La locomotion spontanée a une forte influence sur l’état du réseau hippocampique et joue un rôle crucial lors de l’intégration de l'information spatiale. Différents états d'attention ou de comportement au cours de l'éveil peuvent modifier la réponse des neurones aux stimuli sensoriels ainsi que les performances dans les tâches associées. Au cours du mouvement (mov.) le potentiel de champ local de l’hippocampe est caractérisé par des oscillations de fréquence thêta et les cellules pyramidales (CPs) présentent une décharge spécifique à la localisation de l'animal dans un environnement donné. Cependant, les déterminants intracellulaires liés à l'activation des cellules pyramidales de CA1 pendant du mov. sont peu connus. Dans ce travail de thèse, nous avons enregistré le potentiel de membrane (Vm) des CPs de CA1 chez des souris qui alternaient spontanément entre des périodes de mov. et des périodes d’immobilité lors d’une tâche de navigation spatiale virtuelle. Nous avons trouvé une modulation opposée du Vm entre les CPs de CA1 qui déchargeaient de manière régulière par rapport à celles qui déchargeaient en bouffées de potentiels d’action. Les cellules qui déchargeaient de manière régulière étaient plus dépolarisées et déchargeaient plus pendant le mov.comparé à l’immobilité. Les cellules déchargeant en bouffées de potentiels d’action, préférentiellement inhibées pendant les sharp wave-ripples, étaient hyperpolarisées de façon dépendante à la vitesse pendant le mov.. Cette inhibition dépendante de la vitesse pourrait permettre d’augmenter le rapport signal sur bruit afin de coder l’information spatiale de manière plus efficace pendant le mov.. / Spontaneous locomotion strongly influences the state of the hippocampal network and is critically important for spatial information coding. In neocortex, different attentional or behavioral states during arousal can modify neurons responses to sensorial stimuli and associated task performance. During locomotion, the local field potential of the hippocampus is characterized by theta frequency oscillations (5-12 Hz) and the pyramidal neurons present a specific discharge to the localization of the animal in environments. However, the intracellular determinants of CA1 pyramidal cells activation during locomotion are poorly understood. Here we recorded the membrane potential of CA1 pyramidal cells (PCs) while non-overtrained mice spontaneously alternated between periods of movement and immobility during a virtual spatial navigation task. We found opposite membrane polarization between bursting and regular firing CA1 PCs during movement. Regular firing CA1 PCs were more depolarized and fired at higher frequency during movement compared to immobility while bursting CA1 PCs, preferentially inhibited during sharp wave ripples, were hyperpolarized during movement in a speed dependent manner. This speed-dependent suppression of a subpopulation of CA1 PCs could enhance signal to noise ratio for efficient spatial coding during locomotion.

