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Role of M4 muscarinic receptors in the central nervous system
Křížová, Monika ; Mysliveček, Jaromír (advisor) ; Slepička, Jakub (referee)
Muscarinic receptors type 4 are one of the five subtypes of muscarinic receptors, that are members of cholinergic receptors family, together with nicotinic receptors. An acetylcholine binding on the receptor triggers the receptor activation and the signal transmission into the cell by G-proteins located on plasmatic membrane, in the case of muscarinic receptors type 4 by Gi/o proteins. The M4 receptors are, just like other subtypes on muscarinic receptors, located in central and peripheral nervous system. In the central nervous system, they are mostly located in striatum and hippocampus. The M4 receptors have a whole range of regulative functions, the most significant one is most likely the regulation of a locomotion in striatum by the dopaminergic system. The muscarinic receptors type 4 are involved in many behavioural and cognitive processes and are therefore studied as potential drug target for the treatment of mental disorders. Key words: muscarinic receptors, M4 muscarinic receptors, GPCRs, cholinergic system, central nervous system
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Modulation of central cholinergic neurotransmission
Valušková, Paulína ; Mysliveček, Jaromír (advisor) ; Cendelín, Jan (referee) ; Jakubík, Jan (referee)
Introduction: Central cholinergic system plays a key role in control of different brain functions such as learning, memory, attention, locomotion and rewards. Disrupted integrity, regulation or capacity of cholinergic signalling is closely connected with cognitive symptoms of several neurodegenerative and neuropsychiatric diseases, as Alzheimer disease, Parkinson disease, attention deficit hyperactivity disorder (ADHD), depression, schizophrenia and increased distractibility. The major neurotransmitter of cholinergic neurons is acetylcholine (ACh) and regulation of ACh levels is main pharmacotherapeutic approach to the treatment of diseases associated with central cholinergic system. The aim of the thesis was to study the changes of central cholinergic neurotransmission with respect to various aspects of modulation of ACh levels in the brain by controlling its release through M4 muscarinic receptors (MR), its hydrolysis by acetylcholinesterase (AChE) or butyrylcholinesterase (BChE) and after hydrolysis in the synapse, regulation of the uptake of metabolite choline by high affinity choline transporter (CHT). Methods: Here we used telemetry to measure locomotor activity and body temperature in mice with selective deletion of M4 MR (M4KO) and their wild type (M4WT) controls under the basal conditions...
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Photodynamic Therapy of Xenotransplanted Human Tumours
Sutoris, Karol ; Gürlich, Robert (advisor) ; Mysliveček, Jaromír (referee) ; Šimša, Jaromír (referee)
SUTORIS, Karol. Fotodynamická terapie xenotransplantovaných lidských tumorů. [Photodynamic therapy of xenotransplanted human tumours]. Prague, 2015. 168 pp. Doctoral Thesis. Charles University in Prague, Third Faculty of Medicine, Department of Surgery Faculty Hospital Královské Vinohrady. Lector GÜRLICH, Robert. Language Czech. In todays clinical practice oncological indications of photodynamic therapy (PDT) are limited primarily to palliative treatment and are used as an adjunct to conventional oncosurgical routines with the aim of improving the quality of life and prolonging patient survival. The efficacy of experimental PDT on xenotransplanted human tumours has been proven in our in vivo study on nu/nu mice. One particular cell line of mammary carcinoma (MDA-MB-231) and two biologically different cell lines of prostate carcinoma (LNCaP, PC- 3) were tested. The key aspect of our experiment was the application of newly developed photosensitizer - hydroxy-aluminum phthalocyanine (AlOH-Pc) in the form of liposomal gel designed for locotopical application. Therapy achieved complete remission in 90% of mice with mammary carcinoma xenografts and in 100% of those with prostate carcinoma xenografts. The new photosensitizer, unlike the older ones, has minimal drug-light interval and does not cause...
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Activation mechanisms of muscarinic M1 receptor by atypical agonists
Randáková, Alena ; Doležal, Vladimír (advisor) ; Mysliveček, Jaromír (referee) ; Krůšek, Jan (referee)
Atypical agonists of muscarinic receptors bind to individual receptor subtypes with comparable affinity but activate them selectively to a certain extent. Molecular mechanism underlying this "functional selectivity" is not known and its elucidation may contribute to development of new atypical functionally selective agonists suitable for therapeutic use. Functional selectivity of atypical muscarinic agonists may be caused by a distinct molecular mechanism(s) of how these compounds activate the receptor. Agonist-specific conformations induced by structurally complex atypical agonists may lead to utilization of a parallel activation mechanism that is different than the activation mechanism induced by non-selective classical agonists. In order to examine this possibility we investigated whether the M1 receptor preferring atypical agonists xanomeline and N-desmethylclozapine, and the classical orthosteric agonists carbachol and oxotremorine, activate the M1 receptor through a common cascade of transmission switches. To this end we mutated key amino acids of the M1 receptor that are essential for ligand binding to the orthosteric binding site (D1053.32 , D993.26 ), receptor activation (transmission switch, D712.50 ), or interaction with G-protein (ionic lock switch, R1233.50 D1223.49 ). We compared...
