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Une majorité de neurones du noyau préoptique médian (MnPO) chez le rat sont sensibles aux variations de la concentration extracellulaire de sodium (Na⁺). L'objectif de cette étude était de déterminer le mécanisme cellulaire impliqué dans la détection du Na⁺ dans le MnPO. Pour étudier ce mécanisme, des enregistrements électrophysiologiques ont été réalisés sur des neurones dissociés et des neurones présents dans une tranche de l'hypothalamus. On a d'abord démontré la présence de "senseurs" de Na⁺ dans les neurones du MnPO. Ensuite, on a déterminé que la détection du Na⁺ extracellulaire était spécifiquement attribuée à un flux d'ions Na⁺ à travers un canal de fuite bloqué par le rubidium. Les propriétés de ce canal étaient similaires à celles d'un canal sodique. Même si cela reste à confirmer, la présence d'ARNm codant pour le canal Nax dans le MnPO fait de ce canal un candidat potentiel.

Tableau d’honneur de la Faculté des études supérieures et postdoctorales, 2012-2013. / Les canaux sodiques permettent la génération de potentiels d'action. Ceux-ci sont nécessaires à l'excitabilité cellulaire. Ainsi, les canaux sodiques permettent la génération et la propagation de l'influx nerveux. Ils permettent aussi la contraction des muscles squelettiques et du muscle cardiaque. Un dysfonctionnement dans ces canaux peut être à l'origine d'un vaste spectre de phénotypes pathologiques neuronaux tels que l'épilepsie, la douleur chronique, l'incapacité à ressentir la douleur, neuromusculaires tel la paramyotonie congénitale ou cardiaques tels certaines arythmies cardiaques ou la cardiomyopathie dilatée. L'étude des altérations des propriétés biophysiques des canaux sodiques permet de fournir un point de départ dans l'étude des mécanismes menant aux phénotypes pathologiques en plus d'orienter les traitements. Les travaux présentés dans ce mémoire portent sur une mutation permettant la création d'un pore alternatif dans le canal sodique. Nous avons caractérisé les propriétés du pore et les régions de la protéine où il est possible de créer un tel pore.

Les canaux sodiques voltages dépendants sont responsables de la phase ascendante du potentiel d'action et jouent un rôle important dans l'initiation du potentiel d'action et de sa propagation dans les cellules excitables. Les canaux sodiques sont constitués d'une sous-unité a formant le pore, associée à plusieurs sous-unités p auxiliaire. La sous-unité a est formée de quatre domaines homologues (I-IV), chacun d'eux étant composé de six segments hydrophobiques (S1-S6). Seule la principale sous-unité a est nécessaire à l'expression fonctionnelle, mais les sous-unités p ont une action modulatrice sur la localisation et les propriétés fonctionnelles de la sous-unité a. Suite à des lésions du nerf, les neurones de ganglions dorsaux deviennent hyperexcitables. Les ganglions dorsaux expriment une combinaison de canaux sodiques de cinétique rapide sensibles à la TTX et de canaux de cinétique lente résistants à la TTX. Ces canaux jouent un rôle important dans la douleur chronique. Deux canaux sodiques distincts ont été clones à partir des ganglions dorsaux, Navl .7 et Nav1.8. Ils codent respectivement pour les canaux sensibles et résistants à la TTX. Même si Nav1.8 exprime bien dans les ovocytes de Xenopus laevis, les essais dans les lignées cellulaires de mammifères n'ont pas abouti. Matériels et Méthodes: Dans cette étude, nous avons recherché les déterminants de Nav1.8 dans les cellules de mammifères en utilisant une combinaison d'immunobuvardage et d'électrophysiologie. Résultats: Notre étude révèle que l'expression faible de Nav1.8 dans les cellules de mammifère est reliée à un défaut de trafficking qui emprisonne la protéine canal dans le réticulum endoplasmique. Si on incube les cellules tsA201 avec l'anesthésique local lidocaine, l'expression de Nav1.8 est considérablement augmentée. Les propriétés biophysiques du courant Nav1.8 exprimé dans un système d'expression hétérologue reproduisent celles de la composante du courant Na+ résistant à la TTX mesuré dans les neurones des ganglions dorsaux de rat. Conclusion: Nos résultats indiquent que la lidocaine, un bloqueur du canal sodique, agit comme un chaperon chimique qui stabilise le canal Nav1.8 dans sa conformation native et augmente donc le niveau d'expression de Nav1.8. / Voltage-gated sodium channels are responsible for the rising phase of action potentials. They play an important role in the initiation and propagation of action potential in excitable cells. Sodium channels consist of a pore-forming a subunit, associated with auxiliary p subunit. One a subunit consists of four homologous domains (I-IV), each composed of six hydrophobic segments (S1-S6). Dorsal root ganglion (DRG) neurons express a combination of rapidly gating TTX-sensitive and slowly gating TTX-resistant Na current. Two distinct Na channels have been cloned from the DRG that appear to account for the majority of this Na current. The Nav1.7 and Nav1.8 channels encode for rapidly inactivating TTX-sensitive and slowly inactivating TTX-resistant Na currents respectively. These two channels contribute significantly in pain pathways. Nav1.7 has a highly significant role in determining inflammatory pain thresholds, and Nav1.8 is the decisive role in maintaining the hypersensitivity of primary afferent neurons following nerve injuries. Although the cloned Nav1.8 expresses well in Xenopus oocytes, attempts the express the Nav1.8 channel in mammalian expression Systems have been met with limited success even in the presence of known sodium channel accessory p subunits. Methods: In this study we investigated the important determinants of Nav1.8 expression in mammalian cells using a combination of immunochemistry and patch-clamp technique. Results: Our study revealed that the low expression of Nav1.8 in mammalian cells is related to a trafficking defect that traps the channel protein in the endoplasmic reticulum. Incubating the tsA201 cells with the local anesthetic lidocaine dramatically enhances Nav1.8 expression. The biophysical properties of the heterologously expressed Nav1.8 current accurately reproduce those of the TTX-resistant component of Na current recorded from native DRG neurons. Conclusion: Our data indicates that lidocaine, a sodium channel blocker, can act as a chemical chaperone that stabilizes Nav1.8 channels in its native conformation therefore increases the expression of Nav1.8.

