODUCTION : Clozapine is the most efficacious antipsychotic for treatment-resistant schizophrenia. However, clozapine induced neutropenia warrants treatment discontinuation, hindering recovery. Several case reports describe clozapine rechallenge or continuation despite neutropenia, although many are subject to selective reporting, with incomplete information and short follow-up period. Thus, consecutive case series, devoid of such bias, with long-term comprehensive follow-up are needed to better assess this practice.

OBJECTIF - OBJECTIVE : Describes consecutively the evolution of every patient in the Québec City catchment area in whom clozapine was either reintroduced after neutropenia during a previous clozapine trial or was maintained despite a first neutropenia between January 1, 2000, and October 22, 2017.

MÉTHODES - METHODS : Patients were identified through clozapine’s national hematological monitoring database and their medical records.

RÉSULTATS - RESULTS : Twenty-three patients were identified, 8 patients continued clozapine despite neutropenia, while 15 discontinued clozapine and attempted rechallenge later on. A total of 6 patients experienced further neutropenia episodes. Every patient who had a neutropenia recurrence also had a possible alternative explanation for neutropenia other than clozapine. Six patients discontinued clozapine due to mild neutropenia (absolute neutrophil count between 1.0-1.5 x109/L) (not precluding clozapine cessation ubiquitously). At the end of follow-up, 16 patients were still on clozapine, no deaths occurred and only 3 cases discontinued due to a hematological event.

CONCLUSION: This study suggests that clozapine rechallenge/continuation could be beneficial for good responders, although individual risks must be balanced with advantages of such practice. It also raises questions on pertinence to update Canadian clozapine monitoring guidelines, preventing unnecessary treatment discontinuation.

SCALING BANDIT ONLINE OPTIMIZATION OF SUPER-RESOLUTION OPTICAL MICROSCOPY FOR IMAGING OF NEURONAL STRUCTURES

OBJECTIVE: In order to do quantitative analyses on very small neuronal structures, such as synaptic nanodomains, it is often necessary to use super-resolution (SR) microscopy techniques, which allow increased resolution, down to a few tenths of nanometers, compared to conventional confocal microscopy. SR microscopes are configured by several parameters that need to be tuned properly. This parameter adjustment task is not only typically long and tedious, but also requires a high level of expertise. It has been shown previously that various SR imaging modalities can be efficiently optimized in an online fashion using Thompson sampling (TS) with Gaussian process (GP) regression to model multiple objective functions. However, nonparametric regression such as GP requires a discretization of the input space, which is problematic for large input spaces (e.g. many parameters). The goal of this project is to scale the online optimization of various SR modalities to large and continuous parameter spaces. METHODS: A polynomial (parametric) regression model has been selected for the resolution, signal-to-noise ratio, and photobleaching objectives. The NSGA-II algorithm was used for generating a set of tradeoffs approximating the Pareto front to be presented to the user. In order to obtain a sufficient variety of solutions close to the preferences of the user in the objective space, a weighted preference method was adapted to the four objectives and more than one preferred region case. RESULTS: A set of interactive user interfaces and graphical representations of the parameter and objective spaces were developed to make this optimization approach accessible to microscopists and neurobiologists. Empirical experiments were conducted for a single color STED imaging task in a four objectives setting. Preliminary STED optimization results suggest that at least up to six parameters can be optimized simultaneously. Additionally, the concentration of tradeoffs in specific regions of the objective space makes the choices somewhat easier for the user. CONCLUSION: The resulting online optimization method allows the optimization of multiple  STED microscope parameters. The designed interface facilitates the visualization of the multidimensional space and allows experts to focus on the imaging task, making this machine learning-assisted SR method more accessible. In the future, this approach will be applied to multi-color STED imaging of live cultured neurons and brain slices.

Transcriptional adaptations in the blood-brain barrier underlie pro-resilient effects of enriched environment and physical exercise against chronic stress

OBJECTIVE - OBJECTIVE

Major depressive disorder (MDD) is a leading cause of disease worldwide. Chronic stress is a major risk factor for MDD which can damage the blood brain barrier (BBB), leading to infiltration of immune mediators to the brain and neurological deficits. In mice access to an enriched environment (EE) or physical exercise (PE) can promote resilience to stress, but the biological mechanisms underlying this positive effect remains unclear. We hypothesize that access to EE or PE could prevent stress-induced BBB damage by encouraging adaptations in the mouse neurovasculature promoting resilience to chronic stress.

METHODS - METHODS

Male mice were subject to ten days of chronic social defeat stress (CSDS), during which they had free access to a house, nesting material, and toy (EE) or running wheel (PE). On day 11 a social interaction (SI) test was performed to determine stress susceptible (SS) or resilient (RES) phenotype. Mice were sacrificed to collect brain punches from the nucleus accumbens (NAc) and prefrontal cortex (PFC) for gene expression analysis (qPCR). Results were compared to previous standard environment (SE) CSDS cohorts from our lab.  

RESULTS - RESULTS

Access to EE or PE during CSDS is associated with greater proportion of RES vs. SS mice than SE cohorts. Further, PE and EE positively modulate neurovascular transcription. Both conditions alleviate stress-induced downregulation of tight junction proteins crucial for BBB integrity observed in CSDS with SE. Additionally, stress elicits upregulation of endothelial marker Pecam-1 in the NAc of EE mice, and astrocyte marker Gfap in the NAc of PE mice, suggesting different mechanisms of action. EE was also associated with increased baseline expression of pro-inflammatory genes in the PFC, but not NAc, compared to SE.

CONCLUSION

EE and PE promote resilience to CSDS and positively modulate neurovascular transcription in response to stress. These results clarify environmental contributions to pathophysiology of depression and indicate directions for novel antidepressant therapies.

Modulation of innate immunity as a preventive treatment inAlzheimer’s Disease

Alzheimer’s disease (AD) is a neurodegenerative disease characterized by the accumulation of amyloid beta (Aß) in the brain and within the blood vessels. The most common symptoms are short-term memory loss associated with a change in personality, neuroinflammation and blood brain barrier breakdown. In human and mouse, monocytes play an important role in AD, they are divided into 3 subpopulations, namely inflammatory/classical, intermediate and patrolling/non-classical distinguished by the expression of CD14, CD16 in human and Ly6C in mouse. Patrolling monocytes are of interest since they crawl along the blood vessels wall and are able to uptake amyloid and modulate the inflammation. Inflammatory monocyte is the main subpopulation found in AD, however, Muramyl dipeptide (MDP) can convert the latter into patrolling Ly6Clow. We hypothesized that an increased number of patrolling monocytes could decrease the Aß load in blood vessels and prevent amyloid accumulation in the brain via a well-known mechanism namely the sink effect. Objectif: Our study aims to modulate the innate immune system with MDP to delay or silence AD onset. Methods: We used the Alzheimer’s mouse model APPswe/PS1 and we induced and maintained a high level of patrolling monocytes by a weekly injection of MDP in 3-month-old mice for 3 months. We assessed the effect of MDP on cognition with NOR, quantified amyloid burden within the brain and vasculature using an unbiased stereological count and molecular changes at synapses and BBB level by Westernblot. To understand the molecular pathways triggered by MDP we did in vitro tests using PBMCs from mouse and Alzheimer’s patients. In parallel APP-CX3CR1/GFP mice were used in intravital 2-photon experiences. These mice helped us to demonstrate the conversion of monocytes and showed that patrolling monocytes are the main subpopulation in contact with Aβ and actively remove the latter form vessel. Results: We found that chronic MDP injections delay cognitive decline, decrease Aβ burden and protect the BBB. Furthermore, MDP seems to regulate the inflammation on Alzheimer’s blood in ex vivo assays. Conclusions: MDP is able to delay AD onset and can be considerate as a candidate for a treatment at early stage of the disease.