Tau ist ein Mikrotubuli-assoziiertes Protein, das die Polymerisation von Tubulin fördert und die Mikrotubuli stabilisiert. Folglich wird angenommen, dass Tau essentiell für die neuronale Morphogenese ist, vor allem für die Axonenausbildung und -aufrechterhaltung. Mittels tangentieller Nissl-gefärbter Schnitte von Mäusegehirnen konnte in der vorliegenden Arbeit gezeigt werden, dass Tau-knockout Mäuse die regelhafte thalamokortikale Barthaar-projektion („Barrel“ Konfiguration) entwickeln. Der Einfluss von Tau auf die Entstehung von Dendriten wurde anhand von Golgi-gefärbten Präparaten untersucht. Die Sholl-Analyse der gefärbten CA1-Pyramidenzellen zeigte, dass die Komplexität apikaler Dendriten durch das Fehlen von Tau reduziert wurde, während die Basaldendriten unbeeinflusst blieben. Das Tau-Protein scheint demzufolge unwesentlich für die Entstehung von axonalen Verbindungen im embryonalen Gehirn zu sein, ist aber beteiligt an der Steuerung des dendritischen Verzweigungsmusters. Ferner wurde beobachtet, dass sowohl die adulte Neurogenese, als auch die Mikrotubuli-Stabilität in den Apikal- und Basaldendriten und die Synapsen von dem Fehlen des Tau-Proteins unbeeinflusst blieben. In primären Zellkulturen aus dem Kleinhirn von Tau-knockout und Tau-wildtyp Mäusen konnten zwischen den zwei Genotypen keine signifikanten Unterschiede in der Länge oder im Verzweigungsmuster der Dendriten und der Axone von Körnerzellen beobachtet werden. Die Untersuchung der Effekte einzelner Tau-Isoformen auf die Morphologie von N2A-Zellen zeigte, dass es Unterschiede sowohl zwischen Tau-defizienten und Tau-positiven Zellen, als auch zwischen Zellen mit den verschiedenen Tau-Isoformen gibt. Das Tau-Protein übt demnach in vivo einen wichtigen Einfluss auf die Morphologie der Nervenzellen und besonders der Dendriten aus, welcher in vitro weiter analysiert wurde.:Abkürzungsverzeichnis 1 Einleitung 1 1.1 Das Mikrotubuli-assoziierte Protein Tau 1 1.2 Bedeutung des Tau-Proteins beim Menschen: Tauopathien 3 1.3 Bedeutung des Tau-Proteins beim Menschen: Mikrodeletion des MAPT-Lokus 4 1.4 Ergebnisse aus bisherigen Studien mit Tau-knockout Tieren 6 1.5 Aufgabenstellung 7 2 Material und Methoden 9 2.1 Material 9 2.1.1 Versuchstiere 9 2.1.2 Chemikalien 9 2.1.3 Häufig verwendete Lösungen 10 2.1.4 Geräte 10 2.2 Histologie 11 2.2.1 Fixierung 11 2.2.2 Golgi-Einzelschnittimprägnierung 11 2.2.3 Gefrierschnitte 11 2.2.4 Nissl-Färbung 12 2.2.5 Immunhistochemische Markierungen 13 2.3 Morphometrie 15 2.3.1 Sholl-Analyse 15 2.3.2 Volumenbestimmung 16 2.3.3 Zellzahl (Neurogenese) 17 2.3.4 Synapsenzahl 17 2.4 Proteinbiochemie 19 2.4.1 Proben 19 2.4.2 SDS-Polyacrylamid-Gelelektrophorese 19 2.4.3 Western Blot 20 2.4.4 Immundetektion am Western Blot 21 2.5 Transfektion von Nervenzellen in primärer Zellkultur 25 2.5.1 Primäre Zellkultur 25 2.5.2 Transfektion von primären Zellkulturen 25 2.5.3 Morphometrische Analyse von Körnerzellen des Kleinhirns 26 2.6 Klonierung von Tau-Protein-Isoformen 27 2.6.1 Klonierungsstrategie zur Herstellung eines pIRES-DsRed-Tau Vektors 27 2.6.2 Agarose-Gelelektrophorese 30 2.6.3 Gelextraktion der verschiedenen Tau-Isoform-Sequenzen 30 2.6.4 Herstellung chemisch kompetenter E.coli Zellen 31 2.6.5 Chemische Transformation kompetenter E. coli Zellen 32 2.6.6 Animpfen 32 2.6.7 Plasmid-DNA Purifikation aus 15ml Medium („Miniprep”) 32 2.6.8 Schneiden mit Restriktionsendonukleasen 33 2.6.9 Plasmidpräparation aus 100 ml Medium („Midiprep“) 33 2.6.10 Transfektion von N2A-Zellen mit pIRESRed-Tau 34 2.6.11 Rekonstruktion der transfizierten N2A-Zellen 35 2.7 Statistische Auswertung 36 3 Ergebnisse 37 3.1 Thalamokorticale Projektionen 37 3.2 Komplexität der Dendriten von CA1-Pyramidenzellen 37 3.3 Adulte Neurogenese im Hippocampus 41 3.4 Volumen des Hippocampus 43 3.5 Glia 45 3.6 Synapsen 46 3.7 Stabilität der Mikrotubuli 48 3.8 Mikrotubuli-assoziierte Proteine 50 3.9 Entwicklung in vitro 52 3.9.1 Primärkulturen 52 3.9.2 N2A-Zellkultur 54 4 Diskussion 58 4.1 Diskussion der Methoden 58 4.1.1 Herstellung von Tau-knockout