L’hydrolyse contrôlée et progressive est une des solutions technologiques pour traiter le sodium tritié présent dans divers composants d’un Réacteur à Neutrons Rapides. Une étude expérimentale a été réalisée pour analyser et comprendre les phénomènes physico-chimiques mis en jeu lors de cette hydrolyse, fortement exothermique, et étudier l’influence des paramètres opératoires sur la répartition HT /HTO au sein de l’effluent liquide et de l’effluent gazeux générés. Les deux facteurs prédominants sont l’activité totale du sodium traité et le flux énergétique (J/s) dégagée par la réaction. Un modèle phénoménologique de l’hydrolyse de sodium tritié est proposé pour synthétiser les connaissances acquises et servir d’aide à la prédiction de la composition en tritium dans les effluents générés en vue de leur traitement. / Within the framework of the decommissioning of fast reactors, several processes are under investigation regarding sodium disposal. One of them rests on the implementation of the sodium-water reaction (SWR), in a controlled and progressive way, to remove residual sodium containing impurities such as sodium hydrides, sodium oxides and tritiated sodium hydrides. Such a hydrolysis releases some amount of energy and produces a liquid effluent, composed of a solution of soda, and a gaseous effluent, composed of hydrogen, steam and an inert gas. The tritium, originally into the sodium as a soluble (T-) or precipitate form (NaT), will be distributed between the liquid and gaseous effluent, and according to two chemical forms, the tritium hydride HT and the tritiated water HTO. HTO being 10,000 times more radiotoxic than HT, a precise knowledge of the mechanisms governing the distribution of tritium is necessary in order to estimate the exhaust gas releases and design the process needed to treat the off-gas before its release into the environment. An experimental study has been carried out in order to determine precisely the phenomena involved in the hydrolysis. The influence of the experimental conditions on the tritium distribution has been tested. The results of this study leaded to a phenomenological description of the tritiated sodium hydrolysis that will help to predict the composition of the effluents, regarding tritium.

The practical method of applying the variation principle to the calculation of the energy of an atom demands a trial function which contains variable parameters. The previous work done using this approach was based on the use of some combination of hydrogenic wave functions containing parameters inserted in appropriate places. The present calculation of the energy of the eleven-electron atom has been brought about using this method.