Déficits de production de phrases en contexte d’aphasie : efficacité des traitements et pistes d’intervention

Problématique. Véritable handicap « invisible », l’aphasie est présente chez le tiers des victimes d’accident vasculaire cérébral (AVC), entrainant des difficultés souvent sévères et durables pour l’intégration sociale et la qualité de vie. Les personnes aphasiques peuvent éprouver diverses problématiques associées à la communication, incluant des difficultés de production de phrases (DPP). À ce jour, différents traitements ont été développés pour intervenir auprès des patients ayant des DPP. Toutefois, leur efficacité n’a pas été mesurée de manière constante d’une étude à l’autre, ce qui rend difficile le choix de traitement pour les cliniciens. Objectifs. L’objectif de la présente étude était : (1) analyser l’efficacité des traitements existants – en termes d’amélioration des items entrainés, de généralisation à des items non entrainés, de transfert à d’autres contextes et de maintien des acquis dans le temps – et (2) développer et valider un traitement franco-québécois pour la rééducation des DPP chez les personnes aphasiques post-AVC. Méthodes. (1) Une revue systématique de la littérature a été menée dans trois bases de données : Pubmed, CINHAL et LLBA. La qualité méthodologique des articles a été évaluée. (2) Une étude de cas est réalisée afin de mesurer l’efficacité d’un traitement franco-québécois inspiré du Verb Network Strengthening Treatment et des Mapping Therapies. Résultats et conclusions. (1) Les résultats de la revue de la littérature montrent que les traitements existants permettent une amélioration de ce qui est travaillé et une généralisation à certains items non travaillé. Toutefois, des caractéristiques individuelles des participants (ex. : temps post-AVC, sévérité de l’aphasie) semblent avoir un grand impact sur l’efficacité des traitements. (2) Les résultats préliminaires de l’étude de cas et les pistes futures seront discutées.

Stimulations magnétiques périphériques répétitives chez l’adulte avec paralysie cérébrale: une étude de cas

OBJECTIF - OBJECTIVE

Les stimulations magnétiques périphériques répétitives (rPMS des muscles) sont non invasives et indolores. Leurs mécanismes d’action permettent d’influencer la plasticité cérébrale pour améliorer le contrôle des mouvements chez des enfants avec paralysie cérébrale (PC). Cependant, la littérature reste limitée quant à leurs effets chez des adultes avec PC. La présente étude de cas en PC a donc testé si les rPMS pouvaient encore influencer la plasticité cérébrale à l’âge adulte et améliorer la fonction motrice ainsi que la marche.

MÉTHODES - METHODS

Une femme de 30 ans avec PC mixte (suite hypoxie à la naissance) et marchant avec une aide technique (marchette) a participé à quatre séances de rPMS (une séance par semaine). Les rPMS ont été appliquées en bilatéral sur les muscles des jambes (tibial antérieur-TA, ischio-jambiers) et du tronc (transverse abdominal, multifides paravertébraux). Les données suivantes ont été collectés aux sessions 1 (ligne de base) et 4 (effets) et au suivi (40 jours plus tard) : contrôle sensorimoteur de la cheville (amplitude des mouvements, résistance des muscles à l'étirement), patron de marche (avec/sans marchette) et plasticité cérébrale (testée via stimulation magnétique transcrânienne de la représentation motrice corticale (M1) du TA).

RÉSULTATS - RESULTS

Les rPMS ont amélioré le contrôle de la cheville, avec baisse de la résistance des fléchisseurs plantaires à l'étirement et augmentation de l'amplitude de mouvement en dorsiflexion. Ces améliorations se sont maintenues au suivi. En parallèle, le fonctionnement du M1 controlatéral au muscle TA testé s’est amélioré (et le contrôle anormal par le M1 ipsilatéral s’est retiré) : augmentation de l’excitabilité et de la synchronisation corticospinales et réactivation des processus inhibiteurs impliqués dans la planification motrice. Le patron de marche s’est amélioré en conséquence.

CONCLUSION

Même à l’âge adulte, soit 30 ans après la lésion néonatale, les rPMS ont pu influencer la plasticité cérébrale et améliorer la motricité et la mobilité. Ces résultats originaux sont à reproduire et sont prometteurs pour toute personne adulte ayant eu un AVC périnatal.

Study of the KCC2 chloride transport in different neurodegenerative diseases.

OBJECTIF - OBJECTIVE

The intracellular chloride concentration in neurons plays an important role since it regulates the inhibitory response of the γ-aminobutyric acid type A (GABAA) receptor. In mature neurons, the KCC2 is the principal chloride co-transporter and has for principal function to maintain a low intracellular chloride concentration. This low chloride concentration allows an inhibitory response to GABA release. A loss of activity of the KCC2 already has been associated with many neurologic diseases like epilepsy, stress, anxiety, schizophrenia, and chronic pain. We have shown that KCC2 membrane expression was downregulated in neurons of the medial prefrontal cortex (mPFC) and hippocampus of Alzheimer (AD) mice and the motor cortex of ALS mice. This study as for objective to determine if the downregulation observed is also associated with a decrease of function. 

MÉTHODES - METHODS

We use fluorescence lifetime imaging microscopy (FLIM) combined with a FRET-based chloride sensor to monitor intracellular chloride concentration. The fluorescence lifetime of a fluorophore donor will decrease when its energy is transferred (FRET) to an acceptor. The SuperClomeleon is a FRET-based chloride sensor composed of a mCerulean and a YFP chloride sensitive which is quenched by the binding of the chloride. We injected virus encoding the SuperClomeleon in the brains of mice (mPFC (4 months) and hippocampal (6 months) for AD study & the motor cortex (2 months) of ALS mice). We monitored the intracellular chloride in neurons while increasing the extracellular KCl concentration to reverse the direction of the transport of chloride by the KCC2 and induce activity. 

RÉSULTATS - RESULTS

The entry rate of chloride observed in neurons of AD mice was significantly diminished by ~20% in the mPFC and ~15% in the hippocampus. Similarly, a decrease in the rate of chloride entry by KCC2 was also observed in the ALS mice (~25%), which could be recovered by using a KCC2 enhancer (PCPZ).

CONCLUSION

These results indicate that KCC2 plays an important role in establishing neuronal homeostasis and that it is a promising pharmacological target that could be explored in various neurodegenerative diseases.

pySTED : a STED microscopy simulation tool for machine learning training

OBJECTIF - OBJECTIVE

While super-resolution microscopy techniques such as STimulated Emission Depletion (STED) allow us to resolve nanoscopic structures, they remain challenging to use for non-expert users due to the number of parameters to consider to obtain high quality images. There is a growing interest in applying machine learning (ML) methods to STED microscopy in order to assist microscopists during imaging. However, ML, and more specifically reinforcement learning (RL), approaches are still often data-hungry in training, leading to important biological sample waste. This calls for a way to train RL agents to tackle the application of dynamic STED imaging of neuronal nanostructures.

MÉTHODES - METHODS

We have developed a STED simulation tool (pySTED) that aims to realistically simulate confocal and STED image acquisition of samples through integration of fluorophore mechanics and photon emission simulation. The contributions of pySTED are two fold: i) it allows microscopists to observe the effects that parameter tuning has on the resulting image, and ii) it allows AI methods to be trained in simulation, reducing the requirements for real biological samples. We train RL agents with the goal of optimizing multiple imaging objectives, such as resolution, signal to noise ratio, photobleaching, and synaptic nanodomain (ND) resolvability. RL agents are trained by interacting with their environment (in this case the microscope), in a trial-and-error process, and gain a reward based on their ability to resolve NDs.

