Thesis (PH. D.)--University of Iowa, 2007. / Supervisor: Antoine Bechara. Includes bibliographical references (leaves 174-195).

Ο εγκεφαλικός φλοιός των θηλαστικών σχηματίζεται από το πιο πρόσθιο τμήμα του νευρικού σωλήνα, τον προσεγκέφαλο. Η ανάπτυξη του εγκεφαλικού φλοιού είναι μια διαδικασία που περιλαμβάνει την σωστή οργάνωση των πρόδρομων νευρικών κυττάρων σε συγκεκριμένες περιοχές του αναπτυσσόμενου φλοιού καθώς επίσης και την εμφάνιση τους σε καθορισμένη χρονική στιγμή κατά την πορεία της ανάπτυξης. Κατά την νευρογένεση όλοι οι νευρώνες του εγκεφαλικού φλοιού προέρχονται από το νευροεπιθήλιο που βρίσκεται δίπλα από τις πλευρικές κοιλίες. Τα νευροεπιθηλιακά κύτταρα αρχικά διαιρούνται με σκοπό την δημιουργία ικανού αριθμού πρόδρομων κυττάρων που θα μπορέσουν να δώσουν γένεση στον αναπτυσσόμενο φλοιό. Αργότερα, τα κύτταρα αυτά, διαφοροποιούνται προς τις άλλες κατηγορίες πρόδρομων κυττάρων και προς τους διαφοροποιημένους νευρώνες. Η ανάπτυξη του εγκεφαλικού φλοιού είναι μια διαδικασία που απαιτεί τον έλεγχο της ισορροπίας μεταξύ του πολλαπλασιασμού και της διαφοροποίησης των πρόδρομων νευρικών κυττάρων. Η Geminin έχει δειχτεί ότι ρυθμίζει τον κυτταρικό πολλαπλασιασμό και την διαφοροποίηση αποτελώντας έναν συνδετικό κρίκο μεταξύ των δυο αυτών διαδικασιών. Με σκοπό να διερευνήσουμε τον in vivo ρόλο της Geminin στη διατήρηση και διαφοροποίηση των πρόδρομων νευρικών κυττάρων του εγκεφαλικού φλοιού του μυός, αδρανοποιήσαμε το γονίδιο της Geminin ειδικά στα κύτταρα του αναπτυσσόμενου εγκεφαλικού φλοιού. Τα αποτελέσματα μας έδειξαν ότι η απαλοιφή της Geminin έχει ως αποτέλεσμα την επέκταση της κοιλιακής ζώνης του φλοιού ως συνέπεια της παραγωγής μεγαλύτερου αριθμού πρόδρομων κυττάρων στην κορυφαία περιοχή του φλοιού. Επιπλέον η απώλεια του γονιδίου της Geminin αυξάνει τον καθορισμό των βασικών προγονικών κυττάρων της υποκοιλιακής ζώνης του φλοιού. Ως άμεσο επακόλουθο η αύξηση του αριθμού των πρόδρομων κυττάρων στις νευρογενετικές ζώνες του φλοιού είναι η παραγωγή μεγαλύτερου αριθμού πλήρως διαφοροποιημένων νευρικών κυττάρων στον εγκεφαλικό φλοιό. Η μελέτη μας αναδεικνύει την Geminin ως μόριο κλειδί κατά την πορεία της ανάπτυξης του εγκεφαλικού φλοιού, στον καθορισμό του σωστού αριθμού των πρόδρομων κυττάρων, στην πορεία διαφοροποίησης αυτών και τελικά στον σχηματισμό του σωστού αριθμού των πλήρως διαφοροποιημένων νευρώνων. / The mammalian cerebral cortex originates from the most anterior part of the neural tube, the prosencephalon. The cortical development is a process that involves the correct orchestration of the different neuronal progenitor lineage of the cortex, which involves the regional patterning of progenitors and their temporal specification during neurogenesis. During neurogenesis, the cortical neurons are originated from the neuroepithelium that lies next to the lateral vesicles. At the beginning, the neuroepithelial cells divide in order to expand their population and to create the essential number of progenitor cells that would give rise to the neurons and glia that comprise the cortex, a procedure called self-renewal. Later on, the neuroepithelial cells start to switch from divisions that generate additional progenitor cells to divisions that generate committed progenitors or postmitotic cells. The development of cerebral cortex is a process that requires coordination of proliferation and differentiation of progenitor cells. It has been proposed that Geminin regulates both cell proliferation and differentiation. In order to investigate the in vivo role of Geminin in the maintenance and the differentiation of the neuronal progenitors of the cerebral cortex, we specifically inactivated the mouse Geminin gene in the developing cortex. Our results indicate that deletion of Geminin results in the expansion of the ventricular zone that leads to the production of a higher number of the apical cortical progenitors at the middle stage of cortical neurogenesis. In addition, the depletion of Geminin influences the number of basal progenitor of the subventricular zone. The increase of the apical and basal progenitor cells results in the overproduction of differentiated neurons of the developing cortex. Our work demonstrates Geminin as an important molecule for the development of the mouse cerebral cortex, regulating the correct number of cortical progenitors and the generation of the correct number of neurons and glial cells of the cerebral cortex.

