Introduction: Neurological complications and stroke in association with cardiac surgery is a serious problem. The stroke event can occur during surgery (early stroke) or in the postoperative period with a symptom free interval (delayed stroke). Particle embolization due to aortic manipulation during surgery has been suspected as a mechanism for early stroke. The present thesis address mechanisms and survival effects of stroke both clinically (I-III) and experimentally (IV-V). Methods: Study I) Within a cohort of 2641 consecutive cases, a group of cardiac surgery patients with stroke and evaluated by computed tomography (CT) were studied (n=77). CT-findings were analyzed in relation to stroke symptoms. Study II) Data from 9122 patients undergoing coronary surgery were analyzed. Records of patients with any signs of neurological complications were reviewed to extract 149 subjects with stroke at extubation (early, 1.6%) versus 99 patients having a free interval (delayed, 1.1%). Early and delayed stroke were evaluated separately. Independent risk factors for stroke were analyzed by logistic regression and survival by Cox regression (9.3 years median follow-up). Study III) Patients with early (n=223) and delayed stroke (n=116) were identified among 10809 patients undergoing cardiac and aortic surgery, both groups exposed to cardiopulmonary bypass. Stroke patients were subdivided by the hemispheric location of lesions. Subgroups were compared and their associated pre- and peroperative variables and survival were analyzed. Study IV) Aortic cross-clamp manipulation was studied in a human cadaveric perfusion model. The pressurized aorta was repeatedly cross-clamped and washout samples were collected before and after clamp maneuvers. Particles in the washout samples were evaluated by microscopy and by digital image analysis. Study V) Pig aortas were pressurized and cannulated. Washout samples were collected before and after cannulation (n = 40). Particles were deposited onto a 10-μm filter to be evaluated by microscopy and digital image analysis. Results: Study I) In the group of patients exposed to routine cardiac surgery (i.e., clamping and cannulation) and with early stroke, right-hemispheric lesions were more frequent than of the contra-lateral side (P=0.005). Patients with aortic dissections had a strong dominance of bilateral findings, which was different from the unilateral pattern in the routine-surgery group (P<0.001). Study II) Early and delayed stroke did not share any risk factors. Both early and delayed stroke explained mortality in the early postoperative period (P<0.001, P<0.001 respectively) but also at long term follow-up (P=0.008, P<0.001 respectively). For patients surviving their first postoperative year, delayed but not early stroke influenced long-term mortality (P=0.001 and P=0.695, respectively). Study III) Stroke lesions in association to cardiac surgery were near exclusively ischemic. Early stroke had a preponderance for right-hemispheric lesions (P=0.009). In contrast, patients with early stroke that had undergone surgery of the aorta with circulatory arrest showed a pattern with more bilateral lesions compared to ‘cardiac-type’ operations (P<0.001). Patients with bilateral lesions had a dramatically impaired survival compared to those with unilateral lesions (P<0.001). Study IV) In the cadaveric perfusion model, cross-clamping produced a significant output of particles, which was seen for size intervals of 1 mm and smaller (P=0.002 to P=0.022). In all size intervals the particle output correlated with the degree of overall aortic calcification (P =0.002 to P=0.025). Study V) At cannulation of the pig aorta, more particles were noted after cannulation compared to before the maneuver (P<0.001). This increase included small (<0.1 mm, P<0.001) and intermediate-size particles (0.1-0.5 mm, P< 0.001). Particles above 0.5 mm were few and were not associated with cannulation. Conclusions: The influence of stroke on mortality was devastating, for both early and delayed stroke. These two stroke groups had obvious differences in both their risk factors and their hemispheric distribution. It is here emphasized that early and delayed stroke should be considered as two separate entities with suggested mechanistic differences. Ischemic lesions accounted for near all stroke events seen in association to cardiac surgery. For early stroke, these were mostly located within the right hemisphere. Results from the experimental studies underscore microembolic risks associated with aortic manipulation.

No description available.

