Thesis (MPhil (Physiological Sciences))--University of Stellenbosch, 2006. / The responses of central systems to oxygen deprivation have been well characterised while adaptations in peripheral systems, such as skeletal muscles, have presented confounding variations. Several reasons for these discrepancies are purported, amongst them being the duration of exposure to hypoxia and variations in fibre composition. Moreover, in real-life high altitude situations there may be a combination of factors which have the ability to modify or alter the effect of hypoxia. This study investigates the effect of short duration hypoxia per se on substrate utilisation in different types of skeletal muscles.

Thesis (PhD (Biomedical Sciences. Medical Physiology))--University of Stellenbosch, 2007. / Nitric oxide (NO) is a major signaling molecule in the heart with various biological effects. The putative role of NO as a cardioprotective agent against ischaemiareperfusion injury and in ischaemic preconditioning (IP) has made it one of the fastest growing fields in basic cardiovascular research. However, NO may also be associated with harmful effects, especially when released in excessive amounts. Little is known about the relative contributions to NO-production by the cardiac microvascular endothelial cells (CMECs) and the adjacent cardiomyocytes. Furthermore, the respective roles of endothelial NOS (eNOS) and inducible NOS (iNOS) are not well characterized in these cell types, particularly in hypoxia. In order to gain a better understanding of the role of NO in the hypoxic/ischaemic heart, the aims of this study were to: (1) develop an isolated cardiomyocyte model in which hypoxia and early IP can be induced and the role of NO assessed; (2) measure NOproduction in cardiomyocytes and CMECs under baseline and hypoxic conditions; and (3) evaluate the expression, regulation and activation of eNOS and iNOS in cardiomyocytes and CMECs (baseline and hypoxia) and establish the relationship with NO-production under these conditions. Cardiomyocytes isolated from adult rat hearts and commercially purchased rat CMECs were used as cell models.

Thesis (PhD (Physiological Sciences))--University of Stellenbosch, 2009 / ENGLISH ABSTRACT: Three main morphologies of cell death have been described in the diseased myocardium, type I, better known as apoptotic cell death, which is characterized by cell shrinkage and chromatin condensation, type II, or cell death with autophagy, presents a morphology with intracellular accumulation of autophagic vacuoles and type III, better known as necrosis, is characterized by cellular swelling and rapid loss in cellular membrane integrity. However, recent literature strongly argues against rigid classifications in the context of cell death mechanisms but rather suggests to adopt a view of cell death as a dynamic and integrative cellular response. Furthermore, the contribution of autophagy in cell death or cell survival is still poorly understood. Therefore the aims of this study were twofold: (i) to characterize the contribution of each cell death type in context of the severity and duration of an ischaemic insult and (ii) to determine whether manipulation of the autophagic pathway affects the contribution of cell death and translates into protection of the heart. Rodent derived cardiac myoblast cells were grown in Dulbecco’s Modified Eagle’s Medium (DMEM) supplemented with 10% fetal bovine serum (FBS), and incubated under 5% CO2 conditions. Cells were submitted to protocols of 2, 4 and 8 hrs of simulated ischaemia (SI) under hypoxic conditions in a humidified environment containing 0.1% O2, 5% CO2 and the balance N2, followed by 1 hr of reperfusion respectively. We employed a modified ischaemic buffer containing either 2-deoxy- D-glucose, sodium dithionate or both, with the aim to create an ischaemic insult of mild (mild SI), moderate (moderate SI) and severe (severe SI) character respectively. We evaluated the contribution of each cell death mode using a combination of viability- and ATP assays. Molecular markers for each cell death process such as LC3, PARP and HMGB1 were evaluated using 3-dimensional fluorescence techniques as well as western blot analysis and flow cytometry. Next, autophagy was induced or inhibited prior to the ischaemic insult, using rapamycin and 3MA respectively, and similar parameters were evaluated after 