Parkinson’s disease is an age-related neurodegenerative disorder. The major symptoms of Parkinson’s disease are closely linked to the pathology of the disease. The main pathology of Parkinson’s disease consists of the degeneration of neurons of the substantia nigra pars compacta (SNpc), which leads to reduced amounts of dopamine in the brain. One of the treatment strategies in Parkinson’s disease is to conserve dopamine by inhibiting the enzymes responsible for its catabolism. The monoamine oxidase (MAO) B isoform catalyses the oxidation of dopamine in the central nervous system and is therefore an important target for Parkinson’s disease treatment. Inhibition of MAO-B provides symptomatic relief for Parkinson’s disease patients by increasing endogenous dopamine levels as well as enhancing the levels of dopamine after administration of levodopa (L-dopa), the metabolic precursor of dopamine. Recent studies have shown that phthalide can be used as a scaffold for the design of reversible MAO inhibitors. Although phthalide is a weak MAO-B inhibitor, substitution on the C5 position of phthalide yields highly potent reversible MAO-B inhibitors. In the present study, sesamol and benzodioxane were used as scaffolds for the design of MAO inhibitors. The structures of sesamol and benzodioxane closely resemble that of phthalide, which suggests that these moieties may be useful for the design of MAO inhibitors. This study may be viewed as an exploratory study to discover new scaffolds for MAO inhibition. Since substitution at C5 of phthalide with a benzyloxy side chain yielded particularly potent MAO inhibitors, the sesamol and benzodioxane derivatives possessed the benzyloxy substituent in the analogous positions to C5 of phthalide. These were the C5 and C6 positions of sesamol and benzodioxane, respectively. The sesamol and benzodioxane derivatives were synthesised by reacting sesamol and 6- hydroxy-1,4-benzodioxane, respectively, with an appropriate alkyl bromide in the presence of potassium carbonate (K2CO3) in N,N-dimethylformamide (DMF). 6-Hydroxy-1,4- benzodioxane, in turn, was synthesised from 1,4-benzodioxan-6-carboxaldehyde. The structures of the compounds were verified with nuclear magnetic resonance (NMR) and mass spectrometry (MS) analyses, while the purities were estimated by high-pressure liquid chromatography (HPLC). Sixteen sesamol and benzodioxane derivatives were synthesised. To determine the inhibition potencies of the synthesised compounds the recombinant human MAO-A and MAO-B enzymes were used. The inhibition potencies were expressed as the corresponding IC50 values. The results showed that the sesamol and benzodioxane derivatives are highly potent and selective inhibitors of MAO-B and to a lesser extent MAOA. The most potent MAO-B inhibitor was 6-(3-bromobenzyloxy)-1,4-benzodioxane with an IC50 value of 0.045 μM. All compounds examined displayed selectivity for the MAO-B isoform over MAO-A. Generally the benzodioxane derivatives were found to be more potent inhibitors of human MAO-A and MAO-B than the sesamol derivatives. The reversibility and mode of MAO-B inhibition of a representative derivative, 6-(3- bromobenzyloxy)-1,4-benzodioxane, was examined by measuring the degree to which the enzyme activity recovers after dialysis of enzyme-inhibitor complexes, while Lineweaver- Burk plots were constructed to determine whether the mode of inhibition is competitive. Since MAO-B activity is completely recovered after dialysis of enzyme-inhibitor mixtures, it was concluded that 6-(3-bromobenzyloxy)-1,4-benzodioxane binds reversibly to the MAO-B enzyme. The Lineweaver-Burk plots constructed were linear and intersected on the y-axis. Therefore it may be concluded that 6-(3-bromobenzyloxy)-1,4-benzodioxane is a competitive MAO-B inhibitor. To conclude, the C6-substituted benzodioxane derivatives are potent, selective, reversible and competitive inhibitors of human MAO-B. These compounds are therefore promising leads for the future development of therapy for Parkinson’s disease. / MSc (Pharmaceutical Chemistry), North-West University, Potchefstroom Campus, 2015

