Oxygen can be limited at the environmental (e.g. flood-prone burrows) or cellular (e.g. stroke, heart attack) levels. O2 deprivation in nervous tissue depolarizes cell membranes, incrementing extracellular potassium concentration ([K+]o). Consequently, [K+]o can be used to assess neural failure during anoxia. The effect of temperature on the maintenance of brain [K+]o homeostasis in male and female Drosophila melanogaster (W1118) was assessed during repeated anoxic comas induced by N2 gas. Brain [K+]o was continuously monitored using K+-sensitive microelectrodes while body temperature was gradually increased/decreased using a Peltier plate. Once the desired temperature was reached (16°C/17°C, 23°C or 29°C/30°C), it was maintained for the rest of the experiment and the fly was subjected to repeated anoxic bouts. Repetitive anoxia resulted in a loss of the ability to maintain [K+]o baseline at ~10 mM. In both sexes, the total [K+]o baseline variation (D[K+]o) was augmented at 30°C (D[K+]o male = 119.2 ± 21.9 mM; D[K+]o female = 51.2 ± 8.1 mM), whereas 16°C stabilized [K+]o baseline for the duration of the experiment (D[K+]o male = 17.5 ± 4.1 mM; D[K+]o female = 16.9 ± 6.8 mM). Additionally, D[K+]o in males was significantly greater (114.3 ± 10.5 mM ) than in females (36.1 ± 10.5 mM) at 23°C. Under reduced dehydration, experiments performed only in males showed the same trends although the D[K+]o values where considerably reduced at 17°C (D[K+]o male = -1.0 ± 1.3 mM) and 23°C (D[K+]o male = 17.3 ± 1.5 mM) and increased at 29°C (D[K+]o male = 332.7 ± 83.0 mM). It was concluded that 1) N2-delivery patterns consisting of long anoxia, short normoxia and high cycle frequency increased disruption of brain [K+]o baseline maintenance, 2) males were more susceptible to repeated anoxia than females at room temperature, and 3) hypothermia had a protective effect on brain K+ homeostasis during repetitive anoxia. Male flies are suggested as a useful model for examining deleterious consequences of O2 reperfusion with extensive application on therapeutical treatment of stroke or heart attack. / Thesis (Master, Biology) -- Queen's University, 2012-01-30 13:03:10.913

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An incubation study was conducted to evaluate the effects of added urea, NH$ sb4$Cl, and KCl on fixation of NH$ sb4 sp{+}$ and K$ sp{+}$. A field study was carried out to observe the effects of rates and methods of added urea and KCl on the forms and distribution of N and K in soils, and on yields and nutrient uptake of silage corn. / Generally, the fixation of NH$ sb4 sp{+}$ was increased by added N but reduced by added K and vice versa. One exception was that added K increased NH$ sb4 sp{+}$ fixation when NH$ sb4$Cl was the N source. In comparison with NH$ sb4$Cl, urea resulted in greater K$ sp{+}$ fixation but less NH$ sb4 sp{+}$ fixation. / Band placement of urea and KCl showed apparent accumulation of NH$ sb4$-N and NO$ sb3$-N, and exchangeable K$ sp{+}$ in the fertilizer bands, particularly early in the growing season. On the other hand, band placement resulted in lower levels of available N and K in soil between the bands in comparison with broadcast. / Yields of silage corn and nutrient uptake values were increased by N and K fertilizers, except for one soil in the first year. Band placement of urea was superior in the case of frequent and light rainfall but inferior with dry surface soil conditions followed by a relatively large rainfall 3 or 4 days after application. In the latter condition, banding K resulted in less yields on one soil, and banding the two fertilizers together further reduced N utilization on another soil. / Additions of K suppressed Ca and Mg uptake but did not influence N uptake, whereas K uptake was usually enhanced by added N.

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The effects of added potassium, K, magnesium, Mg and sulfur, S on corn grain and silage yields were investigated in the field for two years. / Due to high nutrient soil, K, Mg and S had no effects on corn grain and silage yields. However, there were trends observed in the treatment effects that were significant. Added K at 240 kg K$ sb2$O/ha increased grain yield over zero kg K$ sb2$O/ha in 1983 and Mg at 60 kg/ha increased grain yield over 30 kg/ha in 1984. In silage, 240 kg K$ sb2$O/ha increased yield over 120 and zero kg K$ sb2$O/ha in 1984. Although there was no K by Mg interaction, there was a need for added Mg to achieve higher yields. / The cumulative effect of fertilizer S at 50 kg/ha significantly reduced silage yield in 1984 over zero S treatment indicating that fertilizer S was unnecessary for corn growth by reason of high levels of soil S and sufficient S provided from precipitation and air dry deposition. S had no interaction with K and Mg because there was no need for fertilizer S to increase yields. / The antagonistic association between K and Mg was seen at the seedling, tasseling and silking stages. / There was an indication of a K by S interaction which was expressed at different stages of growth, but this effect was not evident for dry matter yields.