2 hours of mild or moderate SI. Propidium Iodide exclusion and Fluorescence Resonance Energy Transfer (FRET) in combination with mitochondrial inner membrane depolarization were employed to assess the onset of cell death dynamically. Flow cytometry was employed to evaluate the degree of protection. In addition, the ATP levels and reactive oxygen species (ROS) were evaluated. Our results strongly indicate a differential induction of cell death, which is dependent on the severity and duration of the ischaemic insult. Mild SI led to the induction of autophagy and apoptosis, whilst moderate or severe SI induced both apoptotic and necrotic cell death without an indication of autophagy. Only mild SI, but not moderate and severe SI, resulted in an ATP surge. Moreover, our data provide direct evidence that increased autophagy delays the loss of cellular membrane integrity and delays caspase-3 activation as well as mitochondrial depolarization in ischaemic cardiomyocytes. Our results show a profound effect of increased autophagy on the onset of apoptosis as well as necrosis under simulated ischaemic conditions, providing cellular protection. This ATP surge observed during mild SI was abolished with increased autophagy. Furthermore, our results indicate a profound effect of autophagy on ROS generation. Under normoxic conditions, increased autophagy induced a significant decrease in ROS while the inhibition of autophagy significantly increased ROS generation. However, when increasing or decreasing autophagy prior to the ischaemic insult, ROS increased significantly in both scenarios. The results suggest that the severity of ischaemia determines the mode of cell death differentially. An increase in autophagic responsiveness and flux, as induced through rapamycin treatment, provides a selective advantage for tissue against injury, possibly by maintaining intracellular ATP levels through the provision of metabolic substrates. Autophagy is described as an inherent cellular mechanism v which affects the onset of cell death and exhibits protective effects in the ischaemic myocardium when upregulated prior to the ischaemic insult. The protective effect of increased autophagy was mirrored in the isolated perfused rat heart model, reflected by improved functional recovery during ischaemia/reperfusion. / AFRIKAANSE OPSOMMING: Die drie belangrikste morfologiese beskrywings van seldood in die hart sluit die volgende in: tipe I, beter bekend as apoptose wat gekenmerk word deur selkrimping en chromatienkondensering, tipe II, of seldood deur middel van autofagie wat gekenmerk word deur die intrasellulêre versameling van autofagiese vakuole en tipe III, beter bekend as nekrose wat gekenmerk word deur sel swelling en ‘n vinnige verlies aan membraanintegriteit. Onlangse literatuur waarsku egter teen die onbuigsame klassifikasie van seldoodmeganismes en stel voor dat seldood as ‘n dinamiese proses met integrerende sellulêre meganismes beskou moet word. Die bydrae van autofagie in seldoodmeganismes word ook nog nie goed verstaan nie. Die doel van hierdie studie is dus tweevoudig: (i) om die bydrae van elke tipe seldood te bepaal in konteks van die felheid en tydperk van die iskemiese ingryping en (ii) om te bepaal of the manupilering van autofagie ‘n betekenisvolle bydrae lewer in seldoodmeganismes en sodoende tot beskerming van die hart kan lei. Kardiale mioblaste wat van rotweefsel afkomstig is, is in Dulbecco se gemodifiseerde Eagle medium (DMEM), waarby daar 10% fetale kalfserum gevoeg is en wat onderhewig was aan 5% CO2 toestande, onderhou. Selle was onderhewig aan protokolle van 2, 4 en 8 ure gesimuleerde iskemie (SI) onder hipoksiese toestande in ‘n humiditeitsomgewing wat 0.1% O2, 5% CO2 en die balans N2 bevat. Daarna was die selle onderhewig aan 1 uur reperfusie. ‘n Gemodifiseerde iskemiese buffer wat óf 2-deoksie-D-glukose óf natriumdithionaat, of beide bevat, is gebruik om lig, matig en strawwe iskemiese toestande na te boots. Die bydrae van elke tipe seldood is geëvalueer tydens bogenoemde toestande deur gebruik te maak van ‘n kombinasie van sellewensvatbaarheid- en ATP tegnieke. Molekulêre merkers, wat LC3, PARP en HMGB1 insluit, is gebruik om deur middel van 3-dimensionele fluoresensie tegnieke, westelike kladtegnieke en vii vloeisitometrie die verskillende vorme van seldood te ondersoek. Autofagie is ook geïnduseer en geïnhibeer voor die