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The monitoring of melatonin rhythm in monozygotic tweens as a marker of genetic control of circadian rhythmicity
Spišská, Veronika ; Bendová, Zdeňka (advisor) ; Svobodová, Irena (referee)
In human population, there are individual differences in the length of endogenous period which is generated by transcriptional-translational loops of clock genes in the suprachiasmatic nucleus of the hypothalamus (SCN). On the basis of these differences we divide people into the early, late or neither chronotype. Chronotypes differ from each other mainly in the rhythm of the hormone melatonin, which is used as a marker of endogenous period. They also differ in rhythm of locomotor activity. The aim of this study was to describe a difference in the phase of the circadian system in three pairs of monozygotic twins aged 8-10 years who live together and share the same daily routine. To describe these differences, we used a questionnaire CCTQ, melatonin radioimmunoassay and actigraphy. Acording to the questionnaire, one boy was determined as early chronotype (A3), one girl was determined as late chronotype (B3) and the other children were determined as neither chronotype (A2, B2, C2, C3). Among the twins there were marked differences both in the rhythm of melatonin and in the morning activity. These differences confirmed the chronotype assessed by questionnaire in the boy A3 and the girl B3. On the contrary, differences in melatonin rhythm did not confirm neither chronotype in siblings C2 and C3, and...
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Role of intestinal circadian clock in epithelial transport, proliferation, and tumourigenesis
Soták, Matúš ; Pácha, Jiří (advisor) ; Bendová, Zdeňka (referee) ; Herichová, Iveta (referee)
AABBSSTTRRAACCTT The molecular circadian clock enables anticipation of environmental changes. In mammals, clocks are ubiquitously present in almost all tissues and they are comprised of transcriptional-translational feedback loops of the so-called clock genes. The central clock represents the intrinsic pacemaker which is located in suprachiasmatic nuclei (SCN) of hypothalamus and synchronizes peripheral clocks. Clockwork system in alimentary tract and its regulatory link to intestinal functions are poorly understood. Therefore the objective of the thesis was to characterize molecular clock in particular parts of the rat intestine and to elucidate its link to the intestinal transport, regulation of cell cycle and neoplastic transformation in colonic tissue. We used quantitative RT-PCR (qPCR) to determine circadian profiles of mRNA expression of clock genes in the epithelium of duodenum, jejunum, ileum, and colon of rat. Furthermore, we analysed the expression of genes coding sodium chloride transporters and channels as well as cell cycle regulators in colon. To focus more precisely on different structures of intestinal epithelia we used laser capture microdissection. In addition, we performed Ussing chamber measurements to determine the colonic electrogenic transport. To study the contribution of circadian...
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The effect of melatonin for sleep regulation in humans with autism
Spišská, Veronika ; Bendová, Zdeňka (advisor) ; Kopřivová, Jana (referee)
Autism is, mainly in children, accompanied by a variety of sleep and circadian disorders. These disorders may be partly caused by problems in the synthesis of melatonin. This work is interested in sleep and circadian problems of children with autism and their possible treatment with exogenous melatonin. The paper presents the symptoms of autism and its causes, the basic principles of circadian system, the regulation of melatonin synthesis and the characteristics of sleep. The last chapter describes the changes in melatonin synthesis and disturbances in the production of other hormones (serotonin, cortisol). Sleep and circadian disorders in autistic individuals and their impact on other symptoms of autism such as communication and socialization are also listed in the last chapter. There are also cases of the treatment of sleep disorders in autistic children with melatonin reported. Treatment with melatonin improves sleep disorders and has almost no side effects. Key words: autism, circadian system, melatonin, sleep
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Animal models of schizophrenia based on NMDA receptors
Svojanovská, Markéta ; Stuchlík, Aleš (advisor) ; Bendová, Zdeňka (referee)
Glutamate is the major excitatory neurotransmitter in the mammalian central nervous system responsible for information processing including learning and memory. Its concentration must be maintained at the correct physiological levels because a deficiency or an excess of glutamate can cause variety of disruptions of CNS function. Glutamate acts also via the NMDA receptor, a specific type of ionotropic glutamate receptor. A number of studies have suggested that glutamatergic NMDA receptors play an important role in an ability to learn, memory, brain development and brain function. Importantly, their reduced function is considered as a possible factor for the pathogenesis of schizophrenia. The NMDA receptor hypofunction hypothesis settles on findings that non-competitive NMDA receptor antagonists such as ketamine, phencyclidine and dizocilpine (MK-801) can induce schizophrenic-like symptoms in healthy individuals and exacerbates symptoms in schizophrenic patients; therefore they are often used in animals to induce schizophrenia-like behavior. All these animal models contribute to a better understanding of the etiology and pathophysiology of schizophrenia and may serve as a useful tool in searching for new antipsychotic agents. Key words: glutamate, schizophrenia, memory, NMDA receptors,...
