Epileptic seizures are often un-witnessed and can result in hypoxic brain damage or can be fatal due to injuries, status epilepticus or sudden unexpected death in epilepsy (SUDEP). The first aim of this thesis was to investigate some of the physiological parameters that accompany an epileptic seizure and may be useful in a seizure alarm system. The second aim was to investigate aspects of cardiac dysfunction during clinical and sub-clinical seizures that may be potential contributing factors in SUDEP. Percentage heart rate change and oxygen saturation were studied prospectively during 527 epileptic seizures in 50 patients aged from one-day full term neonate to 60 years with a variety of seizure types (absences, generalised tonic clonic seizures, myoclonic seizures, tonic seizures and focal seizures) and in normal physiological events (e.g. coughing, turning in bed). Higher percentage heart rate change occurred during epileptic seizures (21.8%) than during normal physiological events (16.4%) p<0.001. Diagnostic testing of clinically significant seizures i.e seizures that could potentially lead to serious consequences if left undetected (n=61) had a sensitivity of 91% and specificity of 75% when percentage heart rate change and hypoxaemia parameters were combined. Percentage heart rate change and oxygen saturation could be used as reliable indicators of a seizure when set at specific levels and distinguish clinically significant seizures from normal physiological events. These parameters can now be used to develop a reliable alarm system to detect epileptic seizures at night. Prolongation of QTc and increased vagal tone may be possible mechanisms underlying SUDEP. Corrected Q-T cardiac repolarisation time 5 minutes before and throughout 156 epileptic seizures were analysed using four corrective formulae (Bazett, Hodge, Fridericia and Framingham). All formulae indicated statistically significant lengthening of the corrected Q-T during epileptic seizures (p<0.001) compared to pre-seizure values. All formulae agreed that the greatest lengthening of the corrected QT beyond normal limits occurred during right temporal lobe seizures in two patients. Reflex and tonic vagal activity utilising R-R intervals was assessed in 33 sub-clinical seizures occurring during stages 3 or 4 sleep and was compared to matched counts of R-R interval non-ictal baseline studies from the same stage of sleep in each patient. Altered vagal activity occurred during total sub-clinical seizures compared to baseline studies (p<0.001). Lengthening of the corrected Q-T and changes in cardiac vagal tone during epileptic seizures may have a role in the patho-physiology of SUDEP.

In the pulmonary circulation, alveolar hypoxia contributes to blood flow regulation. Hypoxic pulmonary vasoconstriction (HPV) involves both pulmonary arteries and veins, but little is known of the contractile mechanisms specific to the veins. The aim of these studies was to examine the hypoxic response in small porcine intrapulmonary veins in relation to the arterial response, and investigate the effects of hypoxia on ion conductances in single myocytes from intrapulmonary veins. In wire myography experiments, intrapulmonary veins contracted more than sizematched arteries in response to hypoxia and agonists KCl and PGF2α. Venous contractions were inhibited by removal of extracellular Ca2+ or in the presence of Clchannel blocker NFA, effects not seen in the arteries. To examine the mechanisms of venous contraction at cellular level, single pulmonary vein smooth muscle cells (PVSMC) were freshly isolated and characterised morphologically and electrophysiologically for the first time. In patch-clamp studies, hypoxia reversibly inhibited a whole-cell outward current in the presence of BKCa channel antagonist Penitrem A. By subtracting currents recorded in normoxia and hypoxia, a novel hypoxia-sensitive K+ current (IK(H)) was revealed in PVSMC. IK(H) was a rapidly activating, partially inactivating current and was sensitive to KV channel blocker 4-AP. The biophysical properties of IK(H) revealed the voltage window of current availability with a peak near the resting membrane potential of PVSMC. In conclusion, these findings highlight differences between the contractile properties of veins and arteries and reveal a significant contribution of Ca2+ influx and an NFA-sensitive conductance during venous contraction to agonists and hypoxia. Furthermore, the results suggest that a novel hypoxia-sensitive KV current contributes to membrane potential under resting conditions in PVSMC and its inhibition by hypoxia may contribute to the initiation of HPV in porcine intrapulmonary veins.

