氯胺酮是一種麻醉劑，也是一種濫用藥物。近年來，氯胺酮濫用增長迅速，在香港已經成爲第二大濫用藥物。氯胺酮的短期效果主要導致精神狀態改變，但對其長期效果還了解甚少。研究目的：本研究旨在探討長期使用氯胺酮對中樞神經系統，腎上腺，胰腺和膀胱的影響。研究方法：我們在氯胺酮濫用的動物模型中進行的行爲學，神經化學，組織學和分子生物學研究。在水迷宮中，對這些小鼠學習和記憶能力進行了評估。應用基因芯片評估中樞神經系統的基因表達變化。用聚合酶鏈式反應陣列研究神經遞質及其調控基因表達變化。通過實時定量聚合酶鏈式反應和免疫印迹法檢測-氨基丁酸受體和多巴胺相關基因的基因表達的變化。用酶聯免疫法測定多巴胺含量。用原位末端轉移酶標記技術染色（細胞凋亡），天狼星紅染色（纖維化），免疫組織化學（乳酸脫氫酶，酪氨酸羟化酶，多巴胺β羟化酶）研究腎上腺，胰腺，膀胱癌的組織學變化。研究結果：與對照組相比，氯胺酮組小鼠學習記憶能力下降。基因芯片結果顯示，110個基因表達上調和136個基因表達下調。基因本體分析表明，氯胺酮明顯影響神經遞質和受體的活性。特別地，4-氨基丁酸A受體5型亞基的mRNA和蛋白水平在前額皮層的顯著上調。聚合酶鏈式反應陣列結果表明，氯胺酮顯著改變-氨基丁酸受體，神經肽，多巴胺和膽鹼能系統基因表達。對多巴胺系統的研究結果顯示，中腦多巴胺含量上調，酪氨酸羟化酶的顯著增加。在腎上腺和胰腺，氯胺酮和氯胺酮加酒精組都未觀察到細胞凋亡增加，但是觀察到乳酸脫氫酶的陽性染色。此外，在腎上腺中發現酪氨酸羟化酶和多巴胺β羟化酶下調。在膀胱中，在肌肉層觀察到細胞凋亡和纖維化。結論：本研究的結果研究指出，長期使用氯胺酮能引起中樞神經系統異常的基因表達，還能導致腎上腺，胰腺，膀胱癌的病理性改變。這些結果爲氯胺酮濫用相關的健康風險評估提供了重要的信息。 / Ketamine is an anesthetic agent and a drug of abuse. In recent years, ketamine abuse has been increasing rapidly and it has become the second-most popular abusive drug in Hong Kong. While the acute effects of ketamine are mainly linked to altered mental status, the long-term effects of ketamine are poorly understood. Objectives: The present study was designed to investigate the long-term effects of ketamine on the CNS, adrenal, pancreas and urinary bladder. Methods: Behavioral, neurochemical, histological and molecular studies were performed in a ketamine abuse animal model. Learning and memory ability in these mice were assessed in a morris water maze. An Affymetrix Genechip study was performed to assess the global gene expression changes in the CNS and a PCR-array study focused on the neurotransmitters and regulators was also performed. Gene expression changes for gamma-aminobutyric acid (GABA) receptors and dopamine related genes were assay by real-time PCR and western blot. Dopamine contents were measured by ELISA. Histological changes in adrenal, pancreas and urinary bladder were examined by TUNEL staining (apoptosis), Sirius red staining (fibrosis), and immunohistochemistry. Results: Compared with saline controls, there was a decline in learning and memory performance in the ketamine-treated mice. Genechip results showed that 110 genes were up-regulated and 136 genes were down-regulated in ketamine group. An ontology analysis revealed the most significant effects of ketamine were on neurotransmitter and receptor activities. In particular, there was a significant up-regulation of both mRNA and protein levels of the alpha 5 subunit (Gabra5) of the GABAA receptors in the prefrontal cortex. Results from the PCR-array study revealed significant gene expression changes in the GABA receptors, neuropeptides, dopaminergic and cholinergic system following ketamine treatment. Studies on the DA system revealed significant increase of DA content and up-regulation of Tyrosine Hydroxylase (TH) in the midbrain. In the adrenal and pancreas, no obvious apoptosis was found while lactate dehydrogenase (LDH) positive staining was observed in both ketamine and ketamine plus alcohol treated groups. On top of these, downregulation of TH and DBH were observed. In the urinary bladder, apoptosis and fibrosis were observed in the muscular layer. Conclusion: The present study pointed out that long-term of ketamine use caused aberrant gene expression in the CNS and led to pathological changes in adrenal, pancreas and urinary bladder. These results have provided novel and important insights in evaluating the health risks in ketamine abusers. / Detailed summary in vernacular field only. / Tan, Sijie. / Thesis (Ph.D.)--Chinese University of Hong Kong, 2012. / Includes bibliographical references (leaves 138-154). