Made available in DSpace on 2014-12-17T15:16:18Z (GMT). No. of bitstreams: 1 AngelicaVCS_DISSERT.pdf: 2105616 bytes, checksum: b2b0f7446903336c6b6bfed546f45efd (MD5) Previous issue date: 2012-12-19 / Conselho Nacional de Desenvolvimento Cient?fico e Tecnol?gico / Background: The gait automaticity loss difficults realization of concurrent activities - Dual Task (DT). In these situations, individuals with Parkinson`s disease (PD) show a significant reduction in gait velocity and stride length, as strides variability and asymmetry increased, factors predisposing to falls. However, recent studies have shown that training involving DT may cause subsequent improvements in gait variables with DT in individuals with PD. The treadmill use was adopted by this study, by promoting greater regularity in step and enhance training. Objective:To investigate immediate effects of gait training associated with cognitive tasks on gait in individuals with PD. Methods: Twenty-two volunteers were randomly divided into two groups: control group (n = 11), who performed gait training on a treadmill for 20 minutes, and the experimental group (n = 11), who performed treadmill gait training for 20 minutes associated with cognitive tasks of verbal fluency, memory, and spatial planning. Participants were evaluated in phase on of antiparkinsonian medication as the demographic, clinical and anthropometric (identification form), cognitive status (Montreal Cognitive Assessment - MoCA), executive function (Frontal Assessment Battery), level of physical disability (Hoehn and Yahr Modified), motor and functional status (Unified Rating Scale for Parkinson`s Disease - UPDRS), and kinematics (Qualisys Motion Capture System). Results: There were not differences between groups, but both showed improvement after the intervention. The control group had an increase in velocity (p = 0.008), stride length (p = 0.04), step length (p = 0.02) and decreased double support time(p = 0.03). The experimental group showed an increase in speed (p = 0.002), stride length (p = 0.008), step length (p = 0.02) and cadence (p = 0.01), as well as a decrease in the width stride (p = 0.001) and total support time (p = 0.02). As the angular variables, the experimental group had a significant increase in the initial contact angle of ankle (p = 0.01). Conclusion: The gait training combined with cognitive activities didn`t provide significant improvements in gait variables with DT, but this study was the first to demonstrate that gait training on treadmill as simple task minimized the negative interference of DT in PD / Introdu??o: A perda na automaticidade da marcha dificulta a realiza??o de atividades concorrentes - Dupla Tarefa (DT). Nessas situa??es, indiv?duos com Doen?a de Parkinson (DP) apresentam significativa redu??o na velocidade da marcha e no comprimento do passo, assim como aumento na variabilidade entre as passadas e na assimetria, fatores predisponentes a quedas. No entanto, estudos recentes t?m demonstrado que o treinamento envolvendo DT pode ocasionar posteriores melhoras nas vari?veis da marcha com DT em indiv?duos com DP. O uso da esteira foi adotado por este estudo, por promover maior regularidade no passo e potencializar o treino. Objetivo: Investigar os efeitos imediatos do treino de marcha em esteira associado a tarefas cognitivas na marcha com DT de indiv?duos com DP. Materiais e m?todos: Vinte e dois volunt?rios foram divididos aleatoriamente em dois grupos: Grupo Controle (n=11), que realizou treino de marcha em esteira durante 20 minutos; e Grupo Experimental (n=11), que realizou treino de marcha em esteira durante 20 minutos associado a tarefas cognitivas de flu?ncia verbal, mem?ria e planejamento espacial. Os participantes foram avaliados na fase on do medicamento antiparkinsoniano quanto a dados demogr?ficos, cl?nicos e antropom?tricos (formul?rio de identifica??o), condi??o cognitiva (Montreal Cognitive Assessment - MoCA), fun??es executivas (Bateria de Avalia??o Frontal), n?vel de incapacidade f?sica (Escala de Hoehn e Yahr Modificada), estado motor e funcional (Escala Unificada de Avalia??o para a Doen?a de Parkinson UPDRS), e cinemetria (Qualisys Motion Capture System). Resultados: N?o houve diferen?as entre os grupos, mas ambos apresentaram melhoras ap?s a interven??o. O grupo controle obteve aumento na velocidade (p=0,008), no comprimento da passada (p=0,04), no comprimento do passo (p=0,02) e diminui??o no tempo de duplo suporte (p=0,03). O grupo experimental apresentou aumento na velocidade (p=0,002), no comprimento da passada (p=0,008), no comprimento do passo (p=0,02) e na cad?ncia (p=0,01), assim como diminui??o na largura da passada (p=0,001) e no tempo total de apoio (p=0,02). Quanto ?s vari?veis angulares, o grupo experimental teve um aumento significativo no ?ngulo do contato inicial do tornozelo (p=0,01). Conclus?o: O treino de marcha associado a atividades cognitivas n?o proporcionou melhoras significativas nas vari?veis da marcha com DT, mas este estudo foi o primeiro a demonstrar que o treino de marcha como tarefa simples na esteira minimizou a interfer?ncia negativa da DT na DP