iskemiese ingryping, deur middel van rapamycin en 3MA, respektiewelik om die rol van autofagie tydens seldood te bepaal. Propidium iodite uitluiting en fluoresensie resonansie energie oordrag (FRET) in kombinasie met ‘n merker vir mitochondriale binneste membraan depolarisasie is gebruik om die aanvang van seldood dinamies te ondersoek. Vloeisitometrie is gebruik om die graad van beskerming aan te dui, terwyl intrasellulêre ATP vlakke en reaktiewe suurstof spesies (ROS) ook gemeet is. Ons resultate het getoon dat daar ‘n differensiële indusering van seldood plaasvind wat afhanklik is van die felheid en tydsduur van die iskemiese ingryping. ‘n Ligte graad van iskemie lei tot die indusering van autofagie en apoptose, terwyl matige en strawwe iskemie beide apoptose en nekrose induseer sonder autofagie. Verder het slegs ‘n ligte graad van iskemie ‘n skerp styging in ATP tweeggebring, terwyl dit nie die geval was tydens matige en strawwe iskemie nie. Ons data verskaf ook direkte bewyse dat ‘n toename in autofagie die verlies van sellulêre membraanintegriteit vertraag. Dit het ook ‘n vermindering in caspase-3 aktivering en mitochondriale depolarisasie in iskemiese kardiomiosiete teweegebring. Die data dui aan dat ‘n toename in autofagie ‘n beduidende effek op apoptose en nekrose tydens gesimuleerde iskemiese toestande het om sodoende selbeskerming te verskaf. Die skerp styging in ATP wat tydens die ligte graad van iskemie teweeggebring is, is opgehef met ‘n toename in autofagie. Ons resultate dui ook daarop dat autofagie ‘n beduidende rol in ROS generering speel. Onder normoksiese omstandighede veroorsaak ‘n toename in autofagie ‘n insiggewende afname in ROS generering, terwyl die inhibisie van autofagie ROS generering insiggewend laat toeneem. Wanneer autofagie egter voor die iskemiese ingryping verhoog of verlaag word, vermeerder ROS generering in beide gevalle. Hierdie resultate bewys dat die graad van iskemie ‘n invloed het op die tipe seldood wat geïnduseer word. ‘n Toename in autofagie reaksietyd en vloei, soos viii bewerkstellig deur rapamycin, verskaf ‘n selektiewe voordeel vir weefsel teen beskadiging, heel waarskynlik deur die handhawing van intrasellulêre ATP-vlakke deur die verskaffing van metaboliese substrate. Autofagie word beskryf as ‘n inherente sellulêre meganisme wat seldood beïnvloed en die iskemiese miokardium beskerm wanneer dit opgereguleer word voor die iskemiese ingryping. Hierdie beskermende rol van autofagie wat in die weefselkultuur waargeneem is, is ook in die geïsoleerde geperfuseerde rot hart model waargeneem, waar funksionele herstel verbeter is tydens iskemie/reperfusie.

published_or_final_version / abstract / Surgery / Doctoral / Doctor of Philosophy

The presence of an alpha-adrenoceptor--mediated coronary vasoconstrictor reflex during acute systemic hypoxia was examined in thirteen chloralose-anesthetized dogs. Local vasodilator effects were avoided by perfusing the left common coronary artery (LCC) with normoxic blood, while the dogs were ventilated with 5% 02-95% N2 . Left ventricular afterload was held constant and positive cardiac inotropic responses and beta two-adrenoceptor-mediated coronary vasodilation were blocked by propranolol. Parasympatheticmediated bradycardia and coronary vasodilation were blocked with atropine. Systemic hypoxia decreased LCC flow to normoxic myocardium by 19.4+2.6 %. Although myocardial oxygen extraction increased 9.7+2.9 %, myocardial oxygen consumption decreased 16.5+2.6 %. Intracoronary prazosin prevented the reflex vasoconstriction during repeated hypoxia.

This study investigated possible effects of hypoxic training upon speed of high school sprint freestyle. Thirty-eight subjects, grouped as their two schools, performed identical loads during the ten-week program. The Experimental group used hypoxic techniques for about one-half of each workout. Pretests and posttests conducted for the 50-yard and 100-yard distances yielded highly correlated mean scores, with marked differences between the two groups. Analysis of covariance was used, selecting the .05 level for rejection. The comparison of adjusted group means indicated neither group superior at 50 yards, while the 100-yard F-ratio was significant at the .0047 level favoring hypoxic training. It is recommended that hypoxic techniques be incorporated into existing programs, possibly benefitting other strokes.