Mäusen 58 4.1.2 Golgi-Einzelschnittimprägnierung und die dreidimensionale Zellrekonstruktion 59 4.1.3 Neurogenese 60 4.1.4 Volumenbestimmung von Hippocampus und Gyrus dentatus 61 4.1.5 Synaptische Marker 61 4.1.6 Western Blot 62 4.1.7 Transfektion von primären Zellkulturen 62 4.1.8 Transfektion von N2A-Zellen mit humanen Tau-Isoformen 63 4.2 Vergleich mit bekannten Daten aus der Forschungsliteratur 64 4.2.1 Axonogenese 64 4.2.2 Dendritogenese 64 4.2.3 Mikrotubuli-assoziierte Proteine und Mikrotubuli-Stabilität 67 4.2.4 Neurogenese 67 4.2.5 Synaptogenese 68 4.2.6 Rolle der einzelnen Isoformen 68 4.3 Bedeutung für die Medizin 71 4.4 Fazit 72 5 Zusammenfassung 73 6 Literaturverzeichnis 76 Posterpräsentationen 88 Danksagung 89 Erklärung über die eigenständige Abfassung der Arbeit 90 Lebenslauf 91

Lokale Pyramidenzellen sind die Hauptneurone des Hippocampus und können durch ihre Position und die Morphologie ihrer Dendriten als CA1 und CA3 Pyramidenzellen identifiziert werden. Die Dendriten der exzitatorischen Pyramidenzellen sind mit postsynaptischen Vorwölbungen, den so genannten Spines, bedeckt, welche in einem spezifischen Verteilungsmuster angeordnet sind. Neurotoxine wie das C3 Toxin von Clostridium botulinum sind funktionelle Substanzen, die die neuronale Morphologie verändern und die neuronale Funktion beeinflussen können. In dieser Studie wurden die morphologischen Veränderungen von intrazellulär mit Biocytin gefüllten CA1 Pyramidenzellen qualitativ und quantitativ analysiert. Die hippocampalen Schnittkulturen, in denen sich bekanntermaßen Pyramidenzellen ähnlich entwickeln wie in vivo, wurden dazu herangezogen, die Effekte der C3bot Toxin-Applikation auf die Verzweigung der Dendriten sowie Anzahl und Dichte der dendritischen Spines zu untersuchen. Drei Gruppen von Zellen wurden verglichen: Erstens Neurone, die in serumhaltigem Medium inkubiert worden waren, zweitens Nervenzellen, die in einem Medium ohne Serum inkubiert worden waren und drittens Zellen, die unter Serumentzug dem C3bot Toxin ausgesetzt worden waren. Die Inkubation dauerte 14 Tage, während die Dauer der Toxinexposition zwischen vier und sechs Stunden betrug. Mit Hilfe eines Computers wurden zweidimensionale Nachbildungen der biocytin-markierten CA1 Pyramidenzellen erstellt, und die Gesamtlänge der Dendriten, die Anzahl der dendritischen Verzweigungspunkte und die Gesamtzahl und Dichte der dendritischen Spines gemessen und statistisch ausgewertet. Signifikante Unterschiede wurden zwischen der mit C3 Toxin behandelten Gruppe und der serumhaltig inkubierten Kontrollgruppe beobachtet. Diese signifikanten morphologischen Veränderungen traten selektiv an den Apikaldendriten der toxinbehandelten CA1 Pyramidenzellen auf. Aus der Behandlung resultierte eine Reduktion der Anzahl apikaler Verzweigungspunkte, der Anzahl der apikalen Spines, der Gesamtzahl (basal und apikal addiert) der Spines sowie der Gesamtspinedichte. Im Gegensatz dazu ergaben sich keine signifikanten Unterschiede zwischen der toxinbehandelten Gruppe und der ohne Serum inkubierten Kontrollgruppe, obwohl der Serumentzug im Vergleich zur serumhaltig inkubierten Kontrollgruppe die Entwicklung der Zellen beeinflusste. Auf Grundlage der beobachteten Veränderungen können wir schließen, dass die Behandlung mit C3 bot einen starken Einfluss selektiv auf die Morphologie der Apikaldendriten ausübt. Der Mechanismus, der dieser selektiven Empfindlichkeit der Apikaldendriten gegenüber dem C3 bot Toxin zugrunde liegt, wird Gegenstand weiterer Untersuchungen sein.