Significant advancements in the field of membrane protein crystallography have provided in recent years invaluable images of transporter structures. These structures, however, are static and require complementary kinetic insight to understand how their mechanisms work. Electrophysiological studies of transporters permit the high quality kinetic measurements desired, but there are significant difficulties involved in analyzing and interpreting the data. Current methods allow a variety of kinetic parameters to be measured but there is a disconnect between these parameters and a fundamental understanding of the carrier. The intent of this research was to contribute new tools for studying the electrogenic kinetics of membrane transport proteins, to understand the link between these kinetics and the carrier, and to ultimately understand the mechanisms involved in transport. In this vein, two projects are explored covering two important kinetic time domains, transient and steady-state. The transient project studies the conformational changes of the unloaded carrier of SGLT1 through a multi-exponential analysis of the transient currents. Crystal structures have potentially identified a gated rocker-switch mechanism and the transient kinetics are used to support and study this kinetically. A protocol taking advantage of multiple holding potentials is used to measure the decay time constants and charge movements for voltage jumps from both hyperpolarizing and depolarizing directions. These directional measurements provide insight into the arrangement of the observed transitions through directional inequalities in charge movement, by considering the potential for a slow transition to hide a faster one. Ultimately, four carrier decays are observed that align with the gated rocker-switch mechanism and can be associated one-to-one with the movement of a gate and pore on each side of the membrane. The steady-state project considers a general theoretical model of transporter cycling. Recursive patterns are identified in the steady-state velocity equation that lead to a broad understanding of its geometric properties as a function of voltage and substrate concentration. This results in a simple phenomenological method for characterizing the I–V curves and for measuring the kinetics of rate limiting patterns in the loop, which we find are the basic structures revealed by the steady-state velocity.

The structure of afferent neurons is pseudounipolar. Studies suggest that they relay action potentials (APs) to both directions of the T-junctions to reach the cell body and the spinal cord. Moreover, the somata are electrically excitable and shown to be able to transmit the signals to associated satellite cells. Our study demonstrates that this transmission can go further and pass onto passive neighbouring somata, if they are in direct contact with same satellite cells. The neurons activate the satellite cells by releasing ATP. This triggers the satellite cells to exocytose acetylcholine to the neighbouring neurons. In addition, the ATP inhibits the nicotinic receptors of the neurons by activating P2Y receptors and initiating the G-protein-mediated pathway, thus reducing the signals that return to the neurons that initiated the signals. This “sandwich synapse” represents a unique pathway by the ectopic release between the somata and the satellite cells.

The structure of afferent neurons is pseudounipolar. Studies suggest that they relay action potentials (APs) to both directions of the T-junctions to reach the cell body and the spinal cord. Moreover, the somata are electrically excitable and shown to be able to transmit the signals to associated satellite cells. Our study demonstrates that this transmission can go further and pass onto passive neighbouring somata, if they are in direct contact with same satellite cells. The neurons activate the satellite cells by releasing ATP. This triggers the satellite cells to exocytose acetylcholine to the neighbouring neurons. In addition, the ATP inhibits the nicotinic receptors of the neurons by activating P2Y receptors and initiating the G-protein-mediated pathway, thus reducing the signals that return to the neurons that initiated the signals. This “sandwich synapse” represents a unique pathway by the ectopic release between the somata and the satellite cells.

Significant advancements in the field of membrane protein crystallography have provided in recent years invaluable images of transporter structures. These structures, however, are static and require complementary kinetic insight to understand how their mechanisms work. Electrophysiological studies of transporters permit the high quality kinetic measurements desired, but there are significant difficulties involved in analyzing and interpreting the data. Current methods allow a variety of kinetic parameters to be measured but there is a disconnect between these parameters and a fundamental understanding of the carrier. The intent of this research was to contribute new tools for studying the electrogenic kinetics of membrane transport proteins, to understand the link between these kinetics and the carrier, and to ultimately understand the mechanisms involved in transport. In this vein, two projects are explored covering two important kinetic time domains, transient and steady-state. The transient project studies the conformational changes of the unloaded carrier of SGLT1 through a multi-exponential analysis of the transient currents. Crystal structures have potentially identified a gated rocker-switch mechanism and the transient kinetics are used to support and study this kinetically. A protocol taking advantage of multiple holding potentials is used to measure the decay time constants and charge movements for voltage jumps from both hyperpolarizing and depolarizing directions. These directional measurements provide insight into the arrangement of the observed transitions through directional inequalities in charge movement, by considering the potential for a slow transition to hide a faster one. Ultimately, four carrier decays are observed that align with the gated rocker-switch mechanism and can be associated one-to-one with the movement of a gate and pore on each side of the membrane. The steady-state project considers a general theoretical model of transporter cycling. Recursive patterns are identified in the steady-state velocity equation that lead to a broad understanding of its geometric properties as a function of voltage and substrate concentration. This results in a simple phenomenological method for characterizing the I–V curves and for measuring the kinetics of rate limiting patterns in the loop, which we find are the basic structures revealed by the steady-state velocity.