El NO participa en una amplia gama de procesos biológicos, entre ellos se presume la adaptación a la hipoxia medioambiental. Por tal motivo se determinaron los niveles de nitritos en plasma en un total de 270 ratas albinas machos, divididas en dos ensayos: la mitad sometidos a hipoxia a la altura de 3,320 m.s.n.m confrontados a un grupo control a nivel del mar. La determinación del óxido nítrico (NO) fue indirecta mediante la cuantificación de los nitritos (NO2) y nitratos (NO3) por la reacción de Griess, (1879). Todos los animales fueron muestreados en grupos de 15 los días 1, 2, 5, 7, 14, 21, 28, 35 y 42 respectivamente. La mayor concentración de óxido nítrico en el grupo hipóxia medioambiental se encontró en el día 2 de muestreo ((α = 0.05). Los niveles plasmáticos de óxido nítrico en el grupo hipóxia medioambiental presentaron diferencias significativas con el grupo control (p – valor = 0.001) encontrándose incrementados con respecto al grupo control (α = 0.05). Estos resultados proponen que la hipoxia medioambiental produce un aumento de la síntesis de ON, como consecuencia de procesos adaptativos a la altura, corroborando los resultados de otros trabajos e investigaciones. / NO participate in an ample rate of biological processes, among them presumes the adaptation to the environmental hypoxia. By such reason the plasma levels of nitrates were determined in a total of 270 rats male, divided in two tests: half put under hypoxia to the height of 3.320 mts. confronted to a group control at level of the sea). The determination of nitric oxide (NO) was indirect by means of the cuantification nitrates (NO2) and nitrites (NO3), by the reaction of Griess, (1879). All the animals were tested in groups of 15, on the days 1, 2, 5, 7, 14, 21, 28, 35 y 42 respectively. The greater concentration of NO in the group submissive environmental hypoxia was in day 2 of sampling (α = 0.05). The NO levels in the hypoxia environmental group presented significant differences with the group control (p – value = 0,001) being increased with respect to the group control (α = 0.05). These results propose that environmental hypoxia produces an increase of the synthesis of nitric oxide, consequence of adaptive processes to the height, corroborating the results of other works and investigations.

Se ha estudiado el efecto del ketoprofeno, un antiprostaglandínico bloqueador de la enzima ciclooxigenasa, sobre la presión arterial pulmonar media (PAPm) en 10 terneros Jersey, machos, de 1 a 2 meses de edad, nacidos a nivel del mar y expuestos durante 3 días a una hipoxia ambiental de 3 320m. de altitud; los cuales fueron divididos en 2 grupos de 5 animales cada uno: Grupo Control que recibió placebo y Grupo Tratamiento al que se le suministró ketoprofeno, en dosis de 3mg/kg de peso vivo, por 5 días consecutivos, durante su estadía a nivel del mar. La PAPm se determinó mediante la técnica de cateterismo cardiaco a nivel del mar y al tercer día de su arribo a dicha altitud. Los valores promedio de la PAPm (mmHg) a nivel del mar fueron de 19,00 ± 0,94 para el Grupo Control y 21,00 ± 3,39 para el Grupo Tratamiento, y en la altura de 39,50 ± 6,00 para el Grupo Control y 31,25 ± 5,70 para el Grupo tratamiento. Al no existir diferencia por efecto del ketoprofeno, se analizaron ambos grupos en conjunto obteniéndose los siguientes resultados de la PAPm: A nivel del mar 20,00 ± 2,68 y al tercer día de exposición a la altura 35,28 ± 7,16. Estos resultados nos inducen a sugerir que el ketoprofeno no tuvo efecto significativo (p>0.05) sobre la PAPm en los terneros Jersey sometidos durante 3 días a la hipoxia de la altura; sin embargo sólo la exposición a dicha hipoxia incrementó significativamente (p≤0.05) la PAPm. / The effect of ketoprofen, an antiprostaglandin sustance that inhibits the enzyme ciclooxigenase, had been studied on the mean pulmonary arterial pressure (mPAP) in 10 Jersey, male calves, from 1 to 2 months old. They were born at sea level and exposed during 3 days to a environmental hypoxia of 3 320m. of altitude and were divided in 2 groups of 5 animals each: Control Group which received destiled water as placebo, and Treatment Group which received ketoprofen in dose of 3mg/kg body weight for 5 consecutive days during its permanence at sea level. The mPAP was determinated by cardiac cateterism at sea level and 3 days after arriving to altitude. The mean values of mPAP (mmHg) at sea level were: Control Group 19,00 ± 0,94 and Treatment Group 21,00 ± 3,39; whereas at the altitude were: Control Group 39,50 ± 6,00 and Treatment Group 31,25 ± 5,70. Since there was not significative effect due to ketoprofen, the two groups were joined and analysed together, obtaining the following values for mPAP: At sea level 20,00 ± 2,68 and at day 3 of exposition to altitude 35,28 ± 7,16. These results induce us to sugest that ketoprofen have not significative effect (p>0.05) on the mPAP in Jersey calves exposed during 3 days at altitude hypoxia, however just the effect of this hypoxia increased significatively (p≤0.05) the mPAP.