This thesis aims to explore the two dominant functional roles of the hippocampal formation, in the relational encoding of episodic memory and the neural representation of allocentric space, using a combination of pharmaceutical manipulations and single-unit recording techniques in rodents. The first part of this thesis focuses on episodic-like memory, defined by the original episodic memory triad: ‘what-where-when’ (Tulving 1972), which enables the behavioural aspects of episodic memory to be tested in non-human animals. Permanent neurotoxic lesions of the hippocampus and it’s subregions were induced to assess their role in a putative episodic-like memory task developed by Eacott and Norman (2004). In view of the difficulties encountered in successfully demonstrating the temporal component of episodic-like memory in rats, this task tested integrated memory for ‘what-where-which’, where the temporal component (when) was replaced with another event specifier: context (on ‘which’ occasion). Disruption of the hippocampal circuitry led to a specific impairment in the integration of all three event components, whereas the associative recognition of any combination of these features in isolation was left intact. These results confirm the hippocampal dependence of this episodic-like memory task and further reveals the necessity of both CA3 and CA1, hypothetically due to the underlying autoassociative role of CA3 with CA1 functioning as the vital output pathway for this associated information and/or as a mismatch detector. There has been much debate over the inclusion of the temporal component and sceptics may argue that any such interpretations of task-dependence on episodic-like memory processing are invalid considering the requirement for temporal processing is absent. Due to the proposal that a temporal framework necessarily provides the foundation on which episodic memories are built, the second chapter focuses on the development of a suitable protocol in which integrated memory for the original ‘what-where-when’ episodic memory triad can be reliably tested. The other main function attributed to the hippocampus was brought to light by the fascinating revelation that it’s neurons selectively fire in different regions of an environment, termed ‘place cells’ (O’Keefe and Dostrovsky 1971). From the numerous publications resulting from this discovery it has emerged that place cells not only respond to the spatial features of the environment but are also sensitive to a multitude of non-spatial features. These characteristics support the logical assumption that the primary firing patterns of the hippocampus should underlie it’s main purported roles, leading to speculations that they reflect episodic memory processes. The second part of this thesis aims to examine the relationship between hippocampal cells and behaviour by extending the work of Ainge et al. (2007a), in which a subset of hippocampal place cells were found to encode both current and intended destination in a double Y-maze ‘win-stay’ task. The development of these ‘goal-sensitive’ cells were initially investigated during the learning phase of this task. An exciting pattern of results showed a strong positive correlation between the emergence of goal-sensitive firing and behavioural performance on the task, tempting speculations that these firing patterns may underlie spatial learning and future planning, necessary to support performance. To ensure these firing patterns were not a mere reflection of greater experience on the maze, a second study was conducted in which the task demands changed over set periods of days. A significant increase in the proportion of cells demonstrating goal-sensitive firing was revealed when the protocol shifted to incorporate the spatial memory demands of the ‘win-stay’ task, with all other parameters of the protocol remaining constant. These results support the theory that goal-sensitive firing patterns are specifically related to the learning and memory demands of the spatial task, not a result of increased exploration of the maze. The last of this series of studies assessed hippocampal-dependence of this task and revealed that bilateral hippocampal lesions induced an impairment in spatial ‘win-stay’ performance. Collectively, these experiments demonstrate that goal-sensitive firing of hippocampal cells emerge in line with behavioural performance in a hippocampal-dependent task and the emergence of these firing patterns are specific to the learning and memory demands of a spatial ‘win-stay’ protocol. The functional role of the hippocampus in allocentric spatial processing may thus underpin it’s function in episodic memory and potentially in the imagining and planning of future events, whereby the hippocampus provides a ‘space’ in which retrieved information can be integrated in a coherent context to support the fluent and flexible use of information. This hippocampal function would necessarily require visual information to be accessed, concerning the arrangement of landmarks and cues within the environment, in association with information regarding internal orientation and direction and this leads to the question assessed in the final part of this thesis of where this integration occurs. Based on anatomical evidence and the current literature, the postsubiculum, an input structure to the hippocampus, emerged as a potential site for the convergence of sensory cues into the internally generated head direction cell and place cell networks to enable hippocampal-dependent spatial processing. Thus, the effects of temporary pharmacological blockade of AMPARs and NMDARs in the postsubiculum were assessed on the encoding of spatial memory in an object recognition paradigm. The impairment revealed in the ability to recognise novel object-place configurations demonstrates a key role for NMDAR-dependent plasticity within the postsubiculum itself in the formation of allocentric spatial memory. In summary, the experimental results reported in this thesis further elucidate the critical role the hippocampal formation plays in spatial and episodic memory by combining evidence from cellular physiology and neuroanatomy to the behaving animal and extends these findings to discuss a more general role for the hippocampus in imagining both past and future events, in order to successfully navigate, learn and enable past experience to influence our intended future plans and decisions.