Fundação de Amparo a Pesquisa no Estado do Rio Grande do Sul / Animal models as reserpine are helpful to understand the pathophysiology of several diseases with involuntary movements, as Parkinson s disease (PD), and to search efficient treatments. The present study tested the effects of reserpine on behavioral alterations induced by reserpine in rats and worms, with emphasis in oxidative and dopaminergic parameters, and the effect of the antioxidant gallic acid (GA) in reserpine-exposed rats. As result, reserpine (1mg/Kg, sc, for 3 consecutive days) increased the frequency of vacuous chewing movements (VCMs) in rats in relation to controls, and maintained this increase for at least 3 days after reserpine withdrawal. Treatment with GA (4.5 , 13.5 or 40.5 mg/kg/day, po) for 3 days reverted reserpine-induced increase in VCMs, showing protective effect. Neither reserpine nor GA changed oxidative parameters (TBARS and DCFH-DA oxidation), antioxidant levels (proteic and non-proteic thiol) and the activity of Na+,K+-ATPase (total and α-subunit) in striatum and cortex. Afterward, studies were performed with Caenorhanditis elegans due its several advantages in studies of neurodegeneration and of drugs mechanism of action. L1-larval stage C. elegans were exposed to reserpine (30 ou 60 μM) for different times. Reserpine decreased the survival, development, food intake, locomotor rate on food and dopamine (DA) levels in worms and it had effect on egg laying and defecation cycles. Morphological evaluations of dopaminergic cephalic (CEP) neurons in BY200 worms (with GFP coupled to dat-1 gene) reveled neurodegeneration by: 1) decreased fluorescence intensity, 2) decreased the number of intact neurons, and 3) increased the number of shrunken somas per worm. These effects were unrelated to reserpine s effect on dat-1 gene expression. Interestingly, the reserpine effects on locomotor rate, dopaminergic CEP neurons morphology and dat-1 gene expression were reverted after reserpine withdrawal. Furthermore, reserpine decreased the survival of vesicular monoamine transporter (VMAT) and dat-1 loss-of-function mutant worms, but no of tyrosine hydroxylase (TH, cat-2) and dopaminergic receptors (dop-1, dop-2, dop-3 e dop-4) loss-of-function mutants in relation to wild-type N2 worms. Reserpine also decreased the survival of worms pre-exposed to DA; and it activated SKN-1 detoxification pathway. Moreover, no differences were found in DAT and TH immunoreactivity in striatum of rats treated with reserpine and/or GA. The GA protective effects against reserpine-induced VCMs in rats are probably not related to its antioxidant and antiapoptotic properties or monoamine oxidase (MAO) inhibition. As conclusion, the reserpine decreases DA levels though action on VMAT, and it induces neurotoxicity/neurodegeneration due probably an increase on extracellular DA contents resulted from changes on DAT function. More studies evaluating the reserpine effect on DAT and the GA mechanism of protection are necessary. / Modelos animais como o da reserpina auxiliam no entendimento da fisiopatologia de diversas doenças que se manifestam por movimentos involuntários, como a doença de Parkinson (DP), e na busca por formas de tratamento. O presente trabalho avaliou mecanismos envolvidos na indução de alterações comportamentais induzidas por reserpina em ratos e vermes com ênfase em parâmetros oxidativos e dopaminérgicos, e a ação do antioxidante ácido gálico (AG) em ratos tratados com reserpina. Como resultado, a reserpina (1mg/Kg, sc, por 3 dias consecutivos) aumentou a frequência de movimentos de mascar no vazio (MMVs) em ratos em relação ao controle, e manteve esse aumento por pelo menos 3 dias após o término das administrações da reserpina. O tratamento com AG (4,5 ou 13,5 ou 40,5 mg/kg/day, vo) por 3 dias reverteu esse aumento dos MMVs, mostrando efeito protetor. Nem reserpina nem o AG alteraram os parâmetros avaliados de dano oxidativo (TBARS e oxidação da DCFH-DA), de antioxidantes (tiol protéico e não protéico) e a atividade da Na+,K+-ATPase (total e α-subunidade) no estriado e córtex cerebral. Estudos posteriores foram realizados em Caenorhanditis elegans devido as diversas vantagens oferecidas por esse modelo animal em estudos de neurodegeneração e de investigação do mecanismo de ação de drogas. C. elegans em estágio larval L1 foram expostos a reserpina (30 ou 60 μM) por diferentes tempos. A reserpina reduziu a sobrevivência, o desenvolvimento, a ingestão de alimento, a atividade locomotora na comida e as concentrações de dopamina (DA) nos vermes, e afetou a postura de ovos e tempo entre defecações. Análise da morfologia dos neurônios dopaminérgicos cefálicos (CEP) de vermes BY200 (com GFP acoplado ao gene dat-1) indicam neurodegeneração por: 1) redução da intensidade da fluorescência, 2) redução do número de neurônios intactos e 3) aumento do número de somas atrofiados por verme em relação ao controle. Esses efeitos não estão relacionados a efeitos da reserpina na expressão do gene dat-1. Interessantemente, os efeitos da reserpina na atividade locomotora, na morfologia dos neurônios CEP e na expressão do gene dat-1 foram revertidos após a retirada dos vermes da exposição a reserpina. Em adição, a reserpina reduziu a sobrevivência de vermes deficientes do transportador vesicular de monoaminas (TVMs, cat-1) e do transportador de DA (DAT, dat-1), mas não alterou a sobrevivência de deficientes da tirosina hidroxilase (TH, cat-2) e dos receptores dopaminérgicos (dop-1, dop-2, dop-3 e dop-4) em relação aos vermes selvagens N2. A reserpina também reduziu a sobrevivência de vermes N2 pré-expostos a DA, e ativou a via de detoxificação SKN-1 dos vermes. Alterações na imunoreatividade ao DAT e a TH no estriado de ratos tratados com reserpina e/ou AG não foram encontradas. O efeito protetor do AG nos MMVs induzidos por reserpina em ratos parece não envolver sua atividade antioxidante, antiapoptótica ou na monoaminoxidase (MAO). Como conclusão, a reserpina age no TVMs causando depleção de DA, e causa neurotoxicidade/neurodegeneração dopaminérgica devido provavelmente a um acúmulo de DA no espaço sináptico resultande de uma interferência no funcionamento do DAT. Mais estudos avaliando a ação da reserpina no DAT e o mecanismo de proteção do AG são necessários.

