by Lam Ying Wan. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1991. / Includes bibliographical references. / Acknowledgement --- p.5 / List of Abbreviations --- p.7 / Abstract --- p.10 / Chapter Chapter 1: --- Introduction --- p.13 / Chapter 1.1 --- Distribution and Biosynthesis of Taurine --- p.14 / Chapter 1.2 --- Physiological Functions of Taurine --- p.17 / Chapter 1.2.1 --- Interaction of Taurine and Calcium --- p.17 / Chapter 1.2.2 --- Neuroinhibitory action of Taurine --- p.18 / Chapter 1.2.3 --- Taurine as an Osmoeffector --- p.20 / Chapter 1.2.4 --- Integrative Model of Taurine Action --- p.22 / Chapter 1.3 --- Taurine and Volume Regulation in Astrocytes --- p.22 / Chapter 1.3.1 --- Response of Cells to Anisosmotic Media --- p.22 / Chapter 1.3.2 --- Mechanism of Regulatory Cell Volume Decrease --- p.23 / Chapter 1.3.3 --- Regulatory Volume Decrease (RVD) in Astrocytes --- p.25 / Chapter 1.3.4 --- Taurine and Volume Regulation in Astrocytes --- p.25 / Chapter 1.4 --- Ion Channels and Transporters in Astrocytes --- p.26 / Chapter 1.4.1 --- Potassium Channels --- p.26 / Chapter 1.4.2 --- Sodium Channels --- p.27 / Chapter 1.4.3 --- Chloride Channels --- p.27 / Chapter 1.4.4 --- Stretch-activated Ion Channels --- p.27 / Chapter 1.4.5 --- (KC1 + NaCl) Carrier --- p.27 / Chapter 1.4.6 --- Na+/H+ exchange --- p.28 / Chapter 1.4.7 --- C1-/HCO3- exchange --- p.28 / Chapter Chapter 2: --- Materials and Methods --- p.30 / Chapter 2.1 --- Cell Culture --- p.30 / Chapter 2.1.1 --- Preparation of Culture Medium --- p.30 / Chapter 2.1.2 --- Preparation of Phosphate Buffered Saline --- p.30 / Chapter 2.1.3 --- Cell Counting Method --- p.31 / Chapter 2.1.4 --- Culture of U373MG Human Astrocytoma Cells --- p.31 / Chapter 2.1.5 --- Culture of Primary Astrocytes --- p.32 / Chapter 2.2 --- Taurine Release Experiment --- p.32 / Chapter 2.2.1 --- Preparation of Physiological Salt Solution (PSS) --- p.32 / Chapter 2.2.2 --- Preparation of Hyposmotic Solution --- p.33 / Chapter 2.2.3 --- Preparation of Chloride Free Solution --- p.33 / Chapter 2.2.4 --- Preparation of Sodium Free Solution --- p.33 / Chapter 2.2.5 --- Preparation of Calcium Free Solution --- p.34 / Chapter 2.2.6 --- Preparation of High Potassium Solution --- p.34 / Chapter 2.2.7 --- Preparation of Urea containing PSS --- p.34 / Chapter 2.2.8 --- Assay of [3H]-Taurine Release --- p.34 / Chapter 2.2.9 --- Drug pretreatment --- p.35 / Chapter 2.2.10 --- Data Calculation --- p.35 / Chapter 2.3 --- Volume Determination --- p.36 / Chapter 2.3.1 --- Experimental procedure --- p.36 / Chapter 2.3.2 --- Drug pretreatment --- p.37 / Chapter 2.3.3 --- Data calculation --- p.40 / Chapter 2.4 --- Taurine Influx Experiment --- p.41 / Chapter 2.4.1 --- Experimental Procedure --- p.41 / Chapter 2.5 --- Drug Preparation --- p.42 / Results / Chapter Chapter 3: --- Hyposmolarity-Induced [3H]-Taurine Release --- p.45 / Chapter 3.1 --- Responses of Astrocytes to Hyposmotic Conditions --- p.45 / Chapter 3.1.1 --- Effect of Hyposmotic Medium on the Release of Preloaded [3H]-taurine in U373MG astrocytoma cell --- p.45 / Chapter 3.1.2 --- Time Course of the Hyposmolarity-induced [3H]-taurine Release --- p.49 / Chapter 3.1.3 --- Response of Primary Astrocytes to Hyposmotic Medium --- p.49 / Chapter 3.2 --- Effect of MK196 on Hyposmolarity-Induce Taurine Release --- p.52 / Chapter 3.3 --- Effects of Inhibitors of (NaCl+KCl) Cotransporter and C1- /HCO3- Anion Exchanger on Hyposmolarity-induced [3H]- taurine Release --- p.56 / Chapter 3.3.1 --- Effect of (NaCl + KC1) Cotransporter Inhibitors on Hyposmolarity-induced [3H]-taurine Release --- p.56 / Chapter 3.3.2 --- "Effects of two stilbene derivatives, SITS and DIDS,on hyposmolarity-induced [3H]-taurine release" --- p.56 / Chapter 3.3.3 --- "Effect of a Chloride Channel Blocker, Antracene-9- Carboxylate on Hyposmolarity-induced [3H]-taurine Release" --- p.57 / Chapter 3.3.4 --- Effect of MK473 on Hyposmolarity-induced [3H]-taurine Release --- p.58 / Chapter 3.4 --- Effect of Chloride Depletion on Hyposmolarity-induced [3H]- taurine Release --- p.58 / Chapter 3.4.1 --- Effect of Replacing Chloride with Nitrate --- p.58 / Chapter 3.4.2 --- Effect of Replacing Sodium Chloride with Sucrose --- p.59 / Chapter 3.4.3 --- Effect of Replacing Chloride with Gluconate --- p.59 / Chapter 3.5 --- Investigation of the Transduction Mechanism of Hyposmolarity- induced [3H]-taurine Release --- p.71 / Chapter 3.5.1 --- Effect of Depleting Extracellular Ca2+ --- p.71 / Chapter 3.5.2 --- Effect of Staurosporine on Hyposmolarity-induced [3H]- taurine Release --- p.71 / Chapter 3.6 --- Effect of SITS on the Swelling Process of U373 MG cells --- p.74 / Chapter 3.6.1 --- Regulatory Volume Decrease (RVD) in U373 MG Cells --- p.74 / Chapter 3.6.2 --- Effect of SITS on RVD in U373 MG Cells --- p.74 / Chapter 3.7 --- Effect of Hyposmotic Medium on Sodium-Independent Taurine Uptake in U373 MG Cells --- p.77 / Chapter Chapter 4 : --- Urea-Induced [3H]-Taurine Release --- p.80 / Chapter 4.1 --- Concentration Dependency of Urea-Induced Efflux of [3H]-taurine from U373 MG Cells --- p.80 / Chapter 4.2 --- Effect of MK 196 on the Urea-Induced [3H]-taurine Release from U373 MG Cells --- p.82 / Chapter 4.3 --- Effect of SITS on the Urea-induced [3H]-taurine Release from U373 MG Cells --- p.82 / Chapter Chapter 5: --- High Potassium-Induced Efflux of [3H]-taurine --- p.86 / Chapter 5.1 --- High Potassium Concentration Induced Release of [3H]-taurine from U373 MG Cells --- p.86 / Chapter 5.1.1 --- High Potassium Concentration Induced Release of [3H]- taurine --- p.86 / Chapter 5.1.2 --- Effect of the Concentration of HCO3- on High Potassium Induced Release [3H]-taurine Release --- p.87 / Chapter 5.2 --- Effect of MK 196 on High Potassium Induced [3H]-taurine Release in U373 MG --- p.87 / Chapter 5.3 --- Effect of (NaCl + KC1) Cotransporter Inhibitors on High Potassium Induced Taurine Release from U373 MG Cells --- p.91 / Chapter 5.3.1 --- Effect of Furosemide on High Potassium Induced [3H]- taurine Release --- p.91 / Chapter 5.3.2 --- Effect of Bumetanide on High Potassium Induced [3H]- taurine Release --- p.91 / Chapter 5.4 --- Effect of C1-/HCO3- Anion Exchanger Inhibitors on High Potassium Induced Release of [3H]-taurine from U373 MG Cells --- p.91 / Chapter 5.4.1 --- Effect of SITS on High Potassium Induced [3H]-taurine Release --- p.91 / Chapter 5.4.2 --- Effect of Antracene-9-Carboxylate on High Potassium Induced [3H]-taurine Release --- p.96 / Chapter 5.4.3 --- Effect of MK 473 on High Potassium Induced [3H]- taurine Release --- p.96 / Chapter 5.5 --- Effect of Chloride Depletion on High Potassium-Induced [3H]- taurine Release --- p.96 / Chapter 5.5.1 --- Effect