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese. / Abstract --- p.I / 摘 要 --- p.III / List of abbreviations --- p.IV / Acknowledgements --- p.VI / Contents --- p.VII / Chapter Chapter 1 --- General introduction --- p.1 / Chapter 1.1 --- Ketamine and abuse --- p.1 / Chapter 1.2 --- Pharmacological effects of ketamine --- p.4 / Chapter 1.3 --- Effects of ketamine on the CNS --- p.6 / Chapter 1.4 --- GABA receptors --- p.9 / Chapter 1.5 --- Dopamine system in the CNS --- p.10 / Chapter 1.6 --- The modulation of dopaminergic neurons --- p.12 / Chapter 1.7 --- Toxic effects of ketamine on other organs --- p.14 / Chapter 1.8 --- Thesis outline --- p.17 / Chapter Chapter 2 --- Cognition and GABA receptor expression following long-term ketamine administration --- p.20 / Chapter 2.1 --- Introduction --- p.20 / Chapter 2.2 --- Materials and methods --- p.22 / Chapter 2.2.1 --- Animals and drug administrations --- p.22 / Chapter 2.2.3 --- Dosage Determination --- p.23 / Chapter 2.2.4 --- Morris water maze --- p.24 / Chapter 2.2.5 --- Brain tissue collection and RNA extraction --- p.25 / Chapter 2.2.6 --- Microarray analysis --- p.26 / Chapter 2.2.7 --- Quantitative real-time PCR --- p.27 / Chapter 2.2.8 --- Western blotting --- p.28 / Chapter 2.2.9 --- Statistical analysis --- p.29 / Chapter 2.3 --- Results --- p.29 / Chapter 2.3.1 --- Morris water maze --- p.29 / Chapter 2.3.2 --- Microarray analysis --- p.30 / Chapter 2.3.3 --- Quantitative real-time PCR --- p.30 / Chapter 2.3.4 --- Western blotting --- p.31 / Chapter 2.4 --- Discussion --- p.44 / Chapter Chapter 3 --- PCR-array gene expression profiling on the neurotransmitters following chronic ketamine administration --- p.49 / Chapter 3.1 --- Introduction --- p.49 / Chapter 3.2 --- Materials and methods --- p.50 / Chapter 3.3 --- Results --- p.51 / Chapter 3.4 --- Discussion --- p.63 / Chapter Chapter --- 4 Chronic ketamine administration modulates midbrain dopamine system in mice --- p.67 / Chapter 4.1 --- Introduction --- p.67 / Chapter 4.2 --- Materials and methods --- p.69 / Chapter 4.2.1 --- Cell culture and ketamine treatment --- p.69 / Chapter 4.2.2 --- MTT assay --- p.70 / Chapter 4.2.3 --- Animals and ketamine administration --- p.70 / Chapter 4.2.4 --- Dopamine determination --- p.71 / Chapter 4.2.5 --- Real-time PCR --- p.72 / Chapter 4.2.6 --- Western blotting --- p.73 / Chapter 4.2.7 --- Immunohistchemistry --- p.74 / Chapter 4.2.8 --- Statistical analysis --- p.74 / Chapter 4.3 --- Results --- p.75 / Chapter 4.3.1 --- Effects of ketamine on dopamine concentrations in PC12 cells --- p.75 / Chapter 4.3.2 --- Long-term effects of ketamine on dopamine in mouse brain --- p.76 / Chapter 4.3.3 --- Effects of ketamine on mRNA levels of dopamine related genes --- p.76 / Chapter 4.3.4 --- Long-term effects of ketamine on BDNF protein levels in mouse brain --- p.77 / Chapter 4.3.5 --- Increased TH inmmureactive neurons in midbrain following 3 months ketamine treatment --- p.78 / Chapter 4.4 --- Discussion --- p.90 / Chapter Chapter 5 --- Chronic treatment of ketamine affects adrenal gland and pancreas --- p.95 / Chapter 5.1 --- Introduction --- p.95 / Chapter 5.2 --- Materials and methods --- p.95 / Chapter 5.2.1 --- Grouping of experimental animals and treatments --- p.95 / Chapter 5.2.2 --- Histological studies on pancreas and adrenal --- p.96 / Chapter 5.2.3 --- Immunohistochemistry on pancreas and adrenal --- p.97 / Chapter 5.2.4 --- TUNEL evaluation --- p.99 / Chapter 5.3 --- Results --- p.100 / Chapter 5.4 --- Discussion --- p.114 / Chapter Chapter 6 --- Ketamine Effects on the Urogenital System-Changes in the Urinary Bladder and Sperm Motility --- p.117 / Chapter 6.1 --- Introduction --- p.117 / Chapter 6.2 --- Materials and methods --- p.117 / Chapter 6.2.1 --- Studies of the Bladder --- p.117 / Chapter 6.2.2 --- Studies on Sperm Motility --- p.120 / Chapter 6.3 --- Results --- p.121 / Chapter 6.4 --- Discussion --- p.131 / Chapter Chapter 7 --- Conclusion --- p.134 / Chapter 7.1 --- Conclusion --- p.134 / Chapter 7.2 --- Future studies --- p.137 / Bibliography --- p.138

