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Molecular physiology of opioid receptors
Valný, Martin ; Novotný, Jiří (advisor) ; Hejnová, Lucie (referee)
The opioid receptors (OR) belong to the family of G protein-coupled receptors (GPCRs). ORs mediate the effects of the opioids, leading primarily to inhibition of neuroexcitability, predominantly through the class of the inhibitory G proteins Gi/Go. Cloning of ORs confirmed the existence of four subtypes of ORs, which mediate effects of different classes of opioid ligands. The major aim of this work is to summarize the current knowledge about characteristics and function of ORs at the molecular level. Acute exposition of ORs to their agonists results in activation of the signaling cascades that trigger mechanisms leading to analgesia. Chronic exposition of ORs to their agonists leads to desensitization and internalization of the receptors and induces adaptive changes in signal transduction system that suppresses the opioid action, and may result in the development of opioid tolerance and dependence. Although a big progress has been made in the field of understanding the molecular mechanisms of the OR-mediated signaling, there are still a lot of unresolved questions that are necessary to answer.
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Evolution of olfactory receptors
Klempt, Petr ; Stopka, Pavel (advisor) ; Vinkler, Michal (referee)
This bachelor thesis deals with the largest gene family of mammals which encode olfactory receptors. Olfactory receptors fall in rhodopsin-like GPCRs subfamily, approximately 600 - 800 millions of years old. At least from this time, olfactory receptors play, as a part of one of the oldest senses (smell), fundamental role in detection of chemical cues from water or air. This work summarizes large repertoire of olfactory receptors and its changes during the evolution of important animal taxons with emphasis on number and fraction of functional and nonfunctional olfactory receptor genes. Those values are part of criteria used for olfactory ability of animals. Olfactory receptors are typically placed on surface of sensory neuron placed in olfactory epithelium, where they bind various odorants and triggers signal cascade which leads to neuron's membrane depolarization. Therefore, about one half of this work summarizes knowledge of olfactory receptor's molecular biology like their structure, main parts of signal cascade (Gαolf, ACIII, CNG channel, Ca2+ dependent Cl- channel) just as parts needed for steady-state establishment. Expression of olfactory receptors detected in amount of non-olfactory tissues (mussels, sperm, brain etc.), indicate possibly important biomedical roles of this receptors.
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Classification, structure and function of α-adrenergic receptors.
Makarova, Anna-Marie ; Hejnová, Lucie (advisor) ; Rudajev, Vladimír (referee)
Adrenergic receptors are ones of the most investigated receptors today. Signal transduction by adrenergic receptors is involved in stress response. Stress activates the sympathicus and the hypothalamic-pituitary-adrenal axis of autonomic nervous system. Understanding effects of this activation on adrenergic signalisation is important for affection of the "fight of flight" reaction. Affecting the activity of sympathetic nerve sis important subject of interest in pharmacology and many drugs are developed using this actions. This thesis deals with a group α-adrenergic receptors and its subtypes. One part is devoted to structure which is subject of many explorations recently especially. Next chapter focuses on signal transduction mediated by α-adrenergic receptors. The last section refers to multitude of physiologic functions induced by these receptors. Powered by TCPDF (www.tcpdf.org)
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Effect of cholesterol depletion on signalling cascade initiated with receptors coupled to G protein class Gq/G11
Ostašov, Pavel ; Svoboda, Petr (advisor) ; Teisinger, Jan (referee) ; Hof, Martin (referee)
Membrane domains are an important structure in plasamatic membrane. They concentrate various signaling molecules. Their main structural component is cholesterol and by its removal the membrane domains are disrupted. The aim of our work was to examine the effect of cholesterol depeletion on signaling initiated thyreothropin releasing hormone (TRH). Although its signaling cascade is located within membrane domains the receptor itself is not. We showed that cholesterol depletion by -cyclodextrin caused release of Gq/11 proteins and caveolin 2 from membrane domains. We also discovered that cholesterol depletion decreases potency of TRH to activate G proteins as well as induction of release of intracellular Ca2+ In the last part we investigated the effect of disruption of the cell membrane integrity by cholesterol depletion on thyrotropin-releasing hormone receptor (TRH-R) surface mobility and internalization in HEK293 cells stably expressing TRH-R-eGFP fusion protein. CLSM studies indicated that the internalization of receptor molecules initiated by TRH stimulation was significantly attenuated. The detailed analysis of recovery of TRH-R-eGFP fluorescence in bleached spots of different sizes indicated that cholesterol depletion results in an increase of overall receptor mobility. We suggest that migration of...
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Influence of protein SGIP1 on partners participating in signalization of cannabinoid receptor 1
Pejšková, Lucie ; Blahoš, Jaroslav (advisor) ; Novotný, Jiří (referee)
The G-protein-coupled receptor (GPCR) family represents the largest family of cell surface receptors. GPCRs are activated by endogenous or exogenous ligands, and are targets for more than a quarter of currently used drugs. Activation of receptors initiates intracellular signaling pathways. This way the membrane receptors transfer information from the outside environment into the cell. Based on the signal the cell can respond to the changes of the environment. Key observation important for this thesis is interplay of cannabinoid and opioid signaling in vivo, which can have significant physiological effects1 . Cannabinoid receptor 1 (CB1R) and µ opioid receptor (MOR) belong to the rhodopsin family of receptors, and both are coupled with Gαi/o proteins2 . Both are located in certain areas in central nervous system (CNS) and share a lot of important features. Activation of both of the receptors leads to inhibition of adenylyl cyclase, thus decreasing the level of cyclic adenosine monophosphate in the cell, and modulates extracellular regulated kinase 1 and 2 (ERK1/2)2 . In view of the numerous anatomical, biochemical and pharmacological evidence supporting the existence of the functional interaction between opioid3 and cannabinoid receptor systems this topic became interesting for our research. In our...