Purpose: Monoamine oxidase (MAO) plays a key role in the treatment of Parkinson‟s disease (PD), since it is the major enzyme responsible for the catabolism of dopamine in the substantia nigra of the brain. Inhibition of MAO-B may conserve dopamine in the brain and provide symptomatic relief. The MAO-B inhibitors that are currently used for the treatment of PD, are associated with a variety of adverse effects (psychotoxic and cardiovascular effects) along with additional disadvantages such as irreversible inhibition of the enzyme. Irreversible inhibition may be considered a disadvantage, since following treatment with irreversible inhibitors, the rate by which the enzyme activity is recovered may be variable and may require several weeks. In contrast, following the administration of reversible inhibitors, enzyme activity is recovered when the inhibitor is cleared from the tissues. There exists therefore, a need to develop new reversible inhibitors of MAO-B which are considered to be safer than irreversible MAO-B inhibitors. Rationale: Recently discovered reversible MAO-B inhibitors include safinamide and (E)-8-(3-chlorostyryl)caffeine (CSC). Safinamide has a benzyloxy side chain, which is thought to be important for inhibition of MAO-B. CSC, on the other hand, consists of a caffeine moiety with a styryl substituent at C-8, which is also a critical feature for its inhibitory activity. In a previous study, the caffeine ring and the benzyloxy side chain were combined to produce a series of 8-benzyloxycaffeine analogues which proved to be potent new MAO-B inhibitors. In this study, caffeine was substituted with the phenoxymethyl functional group at C-8, instead of the benzyloxy moiety. The aim of this study was therefore to compare the MAO-B inhibition potencies of selected 8-(phenoxymethyl)caffeine analogues with the previously studied 8-benzyloxycaffeine analogues. In the current study, 8-(phenoxymethyl)caffeine (1) and nine 8-(phenoxymethyl)caffeine analogues (2-10) were synthesized and evaluated as inhibitors of recombinant human MAOA and –B. These analogues only differed in substitution on C3 and C4 of the phenoxymethyl phenyl ring. The substituents that were selected were halogens (Cl, F, and Br), the methyl group, the methoxy group and the trifluoromethyl group. These substituents are similar to those selected in a previous study where 8-benzyloxycaffeine analogues were evaluated as MAO inhibitors. This study therefore explores the effect that a variety of substituents on C3 and C4 of the phenoxymethyl phenyl ring will have on the MAO-A and –B inhibition potencies of 8-(phenoxymethyl)caffeine. Based on the results, additional 8-(phenoxymethyl)caffeine analogues with improved MAO-A and –B inhibition potencies will be proposed for investigation in future studies. Methods: The target, 8-(phenoxymethyl)caffeine, analogues were synthesized by reacting 1,3- dimethyl-5,6-diaminouracil with the appropriately substituted phenoxyacetic acid in the presence of a carbodiimide coupling agent. Ring closure was catalyzed in basic conditions and methylation of the resulting theophyline intermediates at C-7 was carried out with iodomethane. The structures and purities of all the target compounds were verified by NMR, MS and HPLC analysis. All of the 8-(phenoxymethyl)caffeine analogues were subsequently evaluated as MAO-A and –B inhibitors using the recombinant human enzymes. The inhibition potencies of the analogues were expressed as the IC50 values (concentration of the inhibitor that produces 50% inhibition). In addition, the time-dependency of inhibition of both MAO-A and –B was evaluated for two inhibitors in order to determine if these inhibitors interact reversibly or irreversibly with the MAO isozymes. A Hansch-type quantitative structure-activity relationship (QSAR) study was carried out in order to quantify the effect that different substituents on the phenyl ring of the 8-(phenoxymethyl)caffeine analogues have on MAO-B inhibition activity. Results: The results showed that among the test compounds, several analogues potently inhibited human MAO-B. The most potent inhibitor was 8-(3-bromophenoxymethyl)caffeine with an IC50 value of 0.148 μM toward human MAO-B. There were also inhibitors which displayed inhibition activities towards