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No description available.

No description available.

No description available.

No description available.

General anaesthetic agents (GAs) have been used clinically for over a century, yet their mechanisms of actions remain a matter of debate. GAs affect a wide range of membrane lipid components but the actual molecular nature underlying the functional effect is yet to be explained. GAs are believed to alter the flow of sodium ions into neurons through the cell membrane. However, exactly how the anaesthetics achieve this is not yet fully understood.

Malignant hyperthermia (MH) is a pharmacogenetic disorder of skeletal muscle which results from anaesthetic-induced disruption to calcium homeostasis. Mutations in the RYR1 and CACN1AS genes encoding the ryanodine (RyR1) and dihydropyridine receptors (DHPR) Ca2+ release channels represent the causative agent in up to 70% of MH susceptible (MHS) populations. The RYR1 screening project has, however, confirmed that up to 30% of MHS patients carry neither RYR1 nor CACN1AS variants and it has been suggested that some modifier genes may exist. Therefore, the aim of the present study is to investigate the relationships between MH susceptibility and genes encoding skeletal muscle Ca2+ channels or accessory proteins. FKBP1A, CASQ1 and JSRP1 genes encoding FKBP12, Calsquestrin1 and JP45 respectively, were selected as candidates for DNA sequencing to screen for potential MH causative mutations. The exonic sequences of DNA samples from MHS patients in whom RYR1 and CACN1AS variants have been excluded were sequenced. DNA sequencing of JSRP1 revealed two variants in the coding region of JP45: p.P108L and p.G150A. The variant p.G150A demonstrated statistically significant increased frequency in MHS individuals compared to MH normal (MHN) populations (p <0.05). Therefore, we hypothesize that p.G150A is associated with MH phenotype. To test this hypothesis, we performed the transmission disequilibrium test which indicated a significant association between the variant p.G150A and MH susceptibility (p <0.05). Western blotting indicated a significant increase in the expression of JP45 in RYR1-HEK293 cells transfected with mutant JSRP1 compared to cells transfected with wild type protein (p <0.05). Furthermore, calcium release measurements demonstrated significant increase in calcium release levels in the RYR1-HEK293 cells expressing mutant JP45 compared with cells expressing wild type JP45 at 1 and 2 mM caffeine concentrations (p < 0.05 and p < 0.01 respectively). In conclusion, JP-45 functions as a modifier protein in the UK MH population.

This work studies the effect of a range of general and local anaesthetics on model membranes, aiming to establish if the addition of anaesthetic promotes changes in the membranes interfacial region or within their internal structure. The models used for biological membranes were simple surfactantiwater systems, which have been previously well studied and characterised. The phases they form are sensitive to changes in composition and interfacial architecture, though they lack the complexity of real membranes. Previous studies on anaesthetics suggest that their effect on membranes was to make them leaky. Therefore, the starting point of this project was to propose that pore formation might be a possible explanation for the anaesthetic action upon model membranes. The study focused on three model membrane systems and used techniques such as NMR and SAXS to obtain information about any structural changes that may occur as a result of anaesthetic addition. The work started with concentrated surfactant water systems that exhibit bilayer like structures (i.e. larnellar phase L. and porous lamellar phase). Two surfactantlwater systems, the ionic surfactant AOT/ 2H20 and the non-ionic surfactant C16EO/ 2H20, were chosen. AOT/2H20 displays a large lamellar phase region stable over a wide range of temperatures and concentration. It was studied to see if the anaesthetic addition triggers the formation of another phase. It was found that anaesthetics induce dehydration, but no dramatic changes in the phase stability were identified for phases which are not close to a transition boundary. In the C16E06/ 2H20 system there are a porous laniellar phase and a classical lamellar phase, which allowed the effect of anaesthetic on the stability of the existing pores to be studied. In this case the anaesthetics fall into two types with respect to their effect: type I that reduces the curvature in the system and another type II that seems to favour higher curvature in the system at higher temperatures with respect to the binary system. The generality of this observation was then tested upon more biologically realistic systems in terms of surfactant to water ratio. In the non-ionic C 10E03/21-120 system, the effect of anaesthetic was studied on the lamellar phase (La) to the sponge phase (L3) topological phase transition in both unbuffered and buffered solutions (pH = 7.4). In the unbuffered system the results mirrored the behaviour in the concentrated systems, whilst in the buffered system both types of anaesthetic tend to dehydrate the surface and suppress interfacial curvature. It seems that, under physiological conditions (pH = 7.4), all anaesthetics have a tendency to reduce interfacial curvature. This contradicts the original hypothesis that anaesthetics may promote pore formation, i.e. "leaky" membranes.