RÉSULTATS - RESULTS

The pySTED simulator can realistically simulate various environment dynamics, which is important to train robust RL agents. We used pySTED to simulate multiple situations of varying imaging conditions and fluorophore parameters. We then used the pySTED simulator to train an RL agent to resolve synaptic NDs in dendritic spines with optimal STED acquisition parameters. Our results show that RL agents can learn to optimally image and resolve synaptique NDs in simulated dendritic spines.

CONCLUSION

The pySTED simulator allows the development of RL algorithms that can be applied to the imaging of neuronal nanostructures. The next step will be to train agents to image dynamic miniature synaptic calcium transients, adding a temporal challenge to the problem. The final goal will be to deploy the resulting RL agents in order to resolve calcium NDs in living neurons.

Dérégulation de l’axe immun-métabolique en sclérose latérale amyotrophique : étude du rôle de soluble tumor necrosis factor receptor II et du CC-motif chemokine ligand 16

OBJECTIF - OBJECTIVE

La sclérose latérale amyotrophique (SLA) est une maladie neurodégénérative mortelle caractérisée par une vitesse de progression hétérogène. Un mauvais pronostic serait potentiellement lié à un état hypermétabolique, c’est-à-dire à une dépense énergétique de repos supérieure à la population saine. 

L’hypothèse suggérée est qu’une dérégulation de l’immunité créerait un profil immun-métabolique unique aux patients atteints d’une SLA à progression rapide. Il en résulterait un déséquilibre énergétique par l’hyperactivation de la voie de l’AMP-activated protein kinase (AMPK) au sein des adipocytes. Cela entraînerait une réduction de la production de leptine et un hypermétabolisme. 

L’objectif principal est donc de décrire une signature moléculaire associée aux formes plus agressives de la SLA.

MÉTHODES - METHODS

Un modèle in vitro humanisé de l’hypermétabolisme en SLA a été élaboré à partir d’adipocytes. Une fois à maturité, ils ont été traités avec du plasma de patients contrôles et atteints d’une SLA à progression lente ou rapide. Divers traitements ont ensuite pu être effectués avant de récupérer le milieu de culture et de récolter les cellules. Grâce au human obesity cytokine array C1 (RayBio®), 62 cytokines et adipokines ont été évaluées et quantifiées. 

RÉSULTATS - RESULTS

Des résultats préliminaires prometteurs ont permis de noter des niveaux sériques diminués en leptine et augmentés en soluble tumor necrosis factor receptor II (sTNF-RII) et en CC-motif chemokine ligand 16 (CCL16) chez les patients à progression rapide. En in vitro, CCL16 n’a pas eu d’effet significatif sur la sécrétion de leptine par les adipocytes, tandis que sTNF-RII a bloqué la sécrétion de leptine et augmenté la production de AMPK phosphorylé. D’autres traitements avec ces protéines recombinantes, de façon séparée puis combinée, ainsi qu'avec leurs anticorps neutralisants, permettront de mieux cerner leur rôle dans l’axe immun-métabolique en SLA

CONCLUSION

Ces travaux constituent une des premières analyses du rôle de sTNF-RII et de CCL16 dans l’homéostasie métabolique en SLA. Ils démontrent une potentielle signature moléculaire propre aux patients SLA à progression rapide qui permettrait de prédire précocement le rythme d’évolution de la maladie. Au long cours, l’élaboration d’un modèle in vitro humanisé de l’hypermétabolisme en SLA à partir d’adipocytes humains pourrait mener au développement de nouveaux outils diagnostiques, pronostiques et thérapeutiques.

Preventive strategies alleviate depressive-like behavior and improve intestinal barrier integrity

OBJECTIF - OBJECTIVE

Chronic stress is a major contributor of major depressive disorder (MDD), a known risk factor of suicide, and the leading cause of disability worldwide. Throughout life, one in five individuals will be affected. Common antidepressants are only effective in 30 to 50% MDD individuals. This low remission rate suggest that neuron-based pharmacological treatments may benefit from novel therapeutic approaches targeting peripheral biological systems, such as the gut-brain axis, or could be potentiated by preventive strategies.

MÉTHODES - METHODS

To explore these hypotheses, we took advantage of the chronic social defeat stress (CSDS) paradigm, a mouse model of depression. For 10 days, male mice were subjected to a physical encounter with an agonistic aggressor and then left in sensory contact. Thereafter, tested animal went through a social interaction test to determine behavioral phenotypes, namely susceptibility or resilience to stress. Thus, this model allows us to explore stress-induced deleterious alterations but also beneficial biological adaptations. We introduced an enriched environment or physical exercise, two preventive strategies, to verify if it could promote stress resilience via changes in the gut barrier integrity. Intestinal tissue was collected for transcriptional analysis and morphological assessment of tight junction protein levels.

RÉSULTATS - RESULTS

Our results confirmed that introducing nesting material or a running wheel substantially increase the proportion of resilient mice. Interestingly, jejunal tight junction protein expression pattern in these populations of mice support positive molecular adaptations to chronic social stress exposure, suggesting that the gut barrier integrity could play an active role in stress responses at least in mice.

CONCLUSION

We identified changes specific to the mouse susceptible versus resilient phenotype in gut tight junction gene expression which could contribute to exacerbated circulating inflammation leading to maladaptive behaviors. In the next steps, we will investigate peripheral inflammatory cytokine profile and gut microbiota changes in each CSDS-preventive paradigm. Understanding the physiological role of intestinal barrier integrity in a preventive strategies setting will open new opportunities in the treatment of depression.

Contrôle inhibiteur, densité de matière grise cérébrale et perte de poids suivant la chirurgie bariatrique

Objectif : L’obésité est associée à une atrophie de la matière grise (MG) cérébrale dans des régions impliquées dans le contrôle inhibiteur, dont le cortex orbitofrontal (OFC) et le cortex préfrontal ventromédian (vmPFC). Notre objectif est d’examiner le changement du contrôle inhibiteur post-chirurgie bariatrique et de déterminer les associations entre ce changement, les changements de densité de MG et la perte de poids.

Méthodes : Nous avons recruté 82 candidats à la chirurgie bariatrique (âge: 43,5±9,1 ans; IMC: 43,7±3,9 kg/m²) qui sont présentement suivis avant la chirurgie et 4, 12 et 24 mois post-chirurgie (n=82; n=66; n=44; n=29). À chaque session, les participants exécutent la tâche informatisée Stop Signal Reaction Time qui mesure la capacité d’inhiber un mouvement (SSRT). Un modèle linéaire à effets mixtes a été utilisé pour comparer les changements de perte de poids et du SSRT avant et après la chirurgie. La densité de MG des régions d’intérêt (OFC et vmPFC) a été quantifiée à partir des IRM anatomiques obtenues à chaque session.

Résultats : Les participants ont perdu en moyenne 22,4±4,0% de leur poids total à 4 mois, 35,5±7,7% à 12 mois et 33,7±7,8% à 24 mois. Bien qu’aucune différence significative ne soit observée pour les valeurs de SSRT dans le temps (p=0,641), une amélioration du contrôle inhibiteur (réduction du SSRT) est associée à une augmentation de la densité du vmPFC (r=-0,27, p=0,043) et de l’OFC (r=-0,31, p=0,019) 4 mois post-chirurgie. Aucune association n’est observée entre les changements de SSRT et la perte de poids.

Conclusion : Ces résultats préliminaires suggèrent qu’une augmentation de densité de MG dans des régions associées au contrôle cognitif et la prise de décision est associée à un meilleur contrôle inhibiteur 4 mois post-chirurgie. Lorsque nous aurons terminé tous les suivis, nous prévoyons examiner si le contrôle inhibiteur est un prédicteur de la perte de poids à 24 mois.