The guided wave technique is commonly used for rapidly long-range pipeline inspection without removing the insulation of pipes. The torsional mode T(0,1) of the guided waves is usually generated to detect the defects in pipelines, since it has the advantage of being non-dispersive across the whole frequency range. However, a large number of pipelines are carrying fluid, wrapped with the coating material, and supported with clamp for the necessary manufacturing process in refinery and petro-chemical industrials. When these works are employed on the pipeline, the propagating guided waves may vary with the contents of material and how well the material compact on the pipe. Some energy of the incident guided wave in the pipe wall may leak into inside of contents or outside of wrapped materials and reduce the wave propagation distance. The effect of the fluid-filled pipe, the wrapped pipe, and the clamp support mounted on the pipe for guided wave propagation is investigated by both simulative and experimental methods. The wave structure of the T(0,1) mode in the pipes is analyzed by using the DISPERSE software for various cases to evaluate its influence to the guided wave propagation on the pipe. The amplitudes of the reflected signals from various features on the pipe are also measured using pipe screening system for calculating the attenuation of guided waves due to the features. The trend for the results is in good agreement between the experiments andpredictions for all cases of researches in this dissertation. It is found that the low viscosity liquid deposited in the pipe, such as water, diesel oil, and lubricant, has no effect on the torsional mode; while the high viscous of the fuel oil deposited in the pipe attenuates the reflection signal heavily for the pipe carrying fluid. In addition, both the full-filled and half-filled contents in the pipe are also studied in this case. The effects of the half-filled are the same as the full-filled results obtained. For the pipe wrapped with the coated material, the adhesive strength of the coated material is strong, such as bitumen and polyethylene; the attenuation of the guided waves is high; and there is almost no effect for mineral wool coating. Furthermore, the traveling distance of the guided waves in the pipe is also evaluated for various cases of the coated materials. The results indicate that the higher attenuation of the guided waves for the coated material, the shorter of the traveling distance in the pipe. For the clamp support mounted on pipe, the attenuation of the guided waves for the clamp support with a rubber gasket in between the pipe and the clamp is heavier than the case of clamp support without the rubber gasket is. Furthermore, the higher torque setting on the clamp (with or without the rubber gasket), the higher amplitude of the reflected signal is measured for the guided wave propagation. The effect of the frequency excitation is additionally demonstrated in this dissertation. It is noted that the higher amplitude of the reflected signal, the lower frequency excitation; moreover, theresonant effect is observed in the case of the clamp support with rubber gasket during the torque setting in the experiments. Good agreement has been obtained between the experiments and theoretical calculations of this effect.

Pendant les états naturels d'activité in vivo, les neurones neocorticaux sont sujets à une conductance membranaire forte et fluctuante. Cependant, les propriétés intégratives des neurones ne sont pas connues pendant ces états de “haute conductance” (HC). Nous avons (1) caractérisé le lien entre la dynamique des conductances et l'initiation du potentiel d'action (PA) dans les neurones corticaux dans les états HC; (2) comparé différents modèles de réponse de PA (PSTH) pendant ces états. Nous distinguons deux modes de décharge, selon que le PA est évoqué par une augmentation d'excitation ou par une diminution d'inhibition. Nous avons proposé une nouvelle méthode pour calculer les “spike-triggered average” (STA) des conductances à partir du Vm, testé cette méthode numériquement et in vitro, ainsi que appliqué cette méthode aux enregistrements in vivo. Nous démontrons que les PAs inhibiteurs sont majoritaires chez le chat éveillé, ce qui révèle un rôle majeur de l'inhibition.

Thèse d'exercice : Médecine. Biologie Médicale : Paris 12 : 2007. / Titre provenant de l'écran-titre. 55 f. : ill. Bibliogr. f. 50-54.

