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Modulation of synaptic transmission, studies on spinal cord slices in vitro
Mrózková, Petra ; Paleček, Jiří (advisor) ; Krůšek, Jan (referee)
Modulation of a synaptic transmission in the spinal cord dorsal horn plays a key role in nociceptive signalling, especially in states of pathological pain. The goal of this study was to develop a method for calcium imaging in spinal cord slices in vitro. This method allowed us to record changes of intracellular free calcium ions concentration (iCa2+ ), that are a major mediator of neuronal plasticity. In this work, we have focused on application of this method in a conventional fluorescence microscope and on the role of different neuromodulators of synaptic activity. Changes of iCa2+ induced by dorsal root electrical stimulation were recorded altogether in 744 dorsal horn (lamina I and II) neurons. In the first series of experiments, stimulation protocols activating preferentially A and A + C dorsal root fibers were used and long-term stability of the calcium responses was verified. The dorsal root stimulation induced in the neurons fast and delayed type of calcium response. Application of AMPA and NMDA receptors antagonists, CNQX (50μM) and MK801 (45μM), reduced the calcium response amplitude and confirmed the importance of glutamate receptors in synaptic activation. In several experiments the effect of capsaicin a TRPV1 receptors agonist, application was tested. Application of even low...
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Calcium homeostasis and modulation of nociceptive synaptic transmission
Sojka, David ; Paleček, Jiří (advisor) ; Žurmanová, Jitka (referee) ; Krůšek, Jan (referee)
2 SUMMARY OF THE THESIS This study was designed to improve our knowledge regarding mechanisms of nociceptive signaling at spinal cord level. One of the forms of spinal cord synaptic transmission modulation is central sensitization, a manifestation of synaptic plasticity at spinal cord level, which was found to be present at many chronic pain syndromes. This study deals mainly with a development of calcium imaging technique with a final goal to study mechanisms of central sensitization in vitro on population of dorsal horn neurons. We have analyzed synaptically evoked intracellular Ca changes as a result of dorsal root stimulation in a superficial dorsal horn area in spinal cord slices and found two types of Ca responses: one synchronized with electrical stimulation and a second one, delayed response due to Ca release from internal stores. The delayed Ca release was not previously shown to be present in these neurons and it was not dependent on activation of ionotropic glutamatergic receptors, suggesting involvement of metabotropic receptor pathway. The presence of this delayed type of Ca response could have a significant role in the induction of some types of chronic pain syndromes since intracellular calcium increase is thought to be a key trigger point in spinal cord neurons sensitization. An important...
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Cortical electrical stimulation and pain
Rusina, Robert ; Rokyta, Richard (advisor) ; Haninec, Pavel (referee) ; Paleček, Jiří (referee)
The aim of the study was to examine effects of sensorimotor cortex stimulation on pain in animal. A behavioral model investigated pain thresholds in deafferentated rats depending on cortex stimulation and two neurophysiological models studied different components of the jaw opening reflex (JOR) and tooth pulp evoked potentials (TPEPs) following cortical stimulation. The behavioral model used 18 deafferentated (dorsal root rhizotomy) rats and 14 controls. Pain thresholds were measured before and after cortical stimulation using plantar test and tail-flick latencies. In the neurophysiological model, rats were implanted with tooth pulp, cerebral cortex, and digastric muscle electrodes. 15 animals were divided into three groups, receiving 60 Hz, 40 Hz and no cortical stimulation, respectively. TPEPs were recorded before, one, three and fi ve hours after continuous stimulation. 10 other rats were submitted to recordings after a single tooth pulp stimulation, while in 5 more rats we administrated conditioning and test stimulation. TPEPs and digastric EMG were simultaneously recorded. A multiresolution denoising method was used for signal processing. Our results show a similar effect of the stimulation in man and experimental animals despite the differences in the organization of the cerebral cortex. Our results...