Circadian rhythm of tau phosphorylation and secretion is driven by body temperature

OBJECTIF - OBJECTIVE

Aggregates of hyperphosphorylated tau protein are a hallmark of Alzheimer’s disease (AD) and other tauopathies. Moreover, growing evidences suggest that tau spreading can occur via cell-to-cell transfer involving secretion and internalization of pathological forms of tau protein followed by misfolding of normal tau in recipient cells. Previously, we demonstrated that tau phosphorylation level is inversely correlated with body temperature. Cold exposure has been shown to increase tau phosphorylation whereas higher temperature was linked to a reduction of tau pathology. We thus aimed to determine whether tau phosphorylation and secretion are regulated by circadian rhythms, inherently linked to the sleep-wake cycle and body temperature variations.

MÉTHODES - METHODS

First, to answer these questions we analysed by Western blotting tau phosphorylation on relevant epitopes in the brains of awake and sleeping B6 mice, while recording their temperature. Second, we exposed neuronal cells to different physiological temperatures (from 35°C to 39°C) to assess both intracellular tau and tau-secreted species in the cell medium by ELISA and dot blotting.

RÉSULTATS - RESULTS

We found that tau phosphorylation undergoes sleep-driven circadian variations, as it is hyperphosphorylated during sleep, when body temperature is lower. Similar changes in tau phosphorylation were reproduced in neuronal cells exposed to temperatures recorded during the sleep–wake cycle. In addition, we observed that the secretion of tau is temperature-dependent, as higher temperature exposure increased total tau level in the extracellular medium. However, at higher temperatures, tau-secreted species are less phosphorylated on several epitopes. Finally, our results demonstrated that tau cleavage at D421 by caspase-3 regulates its secretion, since the level of extracellular tau was decreased when cells were treated with a selective caspase-3 inhibitor (zDEVD-FMK).

CONCLUSION

Taken together, these data suggest that tau phosphorylation and secretion follow a circadian rhythm driven mostly by body temperature and sleep. Since AD patients are prone to sleep disturbances and thermoregulation deficits, our study provides new outcomes on how tau pathology could spread and affect different brain regions.

Perspective du stress psychologique dans l’étiologie de l’apnée du sommeil

OBJECTIF

L'apnée du sommeil (AS) est un trouble respiratoire multifactoriel qui semble être bien plus qu’un problème anatomique, toutefois ses origines restent peu connues. Le stress chronique induit de l’hypertension et de l’obésité, deux comorbidités fréquentes de l’AS, cependant ses effets sur l'homéostasie cardiorespiratoire sont peu étudiés. L’hypothèse proposée est que chez les rats mâles, l’isolement social (IS) induit une perturbation de la régulation respiratoire au cours du sommeil. Nous avons ensuite comme objectif d’identifier les mécanismes neurobiologiques sous-jacents. 

MÉTHODES

Les rats âgés de 8 semaines ont été soumis à 3 semaines d’IS (hébergement 1 par cage; n=13); les animaux témoins ont été logés en paires (n=12). Les apnées ont été quantifiées par pléthysmographie à corps entier, les mesures cardiovasculaires par pression du brassard de queue et la composition corporelle par résonance magnétique. À la fin du protocole, le sang a été prélevé par ponction cardiaque afin de mesurer le fonctionnement de l'axe neuroendocrinien par dosage immunologique multiplex. Le cerveau a été récolté pour un éventuel marquage par immunohistochimie (Fos-B) des circuits respiratoires.

RÉSULTATS

À la fin du protocole, les rats soumis à l’IS ont un indice d’apnée 27% supérieur aux témoins. Ils sont hypertendus (+15mmHg) et ont un gain de masse grasse et de fluides de, respectivement, 21,3% et 37,6% plus élevés que les témoins. Toutefois, la consommation alimentaire n’est pas différente. Les données préliminaires suggèrent que la corticostérone plasmatique est ~30% plus élevée chez les rats IS. Aucune donnée sur l’identification des structures neuronales n’a été recueillie à ce stade. 

CONCLUSION

Puisque l’isolement induit des troubles cardiorespiratoires significatifs et une perturbation métabolique en lien avec le phénotype observé chez l’humain, nous proposons que le stress psychologique est important dans l’étiologie de l’AS. Les prochains résultats nous permettront d’évaluer les mécanismes neurologiques affectant la fonction respiratoire activés par l’IS.

Exploration of the red blood cells biomechanics with digital holographic microscopy: Towards a methodology to identify cellular phenotypes related to major psychiatric disorders

Major psychiatric diseases (MPDs) including schizophrenia (SZ), are diagnosed very late due to the lack of effective biomarkers. Post-mortem analyzes show that the lipid composition (LC) of neuronal membranes differs in people with SZ. This difference is also reflected in several types of cells, including red blood cells (RBCs). RBCs are known to exhibit spectacular biomechanical properties (BPs) resulting in unique deformability capacity and spontaneous membrane vibrations (MVs) at the nanoscale. It has been reported that the LC of the RBCs membranes affects these BPs. An accurate biomechanical characterization of RBCs could thus reveal MPDs-related phenotypes. Quantitative-phase digital holographic microscopy (QP-DHM), providing images with a nanometric axial sensitivity, represents a highly relevant technique to quantitatively study the RBCs biomechanic MVs.
As a first step to identify MPD-related RBCs phenotypes, we developed a methodology based on QP-DHM to characterize RBCs biomechanics from the dynamics of their MVs. Studies are conducted in different conditions known to specifically impact RBCs BPs. Concretely MVs and RBCs deformations are monitored in environments controlled for temperature, pH, O2 and CO2 partial pressures. We started a biomechanical study of RBCs obtained from mice subjected to a strict diet allowing a specific control of their membrane LC.
After eliminating several potential confounders that could influence the interpretation of our measurements, our preliminary results, obtained on samples of control subjects, show that the spontaneous vibrations of the membranes of the RBCs aren’t affected by the temperature, thus speaking for a non-thermal origin of the latter. Conversely, the morphology of the RBCs seems to be influenced by the temperature.
Aiming at identifying MPDs-related RBCs phenotypes, we will apply this methodology on our clinical data set composed of RBCs samples collected from patients suffering for MPDs.

Modulation of the biological activity of DKK1 decisively influences structural and functional neurovascular recovery after ischemic stroke.

BACKGROUND

Stroke constitutes a major cause of death and disability in adults in Canada, and occurs as a result of a sudden obstruction within a cerebral artery due to an embolus or thrombus. We have previously demonstrated that stroke stressors deregulates the canonical Wnt pathway, which plays key roles in regulating neurovascular functions. Interestingly, Dickkopf-1 (DKK1), a secreted inhibitor of the pathway, is induced after stroke in patients and animal models. Although DKK1 levels have been shown to increase, its implication in stroke pathobiology and therapy remains unknown.

OBJECTIVES

We postulate that DKK1 increased expression impairs structural and functional brain recovery after stroke. To test this hypothesis, our project aims to develop the following specific objectives: 1) investigating the impact of ectopic DKK1 induction in mice on brain functioning, 2) evaluating the consequences of temporal DKK1 induction (early and prolonged induction) in stroke pathobiology, and 3) assessing the potential of DKK1 neutralization on brain recovery after stroke.