human MAO-A with IC50 values ranging from 4.59 μM to 34.0 μM. Compared to the 8-benzyloxycaffeine analogues, that were in general non-selective inhibitors, the 8-(phenoxymethyl)caffeine analogues, evaluated here, were selective for MAO-B. For example, 8-(3-bromophenoxymethyl)caffeine was found to be 141 fold more selective as an inhibitor of MAO-B than of MAO-A. Also, compared to the 8-benzyloxycaffeine analogues, the 8-(phenoxymethyl)caffeine analogues were slightly less potent MAO-B inhibitors. For example, 8-benzyloxycaffeine is reported to have an IC50 value of 1.77 μM for the inhibition of human MAO-B while 8-(phenoxymethyl)caffeine was found to have an IC50 value of 5.78 μM for the inhibition of human MAO-B. This study also shows that two selected analogues bind reversibly to MAO-A and –B, respectively, and that the mode of MAO-B inhibition is competitive for one representative compound. Qualitative inspection of the results revealed interesting structure-activity relationships. For the 8-(phenoxymethyl)caffeine analogues, bearing both the C3 and C4 substituents on the phenyl ring, the MAO-B activity significantly increases with halogen substitution. Furthermore, increased MAO-B inhibition was observed with increased electronegativity of the halogen substituent. To quantify these apparent relationships, a Hansch-type QSAR study was carried out. The results showed that the logarithm of the IC50 values (logIC50) correlated with Hansch lipophilicity (π) and the Swain-Lupton electronic (F) constants of the substituents at C-3 of the phenoxymethyl ring. The correlation exhibited an R2 value of 0.87 and a statistical F value of 13.6. From these results it may be concluded that electron-withdrawing substituents at C3 with a high degree of lipophilicity enhance MAO-B inhibition potency. These results are similar to those previously obtained for the series of 8-benzyloxycaffeine analogues. For this series, the MAO-B inhibition potencies correlated with the Hansch lipophilicity (π) and Hammett electronic (σ) constants of the substituents at C-3 of the benzyloxy ring. Similarly to the 8-(phenoxymethyl)caffeine analogues, electron-withdrawing substituents with a high degree of lipophilicity also enhance the MAO-B inhibition potencies of 8-benzyloxycaffeine analogues. / Thesis (M.Sc. (Pharmaceutical Chemistry))--North-West University, Potchefstroom Campus, 2011

Dopamine is an important neurotransmitter that is involved in several human functions such as reward, cognition, emotions and movement. Abnormalities of the neurotransmitter itself, or the dopamine receptors through which it exerts its actions, contribute to a wide range of psychiatric and neurological disorders such as Parkinson’s disease and schizophrenia. Thus far, despite the great interest and extensive research, the exact role of dopamine and the causalities of dopamine related disorders are not fully understood. Here we have developed multimodal imaging methods, to investigate the release of dopamine and the distribution of the dopamine D2-like receptor family in-vivo in healthy humans. We use the [¹¹C]PHNO PET ligand, which enables exploration of dopamine-related parameters in striatal regions, and for the first time in extrastriatal regions, that are known to be associated with distinctive functions and disorders. Our methods involve robust approaches for the manual and automated delineation of these brain regions, in terms of structural and functional organisation, using information from structural and diffusion MRI images. These data have been combined with [¹¹C]PHNO PET data for quantitative dopamine imaging. Our investigation has revealed the distribution and the relative density of the D3R and D2R sites of the dopamine D2-like receptor family, in healthy humans. In addition, we have demonstrated that the release of dopamine has a functional rather than a structural specificity and that the relative densities of the D3R and D2R sites do not drive this specificity. We have also shown that the dopamine D3R receptor is primarily distributed in regions that have a central role in reward and addiction. A finding that supports theories that assigns a primarily limbic role to the D3R.