Fragile-X syndrome (FXS) is the most common form of inherited intellectual impairment. Caused by the transcriptional repression of the Fmr1 gene on the X chromosome, FXS results in the loss of the Fragile-X Mental Retardation Protein (FMRP). Human female patients with FXS are heterozygous for the Fmr1 mutation whereas males are hemizygous. FXS has been studied far less in females than in males due to a generally less severe clinical phenotype. Previous research has implicated the metabotropic glutamate receptor (mGluR) in synaptic plasticity alterations in the cornu ammonis area 1 (CA1) region of the juvenile male Fmr1 knock-out (KO) hippocampus. In contrast, our investigations into the young adult dentate gyrus (DG) subfield of the hippocampus have revealed N-methyl-D-aspartate receptor (NMDAR)-associated impairments in synaptic plasticity. The current study sought to extend these investigations to the young adult female Fmr1 heterozygous (Het) and Fmr1 KO mouse as well as investigate NMDAR- and mGluR-mediated long-term depression (LTD) in the DG and CA1 of the young adult male Fmr1 KO mouse. Input-output curves and paired pulse measures of short-term plasticity were also evaluated in all genotypes. Field electrophysiology revealed a significant impairment in long-term potentiation (LTP) and LTD in male Fmr1 KO and female Fmr1 Het mice that was associated with NMDAR alteration. A more robust synaptic protocol was not able to rescue LTP in the male Fmr1 KO DG. Paired-pulse low-frequency stimulation and (RS)-3,5-dihydroxyphenylglycine (DHPG)-induced mGluR-LTD was intact in all genotypes and brain regions examined. Although further investigation will be required to expand our understanding of FXS and to fully elucidate the mechanisms behind intact synaptic plasticity in the female Fmr1 KO mouse, our results suggest that NMDARs may be poised as important contributors to hippocampal pathophysiology in FXS. / Graduate

Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2017-03-09T22:59:49Z No. of bitstreams: 1 ArthurSergioCavalcantiDeFranca_TESE.pdf: 14502072 bytes, checksum: 07eea320317f9aa96a67cda6ed40b06a (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2017-03-13T23:54:25Z (GMT) No. of bitstreams: 1 ArthurSergioCavalcantiDeFranca_TESE.pdf: 14502072 bytes, checksum: 07eea320317f9aa96a67cda6ed40b06a (MD5) / Made available in DSpace on 2017-03-13T23:54:25Z (GMT). No. of bitstreams: 1 ArthurSergioCavalcantiDeFranca_TESE.pdf: 14502072 bytes, checksum: 07eea320317f9aa96a67cda6ed40b06a (MD5) Previous issue date: 2016-10-04 / O hipocampo ? relacionado com a forma??o de mem?rias explicitas e com a capacidade de reconhecer novos objetos. No presente trabalho visamos contribuir para uma maior compreens?o do papel da regi?o CA1 do hipocampo nestes processos. Atrav?s da aplica??o de t?cnicas de eletrofisiologia, comportamento animal, psicofarmacologia e optogen?tica em camundongos transg?nicos e selvagens, encontramos que c?lulas OLM?2 do CA1 atuam na codifica??o da representa??o de objetos em uma tarefa de reconhecimento de objetos, e tamb?m influenciam a codifica??o de mem?rias aversivas em uma tarefa associativa de medo ao contexto. Al?m disso, descrevemos uma nova atividade oscilat?ria no potencial de campo local do CA1 na frequ?ncia beta 2 (23-30 Hz), que ? caracteristicamente