Cardiac ischemia, stroke and some neurodegenerative disorders are all characterized by cell damage and death due to low oxygen levels. Comparative studies show that anoxia tolerant model systems present a unique opportunity to study "survival" instead of death in the complete absence of oxygen. The freshwater turtle (Trachemys scripta elegans) is unique in its ability to survive total oxygen deprivation for hours to days, as well as reoxygenation insult after anoxia. The broad objective of this study is to understand the modulation of key molecular mechanisms involving stress proteins and VEGF that offer neuroprotection and enhance cell survival in the freshwater turtle through anoxia and reoxygenation. In vivo analyses have shown that anoxia induced stress proteins (Hsp72, Hsp60, Grp94, Hsp60, Hsp27, HO-1); modest changes in the Bcl2/Bax ratio and no change in cleaved caspase-3 expression suggesting resistance to neuronal damage. These results were corroborated with immunohistochemical evidence indicating no damage in turtle brain when subjected to the stress of anoxia and A/R. To understand the functional role of Hsp72, siRNA against Hsp72 was utilized to knockdown Hsp72 in vitro (neuronally enriched primary cell cultures established from the turtle). Knockdown cultures were characterized by increased cell death associated with elevated ROS levels. Silencing of Hsp72 knocks down the expression of Bcl2 and increases the expression of Bax, thereby decreasing the Bcl2/Bax ratio. However, there was no increase in cytosolic Cytochrome c or the expression levels of cleaved Caspase-3. Significant increase in AIF was observed in the knockdown cultures that increase through anoxia and reoxygenation, suggesting a caspase independent pathway of cell death. / Expression of the master regulator of hypoxia, HIF1 alpha and its target gene, VEGF, were analyzed at the mRNA and protein levels. The results showed no significant increase in HIF-1 alpha levels but anoxia VE GF The levels of stress proteins and VEGF returned to control levels during reoxygenation suggesting robust ROS protection mechanisms through reoxygenation. The present study thereby emphasizes Trachemys scripta as an advantageous model to examine anoxia and reoxygenation survival without major damage to the brain due to it's modulation of molecular mechanisms. / by Shailaja Kesaraju. / Thesis (Ph.D.)--Florida Atlantic University, 2008 / Includes bibliography. / Electronic reproduction. Boca Raton, Fla., 2008. Mode of access: World Wide Web.