Areas of hypoxia are found in coastal areas worldwide, and have become increasingly widespread. These areas vary in their duration and dissolved oxygen concentration from occasional diurnal hypoxia, as found in Corpus Christi Bay, Texas, seasonal hypoxia as in the northern Gulf of Mexico, to continuous hypoxia as found in oceanic oxygen minimum zones. The effects of exposure to low dissolved oxygen (DO) depend on the duration of exposure, the DO concentration and an organism’s tolerance to hypoxic conditions. Most studies have focused on lethal effects of hypoxia by comparing the abundance of benthic organisms and the species composition of benthic communities between hypoxic and normoxic areas. Sub-lethal effects of such as changes in reproduction may occur at less severe hypoxic conditions (by definition), but may still have effects at the population level. The goal of this study is to examine the sub-lethal reproductive effects of low DO on harpacticoid copepods. The life-history traits and reproductive biology of meiobenthic harpacticoid copepods make this group of organisms useful as test organisms to measure the sub-lethal effects of hypoxia on reproduction. It is hypothesized that changes in reproductive traits may be observed at DO concentrations higher than those that cause lethal effects because of the high energetic cost of female harpacticoid reproduction which may result in reduced fitness. Laboratory studies were conducted to examine the effects of low DO concentrations on survival, egg production, and the number and size of eggs. Harpacticoid population abundance, biomass and the abundance of ovigerous females were measured from field samples collected across sites that varied in their degree and duration of exposure to low DO. To contrast the effects of hypoxia in environments with different hypoxic exposures, field studies were carried out in the northwestern Gulf of Mexico in an area with near constant summer hypoxia, and in southeast corner of Corpus Christi Bay, Texas which experiences intermittent hypoxia often for less than one hour in the early morning hours during summer. At both field sites total copepod abundance, biomass and the abundance of ovigerous females were reduced at the most hypoxic site type compared the reference sites. In Corpus Christi Bay, total and ovigerous female abundance varied with exposure to hypoxia, but the relative declines in abundance and biomass were less dramatic than in the northern Gulf of Mexico. In the northern Gulf of Mexico, the percentage of ovigerous females within the total harpacticoids collected was reduced (p<0.001) to 3% at the Intermediate and 1% at the Hypoxic sites during the summers, but in the fall of 2007 at all transects had similar percentages (p = 0.81) of ovigerous females which ranged from 13 to 16%. Reference sites had a higher number of families with ovigerous females, and the relative abundances of those families were more evenly distributed at Reference sites compared to Hypoxic and Intermediate transects. Patterns of ovigerous female abundance with respect to DO concentration were similar across numerous harpacticoid families found at the study sites. Harpacticoid copepods in laboratory studies tolerated near anoxic DO conditions for longer than 120 hours. Average survival rates of the harpacticoids Schizopera knabeni and Nitokra affinis during periods of low DO with elevated ammonium concentrations were less than 50%, while survival rates for the other treatments were near 100%. The formation of an egg mass was reduced in the near anoxia treatment, but egg clutches did not form at all in the treatment that combined near anoxia and 10 micromolar ammonium. The DO concentration and presence of ammonium were important factors to the severity of population decline that were illustrated by population estimate calculations. Population estimates based on laboratory measurements of Nitokra affinis resulted in calculations of reduced survival and reproductive rates related to hypoxic exposure greatly decrease the potential population of the group, which may affect the copepod population’s ability to recover from hypoxic events. The effects of hypoxia on harpacticoid copepods are the result of the length of exposure, the concentration of low dissolved oxygen and exposure to other chemical fluxes that increase during periods of hypoxia. The measurement of reproductive indicators, such as the presence of an egg mass and the proportion of ovigerous females in a population, could be used to indicate the adverse environmental effects of low dissolved oxygen exposure. The studies from this dissertation are the first to document reproductive effects of low dissolved oxygen on harpacticoid populations in the field. Laboratory studies within this dissertation indicate the interaction between ammonium and hypoxia on survival and reproduction in harpacticoids. Future studies are needed to further determine the effects of hypoxia on the whole-life cycle of harpacticoid copepods. / text