In the heart, electrical signaling is responsible for its rhythmicity and is necessary to initiate muscle contraction. The net electrical activity in a cardiac myocyte during a contraction cycle is observed as the action potential (AP), which describes a change in membrane potential as a function of time. In ventricular cardiac myocytes, voltage-gated sodium channels (Nav) and voltage-gated potassium channels (Kv) play antagonistic roles in shaping the AP with the former initiating membrane depolarization and the latter repolarizing it. Functional changes in the primary cardiac Nav isoform, Nav 1.5, or any one of the many Kv isoforms expressed in the ventricle, as evidenced by those characterized in various congenital and/or acquired etiologies, can lead to severe cardiac pathologies. Nav and Kv are large transmembrane proteins that can be extensively post-translationally modified through processes that include glycosylation. The sequential glycosylation process typically ends with negatively charged sialic acid residues added through trans-Golgi sialyltransferase activity. Sialyltransferases belong to a much larger group of glycogene products that number in the hundreds and are responsible for creating a complex and variable glycan profile (glycome) unique to different cell types and tissues. Sialic acids impact Nav and Kv function likely by contributing to the extracellular surface potential and thereby causing channels to gate following smaller depolarizations. Additionally, developmentally regulated sialylation contributes to cardiac myocyte excitability in the neonatal mouse atria. However, little is understood concerning how the glycosylation machinery (glycogene products) influences cell and tissue electrical signaling. The sialytransferase Β-galactoside α-2,3-sialyltransferase 4 (ST3Gal4) adds sialic acids to galactose residues of N- and O-linked glycans through α-2,3-linkgages. ST3Gal4 is uniformly expressed throughout the chambers and developmental stages of the heart and therefore is likely a useful target to question whether and how glycosylation impacts these events. Additionally, diseases of glycosylation often cause symptoms that are consistent with changes in excitability that include arrhythmias and seizures. Congenital disorders of glycosylation lead to variably reduced glycoprotein and glycolipid glycosylation. However, because sialic acids are typically the terminal residues added to glycan structures, disease-related reduced glycosylation often leads to fewer sialic acids being attached. In addition, Chagas disease, which results in pathological changes in cardiac electrical function, may reduce sialic acids directly. Because of this, the ST3Gal4-/- strain was also used to investigate the role of glycosylation in the pathological cardiac electrical remodeling often associated with these diseases. The methodologies included cellular, tissue and whole-animal electrophysiology as well as biochemical assays. The data indicate that deletion of ST3Gal4 significantly affects Nav sialylation and gating with no change in maximum current density or protein expression. ST3Gal4 deletion also depolarizes the activation gating of both voltage-dependent kinetic components of repolarization found in the mouse ventricle: Ito and IKslow; however unlike the effect on INa, ST3Gal4 gene deletion causes a reduction in the peak IK density. Protein expression of the putative Kv isoforms responsible for Ito and IKslow was variably affected by ST3Gal4 gene deletion with Kv1.5 and Kv4.2 demonstrating no differences in protein densities. Contrastingly, a small but significant reduction in Kv2.1 protein from ST3Gal4-/- ventricular tissue was observed. In addition to effects on Nav and Kv activity, ST3Gal4 expression is necessary for normal cellular electrical signaling as demonstrated by a reduction in cellular refractory period and alterations in AP waveforms that include a slowing of cellular conduction and an extension of AP duration in ventricular myocytes from ST3Gal4-/- mice. Concurrent with aberrant excitability at the cellular level, the ST3Gal4-/- left ventricular epicardium demonstrated a reduced refractory period and was more susceptible to arrhythmias as observed through optical mapping studies. Additionally, ECGs of ambulatory ST3Gal4-/- mice demonstrated that deletion of the gene causes modest aberrant conduction under basal conditions and, in preliminary studies, appears to increase susceptibility to arrhythmias following a cardiac challenge, in the form of a low dosage of the Β-adrenergic agonist isoproterenol, suggesting a reduction in repolarization reserve in ST3Gal4 hearts. Based on the data reported here, it is apparent that relatively minor perturbations in the cardiac glycome cause significant changes in cardiac electrical signaling. These data highlight the role of glycosylation in normal physiology and underscore it as an important mediator in diseases where it may be altered.