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Pathological pain states, the role of synaptic modulation at spinal cord level
Nerandžič, Vladimír ; Paleček, Jiří (advisor) ; Krůšek, Jan (referee)
(English) Modulation of synaptic transmission in dorsal horn of spinal cord plays a key role in nociceptive signalling. Recent studies have indicated a great importance of presynaptic TRPV1 receptors (transient receptor potential vanilloid) in spinal cord. These receptors act as molecular integrator of nociceptive stimulation on periphery. The way of their activation and the effect on modulation of the synaptic transmission are not clarified yet. Previous studies demonstrated the influence of many inflammatory mediators and cytokins on TRPV1 receptors. The aim of our research was to show changes in activation of presynaptic TRPV1 receptors in the spinal cord following the application of endogenous agonist N-oleoyl dopamine (OLDA) in a model of peripheral neuropathy, after incubation with cytokine TNFα and to show the effect of precursor of anandamide N-acylphosphatidylethanolamine (NAPE). In our experiments, we have recorded miniature excitatory postsynaptic currents (mEPSC) from neurons of acute spinal cord slices by the patch-clamp method. The first series of experiments tested sensitivity to application of the endogenous agonist OLDA 5 days after evoking peripheral neuropathy. The frequency of mEPSC increased significantly - to 250 % of base level after applying a low concentration of OLDA (0,2...
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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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Modulation of nociceptive synaptic transmission
Nerandžič, Vladimír ; Paleček, Jiří (advisor) ; Krůšek, Jan (referee) ; Hejnová, Lucie (referee)
Modulation of synaptic transmission in the spinal cord dorsal horn plays an important role in development and maintenance of pathological pain states. The indisputable part of this modulation is conducted via activity of the transient receptor potential cation channel subfamily V member 1 (TRPV1) and the cannabinoid receptor 1 (CB1), expressed on presynaptic endings of primary afferents in the superficial spinal cord dorsal horn. Under physiological conditions, activation of TRPV1 receptors is pronociceptive while CB1 receptor activation leads to attenuation of nociceptive signalling. However, both receptors share also one endogenous agonist anandamide (AEA) that may be produced from N-arachidonoyl phosphatidylethanolamine (20:4-NAPE). Main objective of this thesis focuses on the effect of 20:4-NAPE on nociceptive synaptic transmission in spinal cord slices under naïve and inflammatory conditions and consequent on the possible interaction of TRPV1 and CB1 receptors. First, 20:4-NAPE application induced significant release of anandamide from spinal cord slices under in vitro conditions. Next, patch- clamp recordings of excitatory postsynaptic currents (mEPSC and sEPSC) from superficial dorsal horn (DH) neurons in acute spinal cord slices were used. 20:4-NAPE application under the physiological...
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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 TRPV1 receptors in chemokine CCL2 induced modulation of nociceptive synaptic transmission at spinal cord level
Adámek, Pavel ; Paleček, Jiří (advisor) ; Krůšek, Jan (referee)
Modulation of nociceptive synaptic transmission in the spinal cord dorsal horn is a significant mechanism in the development and maintenance of different pathological pain states. Accumulating evidence indicates that the TRPV1 (transient receptor potential vanilloid 1) receptor and chemokine CCL2 (C-C motif ligand 2) may play a critical role in this process. The aim of this diploma thesis was to investigate the CCL2 induced modulation of nociceptive synaptic transmission in the dorsal horn of spinal cord and the role of the TRPV1 receptors. To investigate this aim patch-clamp recordings of spontaneous and miniature excitatory postsynaptic currents (sEPSC, mEPSC) from superficial dorsal horn neurons in acute rat lumbar spinal cord slices were used. After acute application of CCL2 on the slice preparation from naïve animals, a frequency increase of both sEPSC and mEPSC was present. This CCL2 induced increase in both sEPSC and mEPSC frequency was prevented by the TRPV1 receptor antagonist SB366791 application. No changes were observed in the amplitudes of sEPSC or mEPSC after application of the CCL2, SB366791, or co-application of CCL2 and SB366791. This suggests that the observed changes were mediated predominantly by presynaptic mechanisms. The preliminary results indicate that after chronic constriction...
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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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The role of nociceptive synaptic transmission modulation at the spinal cord level in different pain states
Adámek, Pavel
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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Studies on molecular interactions of the mu-opioid and TRPV1 receptors
Melkes, Barbora ; Novotný, Jiří (advisor) ; Blahoš, Jaroslav (referee) ; Krůšek, Jan (referee)
In this work, we investigated the behavior of the -opioid receptor (MOR) and the transient receptor potential vanilloid 1 (TRPV1) ion channel in the plasma membrane and their mutual communication. Both these receptors are implicated in pain perception and analgesia. We observed that the lateral mobility of MOR was strongly affected by different biased opioid agonists. DAMGO and endomorphin-2 display opposite bias towards MOR. According to our results, they also have the opposite effects on the mobility of MOR. Morphine induced only small changes in the mobility of MOR. Moreover, cholesterol depletion and blockage of G protein signaling by pertussis toxin (PTX) affected the ability of different MOR agonists to alter MOR mobility in a unique manner. The effects of DAMGO and endomorphin-2 were compromised under these conditions. On the other hand, we observed increased movement of MOR after the addition of morphine. PTX alone did not affect receptor movement, but it completely disrupted the effect of cholesterol depletion on morphine induced changes the mobility of MOR. Next we studied the mobility of TRPV1. The TRPV1 agonist capsaicin changed the lateral mobility of TRPV1. Surprisingly, after adding the MOR antagonist naloxone, the apparent diffusion coefficient of TRPV1 but to a lower extent than...
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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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The role of nociceptive synaptic transmission modulation at the spinal cord level in different pain states
Adámek, Pavel
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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