METHODS AND RESULTS

For the first objective, using conditional transgenic mice in which DKK1 could be induced upon doxycycline administration (iDKK1), we found that DKK1 induction was sufficient to physiologically deactivate the canonical Wnt pathway in the brain, which was accompanied by a poor motor coordination in absence of macroscopic brain damage. For the second objective, we found that DKK1 induction prior to ischemic stroke, which was induced using middle cerebral artery occlusion (MCAo), worsened structural damage, impaired motor functions, as well as decreased overall survival rate. These changes were associated with an exacerbated neuronal degeneration by increasing FJB+ cell density, and an impaired neurogenesis by decreasing the levels of DCX+ cells in the SVZ and TUJ1+ cells in the perilesional and infarcted areas. Finally, DKK1 induction impaired angiogenesis and cerebral blood flow (CBF) regulation. For the third objective, we found out pharmacological neutralization of DKK1 after MCAo induction  using WAY262611 decreased structural brain damage and restored motor functions 1 week after stroke.  In addition, DKK1 inhibition decreased neuronal degeneration, and stimulated neurogenesis and angiogenesis 1 week after stroke. 

CONCLUSION

Our findings suggest that DKK1 induction is directly implicated in the poor diagnosis after stroke whereas DKK1 neutralization constitutes a very promising and safe therapeutic approach to stimulate brain structural and functional recovery after stroke. Importantly, DKK1 neutralization is now being assessed in various clinical trials for other medical conditions, thus increasing the translational potential of our findings.

The Smart Vivarium: Implementation and validation

Our capacity to consolidate the behavioral strategies to cope with stress are greatly influenced by social interactions. Similarly, chronic mice stress models in rich environments open a wide array of possibilities for the analysis of environmental and social factors using an automated assessment and tracking system. In a large group, mice are influenced by the stress put by other members of the group over time. An evolutive structure is formed by the social hierarchy in the group. For example, in this evolutionary structure, group members interact with each other in a rich and complex environment. They play and fight with each other to prove their dominance to the group. The behavioral ability that each mouse exhibits during daily life to control stress determines its status in the colony. The goal of this project is to develop an automated behavioral platform using machine-learning algorithms and fiducial markers to evaluate the behaviors resulting from daily life in male and female mice ethically. To achieve our goals, we have developed a smart vivarium of large dimensions (1m x1m) equipped with a high-quality camera including various enriching components such as beddings, food, nesting material, games and toys, bridges, and ropes for each group of male and female. We have controlled the behavioral characteristics of different groups up to a maximum of 10 mice for 6 weeks in the vivarium for both male and female groups in parallel and finally archive the corresponding data in the cloud. To that end, we created an extensive library of several thousand images of different mice to automatically detect individuals and identify group behavioral characteristics by tracking them through a hybrid machine system consisting of DeepLabCut with a new MiceTag algorithm. We identify the behaviors identified in sub-categories in behavioral modelings such as combatants, friendly interactions, and group formation, and validate the accuracy of this classification through manual codification. On average, every 2.4 frames the tags are identified in the light cycle and 4.1 in the dark cycle to boost significantly the accuracy of multiple animal tracking and identification by DeepLapCut up to 95% in detection periods.  

Criterion validity of ActiGraph monitoring devices for step counting and distance measurement in adults and older adults: a systematic review

OBJECTIF - OBJECTIVE Wearable activity monitors such as ActiGraph monitoring devices are widely used, especially in research settings. Various research studies have assessed the criterion validity of ActiGraph devices for step counting and distance estimation in adults and older adults. Although several studies have used the ActiGraph devices as a reference system for activity monitoring, there is no summarized evidence of the psychometric properties. The main objective of this systematic review was to summarize evidence related to the criterion validity of ActiGraph monitoring devices for step counting and distance estimation in adults and/or older adults. 

MÉTHODES - METHODS Literature searches were conducted in six databases (Medline (OVID), Embase, IEEExplore, CINAHL, Engineering Village and Web of Science). Two reviewers independently conducted selection, a quality analysis of articles (using COSMIN and MacDermid’s grids) and data extraction.

RÉSULTATS - RESULTS This review included 21 studies involving 637 participants (age 30.3±7.5 years (for adults) and 82.7±3.3 years (for older adults)). Five ActiGraph devices (7164, GT1M, wGTX+, GT3X+/wGT3X+ and wGT3X-BT) were used to collect data at the hip, wrist and ankle to assess various walking and running speeds (ranging from 0.2 m/s to 4.44 m/s) over durations of 2 minutes to 3 days (13hrs30mins per day) for step counting and distance estimation. The ActiGraph GT3X+/wGT3X+ and wGT3X-BT had better criterion validity than the ActiGraph 7164, wGTX+ and GT1M according to walking and running speeds for step counting. Validity of ActiGraph wGT3X+ was good for distance estimation. 

CONCLUSION The ActiGraph wGT3X-BT and GT3X+/wGT3X+ have good criterion validity for step counting, under certain conditions related to walking speeds, positioning and data processing.

Functional dysfunctions of the corticotegmental pathway in a mouse model of chronic stress.

OBJECTIF - OBJECTIVE

Several lines of evidence suggest that cortically driven dysregulation of subcortical circuits triggers depressive-like behaviours in humans and animals. Amongst these circuits, the medial prefrontal cortex (mPFC) sends dense projections to the nucleus accumbens (NAc) and the ventral tegmental area (VTA). Recent findings from our lab showed that chronic stress induces functional and morphological changes affecting the activity of the corticoaccumbal and corticotegmental pathways in a sex-dependent fashion. However, whether these pathway- and sex-specific functional changes result in durable  alterations in the VTA and the NAc of males and females remains unclear.

MÉTHODES - METHODS

Chronic variable stress (CVS) was used to produce a persistent depressive-like phenotype in males and females. We concentrated our investigation on the corticotegmental pathway. A trans-sectional viral strategy was used to expressed channelrhodopsin in the corticotegmental pathway and to label downstream neurons in the VTA. Optogenetic stimulation on live brain slices was used to activate the corticotegmental projections in both stressed males and females. Whole-cell patch-clamp experiments were performed to measure optogenetically-evoked responses (oEPSCs) in VTA neurons and detect pre-and post-synaptic modifications.

RÉSULTATS - RESULTS

21 days of CVS induced a strong depressive-like behavioural phenotype in both males and females. Our viral approach successfully allowed us to optogenetically stimulate cortical projections to the VTA. Analysis of the paired-pulse and AMPA/NMDA ratios from oEPSCs in VTA neurons suggests no effect of chronic stress on pre-and post-synaptic activity, respectively, in both males and females. However, our analysis suggests that chronic stress interferes with the potentiation of mPFC-VTA connections normally observed following repetitive stimulation in control mice, supporting either alterations of the intracellular signaling or an occlusion of such pathway  in both males and females after stress.

CONCLUSION

Our results suggest that CVS interferes with intrinsic plasticity mechanisms at mPFC-VTA synapses in males and females, without affecting unitary evoked responses. Understanding these effects will provide novel insights into the neuronal mechanisms underlying behavioral adaptations to chronic stress in males and females.

Exploring neurodifferentiation and maturation process of iPSCs derived from schizophrenia patients with multimodal optical microscopy

OBJECTIVE -

Schizophrenia (SZ) is a serious mental disorder characterized by hallucinations, delusions, and impaired cognitive behavior. However, substantial heterogeneity in the clinical phenotypes makes it complicated to diagnose and treatment of it. Its pathogenesis results from a complex interaction between genetic vulnerability and environmental factors, altering neuronal development. Several genes and proteins associated with neurite growth, neuronal migration, and synaptic development were found to be altered in SZ. However, the processes of neurodifferentiation and functional maturation of neuronal networks are still unexplored. Therefore, generating in vitro cortical neural cultures using induced pluripotent stem cells (iPSCs) from SZ patients and controls. Further, studying the maturation and organization of these neuronal networks with cutting-edge multimodal optical imaging tools will be helpful to get new insights into the pathogenesis of SZ.