Orientador: Celso Aparecido Bertran / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Quimica / Made available in DSpace on 2018-08-03T21:02:24Z (GMT). No. of bitstreams: 1 Cludi_CassianadeZorzetto_M.pdf: 5355081 bytes, checksum: dd74f392ec23534f8272dc44017bc5c4 (MD5) Previous issue date: 2003 / Resumo: Este trabalho reporta o estudo realizado com 2 tipos de materiais particulados gerados na combustão: Cenosferas, no caso de combustão de óleos com elevada razão C/H e Fuligem no caso de combustão de GLP e misturas de GLP/1-butanol e GLP/Metanol. Foram determinadas a morfologia destes materiais particulados e a formação de HPA (Hidrocarbonetos Policíclicos Aromáticos) nos processos de combustão estudados. A influência da requeima no caso das cenosferas e da adição de álcoois no caso da fuligem sobre estas propriedades, também foram avaliadas. A ação da requeima sobre as cenosferas resultou no aumento da porosidade deste particulado e na diminuição da quantidade de HPA presentes nas cenosferas. A adição de vapor de 1-butanol e de metanol na combustão de GLP resultou no aumento do tamanho dos aglomerados de partículas de fuligem, assim como na diminuição da quantidade de compostos aromáticos presentes neste material. As mudanças observadas nas características da fuligem, coletada na região da chama na qual este material particulado é formado, evidenciaram a ação causada pela adição de vapores de álcool ao GLP sobre o mecanismo de formação de aromáticos e no processo de aglomeração das partículas / Abstract: This work reports the studies performed with two kinds of particulated materiais generated from combustion: cenospheres, in the case of the combustion of oils with high C/H ratio, and soot, in the case of LPG and mixtures of LPG/1butanol and LPG/Methanol combustion. The morphologies of these particulated materiais and the formation of PAH (polycyclic aromatic hydrocarbons) in the combustion processes were determined. The influence of reburning, in the case of cenospheres, and the addition of alcohols, in the case of soot, on these properties were also evaluated. The action of reburning on the cenospheres resulted in an increase of the porosity of this particulated material, as well as in a reduction oh the quantity of P AH present in cenospheres. The addiction of both 1-butanol and methanol vapor in the combustion of LPG resulted in an increase in the size of the soot particle agglomerates as well as in a reduction in the quantity of the aromatic compounds present in this material. The changes observed in the characteristics of the soot collected in the flame region, where this particulated material is formed, demonstrated the effect of the alcohol vapor added to the LPG on the mechanism of aromatic compound formation and on the process of particle agglomeration / Mestrado / Mestre em Química

