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Study of the action of ivermectin on purinergic P2X4 receptor
Jelínková, Irena ; Teisinger, Jan (advisor) ; Šťastný, František (referee) ; Krůšek, Jan (referee)
The P2X4 receptor is ATP-gated cation channel. It is the only mamrnalian purinergic receptor which is modulated by extracellularly applied ivennectin (IVM). rVM is an allosteric modulator that has several effects on receptor [unction: it increases sensitivity to agonists, potentiates maximum current amplitude and prolongs the deactivation kinetics of the channel after agonist washout. The aim of this study was to localize IVM binding site and using its positive allosteric effect to get new inforrnatioll about the structure and function of P2X. receptor. Initially we focused on identification of regions and residues responsible for IVM effect on channel function. We used several chimeras of P2X2 and P2X. receptors and P2X. receptors with single point mutatioll. Experiments with chimeric receptors revealed that extracellular sequence V49-V61 but not tbe sequ nce V64-Y315 is important for the effects af IVM on channel deactivation. Receptor-specific alanine mutations placed in transmembrane domains 029-V61 and N338-L358 showed the importance of residues W50, V61 and V357 for TVM effect Oll channel deactivation. We tested further the irnportance of aH residues in transmembrane domains. Cysteine scanning mutagenesis supported the relevance of previously identified W50 residue and showed the importance ofresidues...
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název v anglickém jazyce není uveden
Kaiser, Miroslav ; Šťastný, František (advisor) ; Marešová, Dana (referee) ; Kršiak, Miloslav (referee)
Although the cause of schizophrenia is not fully elucidated, there is increasing evidence that this severe psychiatric illness may result from a disturbance early in development. Evidence for this so called "neuro- developmental hypothesis" comes from a variety of sources, including epidemiological, gynecological, psychological and pathological data. Fetal and perinatal hypoxia-ischemia belongs to obstetric complications which can increase the risk of schizophrenia. These complications such as premature rupture of fetal membranes, low birth weight, newborn immaturity, forceps delivery and resuscitation at birth are frequently accompanied by hypoxic-ischemic brain damage during delivery. We assume that perinatal hypoxia together with genetic factors influence a susceptibility to schizophrenia and represent significant risk factors of this disease. We used a heuristic model in our study to determine if brain hypoxic-ischemic episode can actually cause not only changes of biochemical markers but also behavioral changes resembling those seen in schizophrenia. Neurodevelopmental processes in neonatal rats continue during the first three weeks of postnatal life and reach the level of those observed in human newborns at the end of the second postnatal week. Therefore, rat pups at postnatal day 12 (PND12) were used...
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Study of the action of ivermectin on purinergic P2X4 receptor
Jelínková, Irena ; Teisinger, Jan (advisor) ; Šťastný, František (referee) ; Krůšek, Jan (referee)
The P2X4 receptor is ATP-gated cation channel. It is the only mamrnalian purinergic receptor which is modulated by extracellularly applied ivennectin (IVM). rVM is an allosteric modulator that has several effects on receptor [unction: it increases sensitivity to agonists, potentiates maximum current amplitude and prolongs the deactivation kinetics of the channel after agonist washout. The aim of this study was to localize IVM binding site and using its positive allosteric effect to get new inforrnatioll about the structure and function of P2X. receptor. Initially we focused on identification of regions and residues responsible for IVM effect on channel function. We used several chimeras of P2X2 and P2X. receptors and P2X. receptors with single point mutatioll. Experiments with chimeric receptors revealed that extracellular sequence V49-V61 but not tbe sequ nce V64-Y315 is important for the effects af IVM on channel deactivation. Receptor-specific alanine mutations placed in transmembrane domains 029-V61 and N338-L358 showed the importance of residues W50, V61 and V357 for TVM effect Oll channel deactivation. We tested further the irnportance of aH residues in transmembrane domains. Cysteine scanning mutagenesis supported the relevance of previously identified W50 residue and showed the importance ofresidues...