La Fibrose Kystique, causée par des mutations du canal CFTR, mène à la dysfonction du transport des fluides et des ions causant la déshydratation du liquide de surface des voies aériennes et ainsi une défaillance de la clairance mucocilliaire. Ce défaut entraine l’accumulation et l’épaississement du mucus au niveau des bronches qui devient alors un environnement idéal pour le développement d’infections chroniques et d’inflammation qui sont associées à la destruction progressive de l’épithélium chez les patients Fibrose Kystique. Même si leur rôle dans les processus lésionnels est très bien connu, l’impact de médiateurs inflammatoires sur la capacité de réparation ne l’est cependant pas. L’objectif de ma maitrise était donc d’étudier la régulation des mécanismes de réparation de l’épithélium bronchique sain et Fibrose Kystique par le facteur de nécrose tumoral (TNF)-alpha, une cytokine pro-inflammatoire cruciale dans l’initiation et la propagation de la réponse inflammatoire chez les patients FK. À l’aide d’un modèle de plaies mécaniques, nous avons montré que le TNF-alpha stimule la réparation de l’épithélium bronchique sain (NuLi-1) et Fibrose Kystique (CuFi-1). De façon surprenante, l’exposition chronique au TNF-alpha augmente cette stimulation tout comme le taux de migration cellulaire pendant la réparation. Cette augmentation de réparation semble être médiée par l’activation de la métalloprotéinase MMP-9, la relâche d’EGF par les cellules épithéliales et ainsi l’activation de la voie d’EGFR. De plus, l’activation de la réparation par le TNF-alpha semble aussi impliquer l’activation des canaux K+, dont nous avons démontré le rôle important dans la réparation. Contrairement à son effet sur la migration cellulaire et sur la réparation, le TNF-alpha diminue la prolifération cellulaire. En somme, en plus de son rôle dans les processus lésionnels, le TNF-alpha semble avoir un rôle complexe dans les processus de réparation puisqu’il stimule la migration et ralentit la prolifération cellulaire. / Cystic fibrosis (CF) pathology, caused by mutations of cftr gene, leads to ion and fluid transport dysfunction that results in mucus thickening and accumulation in the airways. This mucus accumulation promotes bacterial infection and airway inflammation associated with progressive airway epithelial damage in CF patients, unfortunately leading to respiratory failure. However, the effect of inflammatory products on the repair capacity of respiratory epithelia is unclear. Thus, the objective of my project was to study the regulation of normal and CF bronchial epithelial repair mechanisms by tumor necrosis factor-alpha (TNF)-alpha, a major component of inflammation initiation and propagation in CF. With a wound healing model, we observed that TNF-alpha stimulated the non-CF (NuLi-1) and CF (CuFi-1) bronchial wound healing rate. Surprisingly, chronic exposure to TNF-alpha enhanced this stimulation as well as the migration rate during repair. This wound healing rate stimulation by TNF-alpha seems to be due to metalloproteinase MMP-9 activation, EGF shedding by epithelial cells and subsequent EGFR transactivation. Furthermore, we recently reported a crucial relationship between the EGF response and K+ channel function, both controlling bronchial repair. We now show that TNF-alpha wound healing stimulation also implicated KvLQT1 and KATP currents activation. In contrast to its effect on cell migration, TNF-alpha downregulate cell proliferation. Thus, in addition to its recognized role in the inflammatory response leading to epithelial injury, TNF-a could exert complex actions on repair mechanisms of CF airway epithelia by upregulating cell migration while downregulating proliferation.

The developmental profiles of mRNA and protein expression for ornithine decarboxylase (ODC), the Na$ sp+$-dependent glucose co-transporter (SGLT1), sucrase isomaltase (SI), and the Na$ rm sp+K sp+$ ATPase $ alpha sb1$ and $ beta sb1$ subunit isoforms in the postnatal rat small intestine, as well as the effects of exogenous spermidine on their precocious development, were examined. Postnatal age had a significant effect with all enzymes and the nutrient transporter maturing around weaning. Consecutive exposure to exogenous spermidine during suckling precociously induced ODC mRNA, SI protein, and SGLT1 gene expression in the proximal and distal small intestine. Levels of Na$ rm sp+K sp+$ ATPase $ alpha sb1$ and $ beta sb1$ subunit isoform mRNA were precociously induced in the proximal small intestine only. These findings show that exposure to exogenous spermidine can promote precocious alterations in intestinal enzyme and nutrient transporter expression; however, it appears that spermidine must be continuously supplied for these alterations to persist in suckling rats.