[pt] A doença de Parkinson (DP) é considerada a segunda doença neurodegenerativa mais comum e suas características motoras são muito mais conhecidas que as não motoras, mas o comprometimento funcional está presente em quase todos os casos. A Estimulação Cerebral Profunda (ECP), que consiste na estimulação elétrica de estruturas subcorticais para diminuir ou cessar os sintomas motores, tem sido usada como uma ferramenta para maior controle dos sintomas motores e está ganhando terreno em estudos com sintomas não motores. É por esse motivo que foi realizada uma revisão sistemática para conhecer os instrumentos utilizados na avaliação neuropsicológica de pessoas com DP submetidas à cirurgia ECP nos núcleus subtalâmicos (NST), além de investigar também se surgiriam efeitos cognitivos após a cirurgia. Além disso, uma bateria neuropsicológica computadorizada, a CompCog, foi validada para pessoas com DP de um hospital público da cidade do Rio de Janeiro e também foi usada para comparar os estágios ON e OFF de 9 pacientes, de uma clínica privada, que fizeram a implantação da ECP-NST. Com as revisões sistemáticas foi possível elaborar um protocolo de avaliação neuropsicológica, posteriormente utilizado nos estudos empíricos e verificar que a fluência verbal foi o aspecto que apresentou maior diferença entre os estágios ON e OFF dos pacientes com ECP-NST. No estudo de validação clínica da CompCog foi possível estabelecer pontos de corte para as pessoas com DP e no estudo de comparação entre estágios ON e OFF de pessoas com DP e ECP-NST foi possível identificar que as variáveis de tempo como tempo médio de reação e tempo total, foram capazes de diferenciar os dois estágios, ON e OFF, da amostra de 9 pessoas nos subtestes de memória incidental, memória episódica e controle inibitório, além de apresentar uma tendência à diferenciação na atenção, velocidade de processamento e memória episódica. / [en] Parkinson s disease (PD) is considered the second most common neurodegenerative disease and its motor characteristics are much better known than non-motor ones, but functional impairment is present in almost all cases. Deep Brain Stimulation (DBS), which consists of electrical stimulation of subcortical structures to decrease or stop motor symptoms, has been used as a tool for greater control of motor symptoms and is gaining ground in studies with non-motor symptoms. It is for this reason that a systematic review was carried out to find out the instruments used in the neuropsychological assessment of people with PD who underwent DBS surgery in subtalamic nuclei (STN) and to investigate whether cognitive effects would arise after surgery. In addition, a computerized neuropsychological battery, the CompCog, was validated for people with PD from a public hospital in the Rio de Janeiro city and was also used to compare ON and OFF stages of 9 patients, from a private clinic, who did the implementation of the DBS-STN. With systematic reviews, it was possible to develop a neuropsychological assessment protocol, later used in empirical studies and to verify that verbal fluency was the aspect that showed the greatest difference between the ON and OFF stages of patients with ECP-NST. In the clinical validation study of CompCog it was possible to establish cutoff points for people with PD and in the comparison study between ON and OFF stages of people with PD and ECP-NST it was possible to identify that the time variables such as mean reaction time and total time, were able to differentiate the two stages, ON and OFF, of the sample of 9 people in the subtests of incidental memory, episodic memory and inhibitory control, in addition to showing a tendency to differentiate in attention, processing speed and episodic memory.