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Sickness behaviour in the early adjuvant arthritis (role of neuroinflamation and oxidative stress?)
Škurlová, Martina ; Jurčovičová, Jana (advisor) ; Šterzl, Ivan (referee) ; Paleček, Jiří (referee)
Background: Rheumatoid arthritis is an inflammatory autoimmune polyarthritis. Although, it is not a CNS involvement disease, affective disorders and alterations of cognitive functions occur in rheumatic patients and may vary in their relevance from serious psychosis to memory disorders. Aetiology of sickness behaviour in arthritis is not known yet. Aims: The aim of the present work was to study incidence of behavioural components of sickness in the early phase of experimental arthritis, and to confirm an association between behavioural components of sickness and neuro- inflammatory / chemical alterations in the hippocampus in this phase of the disease. Methods: Experimental arthritis was induced to Lewis rats by a single injection of cFA. First four days of experimental arthritis were studied. The body weight and food intake were measured daily. Pain reactivity, behaviour and biochemical analysis in plasma and hippocampus were done on day 2 and on day 4. Pain reactivity was measured separately on limbs and on tail in plantar test. Spatial learning abilities and swim strategies were examined in MWM. Anxiety behaviour was tested in EPM and open field tests. In plasma, concentration of CRP, albumin, ACTH, corticosterone, leptin, ghrelin were estimated. In hippocampus, mRNA gene expression of IL-1β, IL-...
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Neuroinflammation and mechanisms of neuropathic pain development
Kalynovska, Nataliia ; Paleček, Jiří (advisor) ; Krůšek, Jan (referee) ; Hejnová, Lucie (referee)
Neuropathic pain represents a possible outcome of neural tissue injury; it occurs also as a concomitant symptom of different diseases or as a side effect of several treatments. Up to date, it constitutes a great challenge in clinical practice, as currently available treatments are still unsatisfactory. Mechanism-based treatment approaches are promising strategy in neuropathic pain management. However, there is still a lack of information about the exact mechanisms involved in the development and/or maintenance of neuropathic pain. This Doctoral Thesis is aimed to explore the mechanisms underlying the development of neuropathic pain states in different models. The principal part of this work is focused on the study of anti-inflammatory effect of Angiotensin II receptor type 1 (AT1R) blocker, losartan, in two different models of peripheral neuropathy: paclitaxel-induced peripheral neuropathy (PIPN) and spinal nerve ligation (SNL). The work also aimed to access the involvement of spinal transient receptor potential vanilloid type 1 (TRPV1) channels in the process of neuronal activation induced by paclitaxel (PAC) and chemokine CCL2 treatment. In order to fulfil the abovementioned aims, behavioral, immunohistochemical and molecular methods were used. For every model of peripheral neuropathy, the...
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Mechanisms of Activation and Modulation of Ion Channels Specific for Nociceptive Neurones
Touška, Filip ; Vlachová, Viktorie (advisor) ; Paleček, Jiří (referee) ; Tureček, Rostislav (referee)
Human body detects potentially damaging stimuli by specialized sensory nerve endings in the skin, the nociceptors. Their membranes are equipped with ion channels, molecular sensors, coding the outside stimuli into the trains of action potentials and conducting them to the higher brain centers. The most prominent group of transduction ion channels is the transient receptor potential (TRP) channel family followed by ion channels responsible for generation and conduction of action potentials from the periphery to the brain, the voltage-gated sodium channels (VGSCs). Understanding the mechanisms how particular stimulus is encoded and processed is of particular importance to find therapeutics for various types of pain conditions. We characterized the properties of VGSC subtypes NaV1.9 and NaV1.8 at high temperatures. We showed that NaV1.9 undergo large increase in current with increasing temperatures and significantly contribute to the action potential generation in dorsal root ganglion (DRG) neurons. Ciguatoxins (CTXs) are sodium channels activator toxins causing ciguatera fish poisoning, a disease manifested by sensory and neurological disturbances. We elucidated the mechanism of CTX- induced cold allodynia, a pathological phenomenon where normally innocuous cool temperatures are perceived as pain. We...