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The role of protein SGIP1 in regulation of Cannabinoid Receptor 1
Chlupisová, Lenka ; Blahoš, Jaroslav (advisor) ; Novotný, Jiří (referee)
Mutual cell communication in the human body ensures the proper functioning of the essential mechanisms necessary for the life of the individual and preserving the homeostasis of the whole organism. Such communication is established by various types of signal transmission from the recipient cell to the donor cell, depending on the location and type of communicating cells. One such type is signalization through receptor molecules found on the surface or within the cell receiving the signal. These receptors receive the signal molecule in the form of a ligand and bind it to themselves, while activating the receptor and then triggering the intracellular signaling pathways. The most widely represented receptors in the eukaryotic organism include G-protein-coupled receptors, which represent signaling ensured by activation of the intracellular G-protein complex, and one of the main mechanisms occurring in neuronal signaling and signal transmission in the form of a neurotransmitter. Regulation of the amount of receptors on the surface of the cell and transport of the signal molecule into the intracellular spaces of the cell is ensured by the mechanism of endocytosis, whereby internalization of the ligand- bound receptor in the cytoplasm occurs. One of the most researched mechanisms is clatharin-mediated...
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Molecular mechanisms engaged in the development of drug addiction
Pallag, Gergely ; Novotný, Jiří (advisor) ; Nerandžič, Vladimír (referee)
Drug use is part of the human life from the ancient times. Besides their recreational utilization, sustained misuse of these substances can lead to the development of drug addiction especially in susceptible individuals and thus cause serious health and social problems. The aim of this thesis is to briefly introduce brain structures which are affected by addictive substances, and describe some of the mechanisms and molecules that contribute to addiction. A crucial brain structure which plays a role in drug addiction is the reward system, with dopamine as the main neurotransmitter. After repeated use of drugs, in neurons of this system certain molecules and epigenetic changes are accumulating that promote chronic nature of addiction. Especially important is the highly stable transcription factor ΔFosB, which in cooperation with other molecules promotes relapse even after several months or years of the last drug use. Powered by TCPDF (www.tcpdf.org)
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Classification, structure and function of α-adrenergic receptors.
Makarova, Anna-Marie ; Hejnová, Lucie (advisor) ; Rudajev, Vladimír (referee)
Adrenergic receptors are ones of the most investigated receptors today. Signal transduction by adrenergic receptors is involved in stress response. Stress activates the sympathicus and the hypothalamic-pituitary-adrenal axis of autonomic nervous system. Understanding effects of this activation on adrenergic signalisation is important for affection of the "fight of flight" reaction. Affecting the activity of sympathetic nerve sis important subject of interest in pharmacology and many drugs are developed using this actions. This thesis deals with a group α-adrenergic receptors and its subtypes. One part is devoted to structure which is subject of many explorations recently especially. Next chapter focuses on signal transduction mediated by α-adrenergic receptors. The last section refers to multitude of physiologic functions induced by these receptors. Powered by TCPDF (www.tcpdf.org)
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Postnatal development of GABAb-receptors in the frontal rat brain cortex
Kagan, Dmytro ; Svoboda, Petr (advisor) ; Langmeier, Miloš (referee) ; Mareš, Pavel (referee)
In this work, the detailed analysis of GABAB-R/G protein coupling in the course of pre- and postnatal development of rat brain cortex indicated the significant intrinsic efficacy of GABAB-receptors already shortly after the birth: at postnatal day 1 and 2. Subsequently, both baclofen and SKF97541-stimulated G protein activity, measured as the high-affinity [35 S]GTPγS binding, was increased. The highest level of agonist-stimulated [35 S]GTPγS binding was detected at postnatal days 14 and 15. In older rats, the efficacy, i.e. the maximum response of baclofen- and SKF97541-stimulated [35 S]GTPγS binding was continuously decreased so, that the level in adult, 90-days old rats was not different from that in newborn animals. The potency of G protein response to baclofen stimulation, characterized by EC50 values, was also high at birth but unchanged by further development. The individual variance among the agonists was observed in this respect, as the potency of SKF97541 response was decreased when compared in 2-days old and adult rats. The highest plasma membrane density of GABAB-R, determined by saturation binding assay with specific antagonist [3 H]CGP54626AA, was observed in 1-day old animals. The further development was reflected in decrease of receptor number. The adult level was ≈3- fold lower than...
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Muscarinic acetylcholine transmission and Alzheimer's disease
Janíčková, Helena ; Doležal, Vladimír (advisor) ; Blahoš, Jaroslav (referee) ; Rokyta, Richard (referee)
Impairment of the cholinergic neurotransmission system is regularly detected in animal models of Alzheimer's disease as well as in human patients suffering from this serious disease. Moreover, there is increasing amount of evidence suggesting that activation of individual mAChR subtypes specifically influences the cleavage of APP, the precursor for β-amyloid. APP can be processed in an amyloidogenic or non-amyloidogenic pathway and a relative abundance of these patways contributes to establishing the final concentration of neurotoxic β-amyloid in the brain. In this work, I have studied the acute and chronic effects of A β1-42 on binding and functional characteristics of mAChR. I have demonstrated that Aβ1-42 present in cell culture expressing the individual subtypes of mAChR negatively and specifically influences the function of the M1 mAChR subtype. I have also detected a decline in muscarinic receptor-mediated signal transduction in brain tissue of young adult APPswe/PS1dE9 mice, a commonly used animal model of Alzheimer's disease. Demonstration of the impairment of muscarinic transmissin in transgenic mice by soluble β-amyloid that occurs earlier than amyloid pathology and behavioral deficit, and its imitation by soluble Aβ1-42 in vitro lend strong support to the notion of the early involvement...
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