Bien que l’anesthésie générale soit un outil indispensable dans la chirurgie médicale d’aujourd’hui, ses mécanismes sous-jacents précis sont encore inconnus. Au cours de la sédation induite par le propofol les actions anesthésiques à l’échelle microscopique du neurone isolé conduisent à des changements spécifiques à l’échelle macroscopique qui sont observables comme les signaux électroencéphalogrammes (EEG). Pour une concentration faible en propofol, ces changements caractéristiques comprennent une augmentation de l’activité dans les bandes de fréquence delta (0.5-4 Hz) et alpha (8 13 Hz) dans la région frontal, une l’activité augmentée de delta et une l’activité diminuée de alpha dans la région occipitale. Dans cette thèse, nous utilisons des modèles de populations neuronales thalamo-corticales basés sur des données expérimentales. Les effets de propofol sur les synapses et sur les récepteurs extra-synaptiques GABAergiques situés dans le cortex et le thalamus sont modélisés afin de comprendre les mécanismes sous-jacents aux changements observés dans certaines puissances de l’EEG spectrale. Il est démontré que les modèles reproduisent bien les spectrales caractéristiques observées expérimentalement. Une des conclusions principales de ce travail est que l’origine des delta rythmes est fondamentalement différente de celle des alpha rythmes. Nos résultats indiquent qu’en fonction des valeurs moyennes des potentiels de l’état du système au repos, une augmentation ou une diminution des fonctions de gain thalamo-corticale résulte respectivement en une augmentation ou une diminution de alpha puissance. En revanche, l’évolution de la delta puissance est plutôt indépendant de l’état du système au repos; l'amélioration de la puissance spectrale de delta bande résulte de l’inhibition GABAergique synaptique ou extra-synaptique pour les fonctions de gain non linéaire à la fois croissante et décroissante. De plus, nous cherchons à identifier les paramètres d’un modèle de thalamo-corticale en ajustant le spectre de puissance de modèle pour les enregistrements EEG. Pour ce faire, nous considérons la tâche de l’estimation des paramètres dans les modèles qui sont décrits par un ensemble d’équations différentielles ordinaires ou bien stochastiques avec retard. Deux études de cas portant sur des données pseudo-expérimentales bruyantes sont d’abord effectuées pour comparer les performances des différentes méthodes d’optimisation. Les résultats de cette élaboration montrent que la méthode utilisée dans cette étude est capable d’estimer avec précision les paramètres indépendants du modèle et cela nous permet d’éviter les coûts de calcul des intégrations numériques. En considérant l’ensemble, les conclusions de cette thèse apportent de nouveaux éclairages sur les mécanismes responsables des changements spécifiques qui sont observées pendant la sédation propofol-induite dans les modèles de EEG. / Although general anaesthesia is an indispensable tool in today’s medical surgery, its precise underlying mechanisms are still unknown. During the propofol-induced sedation, the anaesthetic actions on the microscopic single neuron scale lead to specific changes in macroscopic-scale observables such as electroencephalogram (EEG) signals. For low concentration of propofol these characteristic changes comprised increased activity in the delta (0.5-4 Hz) and alpha (8-13 Hz) frequency bands over the frontal head region, but increased delta and decreased alpha power activity over the occipital region. In this thesis, we employ thalamo-cortical neural population models, and based on the experimental data, the propofol effects on the synaptic and extrasynaptic GABAergic receptors located in the cortex and thalamus are modelized to understand the mechanisms underlying the observed certain changes in EEG-spectral power. It is shown that the models reproduce well the characteristic spectral features observed experimentally. A key finding of this work is that the origin of delta rhythm is fundamentally different from the alpha rhythm. Our results indicate that dependent on the mean potential values of the system resting states, an increase or decrease in the thalamo-cortical gain functions results in an increase or decrease in the alpha power, respectively. In contrast, the evolution of the delta power is rather independent of the system resting states; the enhancement of spectral power in the delta band results from the increased synaptic or extra-synaptic GABAergic inhibition for both increasing and decreasing nonlinear gain functions. Furthermore, we aim to identify the parameters of a thalamo-cortical model by fitting the model power spectrum to the EEG recordings. To this end, we address the task of parameter estimation in the models that are described by a set of stochastic ordinary or delay differential equations. Two case studies dealing with noisy pseudo-experimental data are first carried out to compare the performance of different optimization methods. The results of this elaboration show that the method used in this study is able to accurately estimate the independent model parameters while it allows us to avoid the computational costs of the numerical integrations. Taken together, the findings of this thesis provide new insights into the mechanisms responsible for the specific changes in EEG patterns that are observed during propofol-induced sedation.