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Proposal of E-commerce
Jakubík, Ján ; Mates, Vojtěch (referee) ; Luhan, Jan (advisor)
This master's thesis deals with the proposal of e-commerce BezPopelniku.cz which sells electronic cigarettes and accessories. The theoretical part explains the concepts and methods that are used in other parts of the thesis. The second part is devoted to analyzing the market of electronic cigarettes in the Czech Republic, SWOT analysis, requirements for the technical features of e-commerce platform, analysis of external and internal environment of the company and ultimately a risk analysis of the project. The third part of the thesis defines procedures for the right proposal of e-commerce, contains proposals for specific competitive advantages and describes the use of marketing tools to promote the store.
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The role of striatal cholinergic signalling in the control of behaviour
Abbondanza, Alice ; Jakubík, Jan (advisor) ; Bellone, Camilla (referee) ; Svoboda, Jan (referee)
Cholinergic transmission regulates many behavioural domains, ranging from motor activity to cognition. Acetylcholine signalling is mediated by muscarinic and nicotinic acetylcholine receptors (mAChRs and nAChRs, respectively). While mAChRs are slow responding metabotropic receptors, nAChRs are ion channels, mediating fast neurotransmission. There is a growing body of evidence suggesting a role of nAChRs as important modulators of behavioural functions. However, as nAChRs consist of many subtypes, depending on their composition in subunits, and as they are expressed by various neuronal populations in different brain regions, their contribution to behavioural control is very complex. To decipher their contribution, it is necessary to selectively target nAChRs expressed not only in particular regions but also by particular neurons with a defined effect on local microcircuits. The goal of the present thesis was to use different genetic strategies to induce regional- and cell-specific deletion of β2-containing nAChRs in the mouse brain, in order to characterize the functional role of these receptors. We focused our work in two brain areas, the striatum and the prefrontal cortex (PFC). In the striatum, we identified the striatal neurons that express one of the most common nicotinic subunits, the β2...
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Circadian changes of the cholinergic system in various areas of the central nervous system
Křížová, Monika ; Mysliveček, Jaromír (advisor) ; Jakubík, Jan (referee)
Muscarinic acetylcholine receptors type 4 are one of the five subtypes of metabotropic muscarinic receptors whose knockout led to disruption of the biorhythm represented by locomotion in female mice. In this thesis, we attempted to determine the biorhythmic activity of selected components of the cholinergic system in structures of the central nervous system which are suspected to regulate circadian locomotion. These selected structures were the striatum, motor cortex, thalamus, intergeniculate leaflet, subparaventricular zone and posterior hypothalamic area. Changes of muscarinic receptor density in wild type and M4 receptor knockout female mice were measured by using autoradiography throughout the day. In the striatum, motor cortex, and thalamus, the density of muscarinic receptors type 1 and the activity of acetyl- and butyrylcholinesterases were also measured by autoradiography and the Ellman's method. Rhythmic oscillations of muscarinic receptor density were observed in all structures selected in both types of mice. The main brain structure responsible for the differences in the biological rhythm of wild type and M4 receptor knockout mice was identified as the striatum and motor cortex region, where the percentage of M4 receptors was highest. Diurnal oscillations were also exhibited by...
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Mobilization of intracellular calcium via muscarinic acetylcholine receptors.
Šantrůčková, Eva ; Jakubík, Jan (advisor) ; Krůšek, Jan (referee)
Muscarinic acetylcholine receptors play important role in many physiological and pathophysiological processes. Muscarinic receptors are metabotropic receptors for acetylcholine. The objective of this thesis is to compare long-term effects of short-term exposure of the muscarinic receptors to xanomeline among subtypes. There are two groups of muscarinic receptors that differ in their extracellular-to-intracellular signal transduction - the odd-numbered ones (M1, M3, M5) preferentially couple to Gq/11 G-proteins, whereas even-numbered ones (M2, M4) couple mainly via Gi/o. CHO cells stably expressing individual muscarinic receptor subtypes were used for all experiments. Cells expressing M2 and M4 receptors were transiently transfected with cDNA for human Gq/16 G-protein to achieve calcium response comparable to the one at oddnumbered subtypes. Calcium level was measured with the fluorescent indicator Fura 2. Xanomeline stimulation induced the fast mobilization of the intracellular calcium at all five receptor subtypes. In accordance with the functional selectivity of xanomeline for M1 and M4 receptors, the immediate calcium response to xanomeline was comparable to that of full agonist carbachol and the elevated calcium level sustained for one hour after xanomeline had been removed. On the other hand,...