Numa primeira etapa, estudamos as conexões da VTA para o complexo VTA-substância negra (SN) utilizando a leucoaglutinina do Phaseolus vulgaris (PHA-L). Estas conexões são substanciais, topograficamente organizadas, com destaque para o pólo caudal da VTA que inerva bilateralmente toda a extensão deste complexo. Numa segunda etapa, estudamos as projeções da VTA para o hipotálamo. A VTA se projeta principalmente para a área pré-óptica lateral e área hipotalâmica lateral, a região subfornical posterior e o núcleo dorsomedial. Foram vistas poucas aposições entre varicosidades PHA-L+ e neurônios imunorreativos para orexina ou para hormônio concentrador de melanina. Por fim, estudamos a colocalização do substrato do receptor de insulina (IRS-1), IRS-1 fosforilado e fosfatidilinositol-3 quinase (PI3K) com tirosina hidroxilase (TH) ou com a subunidade B da toxina colérica (CTb) injetada no estriado. A maioria dos neurônios TH+ da VTA-SN expressa IRS-1; injeções de CTb no estriado resultaram em células duplamente marcadas para CTb/IRS-1, CTb/PI3K e CTb/IRS-1 fosforilado. / In a first step, we studied the connections of the VTA to the complex VTA-substantia nigra (SN) using the Phaseolus vulgaris leucoagglutinin (PHA-L). These connections are substantial, topographically organized, especially the caudal pole of the VTA, which innervates bilaterally throughout the length of this complex. In a second step, we studied the projections of the VTA to the hypothalamus. The VTA projected mainly to the lateral preoptic area, lateral hypothalamic area, posterior subfornical region and dorsomedial nucleus. Were observed few appositions between PHA-L+ varicosities and neurons immunoreactive for orexin or melanin-concentrating hormone. Finally, we studied the co-localization of the insulin receptor substrate-1 (IRS-1), IRS-1-phosphorylated and phosphatidylinositol-3 kinase (PI3K) with tyrosine hydroxylase (TH) or cholera toxin B subunit (CTb) injected into the striatum. Most TH+ neurons of the VTA-SN expressed IRS-1; CTb injections in the striatum resulted in cells double-labeled for CTb/IRS-1, CTb/PI3K and CTb/IRS-1 phosphorylated.

Estudos recentes têm sugerido que as imagens de tensor de difusão podem ser úteis para diagnosticar doença de Parkinson (DP). Nosso objetivo foi estimar a exatidão diagnóstica da anisotropia fracionada da substância negra (FA-SN) para o diagnóstico de DP, em uma amostra mais próxima do cenário clínico, incluindo pacientes com tremor essencial (TE) e voluntários sadios (VS). Para uma compreensão mais profunda de nossos achados, também realizamos uma revisão sistemática da literatura e meta-análise para estimar a mudança média da FA-SN induzida pela DP e a precisão diagnóstica dessa medida. Nossa amostra consistiu de 135 pacientes: 72 pacientes com DP, 21 com TE e 42 VS. Dois exames em RM 3T foram realizados em diferentes locais. Em nossa amostra, não encontramos diferenças significativas entre os grupos e a FA-SN não foi útil para o diagnóstico. O maior componente da variabilidade foi a interação sítio-sujeito. Os resultados desta amostra foram fundidos em uma meta-análise que incluiu 1549 indivíduos compostos de 896 pacientes com DP e 653 VS. Utilizaram-se os modelos bivariados e inversos de variância inversa para resumir as medidas de acurácia diagnóstica e as diferenças de médias, respectivamente. A meta-análise estimou uma pequena diminuição nos valores médios da FA-SN na DP (0,03 menor nos pacientes com DP (IC: 0,01 - 0,06)). Apesar disso, sua capacidade discriminatória para o diagnóstico da DP foi baixa. A sensibilidade e a especificidade combinadas foram, respectivamente, 70% (IC: 65 - 74) e 63% (IC: 57 - 69). Houve alta heterogeneidade entre os resultados dos estudos (I2 = 92%). O estudo de caso-controle e a meta-análise das medidas de anisotropia fracionada na substância negra de pacientes com doença de Parkinson e voluntários sadios demonstram que o seu uso como biomarcador de DP não é confiável / Recent studies have suggested that diffusion tensor images can be useful to diagnose Parkinson\'s disease (PD). Our goal was to estimate the diagnostic accuracy of substantia nigra fractional anisotropy (SN-FA) for PD diagnosis in a sample closer to the clinical setting, including patients with essential tremor (ET) and healthy controls (HC). To a deeper