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Regulace receptorů spřažených s G proteiny Studie muskarinových a β-adrenergních receptorů u M2KO myší
Beneš, Jan ; Mysliveček, Jaromír (advisor) ; Kuncová, Jitka (referee) ; Nováková, Marie (referee)
(in English): The aim of the work was to perform in-depth analysis of M2KO mice both at baseline and upon a challenge with a cold stress and to explore the role of opposing receptors (i.e. adrenoceptors) in adaptation to lacking M2-receptors in the heart. We have performed receptor binding studies, study of receptor gene expression, echocardiography, telemetric monitoring of heart rate, body temperature and activity, heart rate variability and biorhythm analysis, analysis of heart rate response to the application of drugs (carbachol, atropine, isoprenaline, propranolol), assessment of adenylyl cyclase and NO synthase activity, measurement of catecholamine blood concentration and gene expression of catecholamine-synthesizing enzymes. We have found that the disruption of M2-receptor gene caused a compensatory decrease of cardiostimulatory β1-adrenoceptors and β2-adrenoceptors with corresponding down-regulation of the gene expression, M3-receptors were down-regulated as well. Missing M2-receptors were functionally replaced by the main cardioinhibitory β3-adrenoceptors that were up-regulated, not by cardioinhibitory M4-receptors. β3-adrenoceptors were found to signal through adenylyl cyclase instead of NO synthase. All these changes were found in the left ventricle only, so heterologous regulation is...
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The role of whiskers in compensation of visual deficit and the influence of a neurodegenerative disorder on cross-modal compensation in a mousse model of retinal and olivocerebellar degeneration
Voller, Jaroslav ; Vožeh, František (advisor) ; Jagla, Fedor (referee) ; Mysliveček, Jaromír (referee)
Sensory deprivation in one modality can enhance the development of the remaining modalities via mechanisms of synaptic plasticity. Mice of C3H strain suffers from RD1 retinal degeneration that leads to visual impairment at weaning age. Independently on the retinal degeneration there is also present olivocerebellar degeneration caused by Lurcher mutation. This neurodegenerative disorder causes motor deficits, increased CNS excitability as well as changes in synaptic plasticity. The aim of this study was to evaluate a role of whiskers in compensation of the visual deficit and to assess the influence of the olivocerebellar degeneration on this process. To differentiate contribution of the whiskers from other mechanisms that can take part in the compensation, we investigated the effect of both chronic and acute tactile deprivation. We focused on motor skills (rotarod, beam walking test), gait control (CatWalk system), spontaneous motor activity (open field) and the CNS excitability (audiogenic epilepsy). In the seeing mice without olivocerebellar degeneration, the removal of the whiskers had no effect. In the blind animals without olivocerebellar degeneration, chronic tactile deprivation caused changes in gait and impaired the performance in motor tests. Some other compensatory mechanisms were involved but the...
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Adaptace centrálního nervového systému na chybění acetylcholinesterázy
Farár, Vladimír ; Mysliveček, Jaromír (advisor) ; Jakubík, Jan (referee) ; Cordero-Erausquin, Matilde (referee)
Acetylcholinesterase (AChE) effectively hydrolyzes acetylcholine (ACh). The inhibition of AChE is generally lethal and mice without AChE in all tissues (AChE KO) have severe impairments. In the brain, AChE is anchored in the plasma membrane by proline-rich membrane anchor (PRiMA), while in the muscles, AChE is anchored by collagen Q (ColQ) in the basal lamina. We report here that the PRiMA KO mice, in which AChE is essentially eliminated in the brain, show very little changes in behavior despite an excess of ACh in the brain and adaptation of ACh receptors comparable to those seen in AChE KO mice. Moreover, when AChE cannot interact with ColQ and PRiMA, the phenotype resembles that of AChE KO mice, but the biochemical changes in the brain are similar to those in PRiMA KO mice. PRiMA KO mice also differ from other AChE-deficit mice strains in their responses to AChE inhibitor. Our results suggest that AChE in the peripheral tissues is the major target of AChE inhibitors and AChE absence in the peripheral tissues is the leading cause of the phenotype of AChE KO mice.
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Characterisation and regulation of muscarinic and adrenergic receptors Subtitle: The effect of stress on muscarinic and adrenergic receptors in the lung and in the heart
Nováková, Martina ; Mysliveček, Jaromír (advisor) ; Hynie, Sixtus (referee) ; Mravec, Boris (referee)
The aim of this thesis was to clarify the influence of the stress on the adrenergic and muscarinic receptors in the heart and in the lungs. Research was perform on rat hearts and lungs and on the hearts and lungs of the CRH KO mice. First, we assessed mRNA levels of all α- and β-adrenergic receptor and muscarinic receptor subtypes. Subsequently, we performed the radioligand-binding studies to determine densities of these receptors. We identified all three α1-adrenergic receptor subtypes in the rat lungs. In the lungs of WT mice, we found that the amount of α1-adrenergic and muscarinic receptors was sex-dependent. Densities of the former were higher in females and those of the latter were higher in males. There was no difference between males and females in β-adrenergic receptor density. As for CRH KO mice, the basal densities of studied receptors were lower than in CRH WT mice (except β1-adrenergic receptors in females). The main purpose of the thesis was to detect the immobilization-induced changes in the studied receptors in the kontrol (WT) and CRH KO mice. Short-term and long-term immobilization caused decrease in all α1-adrenergic receptor subtypes in females, whereas only α1A-adrenergic receptors decreased in males. The amount of β1-adrenergic receptors decreased in males and remained without...
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