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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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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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Modulatory effect of monovalent ions on δ-opioid receptors
Vošahlíková, Miroslava ; Svoboda, Petr (advisor) ; Jakubík, Jan (referee) ; Kršiak, Miloslav (referee)
The exact role of opioid receptors in drug addiction and modulatory mechanism of action of monovalent cations on these receptors are still not fully understood. Our results support the view that the mechanism of addiction to morphine is primarily based on desensitization of μ- and δ-opioid receptors. Desenzitization of agonist response proceeds already at the level of G protein functional activity. Long-term exposure of rats to morphine resulted in increase of number of δ-opioid receptors and change of their sensitivity to sodium ions. Analysis of the effect of different monovalent ions on agonist binding in δ-OR- Gi1α (Cys351 -Ile351 )-HEK293 cell line confirmed the preferential sensitivity of δ-opioid receptor to sodium ions. We have distinguished the high- and low-affinity Na+ sites. Biophysical analysis of interaction of lithium, sodium, potassium and cesium ions with plasma membranes isolated from HEK293 cells with the help of fluorescent probes indicated that monovalent ions interact, in low-affinity manner, with the polar, membrane-water interface of membrane bilayer. Key words: morphine, forebrain cortex, opioid receptors, G proteins, monovalent ions, plasma membrane, fluorescence spectroscopy.
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The effects of opioid ligands on their receptors in model cells (e.g. lymphocytes, neural cells, cell lines)
Cechová, Kristína ; Svoboda, Petr (advisor) ; Sýkora, Jan (referee) ; Jakubík, Jan (referee)
Morphine and other opioids are powerful drugs used for pain relief, but their clinical use has limitations as prolonged treatment leads to tolerance and addiction. Opioids bind to opioid receptors, and besides pain, they regulate mood and stress. Even though the primary role of opioids and opioid receptors is in the central nervous system, they also affect the immune system. The mechanisms of how they affect the immune system, but also the mechanisms of tolerance and addiction build-out, are not fully understood. These processes may be demonstrated as changes in the opioid receptor expression level, alteration in their function or oligomeric state. The dimerization of opioid receptors is a controversial topic in literature with many contradictory results. In this work, changes in the protein level of opioid receptors in the rat forebrain cortex were not observed after morphine treatment. However, a specific increase in the amount of µ- opioid receptors was determined in rat lymphocytes, using flow cytometry. In other studies, an increased mRNA level of the µ-opioid receptor was connected with immunosuppression. The increased µ-opioid receptor protein level indicates a similar effect. The overall effect of morphine on the protein composition of lymphocytes was determined by proteomic analysis. The...
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Cellular and molecular mechanisms of activation of thermally sensitive TRP ion channels
Máčiková, Lucie ; Vlachová, Viktorie (advisor) ; Anděrová, Miroslava (referee) ; Jakubík, Jan (referee)
The transient receptor potential (TRP) are cation channels mostly permeable to both monovalent and divalent cations. ThermoTRP is a specific group of directly thermally activated TRP channels. The vanilloid transient receptor potential 3 (TRPV3) is an ion channel widely expressed in keratinocytes, that is implicated in the regulation of skin homeostasis, thermo- sensing, nociception and development of itch sensation. Our results show the importance of the cytoplasmic inter-subunit interface in the heat sensitivity of TRPV3. As there is a structural analogy within the vanilloid receptors, our hypothesis of the identified important region is supposed to be valid also for other thermally activated TRPV receptors (TRPV1, TRPV2 and TRPV4). We have proved that TRPV3 is a substrate for ERK1/2 protein kinase (kinase regulated by extracellular signal 1 and 2) and we have identified TRPV3 phosphorylation sites that may be direct targets for ERK1/2. Of these residues, threonine 264 has been shown to be the main phosphorylation site responsible for TRPV3 sensitization mediated by ERK kinase. In human keratinocytes, the phosphorylation might be physiologically and pathophysiologically important in processes of TRPV3 sensitization mediated by MAPK signaling pathway. The transient receptor potential ankyrin 1...
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