of Replacing C1- by NO3- --- p.96 / Chapter 5.5.2 --- Effect of Replacing C1- by Gluconate --- p.96 / Chapter Chapter 6: --- Discussion --- p.102 / Chapter 6.1 --- Hyposmolarity Induced [3H]-taurine Release --- p.103 / Chapter 6.1.1 --- Hyposmolarity is the Key Stimulation for [3H]-taurine Release --- p.103 / Chapter 6.1.2 --- Hyposmolarity Induced [3H]-taurine Release and the C1- /HCO3- anion exchanger --- p.104 / Chapter 6.1.3 --- Comparision of the Hyposmolarity-induced Release of [3H]-taurine in U373 MG cells and primary astrocytes --- p.106 / Chapter 6.1.4 --- Comparision between the Hyposmolarity-induced Taurine Release and the Na+-independent Uptake for Taurine --- p.106 / Chapter 6.1.5 --- Transduction Mechanisms of Hyposmolarity-induced [3H]-taurine Release --- p.107 / Chapter 6.2 --- Urea-Induced Release of [3H]-taurine --- p.107 / Chapter 6.3 --- High Potassium-Induced [3H]-taurine Release --- p.108 / Chapter 6.3.1 --- Pharmacological Properties of High Potassium-induced [3H]-taurine Release --- p.108 / Chapter 6.3.2 --- Effect of Ionic Environment on High Potassium-Induced [3H]-taurine Release --- p.108 / Chapter 6.4 --- Mechanism of Swelling-Induced Taurine Release --- p.109 / Chapter 6.4.1 --- Involvement Stretched Activated Channel (SACs) in Swelling-Induced Taurine Release --- p.109 / Chapter 6.4.2 --- Involvement of the C1-/HCO3- Anion Exchanger in Swelling-Induced Taurine Release --- p.110 / Chapter 6.4.3 --- Possibility of Taurine as a Substrate of the C1-/HCO3- Anion Exchanger --- p.111 / Chapter 6.4.4 --- Conclusion --- p.114 / Chapter Chapter 7: --- Conclusion --- p.116 / References --- p.119

by Chang Chuen Chung. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1993. / Includes bibliographical references (leaves 125-140). / Acknowledgement --- p.4 / List of Abbreviations --- p.5 / Abstract --- p.7 / Chapter CHAPTER I --- INTRODUCTION --- p.10 / Chapter 1.1 --- Astrocytes in the Central Nervous System --- p.10 / Chapter 1.1.1 --- Characteristics of astrocytes --- p.10 / Chapter 1.1.2 --- Functional roles of astrocytes --- p.11 / Chapter 1.1.2.1 --- General functions of astrocytes --- p.11 / Chapter 1.1.2.2 --- Volume regulation of astrocytes in CNS injuries --- p.12 / Chapter 1.1.2.3 --- Immunological functions of astrocytes --- p.13 / Chapter 1.2 --- Taurine in the CNS --- p.15 / Chapter 1.2.1 --- The biochemistry and distribution of taurine --- p.15 / Chapter 1.2.2 --- Physiological functions of taurine in the CNS --- p.19 / Chapter 1.2.3 --- Uptake and release of taurine by cultured astrocytes --- p.20 / Chapter 1.2.3.1 --- Taurine uptake in astrocytes --- p.21 / Chapter 1.2.3.2 --- Taurine release in astrocytes --- p.22 / Chapter 1.3 --- Tumor necrosis factor in the CNS --- p.23 / Chapter 1.3.1 --- Characteristics of tumor necrosis factor --- p.23 / Chapter 1.3.2 --- Sources of TNF in the CNS --- p.25 / Chapter 1.3.3 --- Functions of TNF in the CNS --- p.26 / Chapter 1.3.4 --- TNF and signal transduction --- p.27 / Chapter 1.4 --- cGMP second messenger system in astrocyte --- p.29 / Chapter 1.4.1 --- cGMP as second messenger in astrocytes --- p.29 / Chapter 1.4.2 --- Post cGMP cascade effects --- p.30 / Chapter 1.5 --- The aims of this project --- p.30 / Chapter CHAPTER II --- METHODS --- p.34 / Chapter 2.1 --- Primary astrocytes culture --- p.34 / Chapter 2.1.1 --- Primary