Inanga (Galaxias maculatus) is an amphidromous fish species that is able to successfully inhabit a variety of salinities. Using an integrated approach this thesis has characterised for the first time the physiological characteristics that facilitate acclimation in inanga. Structural studies using scanning electron microscopy (SEM) and laser scanning confocal microscopy (LSCM) revealed freshwater-acclimated inanga have a high density of apical pits and freshwater-type mitochondria-rich cells (MRCs) that can facilitate ion absorption from the hypo-osmotic environment. In seawater, inanga remodel their gills by increased proliferation of seawater-type MRCs to facilitate ion secretion in the hyper-osmotic environment. Concentration-dependent sodium (Na+) kinetic analysis revealed that at a whole body level, inanga regulate Na+ using a saturable, high affinity, low capacity uptake system which makes them extremely adept at extracting Na+ from very dilute freshwater environments. In fact inanga displayed an uptake affinity (Km) of 52 ± 29 µM, which is one of the lowest ever recorded in freshwater fish. The sodium/potassium ATPase transporter (NKA) is central to Na+ regulation within the gill. In high salinties inanga displayed increased NKA activity (6.42 ± 0.51 µmol ADP mg protein-1 h-1) in an effort to excrete the excess Na+, diffusively gained from the hyper-osmotic environment. This increase in NKA was most likely a reflection of the proliferation of NKA-containing MRCs. The NKA activities seen in freshwater- and 50% seawater-acclimated inanga were similar (2.54 ± 0.19 and 2.07 ± 0.22 µmol ADP mg protein-1 h-1 respectively) to each other suggesting the inanga gill is capable of supporting ion regulation in brackish waters without a significant increase in NKA activities, and the energetically-expensive changes in gill structure and function that accompany such a change. Molecular investigation of NKA isoform expression using quantitative PCR (qPCR) showed that inanga displayed salinity-induced changes in the expression of the three α NKA isoform variants investigated. Isoform α1a exhibited a pattern consistent with an important role in freshwater, confirming results from other fish species. While it is generally accepted that α1b isoform is the predominant NKA isoform in seawater, inanga did not display this pattern with a freshwater dominance seen. None of the salinity-induced changes could quantitatively explain the increased NKA activity in seawater suggesting that different isoforms may convey different activities, that there is also regulation of NKA at a post-transcriptional level, and/or other isoforms or subunits may have a significant role. The importance of the osmoregulatory hormone cortisol and prolactin is widely accepted and inanga were treated with cortisol, prolactin and a combination of the two in an effort to further elucidate their role. NKA activity and NKA isoform expression were assessed but no specific patterns were deduced, except for a decrease in both NKA activity and isoform expression in 100% seawater-acclimated inanga treated with cortisol and prolactin. The reasons for this decrease were not evident, although the impact of stress induced by the injection protocol was likely to be a confounding factor. The development of a new confocal-based technique in this study was able to describe, for the first time, intracellular sodium levels ([Na+]i) as a function of salinity in an intact euryhaline fish gill cell. Using the fluorescent Na+ indicator dye CoroNa Green this study demonstrated the ability of inanga gill cells to maintain [Na+]i in the face of environmental change. Freshwater-acclimated inanga displayed basal [Na+]i of 5.2 ± 1.8 mM, with 12 ± 2.3 mM and 16.2 ± 3.0 mM recorded in 50% seawater- and 100% seawater-acclimated cells, respectively. Low [Na+]i is advantageous in hypo-osmotic environments as it provides a gradient between the cell and the blood which is essential for generating electrochemical gradients cell volume regulation and other cellular homeostatic mechanisms. A slightly elevated [Na+]i seen at the higher sanities would help minimise the diffusive gradient for passive influx from the environment which would be of benefit in hyper-osmotic environments. Upon salinity challenge 50% seawater cells were equally adept at maintaining a constant [Na+]i at any salinity, suggesting these cells are have the necessary constituents to regulate Na+ in both lower and higher salinities. This novel LSCM approach is advantageous relative to existing transport models as it will allow the observation of cellular ion transport in real time, within a native filament structure displaying functional interaction of different cell types. The extreme ion uptake characteristics of the inanga and their amenability to in situ confocal-based studies demonstrated in this study, confirm inanga as a valuable model species for future research.

The objective of this thesis is to study the effects of flux deposition non uniformity on spreading of molten metal. Flux deposition non-uniformity here means as to whether the amount of flux deposited in a non-uniform or uniform pattern helps in the better wetting and spreading characteristics of the molten metal or is detrimental to the process. The material selection constraint to the study was imposed by selecting brazing of aluminum i.e., aluminum alloy melting and flow over an aluminum alloy substrate. The study was carried out by conducting a number of Hot Stage microscopy tests using aluminum silicon alloy as the filler metal and Potassium Fluoro Aluminate (Nocolok) as the flux. The flux was applied in different spatial distribution patterns to uncover the varying effects of its distribution on spreading. The uneven pattern of flux deposition indicates the influence on spreading but due to the efficient spreading of flux prior to aluminum melting and associated fuzziness of the achieved coverage distribution the effects are not always conclusive. It has been concluded that non uniform flux deposition does not necessarily mean uneven or less uniform spreading of the molten liquid metal if the spreading of the molten flux is beyond the distance of ultimate metal spreading. This is because, in spite of uneven flux deposition, the flux melts approximately at 560C-570C and spreads on the surface of the metal thereby promoting appreciable spreading and wetting of the molten liquid metal that happens at temperatures above 577C.