METHODS -

The iPSCs lines were derived from the SZ patients and controls in collaboration with the iPSCs Québec Platform. The iPSCs were differentiated into neural progenitor cells (NPCs) and then into mixed cortical neural cells using commercially available protocol from Stemcell Technologies Inc. The characterization was performed by flow cytometry, immunostaining, western blot, patch-clamp techniques, and calcium imaging.

RESULTS - 

We have successfully standardized the mixed neural cortical culture from the controls to explore neurodifferentiation and maturation processes. We have used a multimodal approach combining quantitative phase digital holographic microscopy (QP-DHM) with fluorescence imaging to study the water and calcium movement during neuronal activity. QP-DHM is a high-resolution, non-invasive technique that assesses cell structure and dynamics with axial sensitivity at the nanoscale. Our preliminary molecular and functional study results explore the neuronal cortical network maturation process. Furthermore, we will specifically explore the development of excitatory activity mediated by the NMDA receptors within such iPSC-derived neural networks.

CONCLUSION

The new emerging multimodal imaging approach with human iPSCs disease models coming from deeply phenotyped patients holds great promise to identify cellular risk biomarkers for major neurodevelopmental disorders including schizophrenia.

Investigating the impact of ALS-FUS in mitochondrial health in a neuron-specific mouse model

INTRODUCTION. Mitochondria is an organelle that has multiple roles essential for neuronal function and survival. Mitochondrial alteration and dysfunction have been proposed as a common hallmark in several neurodegenerative diseases, and it is also present in amyotrophic lateral sclerosis (ALS). ALS is a multifactorial disease where alteration of several cellular processes may be present in the neurons, as well as in their neighbouring cell populations. ALS patients and some ALS animal models shows mitochondrial alterations in their brain, spinal cord, and muscles. Most of the ALS cases are sporadic, however mutations in fused in sarcoma (FUS) are causative of the disease. ALS/FUS mice models are characterized by motor dysfunction, a neuro-inflammatory profile, muscle denervation, and neuronal morphological alterations. In addition, it has been proposed that mitochondrial damage can lead to the activation of glial cells, which can contribute to neurodegeneration. In this study, we provide in vivo evidence of the impact of FUS in mitochondrial health and the protective effect of therapeutic approaches that inhibits the development of neuro-inflammation for this organelle.

METHODS. We generated a neuron-specific transgenic mouse expressing the ALS mutation R521G in FUS using the Cre recombinase approach (ALS/FUS). Mice were phenotypically characterised to determine the neuronal contribution of mutant FUS. Mitochondrial health and function were studied by immunolabelling, and the effect of an anti-inflammatory compound (IMS-088) on this organelle was analysed.

RESULTS. ALS/FUS mice present an early and persistent cognitive impairment and a progressive motor dysfunction. In parallel, they present muscular denervation, neuronal morphological alterations and neuroinflammation. A reduction in the number of neuronal mitochondria and their activity is observed, which is exacerbated under stress conditions. Treatment with IMS-088 was able to improve ALS/FUS mice phenotype, as well as partially rescue mitochondrial health.

CONCLUSION. The results provided in this study shows an ALS-FTD phenotype in a neuron-specific mouse model expressing the FUS-R521G mutation. FUSR521G mutant seems to have a negative effect in mitochondrial health which can have an impact in the mice phenotype. Moreover, pharmacological inhibition of the NfkB pathway by IMS-088 can have a potential therapeutic effect in ALS/FUS.

Activation of neurotoxic reactive astrocytes by IL-1α drives the death of oligodendrocytes in the mouse spinal cord

OBJECTIF - OBJECTIVE

Spinal cord injury (SCI) results in loss of communication between the brain and the extremities, thus resulting in loss of sensation and voluntary movements below the site of injury (e.g. quadriplegia, paraplegia). We recently discovered that a small protein named interleukin-1 alpha (IL-1α), normally involved in the regulation of inflammation following infection, is released by spinal cord-resident microglia almost immediately following injury. Our objective is the study of this protein in the spinal cord after inthrathecal administration

MÉTHODES - METHODS

To study the effect of IL-1α in a simplified in vivo model, we took advantage of the LysMeGFP reporter mouse line, which expresses green fluorescence protein in myeloid cells, as well as IL-1R1-restored (Il1r1r/r) mice, which exhibit an IL-1R1 knockout phenotype that can be reversed in a cell-specific manner by Cre-mediated recombination once crossed with mGfapCre or Cdh5CreER mice to restore IL-1R1 expression in astrocytes and endothelial cells (ECs), respectively. We injected recombinant mouse IL-1α (rmIL-1α) intra-cisterna magna (i.c.m) to these mice and examined CNS responses at different time points using in vivo videomicroscopy or immunofluorescence staining in post-mortem tissue sections.

RÉSULTATS - RESULTS

Injection of rmIL-1α to mice induced blood-spinal cord barrier disruption, infiltration of myeloid cells throughout the spinal cord and increased expression of the complement component 3 (C3) protein in astrocytes. Notably, IL-1R1 signalling in ECs was found to drive neutrophil infiltration, while oligodendrocyte cell death was induced through astrocytic IL-1R1. 

CONCLUSION

Il1α produce the infiltration of the neutrophils by the activation of IL-1R1 in endhotelial cells and the death of the oligodendrocytes byt the activation neurotoxic reavtive astrocytes. Thus, blockade of IL-1α could be a potential treatment to limit secondary degeneration after SCI.

Determine the impact of social stress on the expression of anxious-depressive-like behaviors

OBJECTIF - OBJECTIVE. Environmental and social factors are major contributors of stress responses. Social interactions, either positive or negative, play an important role in the way we develop stress coping strategies. These interactions depend on the social status a person has in society. In the wild, mice establish dynamic social hierarchies within complex colonies, representing a good model to reproduce specific human conditions and to study the different biological, genetic, molecular, and functional mechanisms underlying stress susceptibility or resilience in males and females. Therefore, this project aims at defining biosignatures associated with social status and behavioral phenotypes in male mice living in a naturalized environment.

MÉTHODES - METHODS. Ten mice were placed in an environmentally enriched vivarium for 10 weeks in which each mouse was able to freely interact with each other. The tube test was performed to monitor hierarchy; blood and feces samples were taken to analyze biological data (FACS, cytokines, 16S sequencing). Biological samples and hierarchical status were assessed at the beginning, during, and after the 10 experimental weeks. Chronic social defeat stress was used in parallel as a well-known behavioral paradigm inducing depressive-like behaviors through social stress.

RÉSULTATS - RESULTS. Vivarium: we observed a stable hierarchical organization throughout the experimental duration. Interestingly, hierarchical status in the vivarium was associated with specific behavioral outputs. After the experimental 10 weeks, immune profiles vary according to social and emotional status.

CSDS: chronic social stress induced social withdrawal in 66% (susceptible) of our cohort, the remaining 33% (resilient) continuing to interact with social targets. Immune cellular populations and cytokines signatures showed specific effects in susceptible over resilient and control male mice.

CONCLUSION. Our results suggest that social stress induces dynamic variations in the proportion of immune cell populations that associate with social and emotional status in male mice. These biosignatures could serve as biomarkers for preventing and monitoring disease progression and predicting treatment response.

Light-inducible alpha-synuclein aggregation in the midbrain impairs nigrostriatal dopaminergic transmission

OBJECTIF - OBJECTIVE

Parkinson’s disease (PD) is characterized by intracellular inclusions of misfolded alpha-synuclein (alpha-syn), known as Lewy bodies (LBs), and by the loss of dopaminergic neurons in the substantia nigra pars compacta (SNc) that leads to dopamine (DA) depletion at the striatum. Recent studies have been proposed that the aggregation of alpha-syn can lead to synaptic dysfunction, representing an early event in the pathogenesis of PD. However, one of the main hurdles in studying the neurobiology of the disease is that, currently, there are no proper animal models that can reproduce all the characteristics of alpha-syn aggregation and LBs formation found in the brain of PD patients.