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

The goal of this thesis was to investigate the interplay between sodium, glycosaminoglycans, vascular stiffness, and hypertension. In contrast to the traditional view of salt-dependent hypertension, recent studies have found that sodium accumulation can occur without commensurate fluid retention. Researchers hypothesise that this sodium is stored non-osmotically via association with negatively charged glycosaminoglycans (GAGs) in the extracellular matrix. The interaction of sodium and GAGs, the influence of sodium on GAG production, and the ability of GAGs to affect vascular stiffness are of key interest. This thesis first investigates the link between hypertension, vascular stiffness, and GAGs in ex vivo human aortae. Aortae from hypertensive donors were found to be stiffer than normotensive controls even after controlling for both pressure and age, a novel finding in humans. In these aortae, hypertension was associated with GAG remodelling, but not with changes in total GAG content. Next, an interventional rat study is presented to examine the effects of dietary salt on vascular stiffness and GAGs, and to distinguish between salt-dependent and blood pressure-dependent effects. In vivo vascular stiffness was found to be salt-dependent but pressure-independent, with ex vivo stiffness unaffected by salt. Ex vivo stiffness was also independent of aortic GAG content, similar to the human aortae described previously. GAG content in the skin was both salt-dependent and pressure-dependent. Finally, this thesis closes with an interventional study in humans. This study was designed to examine the effects of diuretic-induced salt loss on sodium storage, GAGs, and haemodynamics. An eight-day diuretic course corresponded to a ~10% reduction in skin sodium content, without associated water loss or cardiovascular changes. GAG mRNA expression was decreased in the skin, suggesting reduced GAG content. Pilot work from this study supports the use of 23Na MRI as a non-invasive measurement of skin sodium, but only for pre- vs post-treatment comparisons rather than absolute quantification. In conclusion, this thesis demonstrates that both salt and blood pressure influence GAG accumulation and distribution, but that GAGs do not directly affect vascular stiffness. However, GAGs do play a direct role in osmotically inactive sodium storage, which may modulate development of hypertension.

The epithelial sodium channel (ENaC) made up of α, β and γ subunits is located at the apical membrane of polarised epithelia and mediates transport of sodium ions into the cells. Tight control of ENaC function is essential for maintaining sodium homeostasis, blood volume and blood pressure. Controlling the number of active channels present at the cell surface appears to be critically important in regulating ENaC activity. The neural precursor cell expressed developmentally down-regulated gene 4 (Nedd4) family of proteins (eg. Nedd4-2) ubiquitinate ENaC and decrease its cell surface expression. The activity of Nedd4-2 is modulated by serum and glucocorticoid-induced kinase (SGK), which phosphorylates Nedd4-2 and increases cell surface expression of ENaC. The c̲o̲pper m̲etabolism gene M̲URR1 d̲omain 1 (COMMD1) protein is a recently identified ENaC binding partner and negative regulator of channel activity. Studies by other groups suggest that COMMD1 is also involved in the processes of intracellular protein trafficking and ubiquitin-dependent protein degradation. The aims of this study were 1). To characterise the interactions between COMMD1 and ENaC. 2). To identify the mechanism(s) by which COMMD1 down-regulates ENaC activity. Here protein-protein interaction studies were used to show that a recently identified conserved C-terminal domain (the COMM domain) in COMMD1 is essential for its binding to ENaC. The binding site for COMMD1 in βENaC was found to be located in its N-terminal domain. COMMD1 was shown to down-regulate ENaC by increasing ubiquitin modification of ENaC and by decreasing the cell surface population. COMMD1 was found to interact with SGK and formed a complex with SGK and Nedd4-2. Ussing chamber studies of Na⁺ transport showed that COMMD1 attenuated the stimulation of ENaC by SGK and abolished insulin-stimulated ENaC current in epithelial cells. Conversely, knock-down of COMMD1 increased ENaC current in mammalian epithelial cells. These data suggest that COMMD1 plays a role in regulating ENaC activity in epithelial cells and its effect is likely mediated via SGK. In addition COMMD1 was found to bind to the adaptor protein subunit [mu]2. Mutations in COMMD1 that disrupt its interaction with [mu]2 impair its ability to decrease cell surface expression of ENaC in Cos-7 cells, therefore COMMD1 may also have a role in the endocytosis of ENaC by linking cell surface ENaC to the clathrin-dependent endocytosis machinery. In summary, this study investigated the interactions between COMMD1 and ENaC and identified that the SGK/Nedd4-2 pathway is involved in the COMMD1-mediated ubiquitination and down-regulation of ENaC activity.