Background: Consecutive adult neurogenesis is a well-known phenomenon in the ventricular–subventricular zone of the lateral wall of the lateral ventricles (V–SVZ) and has been controversially discussed in so-called “non-neurogenic” brain areas such as the periventricular regions (PVRs) of the aqueduct and the fourth ventricle. Dopamine is a known modulator of adult neural stem cell (aNSC) proliferation and dopaminergic neurogenesis in the olfactory bulb, though a possible interplay between local dopaminergic neurodegeneration and induction of aNSC proliferation in mid/hindbrain PVRs is currently enigmatic. Objective/Hypothesis: To analyze the influence of chronic–progressive dopaminergic neurodegeneration on both consecutive adult neurogenesis in the PVRs of the V–SVZ and mid/hindbrain aNSCs in two mechanistically different transgenic animal models of Parkinson´s disease (PD). Methods: We used Thy1-m[A30P]h α synuclein mice and Leu9′Ser hypersensitive α4* nAChR mice to assess the influence of midbrain dopaminergic neuronal loss on neurogenic activity in the PVRs of the V–SVZ, the aqueduct and the fourth ventricle. Results: In both animal models, overall proliferative activity in the V–SVZ was not altered, though the proportion of B2/activated B1 cells on all proliferating cells was reduced in the V–SVZ in Leu9′Ser hypersensitive α4* nAChR mice. Putative aNSCs in the mid/hindbrain PVRs are known to be quiescent in vivo in healthy controls, and dopaminergic deficiency did not induce proliferative activity in these regions in both disease models. Conclusions: Our data do not support an activation of endogenous aNSCs in mid/hindbrain PVRs after local dopaminergic neurodegeneration. Spontaneous endogenous regeneration of dopaminergic cell loss through resident aNSCs is therefore unlikely.