Obstructive sleep apnea (OSA) is a common breathing and sleeping disorder, characterized by repeated episodes of airway obstruction during sleep resulting in intermittent hypoxia (IH). From clinical reports, patients with OSA are associated with behavioral and neuropsychological deficits, including impaired spatial learning memory and cognitive deficiencies. Previous studies proposed that reactive oxygen species (ROS) and apoptosis caused by intermittent hypoxia (IH) contributed to this cognitive deficits. However, the exact mechanism is still poorly understood and not settled. / The endoplasmic reticulum (ER) is a cellular organelle in which all secretory and integral membrane proteins are folded and is also the site where proteins are post-translationally modified in ATP-dependent chaperone-mediated processes. In this study, we hypothesized that ER stress in the hippocampus is initiated in the OSA via elevated levels of ROS. Four groups of adult male mice were used, with two of them exposed to normoxia as control, and the other two exposed to IH treatment, each receiving either vehicle or tauroursodeoxycholic acid (TUDCA), an ER stress inhibitor. Eight-armed radial maze was used to investigate the performance of reference memory during the whole IH/normoxia treatment. After behavior test, long-term potentiation (LTP) was measured to investigate synaptic plasticity in hippocampus. Furthermore, ER stress-associated pro-apoptotic effectors were detected by Western blotting, and ultra-structure of rough ER and the morphology of hippocampal dendritic spines and synapses in the hippocampal CA1 area were observed. / LTP was impaired in the hippocampus after IH treatment, which was rescued by TUDCA. Furthermore, ER stress-associated pro-apoptotic effectors, CHOP and caspase-12, were up-regulated after chronic IH treatment and was abolished by co-infusion of TUDCA. Meanwhile, increased cleaved-caspase-3 after chronic IH treatment was reduced by TUDCA via increased expression of Bcl-2. On the other hand, ultrastructural analysis of rough ER in the hippocampal CA1 revealed IH-induced ER luminal swelling, and was attenuated by TUDCA. In addition, the length of synaptic active zone was significantly reduced after chronic IH treatment and was partially rescued by the application of TUDCA. Golgi staining also showed a decrease in mature dendritic spines in IH group, and reversed by TUDCA. In behavioral analysis, the number of reference memory errors significantly increased after IH treatment and rescued by TUDCA injection. Overall, the data suggest a critical role of ER stress underlying the impairment of long-term synaptic plasticity and neurocognitive deficits in chronic IH. Targeting ER stress could be a potential therapeutic strategy for neural dysfunction in OSA. / On the other hand, neuronal firing, especially robust persistent activity of neuron in hippocampus, is critical role in memory formation. Increased ROS induced by IH has been implicated in long-term potentiation of neural activity. IH could be involved in a variety of K⁺ channels which eventually leads to excitotoxicity by increased Ca2⁺-dependent glutamate release. Although the results were just shown in acute IH treatment, the chronic effect of IH on the firing frequency of hippocampus is still unknown. / Therefore, to investigate the effect of chronic IH treatment on firing activities and local field potentials of hippocampal neurons, implantation of multi-channel micro-wires electrode array into