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název v anglickém jazyce není uveden
Kaiser, Miroslav ; Šťastný, František (advisor) ; Marešová, Dana (referee) ; Kršiak, Miloslav (referee)
Although the cause of schizophrenia is not fully elucidated, there is increasing evidence that this severe psychiatric illness may result from a disturbance early in development. Evidence for this so called "neuro- developmental hypothesis" comes from a variety of sources, including epidemiological, gynecological, psychological and pathological data. Fetal and perinatal hypoxia-ischemia belongs to obstetric complications which can increase the risk of schizophrenia. These complications such as premature rupture of fetal membranes, low birth weight, newborn immaturity, forceps delivery and resuscitation at birth are frequently accompanied by hypoxic-ischemic brain damage during delivery. We assume that perinatal hypoxia together with genetic factors influence a susceptibility to schizophrenia and represent significant risk factors of this disease. We used a heuristic model in our study to determine if brain hypoxic-ischemic episode can actually cause not only changes of biochemical markers but also behavioral changes resembling those seen in schizophrenia. Neurodevelopmental processes in neonatal rats continue during the first three weeks of postnatal life and reach the level of those observed in human newborns at the end of the second postnatal week. Therefore, rat pups at postnatal day 12 (PND12) were used...
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A study of the modulation of recombinant and native NMDA receptors by physical and chemical factors
Cais, Ondřej ; Vyklický, Ladislav (advisor) ; Blahoš, Jaroslav (referee) ; Chvátal, Alexandr (referee) ; Šťastný, František (referee)
Excitatory synaptic transmission in mammalian CNS is mostly mediated by ionotropic glutamate receptors. NMDA receptors, one of three subclasses of this ligand-activated ion channels family, are involved in memory formation and learning and also play a role in the pathogenesis of various neurodegenerative diseases. Current knowledge about NMDA receptors function is predominantly based upon results of in vitro experiments conducted at room temperature, far from physiological. The aim of this PhD thesis was to describe the temperature dependence of NMDA receptors. We determined the rate constants that characterise each step in the mechanism of recombinant NR1/NR2B receptors activation in the temperature range 25-45řC. The receptor desensitization, resensitization and glutamate unbinding turned out to be the most temperature sensitive of these processes. In addition to that, we described the temperature dependence of deactivation kinetics in various experimental models of NMDA receptors (both recombinant and native). The second part of the thesis focused on the modulation of NMDA receptors function by steroid compounds derived from pregnanolone sulfate, an endogenously occuring neurosteroid. We tested 21 steroids that showed various degree of ability to inhibit (or, in one case, potentiate) the current...
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název v anglickém jazyce není uveden
Bubeníková, Věra ; Šťastný, František (advisor) ; Mareš, Pavel (referee) ; Rokyta, Richard (referee)
The aim of the study was to create a new animal model of schizophrenia based on a neurodevelopmental hypothesis. The central idea of the project has been that a transient insult at early postnatal age will reveal itself in the form of delayed distinct behavioral changes that can be related to schizophrenia. The experimental design used intracerebroventricular (icv) infusion of N-acetyl-L-aspartyl-L-glutamate (NAAG) to rats at postnatal day 12, followed by combination of histology and quantitative morphology in brain tissue obtained within 24 and 96 hours of the NAAG administration and used to investigate possible neurodegeneration. Finally, a battery of behavioural tests was performed several weeks later (late adolescence/early adulthood). NAAG is the most abundant neuropeptide and interacts with the active site of metabotropic glutamate receptors (mGluR II), however, it is also an agonist at NMDA receptors. Neonatal icv infusion of NAAG resulted in detectable damaged neurones in gyrus dentatus. The damage appeared greater at 24 hours, as compared to 96 hours, after the infusion. The presence of damaged neurons was easily demonstrated by Nissl stain, Fluoro Jade В staining combined with Hoechst 33342 and by TUNEL technique. Neonatal administration of NAAG resulted in the appearance of specific, potentially...
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