No description available.

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

A Doença de Parkinson (DP) é a segunda doença neurodegenerativa mais comum na população, sendo seus mecanismos estudados em modelos animais. Há evidências de que alguns sistemas de comunicação celular podem modular o desenvolvimento da DP. Os canais de potencial transiente (TRPs) e receptores purinérgicos parecem envolvidos com a neurodegeneração e podem contribuir no desenvolvimento da DP, por outro lado, as cininas parecem estar relacionadas com a neuroproteção. O objetivo deste estudo foi avaliar a expressão e o envolvimento destes receptores de membrana no modelo da DP induzida por 6-OHDA em camundongos (C57Bl/6) injetados com agonista (BK) e antagonista (HOE-140) do receptor B2, e antagonista de TRPM7 (Carvacrol); e em nocautes do receptores B2, pelas técnicas de imuno-histoquímica e Western blotting. Os dados revelaram modulação destes receptores no modelo, e neuroproteção após o bloqueio do TRPM7. Assim, podemos sugerir que todos os receptores avaliados podem estar envolvidos na indução do modelo por 6-OHDA, sendo possíveis alvos terapêuticos para a DP. / Parkinson\'s disease (PD) is a second most common neurodegenerative disease in the population, and the mechanisms involved in these are studied in animal models. There is evidence that some systems can modulate the development of PD. Transient potential channels (TRPs) and purinergic receptors seem to be involved in neurodegeneration and may contribute to the development of PD; on the other hand, kininas appear to be related to neuroprotection. The aim of this study was to evaluate the expression and the involvement of these membrane receptors in the 6-OHDA-induced PD model in mice (C57Bl / 6) injected with agonist (BK) and antagonist (HOE-140) of B2 receptor; antagonist of TRPM7 (Carvacrol); and in B2 knockout knockouts, by immunohistochemistry and Western blotting techniques. The data revealed modulation of these receptors in the model, and neuroprotection after TRPM7 blockade. Thus, we can suggest that all the receptors evaluated may be involved in the induction of the 6-OHDA model, and can be possible therapeutic targets for PD.

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.

No description available.

The methods of analysis with a focused ion beam, commonly termed as nuclear microscopy, include quantitative physical processes like PIXE and RBS. The element concentrations in a sample can be quantitatively mapped with a sub-micron spatial resolution and a sub-ppm sensitivity. Its fully quantitative and non-destructive nature makes it particularly suitable for analysing biological samples. The applications in biomedical research are manifold. The iron overload hypothesis in Parkinson\\\'s disease is investigated by a differential analysis of human substantia nigra. The trace element content is quantified in neuromelanin, in microglia cells, and in extraneuronal environment. A comparison of six Parkinsonian cases with six control cases revealed no significant elevation in iron level bound to neuromelanin. In fact, a decrease in the Fe/S ratio of Parkinsonian neuromelanin was measured, suggesting a modification in its iron binding properties. Drosophila melanogaster, or the fruit fly, is a widely used model organism in neurobiological experiments. The electrolyte elements are quantified in various organs associated with the olfactory signalling, namely the brain, the antenna and its sensilla hairs, the mouth parts, and the compound eye. The determination of spatially resolved element concentrations is useful in preparing the organ specific Ringer\\\'s solution, an artificial lymph that is used in disruptive neurobiological experiments. The role of trace elements in the progression of atherosclerosis is examined in a pilot study. A differential quantification of the element content in an induced murine atherosclerotic lesion reveals elevated S and Ca levels in the artery wall adjacent to the lesion and an increase in iron in the lesion. The 3D quantitative distribution of elements is reconstructed by means of stacking the 2D quantitative maps of consecutive sections of an artery. The feasibility of generating a quantitative elemental rodent brain atlas by Large Area Mapping is investigated by measuring at high beam currents. A whole coronal section of the rat brain was measured in segments in 14 h. Individual quantitative maps of the segments are pieced together to reconstruct a high-definition element distribution map of the whole section with a subcellular spatial resolution. The use of immunohistochemical staining enhanced with single elements helps in determining the cell specific element content. Its concurrent use with Large Area Mapping can give cellular element distribution maps.