No description available.

Mémoire numérisé par la Direction des bibliothèques de l'Université de Montréal.

by Ngai Wing Keung Clement. / Thesis (M.Phil.)--Chinese University of Hong Kong, 1992. / Includes bibliographical references (leaves 172-206). / Chapter 1. --- Acknowledgment --- p.iv / Chapter 2. --- Abstract --- p.v / Chapter 3. --- Lists of Tables and Figures --- p.vii / Chapter 4. --- General Introduction --- p.1 / Chapter 5. --- Review of Literature --- p.5 / Chapter 5.1. --- Historical Background of Natriuretic Peptides --- p.5 / Chapter 5.2. --- Secretion of Natriuretic Peptides --- p.8 / Chapter 5.3. --- Structure and Function Relationships --- p.12 / Chapter 5.4. --- Physiological Actions --- p.17 / Chapter 5.5. --- Natriuretic Peptide Receptors and Second Messengers --- p.22 / Chapter 5.6. --- Clinical Implications --- p.28 / Chapter 5.7. --- Review on Astrocytes --- p.31 / Chapter 6. --- General Materials and Methods --- p.43 / Chapter 6.1. --- Sources of Chemicals --- p.43 / Chapter 6.2. --- Culture of Mouse Astrocytes --- p.43 / Chapter 6.3. --- Culture of human astrocytoma cells --- p.57 / Chapter 6.4. --- Protein Determination --- p.57 / Chapter 6.5. --- Gamma Counting --- p.58 / Chapter 6.6. --- Beta Counting --- p.58 / Chapter 7. --- Receptor Identification and Characterization --- p.59 / Chapter 7.2 --- Materials and Methods --- p.60 / Chapter 7.3 --- Results --- p.66 / Chapter 7.4 --- Discussion --- p.81 / Chapter 8. --- Second Messenger Systems --- p.83 / Chapter 8.1 --- Introduction --- p.83 / Chapter 8.2 --- Materials and Methods --- p.85 / Chapter 8.3 --- Results --- p.97 / Chapter 8.4 --- Discussion --- p.109 / Chapter 9. --- Biological Actions of Natriuretic Peptides --- p.113 / Chapter 9.1. --- Potassium Transport --- p.113 / Chapter 9.1.1. --- Introduction --- p.113 / Chapter 9.1.2. --- Materials and Methods --- p.116 / Chapter 9.1.3. --- Results --- p.121 / Chapter 9.1.4. --- Discussion --- p.131 / Chapter 9.2 --- Taurine Release --- p.135 / Chapter 9.2.1. --- Introduction --- p.135 / Chapter 9.2.2. --- Materials and Methods --- p.137 / Chapter 9.2.3. --- Results --- p.139 / Chapter 9.2.4. --- Discussion --- p.143 / Chapter 9.3 --- Thymidine Incorporation --- p.144 / Chapter 9.3.1. --- Introduction --- p.144 / Chapter 9.3.2. --- Materials and Methods --- p.146 / Chapter 9.3.3. --- Results --- p.148 / Chapter 9.3.4. --- Discussion --- p.156 / Chapter 10. --- Interaction with Other Hormonal Systems --- p.160 / Chapter 10.1 --- Introduction --- p.160 / Chapter 10.2 --- Materials and Methods --- p.162 / Chapter 10.3 --- Result --- p.163 / Chapter 10.4 --- Discussion --- p.167 / Chapter 11. --- Conclusion --- p.169 / Chapter 12. --- References --- p.172 / Chapter 13. --- Appendix --- p.208

