Browsing by discipline "Farmakologi och toxikologi"
Now showing items 1-4 of 4
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(2010)Neuronal nicotinic acetylcholine receptors are ligand-gated ion channel receptors that consist of five transmembral receptor subunits. They can form a functional receptor subtype solely out of alfa-subunits or out of a combination of alfa- and beta-subunits. The number of potential assembly of nicotinic receptor subunits is high because at least nine alfa-subunits (α2-α10) and three beta-subunits (β2-β4) have been recognized. The composition, location and pharmacological properties of different subtypes have not yet been fully characterized. However, it has been shown that the neuronal nicotinic receptors are involved in a wide range of physiological and pathofysiological processes especially in the central nervous system. Toxins from snakes and Conus -sea snails have proved to be important tools in neuropharmacological research, from which considerable information on structure and subunit combinations of the nicotinic receptors have been received. Receptors binding assays or autoradiographic experiments exploiting toxins have also been useful methods to locate the neuronal nicotinic receptors in mammalian brain. The aim of the experimental part of the Pro gradu was to characterize in vitro binding affinities of α-contoxin MII, α-conotoxin Vc1.1, neurotoxin II, α-cobratoxin and weak-toxin synthetized in Moscow and of well-known receptor ligands cytisine and methyllycaconitine to neuronal nicotinic receptors in SH-SY5Y- and SH-EP1-hα7-cell membrane preparations. SH-SY5Y-cells are known to express various neuronal receptor subtypes (α3* or α7) endogenously. For the SH-EP1-hα7-cells part, the cell line has been transfected with α7 nAChR-genes and it expresses only α7 receptor subtypes. Receptor competition studies were performed with [3H]-epibatidine (400 pM SH-SY5Y, 2000 pM SH-EP1-hα7) and the radioactivity was measured with a Microbeta- scintillation counter. [3H]-epibatidine was perceived to be displaced almost completely by cytisine and methyllycaconitine in both cell lines. In addition to this, the toxins were shown to bind to two distinct receptor-binding sites in SH-SY5Y-cells. Also α-contoxin MII and α-conotoxin Vc1.1 inhibited [3H]-epibatidine binding by biphasic manner, but the maximal displacement failed to be complete. From α-conotoxins only MII had affinity to α7 receptors in SH-EP1-hα7-cells. Neurotoxin II, α-cobratoxin and weak-toxin were not found to compete with [3H]-epibatidine for the same binding sites in SH-SY5Y-cells. The results confirm the assumption that cytisine and methyllycaconitine label various nAChR-subtypes. Instead, based on SH-SY5Y-cell assays α-conotoxins used in this study would seem to label spesifically only particular nAChR-subtypes. The receptor competition studies also confirm the prevalent conception that neurotoxin II, α-cobratoxin and weak-toxin do not bind to neuronal nicotinic receptor subtypes containing α3-subunits.
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(2010)Glutamate is the principal excitatory neurotransmitter in the central nervous system. Glutamatergic neurotransmission plays a central role in the development and maintenance of drug addiction. Glutamate interacts with other neurotransmitters such as dopamine in the actions concerning addiction. During the development of drug addiction, plastic changes in the neuronal connections related to memory and learning occur for example in the amount of synapses and in the efficacy of their action. Glutamatergic AMPA receptor and especially its GluA1 subunit are thought to be included in the neurobiological mechanisms related to drug addiction. Compulsive drug craving and relapses to drug use after a period of abstinence are central problems among people suffering drug addiction. Conditioned place preference is a technique that is used to study motivational properties of drugs in experimental animals. The aim of this master's thesis was to examine the importance of glutamatergic AMPA receptor GluA1 subunit in the morphine-induced place preference and in its extinction and reinstatement behaviour. Locomotor activity of mice was studied during all the phases of experiment. Glutamatergic AMPA receptor GluA1 subunit-deficient (GluA1-/-) and their control (wildtype) mice, based on C57BL/6J mouse strain, were used in the experiments. During the conditioning phase, the mice were trained to associate the effects of morphine (20 mg/kg) with a specific environment. After conditioning, the extinction with morphine paired conditioning environment was assessed by giving saline (0,9 % NaCl solution) to mice. The extinction phase was followed by reinstatement test, in which mice were given morphine (20 mg/kg). The seeking of animals with morphine paired conditioning environment described drug-seeking during different phases of experiment. GluA1-/- mice were more hyperactive when placed in the testing environment compared to the wildtype mice. However, the morphine-induced locomotor activity did not differ between genotypes. Locomotor activity of both genotypes was sensitized equally in consequence of repeated morphine exposures. Morphine induced place preference in both genotypes. Furthermore, the extinction of morphine place preference happened in both genotypes. However, the results of reinstatement test differed partly between genotypes. The place preference was reinstated by morphine in wildtype mice, but not in GluA1-/- mice, when using repeated testing extinction method. Instead of place preference, wildtype mice exhibited place aversion, when extinction method was saline conditioning. As a result of these experiments, extinction method can have an impact on the results of reinstatement test and conclusions cannot be done on the importance of GluA1 subunit in morphine reinstatement. In conclusion, the results of place preference experiments support the conception that GluA1 subunit is not significant in morphine conditioning. However, based on these experiments, GluA1 subunit is not important in morphine extinction, as one might assume on the basis of literature. GluA1 subunit may have an importance in morphine reinstatement, although the results of reinstatement test were partly contradictory.