understanding of our findings, we also performed a systematic literature review and meta-analysis to estimate mean change of SN-FA induced by PD, and diagnostic accuracy of this measurement. Our sample consisted of 135 subjects: 72 PD and 21 ET patients and 42 HC. Two 3T MRI scans were performed in different sites. In our sample, we did not find significant mean difference between groups and SN-FA was useless for diagnosis. The largest component of explained variability of SN-FA was site-subject interaction. MRI results of this sample were merged in a meta-analysis that included 1549 subjects composed of 896 PD patients and 653 HC. The normal bivariate and the inverse-variance random-effect models were used to summarize diagnostic accuracy measures and mean differences respectively. Meta-analysis estimated a small decrease in mean SN-FA values in PD (0.03 lower in PD patients (CI: 0.01 - 0.06)). Despite this fact, its discriminatory capability to diagnose PD was low. The pooled sensitivity and specificity was respectively 70% (CI: 65 - 74) and 63% (CI: 57 - 69). There was high heterogeneity between studies results (I2 = 92 %). This case-control study and meta-analysis of substantia nigra fractional anisotropy measurements in Parkinson\'s disease and healthy volunteers demonstrate that their use as a PD biomarker is not reliable

La caractéristique neuropathologique majeure de la maladie de Parkinson (MP) est la perte progressive des neurones dopaminergiques (DA) de la substance noire (SN). Nous avons examiné si un dysfonctionnement aigu des EAATs pourrait contribuer au cercle vicieux entretenant la progression des pertes DA. Les effets du PDC, un inhibiteur substrat des EAATs, ont été analysés chez le rat. L'analyse cinétique (4-120 jours) des effets d'une seule injection intranigrale de PDC montre une perte progressive spécifique des neurones DA, avec une évolution unilatérale vers bilatérale et caudo-rostrale. Le processus dégénératif associe déplétion en glutathion et augmentation de l'activité de la γ-glutamyltranspeptidase, stress oxydatif, processus excitotoxiques, autophagie et réactivités gliales. L'antioxydant N-acétylcystéine et les antagonistes des récepteurs NMDA ifenprodil et mémantine exercent un effet neuroprotecteur. Des effets compensatoires transitoires au niveau de marqueurs de la fonction DA dans la SN et le striatum accompagnent la perte cellulaire et des dystrophies axonales. Des troubles moteurs apparaissent de façon tardive lorsque la perte neuronale ipsilatérale avoisine les 50%. Ces résultats montrent un lien fonctionnel entre dysfonctionnement des EAATs et plusieurs mécanismes pathogéniques ainsi que des caractéristiques neuropathologiques majeures de la MP, et fournissent le premier modèle progressif de la maladie induit de façon aiguë. / Parkinson's disease (PD) is characterized by the progressive degeneration of substantia nigra (SN) dopaminergic neurons. Central players in PD pathogenesis, including mitochondrial dysfunction and oxidative stress, might affect the function of excitatory amino acid transporters (EAATs). Here, we investigated whether acute EAATs dysfunction might in turn contribute to the vicious cycles sustaining the progression of dopamine neuron degeneration. PDC application on nigral slices triggered sustained glutamate-mediated excitation selectively in dopamine neurons. In vivo time-course study (4-120 days) revealed that a single intranigral PDC injection triggers progressive degeneration of exclusively dopamine neurons with unilateral to bilateral and caudorostral evolution. This degenerative process associates GSH depletion and specific increase in γ-glutamyltranspeptidase activity, oxidative stress, excitotoxicity, autophagy and glial reaction. The anti-oxidant N-acetylcysteine and the NMDA receptor antagonists ifenprodil and memantine provided significant neuroprotection Transient compensatory changes in dopamine function markers in SN and striatum accompanied cell loss and axonal dystrophy. Motor abnormalities (hypolocomotion and forelimb akinesia) showed late onset, when ipsilateral neuronal loss exceeded 50%. These findings outline a functional link between EAATs dysfunction and several PD pathogenic mechanisms and pathological hallmarks, and provide the first acutely-triggered rodent model of progressive parkinsonism.