hippocampus of OSA model rat was performed to monitor spontaneous discharge. The results were shown the firing frequency of pyramidal neurons (PNs) was significantly elevated after 8 hours IH in second and third days, on the other hand, interneurons (INs) seem to be more sensitive to intermittent hypoxia since the higher firing frequency was sustained from third day to seventh day after 8 hours IH, however, at the end of 14 days IH treatment, the firing frequencies of PNs and INs are all both dramatically reduced. Meanwhile, the results in this part will enable us to understand the exact change of firing pattern and local field potential during intermittent hypoxia. The percentage of complex burst spikes was decreased after 14 days IH in PNs and the power of theta rhythms was also impaired. It suggests that the disorder of neuronal pattern and the change of local field potential are associated with cognitive impairment in OSA model. After 1 week recovery, the firing frequency of PNs was rescued again, but not for that of INs. We also found that the power of theta rhythms which had an important role in memory formation was weaker after 2 weeks IH treatment, however, the precise mechanism was still unknown. From the effect of intermittent hypoxia on spontaneous discharges and LFP of hippocampal neurons in free moving rat, it may reveal some roles of IH in cognitive impairment via disorder neuronal function in CA1 region. / 阻塞性睡眠呼吸暫停(OSA) 是一種常見的睡眠障礙疾病，這種疾病的主要特徵是在睡眠過程中反復發作的氣道阻塞，從而導致间歇性缺氧(IH)。從臨床報導中發現，OSA患者表現出行為和神經心理缺陷，包括空間學習記憶的受損和認知缺陷。通過之前的研究表明，活性氧(ROS)的增多和細胞凋亡是間歇性缺氧所引起認知功能障礙的主要機制之一，然而，其具體的機制仍不清楚。 / 作為細胞重要的細胞器，內質網是分泌蛋白和膜蛋白折疊組裝的主要場所，同時，由ATP依賴的分子伴侶所介導的蛋白質翻譯後修飾這一過程也主要在內置網中完成。在本課題中，我們假設在OSA模型的海馬組織中，內質網應激的啟動是由於缺氧引起的與活性氧(ROS)的升高。在本課題中，我們使用了四組成年雄性小鼠，其中兩組作為正常對照組，分別接受生理鹽水和牛磺去氧膽酸（一種常用的內質網抑制劑）的腹腔注射，另外兩組接受缺氧處理，同時也分別接受照生理鹽水和牛磺去氧膽酸注射。八臂放射迷宮被用來研究參考記憶的表現。行為學結束之後，長時程增強(LTP)用來測定海馬的突觸可塑性。用免疫印跡的方法檢測內質網應激的相關凋亡蛋白的表達情況，並且觀察海馬CA1區域中，內質網超微結構和海馬樹突棘數目及突觸形態的變化。 / 從實驗結果中，LTP在缺氧後減弱，而TUDCA能夠部分恢復由於缺氧所導致的LTP的降低。除此之外，內質網應激相關的促凋亡蛋白(CHOP和caspase-12)在缺氧組中表達升高，但是在TUDCA組中有所減低，同時，我們還發現，TUDCA也能夠減低缺氧組中cleaved-caspase-3的表達，而這一作用，可能與提高Bcl-2蛋白的表達（一個可標記的抗凋亡蛋白）有關。在間歇性缺氧組的海馬CA1區域中，粗面內質網出現管腔的腫脹，這一超微結構的變化表明在內質網出現官腔中有的許多未折疊蛋白聚集，並通過TUDCA的注射能夠降解未折疊蛋白來緩解這一現象的發生。同時，在IH處理後，突觸超微結構也發生了形態上的變化。突觸活性區的長度在IH處理組中顯著減少，但是在TUDCA組中有一定程度的恢復。高爾基染色顯示，成熟樹突棘（海馬突觸可塑性的結構基礎）的數目在間歇性缺氧組中有所下降，而在TUDCA治療後，成熟樹突棘的數目有所上升。我們發現參考記憶錯誤次數在缺氧後都有明顯的升高，而在注射TUDCA後，參考記憶錯誤次數都有所降低。總之，這些結果證明，內質網應激在間歇性缺氧的所引起的長時程突觸可塑性減弱和神經認知功能的損傷起到關鍵的作用，而抑制內質網應激對OSA中的出現神經功能紊亂起到一定的預防和治療效果。 / 而另一方面，神經元的放電，特別是海馬中神經元穩定持久的放電形式，對記憶的形成起到關鍵的作用。間歇性缺氧所引起的ROS的升高對於長時程增強的神經活動存在一定的關係，因為，通過以往的研究發現，間歇性缺氧可以通過多種鉀離子通道的啟動，最終由於鈣離子依賴的谷氨酸釋放的增多从而導致興奮性毒性的神經遞質的釋放。而這些結果只在急性缺氧模型中發現，慢性的間歇性缺氧對海馬的放電頻率的影響仍是未知之數。 / 因此，為了探討長時程的間歇性缺氧對海馬神經元的放電頻率和局部場電位的影響，多管道微絲電極陣列植入OSA大鼠的海馬中來監控自發放電的影響。結果表明，錐體細胞的放電頻率在第二天和第三天的8小時的間歇性低氧後明顯的升高了。另一方面，我們觀察到中間神經元似乎對間歇性缺氧更敏感，因為，從第三天到第七天缺氧8小時後，神經元的放電頻率都明顯的增高。但是在間歇性缺氧14天后，錐體細胞和中間神經元的放電頻率都所有顯著性的減少。同時，這一部分結果準確表明了海馬神經元的放電模式和局部場電位在間歇性缺氧的模型的是如何變化的。我們發現錐體細胞所具有的複合簇狀放電的比例減少，同時，theta波(與記憶的形成有關)的能量也有所減低。而這種神經元活動和局部的場電位的異常變化可能與OSA模型中出現的總認知功能障礙有關。在恢復一周後，錐體細胞的放電頻率有所增加，基本上可以恢復到缺氧前的狀態，但是中間神經元的頻率卻沒有多大的改變, 但是，其具體機制仍不清楚。從間歇性缺氧對大鼠海馬神經元自發放電和場電位影響的結果，它揭示了間歇性缺氧通過擾亂海馬CA1區域神經元的功能從而導致認知功能損傷。 / Xu, Linhao. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2014. / Includes bibliographical references (leaves 167-199). / Abstracts also in Chinese. / Title from PDF title page (viewed on 03, October, 2016). / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only. / Detailed summary in vernacular field only.