CoordenaÃÃo de AperfeiÃoamento de Pessoal de NÃvel Superior / In last years, considerable efforts have been made to identify neuroactive and neuroprotective peptides derived from the venom of different natural species. In this work, were studied the activity of Dinoponera quadriceps native (DqV) and denatured (DqDV) venom on chemically induced seizures models in vivo and on in vitro cortical astrocytes viability. Male Swiss mice (28- 33g) were pretreated with DqV (0.1 or 0.5 mg/kg, e.v., n = 6-8), DqDV (0.5 or 2.0 mg/kg, i.p., n = 6-8) or DqDV (0.1 or 0.5 mg/kg, e.v., n = 6-8). 30 minutes after the intraperitoneal pretreatment or ten minutes after intravenous pretreatment with the venom was induced seizures in all animals by the administration of pentylenetetrazole (80 mg/kg) pilocarpine (400 mg/kg) or strychnine (3.0 mg/kg). In behavioral analysis, we recorded the time to the first seizure and to death and the percentage of survival. To determine the parameters of oxidative stress was dissected three brain areas (prefrontal cortex, hippocampus and striatum) of animals used in behavioral analysis, in order to determine the degree of lipid peroxidation, by measuring the levels of malondialdehyde (MDA), and the content of nitrite. In in vitro assay, cell viability of cortical astrocytes was determined after treatment with different concentrations of DqV, PTZ and DqV + PTZ. The data were analyzed by ANOVA followed by a Student Newman-Keuls (for in vivo tests) or Bonferroni (for in vitro experiments) as post hoc tests. It was observed that the DqV had effect only on the PTZ model, both in behavioral analysis as for the determination of the oxidative parameters. Pretreatment with DqV significantly reduced the time until the occurrence of first seizure (0.1 mg/kg: 77.83 Â 5.27 compared to the 101.0 Â 3.31 seconds in the control group; 0.5 mg/kg: 74.43 Â 3.94 compared to the 101.0 Â 3.31 seconds in the control group), while DqDV caused an increase in the percentage of survival when pretreated by i.p. (0.5 mg/kg: 25% of survival compared with 0% in the control group, 2.0 mg/kg: 62.5% of survival compared with 0% in control group) and e.v (0.5 mg/kg: 28.57% of survival compared with 0% in control group). routes. In relation to the oxidative stress parameters, both pretreatments with DqV and with DqDV caused increases of MDA levels in all three brain areas analyzed. The nitrite content also increased after pretreatment with DqV in the three areas of the brain and after pretreatment with DqDV via e.v., only in the hippocampus. About cell viability assays, were observed that DqV was not able to change this parameter. The PTZ reduced the cell viability of astrocytes in a dose-dependent way, with an IC 50 (cytotoxicity index to 50% of the cell population under study) corresponding to 33.12 mM. The combined treatment of DqV (100 Âg/mL) and PTZ (IC 50) also caused a reduction in cell viability. The results suggest that the DqV, probably, has both neurotoxic and neuroprotective components, and that the astrocytes should be the cells involved in the venomâs neurotoxicity. / Nos Ãltimos anos, esforÃos considerÃveis tÃm sido feitos no sentido de identificar peptÃdeos naturais neuroativos e neuroprotetores derivados do veneno de diferentes espÃcies animais. Nesse trabalho foi estudada a atividade do veneno de Dinoponera quadriceps nativo (vDq) e desnaturado (vdDq) em modelos animais de convulsÃo quimicamente induzidos in vivo e sobre a viabilidade de astrÃcitos corticais in vitro. Camundongos Swiss machos (28-33g) foram prÃ-tratados com o vDq (0,1 ou 0,5 mg/kg, e.v, n= 6-8), vdDq (0,5 ou 2,0 mg/kg, i.p., n= 6-8) ou com vdDq (0,1 ou 0,5 mg/kg, e.v., n= 6-8). Meia hora apÃs o prÃ-tratamento intraperitoneal ou dez minutos apÃs o prÃ-tratamento endovenoso com o veneno foi induzida a convulsÃo em todos os animais atravÃs da administraÃÃo de pentilenotetrazol (80 mg/kg), pilocarpina (400 mg/kg,) ou estricnina (3,0 mg/kg). Na anÃlise comportamental, foram registrados os tempos para ocorrÃncia da primeira convulsÃo e morte e o percentual de sobrevida. Para determinaÃÃo dos parÃmetros de stress oxidativo foram utilizadas trÃs Ãreas cerebrais (cÃrtex prÃ-frontal, hipocampo e corpo estriado) de animais utilizados na anÃlise comportamental, a fim de se determinar o grau de peroxidaÃÃo lipÃdica, pela mensuraÃÃo dos nÃveis de malondialdeÃdo (MDA), e o conteÃdo de nitrito. No ensaio in vitro, foi determinada a viabilidade celular de astrÃcitos corticais apÃs o tratamento com diferentes concentraÃÃes de vDq, PTZ e vDq + PTZ. Os dados foram analisados por ANOVA e Student-Newman-Keuls como pÃs-teste, para os ensaios in vivo, e ANOVA seguido pelo pÃs-teste de Bonferroni, para as experimentaÃÃes in vitro. Foi observado que o vDq apresentou efeito apenas no modelo de PTZ, tanto na anÃlise comportamental quanto na determinaÃÃo dos parÃmetros oxidativos. O prÃ-tratamento com vDq reduziu significativamente o tempo para ocorrÃncia da primeira convulsÃo (0,1 mg/kg: 77,83 Â 5,27 comparado com 101,0 Â 3,31 segundos no grupo controle; 0,5 mg/kg: 74,43 Â 3,94 comparado com 101,0 Â 3,31 segundos no grupo controle), enquanto que o vdDq causou aumento do percentual de sobrevida quando prÃ-tratado por via i.p. (0,5 mg/kg: 25% de sobrevida comparado com 0% no grupo controle; 2,0 mg/kg: 62,5% de sobrevida comparado com 0% no grupo controle) e e.v (0,5 mg/kg: 28,57% de sobrevida comparado com 0% no grupo controle). Em relaÃÃo aos parÃmetros de estresse oxidativo, tanto o prÃ-tratamento com o vDq quanto com o vdDq causaram aumentos dos nÃveis de MDA nas trÃs Ãreas cerebrais analisadas. O conteÃdo de nitrito tambÃm sofreu elevaÃÃo apÃs prÃ-tratamento com vDq, nas trÃs Ãreas cerebrais, e apÃs o prÃ-tratamento com vdDq, via e.v., apenas no hipocampo. Quanto aos ensaios de viabilidade celular, foi observado que o vDq, isoladamente, nÃo foi capaz de alterar esse parÃmetro. O PTZ causou reduÃÃo da viabilidade de astrÃcitos de modo dose-dependente e com uma IC 50 (Ãndice de citotoxicidade para 50% da populaÃÃo celular em estudo) correspondente a 33,12 mM. O tratamento combinado entre vDq (100 Âg/mL) e PTZ (IC 50) tambÃm causou reduÃÃo da viabilidade das cÃlulas. Os resultados sugerem que o vDq possivelmente apresenta ambos componentes neurotÃxicos e neuroprotetores, e os astrÃcitos devem ser uma das cÃlulas envolvidas na neurotoxicidade do veneno.