<p>The developing nervous system depends largely on extracellular cues to shape its complex network of neurons. Classically, neurotrophins are known to be important mediators in this process. More recently, Bone Morphogenetic Proteins (BMPs), belonging to the Transforming Growth Factor beta (TGFβ) superfamily of secreted cytokines, have been shown to exert a wide range of effects, such as cellular growth, differentiation, survival and apoptosis, both in the developing and adult nervous system. They signal via serine/threonine kinase receptor essentially to the Smad pathway, which carries the signal to the nucleus where the transcription of target genes is regulated.</p><p>This thesis investigates the functions of BMPs in the nervous system, using a set of different models. Firstly, a targeted deletion of GDF10 (BMP3b) in the mouse was established to evaluate the role of this growth/differentiation factor in the hippocampal formation, a brain area known to be involved in memory processing. Other members of the TGFβ superfamily likely compensate for the lack of GDF10, since no detectable alterations in hippocampal function or gene transcription profile have been found. Secondly, a mouse model was set up, with the aim to study impaired BMP-signalling in dopaminergic neurons. The tyrosine hydroxylase (TH) locus was used to drive the expression of dominant negative BMP receptors by means of bicistronic mRNAs. TH is the rate-limiting enzyme in the biosynthesis of catecholamine and the mice described, show a graded decrease of TH-activity resulting in severe to mild dopamine deficiency. The contribution of the dominant negative BMP receptors to the phenotype is however secondary to the apparent TH hypomorphism. The final theme of this thesis is the potentiating effects of BMPs on neurotrophin-induced neurite outgrowth as studied in explanted ganglia from chick embryos and in the rat phaeochromocytoma cell line PC12. A number of pharmacological inhibitors of intracellular signalling kinases were applied to the cultures in order to reveal the contribution of different pathways to the enhanced neurite outgrowth. We made the unexpected finding that inhibition of MEK signalling mimicked the potentiating effects of BMP stimulation in the chick system. The underlying mechanisms for the synergistic effects, however, are still an enigma.</p>

The developing nervous system depends largely on extracellular cues to shape its complex network of neurons. Classically, neurotrophins are known to be important mediators in this process. More recently, Bone Morphogenetic Proteins (BMPs), belonging to the Transforming Growth Factor beta (TGFβ) superfamily of secreted cytokines, have been shown to exert a wide range of effects, such as cellular growth, differentiation, survival and apoptosis, both in the developing and adult nervous system. They signal via serine/threonine kinase receptor essentially to the Smad pathway, which carries the signal to the nucleus where the transcription of target genes is regulated. This thesis investigates the functions of BMPs in the nervous system, using a set of different models. Firstly, a targeted deletion of GDF10 (BMP3b) in the mouse was established to evaluate the role of this growth/differentiation factor in the hippocampal formation, a brain area known to be involved in memory processing. Other members of the TGFβ superfamily likely compensate for the lack of GDF10, since no detectable alterations in hippocampal function or gene transcription profile have been found. Secondly, a mouse model was set up, with the aim to study impaired BMP-signalling in dopaminergic neurons. The tyrosine hydroxylase (TH) locus was used to drive the expression of dominant negative BMP receptors by means of bicistronic mRNAs. TH is the rate-limiting enzyme in the biosynthesis of catecholamine and the mice described, show a graded decrease of TH-activity resulting in severe to mild dopamine deficiency. The contribution of the dominant negative BMP receptors to the phenotype is however secondary to the apparent TH hypomorphism. The final theme of this thesis is the potentiating effects of BMPs on neurotrophin-induced neurite outgrowth as studied in explanted ganglia from chick embryos and in the rat phaeochromocytoma cell line PC12. A number of pharmacological inhibitors of intracellular signalling kinases were applied to the cultures in order to reveal the contribution of different pathways to the enhanced neurite outgrowth. We made the unexpected finding that inhibition of MEK signalling mimicked the potentiating effects of BMP stimulation in the chick system. The underlying mechanisms for the synergistic effects, however, are still an enigma.