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The most widely accepted hypotheses suggest that general anaesthetics interrupt conscious processes in the brain by decreasing synaptic excitation or by potentiating synaptic inhibition, especially in the neocortex. The putative transmitters in the neurological systems that generate neocortical arousal include acetylcholine, glutamate and γ-aminobutyrate (GABA). The primary objective here was to determine the neuronal mechanisms by which anaesthetics may obtund this arousal. The majority of the investigations were carried out on pyramidal neurons in layers IV and V of guinea pig neocortex (in vitro slices), using intracellular recording and pharmacological, including microiontophoretic, techniques. Bath applications of structurally dissimilar anaesthetics, isoflurane - a halogenated ether, and Althesin - a steroidal preparation, in concentrations of 0.5-2.5 minimum alveolar concentration (MAC) and 10-1300 μM, respectively, produced a small hyperpolarization (3-5 mV) which was associated with an increase in input conductance (10-30%). The lower concentrations (0.5-1.5 MAC and 10-200 μM) of these agents which are most relevant to the production of unconsciousness did not significantly affect the passive membrane properties. However, they produced striking decreases in spontaneous activities and the repetitive spike firing evoked by orthodromic (electrical) stimulation or intracellular current injections. Because the observed changes in membrane properties could not explain the reduction in neuronal excitability, the effects of anaesthetics were investigated extensively on excitatory and inhibitory postsynaptic potentials (EPSPs and IPSPs). The application of isoflurane or Althesin induced a dose-dependent, reversible depression in the amplitude of EPSPs, with EC₅₀s of 1 MAC and ~50 μM, respectively. The IPSPs also were reduced in a dose-dependent manner. In order to eliminate possible shunting of the EPSPs by the GABA-activated Cl-conductance that produces the IPSP in the observed EPSP-IPSP sequence, a GABA[symbol omitted]-antagonist, bicuculline, was additionally applied. Despite this IPSP-blockade, the anaesthetics strongly depressed the EPSPs as well as epileptiform activities evoked by subpial electrical stimulation. In cognizance of the possibility that a postsynaptic attenuation of responsiveness to transmitter substances may be involved in the EPSP depression, the neuronal sensitivities to acetylcholine, glutamate. and GABA were determined. Anaesthetic administration markedly reduced the depolarizations and associated conductance changes evoked by dendritic applications of acetylcholine, glutamate and N-methyl-D-aspartate (NMDA). The hyperpolarizing responses to somatic applications of GABA were not affected significantly whereas the depolarizing effects observed with its dendritic application were slightly depressed. Same degree of selectivity also was evident from the lower EC₅₀s for the isoflurane- and Althesin-induced depressions of responses to acetylcholine compared with glutamate. Under in vitro conditions of hypomagnesia the responses to acetylcholine were totally blocked and the order of depression in the responses to GABA and glutamate was reversed; this may be of importance in the mechanism for the known increase in anaesthetic requirements in clinical syndromes associated with hypomagnesaemia. Because the genesis of synaptic transients is affected by Ca²⁺ influx or disposition, the interactions of anaesthetics were investigated on spike afterhyperpolarizations (AHPs). The AHPs which are produced specifically by a Ca²⁺ -activated K⁺ -conductance were suppressed by the anaesthetics in a dose-dependent manner under conditions where contaminating IPSPs had been blocked by bicuculline. Since the passive membrane properties were unaffected, an interference with a transmembrane Ca⁺ -influx may be involved in the anaesthetic actions. The effects of anaesthetics on glutamate-induced and voltage-dependent increases in intraneuronal Ca²⁺ ([Ca²⁺]i) were determined in cultured hippocampal neurons with a Ca-sensitive probe (Fura-2) and microspectro- fluorometric techniques. Isoflurane application depressed the increases in [Ca²⁺]i. produced by application of glutamate under conditions where its actions would be favoured at NMDA- and quisqualate-subtypes of receptors. K⁺ -induced increases in [Ca²⁺]i also were reduced by application of isoflurane, probably due to actions on voltage-dependent Ca-channels in the membrane. These investigations have provided evidence for the first time that excitatory transmitter actions in neocortex are selectively depressed by anaesthesia. A plausible mechanism would include suppression of the postsynaptic Ca-conductances associated with the AHPs and glutamatergic, as well as cholinergic interactions at pre- and post-synaptic sites on neurons involved in neocortical arousal. / Medicine, Faculty of / Anesthesiology, Pharmacology and Therapeutics, Department of / Graduate

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The brain is a complex system which consists of billions of neuron cells and gives rise to diverse neural dynamics spatially and temporally. Spontaneous neural activities construct the foundation for various cognitive processing. However, caused by the limitation spatiotemporal resolution and coverage of recording methods in experiments, the organization of spatiotemporal dynamics of the self-organized brain activity remains largely unknown. Current experimental technique can optically image population voltage transients generated by pyramidal neurons across cortical layer 2/3 of the mouse dorsally with a genetically encoded voltage indicator. Such data provided unique opportunities to investigate the structure- dynamics relationship to elucidate the mechanisms of spontaneous brain activity. The aim of this thesis is to develop a systematic understanding of spatiotemporal mechanism in the mouse cortex by analyzing voltage imaging data, in collaboration with neuroscientist Dr. Knöpfel from the Imperial College London. Local oscillation properties such as duration, amplitude and oscillation forms were studies on the cortex-wide scale and be compared among brain states. Wakefulness modulated the excitability of the neural activity which influenced the duration of the oscillation and the transition of different half wave types. Relatively larger amplitude of parietal cortex reflected stronger neural activity determined by structural hierarchy. Motifs of the oscillations showed consistency in different brain states which indicated typical pathways of the wave propagations. Dynamical properties of various waves and their interactions in sedated mice were investigated. Based on phase velocity fields, there were only a small number of large-scale, cortex-wide plane wave and synchrony (standing wave) patterns during Up-Down states. Interactions of local sources and sinks can generate saddles, and interactions of local wave patterns with large plane waves can induce a change of their wave propagating direction. Local wave patterns emerged at preferred spatial locations. Specifically, sources were predominantly found in cortical regions with high cumulative input through the underlying connectome. The findings revealed the principled spatiotemporal dynamics of Up-Down states and associated them with the large-scale cortical connectome. Waking from deep anesthesia to consciousness increased the number of local wave patterns and made the spatiotemporal dynamics more complex. Although the active state increased the wave propagation speeds, the average speed decreased because of the interaction and collapse of wave patterns. Not affected by the brain states, the two principal modes with the highest variance remained stable. The first mode represented the large waves spreading across the cortex forward or backward while the second mode corresponded to the waves propagating in opposite direction in the frontal and parietal cortex. An infra-slow frequency of the wave number might reflect the bold flow and oxygenation. The characterizations presented in this thesis can be used to predict and guide measurement and analysis of large-scale brain activity. The analysis of cortex-wide neural dynamical patterns builds foundation for further investigation of their functional implications.

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