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Relationship shooting velocity and accuracy in handball on beginners level
Paleček, Jan ; Tůma, Martin (advisor) ; Pravečková, Petra (referee)
Title: Relation between throwing velocity and accuracy in handball on elementary level Goal: Throwing effectiveness in handball can be influenced by many factors. Throwing accuracy and velocity are considered to be the determinative factors. The aim of this bachelor thesis was to explore the mutual relation of these parameters in case of handball players on elementary level. The field research was realized in the handball club Lovci Lovosice in age category of 12-13 years old players. Methodology: In terms of this bachelor thesis, two field researches in the interval of two months were realised. These field researches were realized in the handball club Lovci Lovosice in the age category of 12-13 years old players by using a hand-operated radar device The Stalker Pro 2. The research was performed only for one type of throw, specifically the one-handed overarm on the spot throw. Both throwing velocity and accuracy were traced within the research mentioned above. Statistical data processing was performed in Microsoft Excel 2010 using one-tailed statistical test. Results: The results of the research in this bachelor thesis show a very close relation between throwing velocity and accuracy in handball. The players hit more targets when they did not perform such high throwing velocity. While in the first...
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Modulation of synaptic transmission in the development of painful states
Slepička, Jakub ; Paleček, Jiří (advisor) ; Hejnová, Lucie (referee)
My thesis introduces the topic of nociceptive signalisation and processes involved in the formation and spreading of neuropathic pain. This study focuses on the mechanisms of nociceptive synaptic transmission mechanisms in the level of spinal dorsal horn and its modulation by paclitaxel, a chemotherapeutic drug inducing neuropathic changes. The attention is put especially on the possibility of glial activity participation in paclitaxel side effects. This idea stems from the existing hypothesis of the functional connection between TLR4 and TRPV1 receptor activity. TRPV1 is well known for its participation in chemical, thermal and nociceptive sensory transmission. Minocycline antibiotic is considered as an inhibitor of microglial activation therefore it was used for blocking neuroinflammation. The experimental part is comparing an impact of substances applied to the model of tachyphylaxis used for monitoring of nociceptive transmission changes according to decreasing activity of TRPV1 receptors. Electrophysiological recording of miniature excitatory postsynaptic currents from neurons in the Rexed laminae I. and II. of spinal dorsal horn was used. The results of my measurements show that minocycline is able to suppress acute effects of paclitaxel application in vitro if the spinal slice is incubated...
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The role of nociceptive synaptic transmission modulation at the spinal cord level in different pain states
Adámek, Pavel ; Paleček, Jiří (advisor) ; Vaculín, Šimon (referee) ; Vlachová, Viktorie (referee)
Pain is a common symptom of many clinical syndromes and diseases. In particular, the treatment of neuropathic pain represents a serious public health issue because currently available analgesia is ineffective in many cases or it has adverse effects. Treatment of pain-related suffering requires knowledge of how pain signals are initially generated and subsequently transmitted by the nervous system. A nociceptive system plays a key role in this process of encoding and transmission of pain signals. Modulation of the nociceptive synaptic transmission in the spinal cord dorsal horn represents an important mechanism in the development and maintenance of different pathological pain states. This doctoral thesis has aimed to investigate and clarify some of the mechanisms involved in the modulation of the spinal nociceptive processing in different pain states. The main attention was paid to study the following issues: (I.) Which is the role of Transient Receptor Potential Vanilloid type 1 channels (TRPV1), Toll-Like Receptors 4 (TLR4), and phosphatidylinositol 3-kinase (PI3K) in the development of neuropathic pain induced by paclitaxel (PAC) chemotherapy in acute in vitro, and subchronic in vivo murine model of PAC-induced peripheral neuropathy (PIPN)? (II.) How is affected spinal inhibitory synaptic control...
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