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Circadian system in bulbus olfactorius
Kyclerová, Hana ; Bendová, Zdeňka (advisor) ; Polidarová, Lenka (referee)
The mammalian circadian system is composed of major circadian pacemaker located in the hypothalamic suprachiasmatic nuclei and peripheral circadian oscillators. Molecular mechanism of interlocked transcription-translation feedback loops, as a characteristic of each circadian oscillator, results in circadian rhythms. Peripheral oscillators in other brain parts and body organs are driven by the signals from the suprachiasmatic nuclei. Some structures however generate oscillations independently from the major circadian pacemaker. The best described of them resides in the mammalian retina. The retinal circadian oscillator regulates local rhythmic synthesis of melatonin, retinal pH and photoreceptors viability. Recently, some studies occur charactering the existence of such an independent circadian oscillator also in bulbus olfactorius. For example, the olfactory bulb circadian oscillator drives olfactory sensitivity during the day. Studies about development of the mamalian circadian system have shown that the olfactory bulb circadian oscillator matures earlier than the major circadian pacemaker and represents its function during early development. Powered by TCPDF (www.tcpdf.org)
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Expression of G-protein Dexras1 in the rat suprachiasmatic nucleus.
Marschnerová, Tereza ; Bendová, Zdeňka (advisor) ; Ostašov, Pavel (referee)
The main mammalian circadian oscillators, suprachiasmatic nuclei (SCN), are stored in the hypothalamus along the third ventricle. They are responsible for time synchronization of behavioral and physiological processes in the body. The endogenous period and phase are controlled by rhythmic changes in the external environment, especially by changing of light and darkness. The light pulse during the night activates NMDA type glutamate receptors, neuronal NO synthase kinase and MAPK ERK1 / 2. This signaling pathway affects the transcription of clock genes and thus the mechanism generating circadian oscillations. Protein Dexras1 acts as a nucleotide exchanger monomer and heterotrimer Gi/o protein and is considered a connecting link between the activation of NMDA glutamate receptor agonist, nNOS signaling and phosforylation of ERK1 / 2. It is rhythmically expressed in the suprachiasmatic nucleus and its genetic deletion leads to abnormal light synchronization.This may indicate its role in the signaling cascade that results in the information concerning the light pulse from the retina to the SCN. The aim of this thesis was to describe the expression of mRNA and protein Dexras1 in the development of the rat suprachiasmatic nuclei and correlate it with the maturation mechanism of light synchronization. Our...
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The influence of endotoxin lipopolysaccharide on the circadian system
Štěrbová, Petra ; Bendová, Zdeňka (advisor) ; Svobodová, Irena (referee)
The biological clock are responsible for entraining of circadian rhytms in many physiological and behavioral functions, including the activity of immune system. In mammals, the biological clock are located in the suprachiasmatic nuclei of the hypothalamus and also on the periphery. The main oscillator in suprachiamatic nuclei synchronizes the phases of the peripheral oscillators in tissues and the other brain areas and it also drives synthesis of melatonin in the pineal gland. Its endogenous period Tau (τ) is being synchronized with external 24-hour day under normal conditions, mostly by the light-dark cycles during the day and night and also by non-photic stimuli. By the application of endotoxin, disruption of normal endogenous conditions in organism causes activation of the immune response, behavioral changes and phase shifts of the circadian rhythmicity. This work examines effect of acute application endotoxin lipopolysaccharid on the cellular signaling cascades in the cells of suprachiasmatic nuclei and pineal gland. Our data indicate, that levels of phosphorylated forms of kinases GSK3β, ERK1/2 and levels of phosphorylated forms transcription factors STAT3 are substantially modificated after administration of lipopolysaccharid in the suprachiasmatic nuclei and especially in the pineal gland of...
