Thesis (MSc)--Stellenbosch University, 2003. / ENGLISH ABSTRACT: Chapter 1 presents a general literature review on the acute isocapnic hypoxic ventilatory response (HVR). The main findings from Chapter 2 indicate that our modified breathing circuit effectively measured the HVR while maintaining isocapnia. The measured ventilatory variables changed significantly with repeated short-term exposure to hypoxia over a 30-minute period, and the within- and between-day variability did not differ significantly. Furthermore, the variability in the HVR response (as measured by the coefficient of variation, (CV» amounted to approximately 27% between tests in both parameters. Repeated measures are recommended in future determinations of the HVR. In Chapter 3 the main findings were that hypoxic sensitivity does not differ between Caucasian and Xhosa sea-level populations in South Africa, and that ventilatory components in both normoxia and hypoxia differed between these two populations. Two distinct patterns of breathing were evident: shallow, rapid breathing among Xhosa subjects, and deeper, slower breathing among Caucasians. Moreover, lower arterial oxygen saturation levels during hypoxia among Xhosa subjects suggest that these two patterns of breathing differ in the effectiveness with which they oxygenate the blood. Inter-individual variation in HVR within each population is of the same high magnitude as that reported in the literature (Beall et al., 1997), further supporting the use of repeated measures in future studies. As previously reported (Sahn et al., 1977, Reeves et al., 1993), in Chapter 3 I document a significant correlation between HVR and partial pressure of end-tidal CO2 (PETCO). Future studies of HVR should consider PETCO2 as a covariate, despite the fact that my analyses of covariance (ANCOV A) showed no inter-population differences in HVR. In Chapter 4 I report that regression analysis shows that the HVR of parents is not a predictor of that of their offspring. No significant heritability was evident for any of the additional key variables of hypoxic VE ,hypoxic Sa02, and the CV for HVR, but a priori analyses showed that I tested too few subjects to be able to demonstrate heritability (or the lack thereof) conclusively by means of regression analyses. Importantly, repeatability estimates within populations (86 %) revealed that despite its high variability, the HVR is highly repeatable, and therefore remains a useful comparative research tool for studies of human adaptation to hypoxia. / AFRIKAANSE OPSOMMING: Hoofstuk 1 gee 'n algemene literatuuroorsig van die akute isokapniese hipoksiese ventilatoriese reaksie (HVR). Die hoofbevindinge uit Hoofstuk 2 dui aan dat ons gemodifiseerde asemhalingsbaan HVR effektief meet terwyl isokapniese toestande gehantaaf word. Die ventilatoriese veranderlikes gemeet, het betekenisvol verskil met herhaalde korttermyn blootstelling aan hipoksie in a 30-minuut periode, en die binne- en tussen-daagse afwykbaarheid het nie betekenisvol verskil nie. Verder het die afwykbaarheid van die HVR reaksie (soos bepaal deur die koëffisiënt van variasie (KV)) ongeveer 27 % beloop tussen toetse van beide parameters. Herhaalde metings word vir toekomstige bepalings van die HVR voorgestel. In Hoofstuk 3 was die hoofbevindinge dat hipoksiese sensitiwiteit nie verskil tussen Kaukasiese- en Xhosa- seevlak populasies in Suid-Afrika nie, en dat ventilatoriese komponente in beide normoksie en hipoksie verskillend was tussen hierdie twee populasies. Twee definitiewe asemhalingspatrone was duidelik merkbaar: vlak, vinnige asemhaling in Xhosa proefpersone, en dieper, stadiger asemhaling in Kaukasiërs. Verder het laer arteriële suurstof versadigingsvlakke gedurende hipoksie in Xhosa proefpersone daarop gedui dat hierdie twee asemhalingspatrone moontlik verskil in hul effektiwiteit om die bloed met suurstof te verryk. Inter-individuele variasie in HVR binne elke populasie was van dieselfde groot omvang as wat in die literatuur gerapporteer word (Beall et al., 1997), wat die gebruik van herhaalde metings in toekomstige studies verder ondersteun. Soos voorheen gerapporteer (Sahn et al., 1977, Reeves et al., 1993), dokumenteer ek in Hoofstuk 3 'n merkbare korrelasie tussen HVR en parsiële druk van eind-tidale CO2 (PET CO2 ). Verdere HVR studies behoort PET CO2 as a kovariant te beskou, ten spyte van die feit dat my analise van kovariansie (ANCOV A) geen inter-populasie verskille in HVR getoon het nie. In Hoofstuk 4 rapporteer ek dat regressie analise bewys dat die HVR van ouers nie 'n voorspeller van dié van hul kinders is nie. Geen betekenisvolle oorerflikheid was duidelik vir enige van die addisionele sleutelveranderlikes van hipoksiese VE ,hipoksiese Sa02, of die KV van HVR nie, maar 'n vorige analise het getoon dat ek te min proefpersone getoets het om oorerflikheid (of die gebrek daaraan) m.b.v. regressie analises te kan demonstreer. Dit is belangrik dat intra-populasie herhaalbaarheidsskattings (86 %) getoon het dat ten spyte van sy hoë afwykbaarheid, die HVR hoogs herhaalbaar is, en daarom 'n nuttige vergelykende navorsingshulpmiddel is vir studies rakende menslike aanpassing by hipoksie.