α-synuclein is an abundant and conserved presynaptic brain protein (Uversky 2007). It has received extensive attention since its aggregation was identified as the main component of Lewy bodies and Lewy neurites, which is the pathological hallmark of several neurodegenerative diseases, collectively known as synucleinopathies, including Parkinson's Disease (PD) (Uversky 2007). Considerable information has been collected about the structural properties and conformational behavior of α-synuclein, although the precise function is still under investigation. Metal ions such as copper and iron, can accelerate the aggregation and fibrillation of α-synuclein. Metal ions may exert their dual physiopathological properties through the interaction with α-synuclein, converting protein structure and/or inducing oxidative stress. In this study, isothermal titration calorimetry and electron paramagnetic resonance were used to determine the metal-binding property of the synuclein proteins, proving the presence of four Cu(II) binding sites per molecular of α-synuclein, with the coordination modes of 1N3O and 2N2O. Furthermore, α-synuclein has a catalytic action on the redox cycling of Cu(II), which was assessed by the application of cyclic voltammetry. However, this property is absent on β-synuclein and γ-synuclein, which belong to the synuclein family and have been suggested to be the physiological regulators of α-synuclein expression. In vivo, immunofluoresence studies revealed that Cu(II) increases the aggregates formation in mammalian doperminergic neuron cells overexpressing α-synuclein and the PD-associated mutants, while no aggregates have been found in cells overexpressing β-synuclein and γ-synuclein.