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The mechanism of regulation of NMDA receptors transport to the cell surface
Lichnerová, Katarína ; Horák, Martin (advisor) ; Bendová, Zdeňka (referee)
N-methyl-D-aspartate (NMDA) receptors are a class of ionotropic glutamate receptors, involved in excitatory synaptic transmission, synaptic plasticity and excitotoxicity. They form heterotetrameric complexes composed of GluN1, GluN2A-D and/or GluN3A-B subunits that are activated by glutamate and glycine. Previous reports showed that different subunits of NMDA receptors, especially the GluN2 subunits, confer different functional and pharmacological properties on the receptor complexes. However, the subunit-dependent differences in the regulation of intracellular processing and transport of NMDA receptor subtypes has not been clearly elucidated. The aim of this work was to clarify the mechanisms of regulation of the NMDA receptor transport. In our experiments we performed immunocytochemistry of receptors on heterologous COS-7 cells and cultured cerebellar granule cells (CGC), both expressing recombinant NMDA receptors. The results of my work show that the transport of NMDA receptors is regulated by presence of GluN2A and GluN2B subunits. Our results further showed that transport of the GluN1/GluN2C receptors is regulated by three specific areas of the GluN2C subunit: i) the A2 segment within the amino- terminal domain, ii.) the M3 domain, and iii.) the proximal part of the C-terminus containing the...
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Struktura a funkce rekombinantního P2X4 receptoru
Rokič, Miloš ; Zemková, Hana (advisor) ; Vlachová, Viktorie (referee) ; Bendová, Zdeňka (referee)
4 Abstract Purinergic P2X receptors are membrane ion channels activated by extracellular ATP. There are seven isoforms of mammalian P2X receptors designated as P2X1-7, which according to their structure represent a specific family of ligand gated ionic channels, with extraordinary structural/functional properties. The P2X receptor consists of three subunits and each subunit has two transmembrane domains. Crystalographic data demonstrate that ionic channel pore is situated between the second transmembrane domains. Crystal structure of P2X4 receptor from the zebrafish (Danio rerio) is available in both open and closed state of the channel and the exact structure of ATP binding site is solved. The aim of this thesis was to study the structure-function relationships in a model of recombinant P2X4 receptor of the rat. By employing the point mutagenesis and electrophysiological recording, the functional importance of conserved cysteine residues in the ectodomain and amino acid residues which form the extracellular vestibule was investigated. All ten cysteins were substituted one by one with alanine or threonine and ATP-induced currents were measured from HEK293T cells expressing wild type (WT) and mutated P2X4 receptors. The results indicate that C116A, C126A, C149A and C165A mutations disrupt two disulfide bonds...
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Clock genes in mammalian circadian pacemaker
Cimerová, Veronika ; Bendová, Zdeňka (advisor) ; Polidarová, Lenka (referee)
The behavior of mammals and their physiological processes are dependent on the daily rhythms. These rhythms are controlled by an endogenous circadian clock that responds to light/dark cycles of environment. In mammals, the circadian clocks, are arranged hierarchically, and work in almost all cells and tissues. Suprachiasmatic nuclei (SCN) in the hypothalamus are at the top of the hierarchy and work as a major circadian pacemaker. This work presents the clock genes that have been discovered in the last twenty years in the SCN of mammals. The first chapter focuses on the general mechanism of circadian rhythms and structure of the SCN. The second chapter describes the transcriptional-translational feedback loops that are an essential part of the proper function of the circadian clock, and function of individual genes is briefly introduced in this chapter. The following chapters already include a chronological characterization of Clock, Bmal1, Period and Cryptochrome genes, as they were discovered. If the clock gene expresses different function from the clockwork mechanism, it is briefly introduced in the final paragraph on the chapter. The final chapters deal with other molecules, casein kinases, ROR and REV-ERB receptors that affect its function expression and degradation of circadian genes in the...
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