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(2003)Epidemiological data suggest an important role of perinatal viral infections in the etiology of schizophrenia. In this thesis the connection between neonatal viral brain infection and its consequences to the development of central nervous system was studied. In schizophrenia the symptoms are divided into three categories as positive, negative and cognitive ones. Positive symptoms refer to hallucinations and delusions, negative symptoms are defined as social withdrawal, apathy and poor motivation and cognitive symptoms include deficits in abstraction and paying attention into subjects. Symptoms suggest that in schizophrenia the received information can not be filtered properly in central nervous system, but comes into patients senses in excess i.e. there are defects in sensorimotor gating. Sensorimotor gating was studied by prepulse inhibition of acoustic startle -phenomenon. Prepulse inhibition refers to the inhibition of the startle reflex by weak prepulse presented before the startling stimulus. In schizophrenic patients prepulse inhibition is decreased and in addition to that psychotomimetic drugs disrupt prepulse inhibition in humans as well as in experimental animals. Sensorimotor gating ability is developed under neuronal development and it can be affected by several neurodevelopmental disturbances. In the present study rats were infected with herpes simplex type 1 virus at neonatal age and later challenged to dopaminergic and glutamatergic systems. Results show controversial data of effects on prepulse inhibition, still some attenuation can be seen. Challenge studies did not show clear and persistent effect either in dopaminergic or glutamatergic tests. Corticosterone, naturally occurring hormone in rats, was administered to rat mothers under gestation until weaning in terms to clarify its effects to neuronal development. Administration was carried out by implanted pellet as well as by drinking water. The latter was found to work out better as it releases corticosterone in pulsatile manner. Corticosterone was administered also in acute test to drug naïve animals. This test showed significant decrease on prepulse inhibition. The same could not be repeated in corticosterone challenge test after perinatal treatments. Nitric oxide synthase inhibitor L-NMMA was administered to neonates under days 5-9 after partus. This was supposed to prevent neonates from neurodevelopmental disturbances affected by virus and corticosterone. Despite various dose levels used, any clear effect could not be seen. In summary, the studies show some effect of treatments on neuronal development and sensorimotor gating measured by prepulse inhibition. In the test groups inspected many treatments showed effect at first, but those effects disappeared at later tests as rats grew up. This might be an outcome of the potential compensatory mechanisms of the central nervous system to counteract harmful neurodevelopmental events.
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(2010)Prolyl oligopeptidase (POP, E.C. 3.4.21.26) cleaves short peptides, of less than 30 amino acid long, at the C-side of an internal proline. It has been associated with many pathophysiological processes, such as neurodegeneration and inflammation. At the moment there are no studies that have been focused on POP function in multiple sclerosis (MS). A preliminary study in a Spanish cohort reported altered POP activity in plasma samples of patients with relapsing-remitting multiple sclerosis (RR-MS) compared with healthy controls. Also they observed increased levels of the endogenous POP inhibitor in plasma samples of patients with RR-MS. The first objective of this study was to evaluate the POP activity levels in serum and cerebrospinal fluid (CSF) samples from RR-MS patients and healthy controls in a Finnish population using a kinetic fluorescence assay. The seral levels of the endogenous POP inhibitor were also investigated by preincubating recombinant porcine POP (rPOP) with serum and determining the percentual decrease of POP activity compared to basal rPOP values (inhibitory capacity %). The second objective of this study was to purify and characterize the endogenous POP inhibitor in serum. In order to accomplish this goal, different biochemical and biophysical features, such as temperature resistance and filtering cut-off were tested. Also a combination of chromatographic approaches (affinity/anion exchange/hydrophobic interaction chromatography) with polyacrylamide gel electrophoresis and protein staining was used. All the differences observed in POP activity/inhibitor levels (serum, serum with DTT, CSF) between healthy controls and patients with RR-MS in this study did not reach statistical significance due to low values in all the samples. However, the trends in all the measured parameters were similar to the preliminary study in a Spanish cohort. Thus, the data supports further, more comprehensive, studies on the role of POP in MS. After series of chromatographic runs, a mass spectrometry analysis revealed the endogenous POP inhibitor to be α2-macroglobulin, a panprotease inhibitor in serum. α2-Macroglobulin has also been associated with MS, thus this finding substantiate the relationship between POP and MS.
Now showing items 1-4 of 4