No description available.

<p>Two thirds of patients with diabetes avoid regularly monitoring their blood glucose levels because of the painful and invasive nature of current blood glucose detection. As an alternative to blood sample collection, exhaled breath condensate (EBC) has emerged as a promising non-invasive sample from which to monitor glucose levels. However, the inconsistency in the methods used to collect EBC significantly impacts the reliability of reported analyte concentrations in EBC. Furthermore, this dilute sample matrix requires a highly sensitive glucose biosensor to enable robust and accurate glucose detection at the point-of-care. Together, a reliable collection method and sensitive detection system can enable accurate modeling of glucose transport from blood to breath that is reflective of airway glucose homeostasis.</p> <p> I address this research gap by simultaneously designing a standardized EBC collection method that allows for separation of dead space and alveolar air and developing a competitive fluorescent biosensor that can resolve micromolar glucose concentrations changes. First, I develop a low-cost, automated condenser that selectively collects exhaled breath that has been exchanged with lung fluid based on the detection of higher breath temperatures that are characteristic of the lower respiratory regions. Using this device, I investigate the relationship between blood and EBC glucose in diabetic and normoglycemic human subjects. Next, I engineer the exquisitely sensitive <i>E. coli</i> glucose binding protein (GBP) with a chemo-enzymatic tag to selectively conjugate it to highly photostable quantum dots (QDs). Finally, I take advantage of the competitive binding of glucose (K_D=0.35 µM) and galactose (K_D=1.4 µM) to GBP to develop a fluorescent glucose biosensor using the GBP-QD conjugate.</p>

L’hyperglycémie est détectée et intégrée au niveau de l’hypothalamus médio-basal (MBH) qui inhibe la prise alimentaire et déclenche la sécrétion d’insuline. Le MBH renferme des neurones spécialisés gluco-sensibles (GS) qui détectent directement ou indirectement des variations de la concentration extracellulaire en glucose. Dans une première étude, nous suggérons que la détection indirecte du glucose par les neurones GS hypothalamiques repose sur la libération d’endozépines par les astrocytes, un gliotransmetteur connu pour inhiber la prise alimentaire en réponse à l’hyperglycémie. Nous travaux montrent que les endozépines activent spécifiquement les neurones à pro-opiomélanocortine (POMC) du MBH pour générer leur effet anorexigène. Dans une seconde étude, nous montrons que la détection directe de l’hyperglycémie implique les neurones hypothalamiques dits « high gluco-excited » (HGE). Grâce à des approches pharmacologiques et génétiques, nous mettons en évidence que les canaux redox sensibles Transient Receptor Potential Canonical 3 et 4 (TRPC3/4) sont fondamentaux pour la détection du glucose par les neurones HGE in vitro, la stimulation de la sécrétion d’insuline et la diminution de la prise alimentaire en réponse à l’hyperglycémie cérébrale in vivo. De plus, nos travaux démontrent que les canaux TRPC3 du MBH jouent un rôle clef dans le contrôle de l’homéostasie énergétique. Les travaux de cette thèse permettent de mettre en évidence deux nouveaux mécanismes de détection hypothalamique de l’hyperglycémie : l’un reposant sur l’implication des canaux TRPC3/4 dans les neurones HGE et l’autre proposant les endozépines astrocytaires comme relai du signal « glucose » aux neurones POMC. / Hyperglycemia is detected and integrated by the mediobasal hypothalamus (MBH) which, in turn, inhibits food intake and triggers insulin secretion. The MBH houses specialized glucose-sensitive (GS) neurons, which directly or indirectly modulate their electrical activity in response to changes in glucose level. In a first study, we hypothesized that indirect detection of glucose by MBH GS neurons involves the secretion of endozepine by astrocytes, a gliotransmitter known to inhibit food intake in response to hyperglycemia. The present work shows that endozepines selectively activate anorexigenic MBH pro-opiomelanotortine (POMC) neurons. In the second study, we show that the direct detection of increased glucose level involves hypothalamic glucose-excited (HGE) neurons. Using pharmacological and genetic approaches, we demonstrate that the redox-sensitive Transient Receptor Potential Canonical 3 et 4 (TRPC3/4) channels are involved in MBH HGE response to glucose in vitro and increased insulin secretion and decreased food intake in response to cerebral hyperglycemia in vivo. We also obtained evidences that MBH TRPC3 channel is a critical new player for energy homeostasis. This thesis work identifies two new mechanisms involved in hypothalamic detection of hyperglycemia: the first based on the involvement of TRPC3/4 channels in HGE neurons and the second highlighting the astroglial endozepines as a relay of the “glucose” signal to POMC neurons.