by Cui Yanyan. / "June 1999." / Thesis (Ph.D.)--Chinese University of Hong Kong, 1999. / Includes bibliographical references (p. 154-163). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Mode of access: World Wide Web. / Abstracts in English and Chinese.

Toxicity of organic solvents to microbial hosts is a major consideration in the economical production of biofuels such as ethanol and especially butanol, with low product concentrations leading to high recovery costs. The key to rational engineering of solvent tolerant microorganisms for such processes lies in obtaining appropriate tolerance genes (modules) suited for different compounds. In this project, a synthetic biology approach was adopted to generate a library of standardised BioBrick parts involved in different stress responses. Using a multiple-assay approach, including a bioluminescence assay, these stress response genes were tested individually and in combination to determine their effects on survival in ethanol, nbutanol, acetone and fermentation inhibitors produced by biomass pre-treatment. A set of tolerance modules was obtained for ethanol and n-butanol. Proof-of-concept tests suggested that ethanol and n-butanol toxicity was mainly due to damage to membrane, cellular proteins and DNA possibly by oxidative stress. No synergistic interactions were observed from a combination of different tolerance genes. Further tests carried out using enzyme and fluorescence-based assays to elucidate the effect of n-butanol on the cell envelope showed that the solvent released lipopolysaccharides from the outer membrane of E. coli and also caused both outer and inner membranes to be leaky. Very high n-butanol concentrations resulted in an altered cell shape and bleb formation suggesting an impairment in cell division and peptidoglycan biosynthesis. The cell membrane was modified by cis-trans isomerisation of unsaturated fatty acids in the phospholipids resulting in a reduction of membrane leakage which in effect, increased n-butanol tolerance in E. coli. In conclusion, results from this research suggest that strategies to protect the membrane and cellular proteins should be included in rational engineering of n-butanol tolerant bacteria.

Protein deposition in the form of amyloid fibrils is the hallmark of more than 40 human pathologies, including Alzheimer's disease (AD) and Parkinson's disease (PD). Misfolded protein oligomers formed as intermediates during the aggregation process have been strongly implicated in the onset and progression of these diseases. In this thesis, I describe our efforts to uncover molecular agents that can reduce the toxicity caused by protein aggregation via targeting the generation, the physiochemical properties or the membrane affinity of oligomeric species. We employed an integrative approach combining in vitro techniques, including chemical kinetics, atomic force microscopy, and biophysical measurements, and in vivo methods, including neuroblastoma cells and C. elegans models of AD and PD, to identify a range of small molecules and antibodies that can suppress the toxicity related to protein aggregation through a variety of mechanisms. In Chapter 3, we show that the deleterious effects of protein aggregation can be suppressed in AD and PD worms by interfering with the aggregation rates of the amyloid-β peptide (Aβ) and the α-synuclein protein (αS). In Chapter 4, we resolve the mechanism of action for a molecule that enhances the rate of Aβ42 aggregation in AD worms with the result that toxicity is reduced, and find that it potentiates the secondary nucleation microscopic step in vitro. In Chapter 5, we characterize molecules and antibodies that modify the physiochemical properties and self-association of oligomers comprised of several proteins into clusters with reduced diffusibility. In Chapter 6, we classify a family of molecules that protect the cell by displacing several types of oligomeric species from the membrane through a generic mechanism. These results demonstrate strategies by which one can target the aggregation process to alter its resulting toxicity, provide insight into modifying the properties of the most deleterious species associated with protein aggregation and suggest that the protection of the cell from the oligomer-induced cytotoxicity associated with numerous protein misfolding diseases is a promising strategy to combat protein misfolding diseases.

Key understanding of potential transformations that may occur on silver nanoparticle (AgNP) surface upon interaction with naturally ubiquitous organic ligands (e.g., −SH (thoil), humic acid, or −COO (carboxylate)) is limited. Herein we investigated how dissolved organic carbon (DOC), −SH (in cysteine, a well-known Ag+ chelating agent), and −COO (in trolox, a well-known antioxidant) could alter the colloidal stability, dissolution rate, and toxicity of citrate-functionalized AgNPs (citrate–AgNPs) against a keystone crustacean Daphnia magna. Cysteine, DOC, or trolox amendment of citrate–AgNPs differentially modified particle size, surface properties (charge, plasmonic spectra), and ion release dynamics, thereby attenuating (with cysteine or trolox) or promoting (with DOC) AgNP toxicity. Except with DOC amendment, the combined toxicity of AgNPs and released Ag under cysteine or trolox amendment was lower than of AgNO3 alone. The results of this study show that citrate–AgNP toxicity can be associated with oxidative stress, ion release, and the organism biology. Our evidence suggests that specific organic ligands available in the receiving waters can differentially surface modify AgNPs and alter their environmental persistence (changing dissolution dynamics) and subsequently the toxicity; hence, we caveat to generalize that surface modified nanoparticles upon environmental release may not be toxic to receptor organisms.