Der Morbus Parkinson ist durch den selektiven Zelltod der dopaminergen Neurone der Substantia nigra pars compacta gekennzeichnet. Hierbei sind die verschiedenen Populationen pigmentierter Neurone innerhalb der SNc unterschiedlich stark betroffen. Die Ursachen für diese unterschiedliche Schädigung sind noch nicht bekannt. Möglicherweise besteht aber ein Zusammenhang mit der Verteilung der extrazellulären Matrix innerhalb der Substantia nigra. Für die Untersuchung wurden immunhistochemische Methoden an Hirnschnittserien von menschlichen Kontrollgehirnen angewandt. Zur Darstellung von Komponenten der extrazellulären Matrix wurden drei verschiedene Antikörper genutzt. Dazu gehörten anti- CRTL-1, welcher das Link- Protein 1 von CSPGs detektiert, ein Aggrecan- Antikörper ( Klon HAG7D4), welcher an das Kern- Protein menschlichen Aggrecans bindet, sowie anti- Proteoglykan- Di-0S (Klon 1B5), der die Reste der Chondroitin- Sulfat- Seitenketten verschiedener Proteoglykane detektiert, die nach Verdau mit Chondroitinase ABC übrigbleiben. Zur räumlichen Orientierung und strukturellen Gliederung der Substantia nigra nach der von Damier et al. ( 1999) beschriebenen Calbindin- Methode, auf deren Grundlage die SNc in eine Calbindin-reiche Matrix und Calbindin- arme Bereiche, die sogenannten Nigrosomen, gegliedert wird, wurden benachbarte Hirnschnitte mit anti- Calbindin D₂₈K behandelt. Es zeigte sich, dass extrazelluläre Matrix in Form von perineuronalen Netzen nur an den nicht pigmentierten Neuronen der SNr und SNl vorkommt, während die pigmentierten Neurone der SNc keine perineuronalen Netze besitzen, aber von einer Vielzahl von ACs kontaktiert werden. Deren Dichte war an großen, stark Melanin- haltigen Neuronen am höchsten, sodass in der dorsalen Schicht der SNc, also in den Nigrosomen 3 und 4, besonders viel extrazelluläre Matrix detektiert werden konnte. Im ventralen Anteil der SNc war entsprechend der unterschiedlichen Zellgrößen, insbesondere in Nigrosom 1, eine heterogene Verteilung der extrazellulären Matrix festzustellen. Zur Untersuchung über mögliche Veränderungen der extrazellulären Matrix im Verlauf des Morbus Parkinson wurden Hirnschnitte menschlicher Gehirne mit diagnostiziertem Morbus Parkinson ebenfalls mit den drei Antikörpern zur Darstellung der extrazellulären Matrix behandelt. Dabei zeigte sich, dass insgesamt die Menge extrazellulärer Matrix verringert scheint. Eine Darstellung der perineuronalen Netze mit anti- Proteoglykan- Di-0S (Klon 1B5) war nicht mehr möglich. Wie bereits in früheren Studien verschiedener Autoren festgestellt, waren die stärksten Auswirkungen der neurodegenerativen Prozesse im ventralen Anteil der SNc, vor allem in Nigrosom 1, auszumachen, während die Neurone der Nigrosomen 3 und 4 im dorsalen Anteil weniger vulnerabel erscheinen. Diese Ergebnisse verstärken die Annahme, dass die extrazelluläre Matrix eine protektive Funktion für bestimmte Neuronengruppen besitzt. Bei der Parkinsonschen Erkrankung wird möglicherweise zuerst dieses Schutzsystem zerstört bevor es zum progressiven Neuronenverlust kommt. Ungeklärt bleibt weiterhin was die Ursachen dafür sind.

A recent model of Parkinson’s disease (PD) suggests that the neuropathological, behavioural and cognitive symptoms progress in stages. There is substantial evidence for a prodromal stage of PD, during which time pre-motor symptoms develop. Rapid eye movement (REM) sleep behaviour disorder (RBD) is a risk factor for developing PD and may be part of the pre-motor stage. In both disorders, neuropathological α-synuclein aggregates are thought to be a direct cause of the resulting symptoms. One model has shown that in rats, proteasome inhibition produced by systemic exposure to environmental toxins results in α-synuclein pathology and motor behaviour dysfunction that mimics the progression of PD in humans. The present study examined the hypothesis that the systemic proteasome inhibition model would produce pre-Parkinsonian RBD-like pathology in rats. It was expected that sleep disturbances would be seen prior to behavioural disturbances in rats treated systemically with PSI (a proteasome inhibitor). Following baseline sleep recording and training on the inclined beam-traverse task, rats were injected with PSI (a proteasome inhibitor) or ethanol (control), 6 times over 2 wk. Sleep recording over 8 wk and behavioural testing over 16 wk provided no evidence of sleep disturbances or motor dysfunction. Post-mortem immunohistochemical analyses of brain tissue provided no evidence of PSI-associated α-synuclein aggregates in the locus coeruleus, subcoeruleus (dorsal part), or substantia nigra (areas involved in RBD and/or PD). These results did not provide support for RBD as a prodromal phase of PD within the systemic proteasome inhibitor-based model and add to a growing body of research reporting inconsistent findings using this model. We suggest that systemic PSI exposure in rats does not produce a viable model of RBD or PD. Whether RBD is an early symptom in the progression of PD remains to be established. / Thesis (Master, Neuroscience Studies) -- Queen's University, 2010-04-28 12:04:50.613