In neurodegenerative diseases, the CNS becomes inflamed through activation of pathways, including the NF-B pathway. Some of the therapies for those diseases target neuroinflammatory pathways. Here, we explore the mechanisms for the upregulation of a subset of genes following a restimulation of the NF-B pathway. We discover that this upregulation occurs independent of IRF1 expression and type 1 interferon signaling. A knockdown of IRF1 using siRNA and an inhibition of JAK proteins using inhibitor AG490 both had no effect on priming. A secreted factor was found to upregulate the expression of both this subset of genes and genes encoding pro-inflammatory cytokines induced by NF-B activation. We also explored the role of IRF1 in a mouse model of multiple sclerosis. We found that the deletion of IRF1 from oligodendrocytes diminished EAE severity. A deletion of IRF1 from myeloid cells within mice did not diminish EAE severity, however showed a promising decrease in the expression of certain inflammatory genes. Thus, IRF1 plays a critical role in fine-tuning inflammatory responses in the brain.

Abstract not available

There is evidence that the excitotoxin quinolinic acid (QUIN) synthesized through the kynurenine pathway (KP) by activated microglia may play a role in the pathogenesis of several major neuroinflammatory diseases and more particularly in Alzheimer??s disease (AD). The hypothesis of this project is QUIN affects the function and morphology of astrocytes. In this study I used human foetal astrocytes stimulated with AD associated cytokines including IFN-gamma, TNF-alpha, TGF-alpha and different concentrations of QUIN ranging from low physiological to high excitotoxic concentrations. I found that QUIN induces IL-1beta expression in human astrocytes and subsequently, contribute to the inflammatory cascade that is present in AD pathology. Glial fibrillary acid protein (GFAP) and vimentin protein expression were complementary in expression to each other after 24 hr stimulation with different QUIN doses. However, there were marked increases in GFAP levels and reduction in vimentin levels compared to controls with QUIN treatment indicating that QUIN can trigger astrogliosis in human astrocytes. Glutamine synthetase (GS) activity was used as a functional metabolic test for astrocytes and I found a dose-dependent inhibition of GS activity by QUIN. This inhibition was inversely correlated with iNOS expression whereby reduced GS activity is accompanied with an increase expression of iNOS in human astrocytes. These results suggest that reduction in GS activity can lead to accumulation of extracellular glutamate then leading to exacerbated excitotoxicity via NMDA receptor over-activation and ultimately neuronal death. PCR array results showed that at least four different pathways were activated with pathological concentration of QUIN including p38 MAPK that is associated with pro-inflammatory cytokine production, ERK/MAPK growth and differentiation that can modulate structural proteins, mitochondrial-induced apoptotic cascade and cell cycle control pathway. QUIN-induced astrogliosis and excitotoxicity could lead to glial scar formation and prevention of axonal growth thus exacerbation of neurodegeneration via synaptosomal NMDA receptor over-activation. All together, this study showed that, in the context of AD, QUIN is an important factor for astroglial activation, dysregulation and death, which can be mediated by the previously mentioned pathways.