Endometrial cancer is one of the most significant gynaecological malignancies that affect women from New Zealand and the rest of the world. One of the critical stages in the development of a tumour is the onset of hypoxia. The malignancy responds by having raised levels of Hypoxia Inducible Factor (HIF) that in turn induces increased production of Vascular Endothelial Growth Factor (VEGF). VEGF is a potent angiogenic factor that will mediate vascular supply of nutrients and oxygen to the developing tumour. The aim of this study was to investigate whether two compounds found in extracts of plant materials, Resveratrol (Resveratrol) and Epigallocatechin gallate (EGCG), altered the levels of VEGF in the supernatant of cultured endometrial cancer cells. Resveratrol is a phytoalexin that is found in many foods, such as grapes, nuts and berries, as well as in high concentrations in some red wines. 100 µM of resveratrol was added to cell cultures for 24 hours. VEGF levels in the supernatant were then analysed using ELISA. Resveratrol was found to have significant inhibitory effects in both primary endometrial cancer cell cultures and immortalised endometrial cancer cell cultures. Resveratrol was also shown to reverse the increase in VEGF caused by the hypoxia mimic cobalt chloride (CoCl₂). Epigallocatechin gallate (EGCG) is an antioxidant catechin extracted from green tea. The effect of EGCG was analysed using the same method as for resveratrol. 100 µM of EGCG was also shown to have a significant inhibitory effect on the level of VEGF in the supernatant of cultured endometrial cancer cells, as well as reducing the effect of CoCl₂. These results suggest that selected food compounds, resveratrol and EGCG, can reduce VEGF levels by inhibiting HIF. Further investigation This may have anti-tumour effects in women with endometrial cancer.

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Acute kidney injury (AKI) may result from perioperative renal medullary hypoxia. Despite high oxygen delivery to the kidney, the medullary thick ascending limb (mTAL) in the outer renal medulla is susceptible to hypoxia because of its high oxygen consumption and relatively low rate of blood flow. The objective of this study was to evaluate the effect of a low pO2 (8% FiO2 for 5 h) on the major function of the mTAL and to develop a strategy to protect the mTAL in this setting. Evidence of hypoxia-induced reduction in mTAL function included low interstitial and urine osmolality but only a minimal rise in Na+ excretion; this was prevented by pre-treatment with desmopressin acetate (dDAVP), a vasopressin analogue which may increase tissue pO2. A decrease in urine osmolality may be of diagnostic value for hypoxic renal damage and dDAVP may prevent acute kidney injury in the perioperative setting.

Acute kidney injury (AKI) may result from perioperative renal medullary hypoxia. Despite high oxygen delivery to the kidney, the medullary thick ascending limb (mTAL) in the outer renal medulla is susceptible to hypoxia because of its high oxygen consumption and relatively low rate of blood flow. The objective of this study was to evaluate the effect of a low pO2 (8% FiO2 for 5 h) on the major function of the mTAL and to develop a strategy to protect the mTAL in this setting. Evidence of hypoxia-induced reduction in mTAL function included low interstitial and urine osmolality but only a minimal rise in Na+ excretion; this was prevented by pre-treatment with desmopressin acetate (dDAVP), a vasopressin analogue which may increase tissue pO2. A decrease in urine osmolality may be of diagnostic value for hypoxic renal damage and dDAVP may prevent acute kidney injury in the perioperative setting.

Hypoxic responses in quail development differ depending upon stage, duration and level of oxygen partial pressure of embryo. Incubation was switched to/from 110mmHg partial pressure (hypoxia), to/from 150mmHg (normoxia) during different stages in development, and control was incubated in normoxia throughout. Hatchability and embryo survival resulted in no hatchlings in continuous hypoxia. Responses to various hypoxic exposures throughout development resulted in recovery/repair of hypoxic damage by hatch. Heart and body mass, beak and toe length, hemoglobin, and hematocrit were measured to determine embryo responses to hypoxia during development at days 10, 15, and hatch. Hypoxia seemed to have the most deleterious effects on eggs in continuous hypoxia. Collectively, data indicate critical developmental windows for hypoxia susceptibility, especially during mid-embryonic development.