Ventilatory drive is mediated by respiratory central pattern generators that are located in the brainstem, which are continuously modulated by specialised peripheral and central chemoreceptors to adjust ventilatory patterns according to changes in arterial PO2. These specialised oxygen-sensing chemoreceptors are activated in response to acute reductions in arterial PO2 and ultimately trigger a respiratory response that acts to restore oxygen-levels. However, the molecular mechanism by which mammals are able to regulate their breathing pattern in such a manner during hypoxia remains controversial. Therefore, the studies performed in this thesis aimed to investigate the possibility that this process may be mediated by the liver kinase B 1 (LKB1)/ AMP-activated protein kinase (AMPK) signalling pathway, which is central to cellular adaptations to metabolic stress. This first involved the development of transgenic mice in which Lkb1 or AMPK were deleted. Global knockout of Lkb1 (Sakamoto, 2006) or AMPK activity (Viollet et al., 2009) are embryonic lethal. Thus, the Cre/loxP system was used to develop transgenic mice that had either Lkb1 or both isoforms of the AMPK catalytic α- subunit (α1 and α2) conditionally knocked out in catecholaminergic cells (including therein hypoxia-activated cells of the brainstem and carotid body) by driving Cre expression through a tyrosine-hydroxylase-specific promoter region. The consequent effects on the ventilatory response to hypoxia were then examined using unrestrained whole-body plethysmography. This demonstrated that, in contrast to the hyperventilation evoked in controls, increased ventilation was virtually abolished in the Lkb1 and AMPK α1 and α2 double knockouts during hypoxia. Both knockout mice also exhibited periods of hypoventilation with frequent apnoeas during hypoxia. Additionally, studies on single AMPK α1 and AMPK α2 knockouts identified that the ventilatory dysfunction in AMPK α1 and α2 double knockouts was primarily caused by AMPK α1 deletion. In contrast, the severe ventilatory abnormalities exhibited during hypoxia following the deletion of Lkb1 and AMPK in catecholaminergic cells were mostly reversed upon exposure of mice to hypoxia with hypercapnia. Also, the ventilatory response to hypercapnia alone was without any major effect as a result of Lkb1 deletion or the dual-deletion of AMPK α1 and α2 catalytic subunits in catecholaminergic cells. This thesis therefore demonstrates, for the first time, that the LKB1-AMPK signalling pathway is key to respiratory adaptations during hypoxia, by regulating catecholaminergic oxygen-sensing cells, thus protecting against hypoventilation and apnoeas. The LKB1-AMPK signaling pathway can thereby determine oxygen and energy supply at both a cellular and whole-body level.

This thesis describes experiments to measure the ventilatory response to hypoxia at a constant (isocapnic) level of CO2 (HVR) in 18 subjects. So as to provide a complete picture of the autonomic responses, middle cerebral artery velocity, a surrogate for cerebral blood flow (CBF), as well as finger plethysmography blood pressure (BP) were also measured. Ventilatory responses have been previously described only in terms of an acute peak followed by a decline. However, rather than a single type of response, I found four types categorized as: Decline, Double, Plateau, or No response. The Double pattern, characterized by a second peak of response was the most common, yet is described here for the first time. These patterns are also characteristic of the CBF and BP responses. Furthermore the temporal correlations between these brainstem-controlled responses are also reported here for the first time.