Thesis (Ph. D.)--University of Missouri-Columbia, 2006. / The entire dissertation/thesis text is included in the research.pdf file; the official abstract appears in the short.pdf file (which also appears in the research.pdf); a non-technical general description, or public abstract, appears in the public.pdf file. Title from title screen of research.pdf file viewed on (March 3, 2007) Vita. Includes bibliographical references.

Astrocytes, the major type of non-neuronal cells in the brain, play an important functional role in the brains extracellular potassium (K+) and pH homeostasis. Pathological brain states have been shown to cause astrocyte swelling. However, these volume changes have never before been verified to occur in response to physiological activity. In the present thesis, two-photon laser scanning microscopy was used to visualize real-time astrocyte volume changes in the stratum radiatum of the CA1 region of the hippocampus. Astrocyte somas and primary processes were observed to swell by 19.0±0.9% in response to a physiological (3 mM) increase in the concentration of extracellular K+. Astrocyte swelling was partially mediated by K+ influx through inwardly rectifying K+ channels (Kir), as their inhibition resulted in a significant decrease of the increased K+ induced astrocyte swelling (13.9±0.9%). In addition, the bicarbonate ion (HCO3-) was found to play a significant role in the increased K+ induced astrocyte swelling. The astrocyte swelling was significantly decreased when the influx of HCO3- was decreased in 1) a HCO3- free extracellular solution (5.4±0.7%), 2) in the presence of an extracellular carbonic anhydrase inhibitor (11.4±0.6% ), and 3) when the activity of the sodium-bicarbonate cotransporter (NBC) was blocked (8.3±0.7%) . Conversely, astrocytes were found to shrink by 7.7±0.5% in response to ã-Amino-butyric Acid (GABA) receptor activation. GABAA receptor mediated astrocyte shrinkage was significantly decreased to 5.0±0.6% when HCO3- efflux was reduced. Furthermore, in this thesis it was shown for the first time that astrocytes swell in response to neuronal stimulation (4.0±0.4%). This activity induced astrocyte swelling was significantly decreased to 1.5±0.2% in a HCO3- free extracellular solution. These astrocyte volume changes may have important implications for the regulation of brain activity under both physiological and pathological brain states.

Astrocytes, the major type of non-neuronal cells in the brain, play an important functional role in the brains extracellular potassium (K+) and pH homeostasis. Pathological brain states have been shown to cause astrocyte swelling. However, these volume changes have never before been verified to occur in response to physiological activity. In the present thesis, two-photon laser scanning microscopy was used to visualize real-time astrocyte volume changes in the stratum radiatum of the CA1 region of the hippocampus. Astrocyte somas and primary processes were observed to swell by 19.0±0.9% in response to a physiological (3 mM) increase in the concentration of extracellular K+. Astrocyte swelling was partially mediated by K+ influx through inwardly rectifying K+ channels (Kir), as their inhibition resulted in a significant decrease of the increased K+ induced astrocyte swelling (13.9±0.9%). In addition, the bicarbonate ion (HCO3-) was found to play a significant role in the increased K+ induced astrocyte swelling. The astrocyte swelling was significantly decreased when the influx of HCO3- was decreased in 1) a HCO3- free extracellular solution (5.4±0.7%), 2) in the presence of an extracellular carbonic anhydrase inhibitor (11.4±0.6% ), and 3) when the activity of the sodium-bicarbonate cotransporter (NBC) was blocked (8.3±0.7%) . Conversely, astrocytes were found to shrink by 7.7±0.5% in response to ã-Amino-butyric Acid (GABA) receptor activation. GABAA receptor mediated astrocyte shrinkage was significantly decreased to 5.0±0.6% when HCO3- efflux was reduced. Furthermore, in this thesis it was shown for the first time that astrocytes swell in response to neuronal stimulation (4.0±0.4%). This activity induced astrocyte swelling was significantly decreased to 1.5±0.2% in a HCO3- free extracellular solution. These astrocyte volume changes may have important implications for the regulation of brain activity under both physiological and pathological brain states.