In this context, our laboratory has recently developed an in vitro and in vivo model of PD based on the optogenetics technology named LIPA (light-inducible protein aggregation) that controls the aggregation of alpha-syn under the control of blue light. We showed that LIPA mimics the histopathological characteristics of PD, and allow thus to study how the aggregation of alpha-syn in dopaminergic cells of SNc can cause a progressive disruption in the nigrostriatal pathway.

MÉTHODES - METHODS

To investigate the impact of LIPA-induced alpha-syn aggregation on the dopaminergic projections, we assessed the activity of striatal cells. Briefly, we implanted mini-endoscopes coupled with an optic fiber to induce alpha-syn aggregation in the SNc, and analyzed the neuronal activity using the calcium indicator GCaMP6s in the striatum of freely moving mice.

RÉSULTATS - RESULTS

Our results show a progressive decrease in the synchronized activity and frequency of striatal cells caused by the aggregation of alpha-syn.

CONCLUSION

Altogether, our data showed that the use of this new LIPA-alpha-syn system offer a unique tool to elucidate the morphological, and physiological changes occurring in the dopaminergic projections in the context of PD at early stages after alpha-syn aggregation.

Dans quelles conditions neurologiques les difficultés à comprendre et à exprimer les émotions par la voix se présentent-elles ?

OBJECTIF: Pour comprendre les émotions des autres et exprimer les nôtres, nous utilisons la prosodie affective (c’est-à-dire l’intonation et le rythme de la parole). La compréhension et la production de la prosodie affective nécessitent de nombreuses habiletés, dont celles liées au langage et à la cognition sociale, et requièrent des interactions entre plusieurs régions corticales et sous-corticales des deux hémisphères. Par conséquent, l’intégrité des habiletés prosodiques affectives est très vulnérable aux lésions cérébrales et de nombreux troubles neurologiques sont susceptibles d’engendrer des déficits prosodiques affectifs. Cependant, les populations cliniques à risque de présenter des difficultés et leurs profils d’atteintes sont méconnus. 

Cette étude vise à résumer les conditions neurologiques pouvant conduire à des difficultés à comprendre ou à exprimer les émotions par la voix et à caractériser différents profils d’atteintes de la prosodie affective chez les adultes avec un trouble neurologique. 

MÉTHODES: Une recherche systématique a été menée dans 5 bases de données avec un devis d’examen de la portée suivant les lignes directrices PRISMA. 

RÉSULTATS: Parmi les 94 études analysées, les difficultés de prosodie affective ont été répertoriées dans 17 conditions neurologiques. Les conditions neurologiques à risque d’entrainer un trouble de la prosodie affective sont hétérogènes quant à leurs manifestations cliniques (p. ex., aphasie, troubles cognitifs et troubles moteurs) et leurs lésions cérébrales (d’origines subites ou progressives; pouvant affecter les régions corticales, sous-corticales, la matière blanche des deux hémisphères). De plus, les profils d’atteintes de la prosodie affective se distinguent entre les groupes cliniques en fonction des demandes associées à la tâche d’évaluation. 

CONCLUSION: Cet examen de la portée suggère que les troubles de la prosodie affective sont communs à divers groupes cliniques de la population adulte avec lésions cérébrales acquises et que les difficultés puissent résulter d’atteintes à différents systèmes impliqués dans le traitement de la prosodie affective. 

Whole-brain morphometry in Canadian soldiers with posttraumatic stress disorder

OBJECTIVE: Patients with posttraumatic stress disorder (PTSD) display several structural brain differences when compared to healthy individuals. However, findings are particularly inconsistent for soldiers with PTSD. METHODS: Here, we characterized brain morphometry of 37 soldiers from the Canadian Armed Forces with adulthood war-related PTSD using structural magnetic resonance imaging. We assessed time since trauma, as well as PTSD, depressive and anxiety symptoms with the Modified PTSD Symptoms Scale, Beck Depression Inventory and Beck Anxiety Inventory, respectively. Whole-brain morphometry was extracted with FreeSurfer and compared with a validated normative database of more than 2700 healthy individuals. RESULTS: Volume and thickness from several regions differed from the norms. Frontal regions were smaller and thinner, particularly the superior and rostral middle frontal gyri. Further, smaller left rostral middle frontal gyrus, left pericalcarine cortex and right fusiform gyrus were associated with more recent trauma. All subcortical structures were bigger, except the hippocampus. CONCLUSION: These findings suggest a particular brain morphometric signature of PTSD in soldiers. Smaller and thinner frontal and larger subcortical regions support impaired top-down and/or down-regulation of emotional response in PTSD. Finally, the correlation of smaller frontal, temporal and occipital regions with more recent trauma might inform future therapeutic approaches.

Development of a live-cell multi-modal microscopy and spectroscopy platform to study endogenous proteins in the tight and adherens junctions

OBJECTIVE

Increasing scientific evidence suggests that diet and lifestyle can impair the tight and adherens junction (TJ/AJ) structures involved in gut membrane permeability, to the extent that food or bacterial antigens can access the underlying immune system more freely. To study these processes, we will develop a live-cell multi-modal microscopy and spectroscopy platform. This will allow precise measurements of TJ/AJ protein dynamics at the nanoscale combined with label-free Surface-Enhanced Raman Scattering (SERS) & Raman imaging of paracellular molecules. 

METHODS

We are developing a multimodal microscope including five complementary modalities: 1)2-color parallelized REversible Switchable OpticaL Fluorescence Transitions microscopy (RESOLFT), 2)3-color Structured Illumination Microscopy, 3)Single Molecule Localisation Microscopy, 4)Parallelized Raman microscopy, and 5)Structured SERS microscopy. RESOLFT microscopy uses reversibly switchable fluorescent proteins (rsFPs) and can be parallelized to perform high-speed imaging with optimized resolution and contrast. In order to avoid artifacts resulting from overexpression, the proteins of the TJ and AJ will be labelled with rsFPs using CRISPR-Cas9 methods, allowing super-resolution imaging of endogenous protein organization. RESOLFT and 3D-SIM microscopy are combined, giving us the possibility to characterize cell membrane morphology and integrity simultaneously. We will also include SERS and parallelized Raman microscopy, label-free molecular identification techniques, to monitor the leaking of molecules through the TJ/AJ.

RESULTS 

We present the design and preliminary integration of different modalities to the multimodal imaging platform. Using plasmids that express fusion proteins with red and green rsFPs, we performed preliminary RESOLFT experiments to characterize the photophysical properties of the rsFPs. Concurrently, the label-free imaging modalities were characterized on a point-scanning Raman microscope to calibrate the experimental conditions to be used for the implementation of the parallelized acquisition schemes.

CONCLUSION

Once the microscopy platform is complete, it will be possible to simultaneously and in real-time; 1) locate specific TJ/AJ proteins along with cell morphology using RESOLFT and SIM and 2) monitor the components crossing the epithelial membrane using SERS microscopy. This platform will allow us to compare the structure and function of the TJ/AJ proteins in brain and gut tissues.

Résultats dans la vraie vie du passage d'une formulation injectable au mois vers une formulation injectable à action prolongée aux trois mois, Invéga TrinzaMD (étude SeASONS)

OBJECTIF - OBJECTIVE

L’adhésion et la poursuite du traitement à long terme sont des objectifs critiques pour le rétablissement des patients, nécessitant des médicaments efficaces, bien tolérés et le soutien d’une équipe traitante. En 2016, une formulation injectable à action prolongée trimestrielle de palmitate de palipéridone (Invéga TrinzaMD) a été introduite comme un nouvel outil thérapeutique pour le traitement de la schizophrénie. Cette formulation peut favoriser la poursuite et l’engagement du traitement. Pourtant, l’évaluation des résultats cliniques des patients traités avec Invéga TrinzaMD dans des contextes de la vraie vie présentant des comorbidités et un profil clinique parfois plus complexe est justifiée. 