This thesis describes experiments to measure the ventilatory response to hypoxia at a constant (isocapnic) level of CO2 (HVR) in 18 subjects. So as to provide a complete picture of the autonomic responses, middle cerebral artery velocity, a surrogate for cerebral blood flow (CBF), as well as finger plethysmography blood pressure (BP) were also measured. Ventilatory responses have been previously described only in terms of an acute peak followed by a decline. However, rather than a single type of response, I found four types categorized as: Decline, Double, Plateau, or No response. The Double pattern, characterized by a second peak of response was the most common, yet is described here for the first time. These patterns are also characteristic of the CBF and BP responses. Furthermore the temporal correlations between these brainstem-controlled responses are also reported here for the first time.

No description available.

No description available.

Multicellular organisms typically depend on O₂ for energy production to maintain normal cellular function, and even brief periods of O₂ deprivation may have fatal consequences. The aqueous environment is prone to changes in ambient water O₂ tension (PO₂) and thus the ability of fish to sense changes in water PO₂ and to elicit appropriate physiological responses is essential for their survival. Studies on fish O₂ chemoreception have identified neuroepithelial cells (NECs), which are characterized as having dense-cored vesicles containing serotonin (5-HT), as peripheral O₂ chemoreceptors. Upon exposure to hypoxia, isolated and cultured NECs in vitro depolarize, likely resulting in neurotransmitter release. However, to date there is no evidence that NECs are activated by hypoxia in vivo to initiate physiological responses such as the hypoxic ventilatory response (HVR), which is the focus of this thesis. Initial findings demonstrated that larval zebrafish fine-tune the HVR as early as 4 days post fertilization (dpf) and by 7 dpf, the HVR aids in O₂ uptake under hypoxic conditions. In addition, the HVR is multiphasic, with an initiation phase followed by a decline phase that gradually stabilizes above normoxic baseline values (Chapter 2). In the absence of tools to probe the hypoxia sensitivity of NECs in vivo, research focused on Merkel-like cells (MLCs), a newly proposed O₂ chemoreceptor in larval zebrafish. Using in vivo calcium imaging it was shown that MLCs are stimulated by hypoxia. Data suggest that MLCs are responsible for the initiation phase of the HVR, while peripheral sensory neurons (PSNs)/peripheral sensory ganglia (PSG) that innervate MLCs play a more important role in reducing ventilation during the decline phase of the HVR (Chapter 4). Attempts at identifying the putative neurotransmitter(s) involved in the O₂ signal transduction pathway revealed that adrenaline (AD), serotonin (5-HT), and dopamine (DA) are probable candidates (Chapter 4), though the presence of AD and DA within MLCs is yet to be confirmed. In addition, 5-HT likely plays a role in the central nervous system (CNS), integrating peripheral signals resulting in the final HVR (Chapter 3). Taken together, this thesis provides the first evidence of putative O₂ chemoreceptors responding to hypoxia in vivo and thus significantly advances models for O₂ signal transduction in larval zebrafish.

Methadone is a long acting mu-opioid and is the most effective treatment for heroin addiction. However, opioids depress respiration and methadone maintenance treatment (MMT) patients have a higher mortality rate than the general population. Teichtahl et al conducted a pilot study and found 6 out of 10 MMT patients had central sleep apnea (CSA). But no definite conclusions were made regarding the prevalence and possible pathogenesis of CSA in the patients due to the small sample size and lack of blood toxicology data. The present project aims to confirm the preliminary results and further quantify the sleep disordered breathing (SDB) in stable MMT patients and to delineate the pathogenesis involved. (For complete abstract open document)

The 'exponential peeling' technique has been applied to minute ventilation and tidal volume transients occurring after the abrupt removal of 7,6 and 5% CO2 in inspired air. These transients, in many cases, were found to be composed of three exponential components, each contributing to the total ventilatory response and each having individual time responses. Gelfand and Lambertsen (1973) have attributed these components to the peripheral chemoreceptors as a group and to two central chemoreceptors. Statistical analysis to determine the constancy of the contribution of the three components over the the range of CO2 values studied showed that, although the values for each at the different stimulus levels were not significantly different, the great subject-to-subject variation in the data precluded a firm conclusion about the constancy of the components. Because of a number of considerations it was concluded that exponential peeling of respiratory transients following abrupt removal of CO2 inhalation is not a satisfactory way to approach the problem of the numbers, relative contributions and time responses of the various receptor groups comprising the respiratory controller.

No description available.