rat astrocytes culture --- p.34 / Chapter 2.1.2 --- Primary mouse astrocytes culture --- p.36 / Chapter 2.1.3 --- Culture of rat C6 glioma cell line --- p.36 / Chapter 2.1.4 --- Subculture of astrocytes in different media --- p.37 / Chapter 2.2 --- Taurine uptake and release assay --- p.39 / Chapter 2.2.1 --- Taurine uptake assay --- p.39 / Chapter 2.2.2 --- Taurine release assay --- p.41 / Chapter 2.3 --- The effects of TNF on taurine transport --- p.42 / Chapter 2.4 --- The effects of TNF on cell volume in astrocytes --- p.43 / Chapter 2.5 --- "The effects of TNF on amino acids, glucose and neurotransmitters uptake" --- p.43 / Chapter 2.5.1 --- The effects of TNF on amino acids uptake --- p.43 / Chapter 2.5.2 --- The effects of TNF on glucose uptake --- p.44 / Chapter 2.5.3 --- The effects of TNF on neurotransmitters uptake --- p.45 / Chapter 2.6 --- The effects of LPS on taurine uptake in astrocytes --- p.46 / Chapter 2.7 --- The effects of IFN-¡’ on taurine uptake in astrocytes --- p.46 / Chapter 2.8 --- The effects of PMA on taurine uptake in astrocytes --- p.47 / Chapter 2.9 --- "The effects of TNF on thymidine, uridine and leucine incorporation in astrocytes" --- p.47 / Chapter 2.10 --- The effects of TNF on basal level of cGMP in astrocytes --- p.48 / Chapter 2.11 --- The effects of TNF on protein phosphorylation in astrocytes --- p.49 / Chapter 2.12 --- The effects of TNF on calcium uptake in astrocytes --- p.50 / Chapter CHAPTER III --- RESULTS --- p.51 / Chapter 3.1 --- The effects of TNF on taurine transport in cultured rat astrocytes --- p.51 / Chapter 3.1.1 --- The effects of TNF on [3H]-taurine uptake -time course study --- p.52 / Chapter 3.1.2 --- The effects of TNF on the kinetic parameters of the taurine uptake system --- p.54 / Chapter 3.1.3 --- The effects of TNF concentration on taurine uptake --- p.63 / Chapter 3.1.4 --- The effects of TNF exposure time on taurine uptake --- p.65 / Chapter 3.1.5 --- The effects of TNF on cell volume change in astrocytes --- p.67 / Chapter 3.1.6 --- "Comparison of the effects of TNF on taurine uptake amongst cultured primary rat astrocytes, primary mouse astrocytes and C6 glioma cell line" --- p.69 / Chapter 3.1.7 --- The effects of TNF on taurine release --- p.71 / Chapter 3.1.8 --- The specificity of the effects of TNF on taurine uptake --- p.74 / Chapter 3.1.8.1 --- The effects of TNF on the uptake of amino acids and glucose in primary rat astrocytes --- p.79 / Chapter 3.1.8.2 --- The effects of TNF on neurotransmitters uptake --- p.87 / Chapter 3.1.9 --- The effects of LPS on taurine uptake in astrocytes --- p.92 / Chapter 3.1.10 --- The effects of IFN-¡’ on taurine uptake in astrocytes --- p.97 / Chapter 3.1.11 --- The effects of PMA on taurine uptake --- p.99 / Chapter 3.2 --- The effects of TNF on cell metabolism in rat astrocytes --- p.102 / Chapter 3.2.1 --- The effects of TNF on astrocyte proliferation --- p.102 / Chapter 3.2.2 --- The effects of TNF on RNA synthesis --- p.103 / Chapter 3.2.3 --- The effects of TNF on protein synthesis --- p.106 / Chapter 3.2.4 --- The effects of TNF on basal level of cGMP --- p.108 / Chapter 3.2.5 --- The effects of TNF on protein phosphorylation --- p.111 / Chapter 3.2.6 --- The effects of TNF on calcium uptake --- p.113 / Chapter Chapter IV --- DISCUSSION AND CONCLUSION --- p.116 / References --- p.125