No description available.

Carbaryl toxicity, metabolism, and distribution in adults of three species of grass bugs _from the genus Labops were studied in relation to species, sex, and treatment with piperonyl butoxide. Lc50 values for 8 hour exposure periods ranged from 0.02-0.14, 0.03-0.3, and 0.2-0.7 μg carbaryl/vial for L. utahensis, L. hirtus, and L. hesperius respectively. The males were more susceptible to carbaryl than females. Males of L. utahensis were more susceptible than L. hirtus and L. hesperius respectively. The synergist difference values (Lc50 of carbaryl alone - Lc50 values of carbaryl after piperonyl butoxide treatment) were measured. The percent dependency of these _insects on MFOs for detoxifying carbaryl was estimated based on the theoretical synergist difference which was calculated by the equation Log LC50 = 1.014 log SD - 0.009. The percent dependency values were 38-59, 25-46, and 13-33% for L. hesperius, L. hirtus, and L. utahensis, respectively. Males of L. utahensis had the lowest percent dependency upon MFOs in detoxifying carbaryl (13%) indicating the possibility that carbaryl toxicity may be controlled by other potential defense mechanisms which are relatively ineffective themselves in view of the low tolerances of the insects to carbaryl. Treatment with piperonyl butoxide resulted in greater enhancement of carbaryl toxicity against L. hesperius (synergized Lc50 0.1-0.26 μg carbaryl/vial) while it showed a moderate effect on L. hirtus (synergized Lc50 0.02-0.16 μg carbaryl/vial). Piperonyl butoxide's effect was less pronounced in the case of L. utahensis (synergized Lc50 0.013- 0.09 μg carbaryl/vial. Unmetabolized carbaryl was the principle compound isolated from the bugs after 6 hours from treatment, being more prominent in males of L. hirtus (71.1% of the total metabolites) and less prominent in females hesperius (36.7% of the total metabolites). The mechanism of detoxication appeared to include ring hydroxylation for both species and sexes. 4 and 5-hydroxycarbaryl were the only metabolites associated with the degrading of carbaryl by the bugs, since the levels of metabolites obtained were too low for accurate quantitation. Pretreatment with piperonyl butoxide prevented the appearance of both carbaryl metabolites in the organosoluble fraction and increased the accumulation of unmetabolized carbaryl. This effect was probably due to inhibition of the insect's MFO system. Generally, this study showed a good correlation between the bioassays and the metabolic studies, thus reflecting the effectiveness of the bioassays along with synergist difference (SD) and percent dependency concepts in establishing some conclusions regarding the MFOs of Labops bugs. Further application of these techniques with agricultural insects should provide a practical means of characterizing field populations for insecticide tolerance, relative levels of MFOs and their role as a defense mechanism.

The basic mineral nutritional requirements of Proteaceae are not well understood.They are generally assumed to require low levels of nutrients and be susceptible to nutrient (especially Phosphorus) toxicity.This project aimed to estimate the general nutritional requirements of Proteaceae for optimum growth, with special emphasis on the Phosphorus requirement. Potted plants were grown in soilless growth media with controlled release fertiliser and were watered according to need in a naturally lit greenhouse. The nutrient requirements of Proteaceae were found to vary among species but were not lower than the reported requirements for the Ericaceae, another heath family. The variables of growth media and plant development were found to be important factors affecting the critical Phosphorus concentration and need consideration in the derivation of the Phosphorus requirement of Banksia ericifolia. / Doctor of Philosophy (PhD)

Thesis (Ph. D.)--University of Washington, 1984. / Vita. Bibliography: leaves [189]-210.