MÉTHODES - METHODS

Cette revue rétrospective des dossiers a examiné les premières utilisations d’Invéga TrinzaMD à travers le Canada depuis 2016. De larges critères d’inclusion/exclusion ont permis l’inclusion de patients généralement exclus des essais contrôlées randomisés. Le résultat principal était un composite incluant à la fois l’arrêt du traitement et la rechute psychotique.

RÉSULTATS - RESULTS

La population étudiée comprend 180 patients, majoritairement des hommes et caucasiens. À 12 mois, il y avait 38 évènements composites, dont 15 arrêts de traitement et 23 rechutes psychotiques. Pendant la période de suivi, il y a eu 71 évènements. Par rapport aux patients qui étaient encore sous Invéga TrinzaMD à la fin du suivi (n = 109), ceux qui ont rechuté (n = 52) présentaient plus fréquemment un trouble d’usage de substances. D’autres caractéristiques, telles que l’origine ethnique, la durée de la maladie psychotique, le diagnostic psychiatrique principal et la gravité de la psychopathologie, étaient similaires dans les deux groupes.

CONCLUSION

Ces résultats soulignent l’importance de reconnaître les écarts entre les patients inclus dans les essais contrôlés randomisés et ceux traités dans nos cliniques. Ils soulignent aussi la nécessité d’études plus naturalistiques pour éclairer le processus de prise de décision partagée.

Assessing mitochondrial health and morphology in Parkinson’s disease with super-resolution microscopy

OBJECTIF - OBJECTIVE

Mitochondria supply the majority of the energy required for metabolism in the brain, and it has become increasingly clear that mitochondrial dysfunction is implicated in Parkinson’s Disease (PD). In this project, we have two primary objectives: (1) observe mitochondrial interactions with other proteins at the nanoscale in order to understand the mechanisms of mitochondrial dysfunction in PD,  and (2) correlate mitochondrial health and morphology to determine whether or not dysfunctional mitochondria tend to have aberrant morphologies.

MÉTHODES - METHODS

We are using optical microscopy to observe mitochondrial interactions, function, and morphology in dopaminergic neurons (DAns). Importantly, these DAns are differentiated from human induced pluripotent stem cells (hiPSCs) that we obtained from a PD patient with a triplication of the SNCA locus (3x SNCA) along with an isogenic control. We used confocal microscopy to characterize mitochondrial health by observing functional markers that measure mitochondrial membrane potential and reactive oxygen species levels.  We characterize mitochondrial morphology with fixed cells by visualizing the mitochondrial outer membrane protein Tom20 with super-resolution STimulated Emission Depletion (STED) microscopy. In addition, we also use STED microscopy in live-cell imaging experiments with STED-compatible mitotracker dyes in order to visualize how mitochondrial dynamics differ between 3xSNCA and control groups.

RÉSULTATS - RESULTS

Preliminary results show that human 3x SNCA DAns have a reduced mitochondrial membrane potentials and higher levels of reactive oxygen species when compared to the isogenic control DAns. Furthermore, our super-resolution images show that the mitochondria in 3xSNCA DAns are smaller and more circular than those in the isogenic control DAns. These data suggest that the mitochondria in this in vitro model of 3xSNCA PD are more fragmented and less healthy than mitochondria in control DAns.

CONCLUSION

The characterization of mitochondrial health, interactions, and morphology in human DAns with super-resolution microscopy will help uncover the mechanisms of neuronal cell death in PD and elucidate the correlation between mitochondrial dysfunction and morphology. Concurrently, the data acquired here will be used to develop an analysis pipeline capable of characterizing cell health from mitochondrial images, and will allow us to conduct high-throughput super-resolution tests of novel neuroprotective treatments for PD.

NOVEL BIOMARKERS FOR THE IDENTIFICATION OF PRODROMAL PARKINSON'S DISEASE

OBJECTIVE

Parkinson's Disease (PD) is correlated with the degeneration of nigral dopaminergic neurons and accumulation of toxic phosphorylated α-synuclein protein aggregates. With multifactorial causes and manifestations, present diagnostic methods identify PD only after the apparition of motor dysfunction, once the dopaminergic neurons are severely affected. Therefore, we undertake an interdisciplinary collaboration to test the electroretinography (ERG) and compositional changes in oral microbiota, which are non-invasive peripheral methods to measure central monoaminergic fluctuations indirectly.

METHODS

An inclusive model of PD, M83 homozygous transgenic mice overexpressing human A53T variant α-synuclein, progressively develops α-synuclein aggregate inclusions parallel to the motor phenotype. These M83 mice and their recommended B6C3F1/J controls undergo a 3-day battery of tests every 2 months for up to 10 months, i.e., motor abnormalities on the accelerating rotarod test, pole test, and an open-field test (n=16). We use the swab technique to sample the oral microbiome and Next-Generation Sequencing to analyze compositional changes. Finally, we measure retinal electrical function via modified ERG protocols and harvest tissue for histological analysis of central and retinal abnormalities. 

We likewise recruit patients with primary, familial, or sporadic PD of different duration, medication treatment, genders, and ages 40-75 years old (n=50). They attend a one-day visit layout with healthy participants to cover the ERG, unstimulated 3 mL saliva collection, oral bacteria Salivette swabs, and complementary pupillometry testing. 

RESULTS

Preliminary data from our PD mouse models revealed that ERG activity and oral microbiota composition are affected. Inner retinal functional changes arise along with α-synucleinopathy within interneurons. Bacteria families that show drastic changes have previous ties in microbiota studies of PD patients (i.e., Comamonadaceae, Carnobacteriaceae, and Propionibacteriaceae). In humans, the ERG also shows a divergence from healthy expectations. 

CONCLUSION

Both biomarkers can detect changes specific to PD model induction in mice. We expect to find similar ERG activity and oral microbiota composition changes in PD patients and control individuals. Although these biomarkers are not meant to be used for neuroprotection, early detection possible through them can provide a wider window for neuroprotective therapeutic intervention in the future. 

Episodic-like memory related activity of hippocampal VIP interneurons in freely behaving mice

Objective - 

Most of the hippocampal vasoactive intestinal polypeptide (VIP) expressing interneurons (INs) specifically target inhibitory neurons, providing a disinhibitory mechanism for ungating the external inputs onto excitatory neurons. CA1 VIP INs have been shown to aid spatial learning and memory but their role in episodic memory is yet unknown.

Methods - 

To address this subject, first, we employed a series of behavioral paradigms to assess various aspects of episodic memory and a unified paradigm to examine What-Where-Which episodic-like memory. Second, we simultaneously performed in vivo calcium imaging of CA1 VIP INs activity in freely behaving mice using wireless fiber photometry.

Results - 

We found that VIP INs were routinely active in all memory tasks throughout the entire period of exploration and exhibited object-modulated activity. Further examination of VIP INs’ activity during various behavioral states revealed that calcium transients (CaTs) during rearing periods were higher compared to object explorations but lower than grooming periods. Additionally, grooming-associated CaTs showed distinct activity pattern and a robust correlation with the duration of grooming.

Conclusion - 

Taken together, these results indicate that VIP INs help to encode object features during episodic-like memory and certain aspects of behavioral states. Ongoing analysis of VIP INs’ activity and experiments using behavioral state-dependent optogenetic perturbations will help us better understand the contribution of disinhibitory interneurons in hippocampal memory processing.