transit?ria e ligada ? detec??o de novos objetos durante uma tarefa de reconhecimento de objetos. Estes resultados sugerem potenciais mecanismos celulares e de rede neuronal na regi?o CA1 subjacentes ao seu papel na forma??o de mem?rias e na detec??o de novidade. / The hippocampus is associated to novelty detection and formation of explicit memories. The present work aims at better understanding the role of the CA1 region of the hippocampus in these processes. By employing electrophysiology, animal behavior, psychopharmacology and optogenetic techniques in transgenic and wild-type mice, we found that CA1 OLM?2 cells influence the formation of new object representations in an object recognition task, as well as the encoding of aversive memories in a contextual fear memory task. Furthermore, we characterized a new oscillatory activity in the local field potential of CA1 at beta 2 frequency (23-30 Hz), which was typically transient and linked to the amount of novelty in an object recognition task. These results suggest potential cellular and network mechanisms that underlie the role of CA1 in memory formation and novelty detection.

Submitted by Automa??o e Estat?stica (sst@bczm.ufrn.br) on 2017-07-17T13:25:04Z No. of bitstreams: 1 RobsonSchefferTeixeira_TESE.pdf: 28541390 bytes, checksum: 963c2c435bf67ce018fa618617a1a0a6 (MD5) / Approved for entry into archive by Arlan Eloi Leite Silva (eloihistoriador@yahoo.com.br) on 2017-07-19T14:26:40Z (GMT) No. of bitstreams: 1 RobsonSchefferTeixeira_TESE.pdf: 28541390 bytes, checksum: 963c2c435bf67ce018fa618617a1a0a6 (MD5) / Made available in DSpace on 2017-07-19T14:26:40Z (GMT). No. of bitstreams: 1 RobsonSchefferTeixeira_TESE.pdf: 28541390 bytes, checksum: 963c2c435bf67ce018fa618617a1a0a6 (MD5) Previous issue date: 2016-04-07 / An?lises espectrais de registros eletrofisiol?gicos extracelulares t?m revelado que a atividade el?trica produzida pelo c?rebro ? comumente organizada em padr?es r?tmicos, conhecidos como oscila??es neuronais. Mais recentemente, descobriu-se que as oscila??es neuronais de frequ?ncias distintas n?o s?o independentes, mas podem interagir entre si. Ao longo das ?ltimas duas d?cadas, diversas ferramentas de an?lises foram desenvolvidas, amadurecidas e incorporadas de outras ?reas para se estudar os chamados acoplamentos entre frequ?ncias de oscila??es neuronais observadas nestes registros. Oscila??es neuronais s?o ditas acopladas se houver uma rela??o de depend?ncia entre suas caracter?sticas, como fase, amplitude ou frequ?ncia instant?neas. Dentre elas, o acoplamento fase-amplitude ? caracterizado por um aumento da amplitude instant?nea de uma banda de frequ?ncia condicionado a uma fase instant?nea de uma oscila??o de outra banda, enquanto que o acoplamento fase-fase do tipo n:m ? caracterizado pela rela??o fixa entre m ciclos de uma frequ?ncia em nciclos de outra. O hipocampo ? uma regi?o cerebral envolvida na forma??o de mem?rias e navega??o espacial. Assim como em outras estruturas, as redes neuronais do hipocampo produzem diversos padr?es oscilat?rios, que variam de acordo com os estados do ciclo sono-vig?lia. Entre estes padr?es, classicamente destacam-se os ritmos teta (4-12 Hz) e gama (30-100 Hz), que caracterizam estados comportamentais de locomo??o e sono REM. No entanto, o estudo dos padr?es de acoplamento oscilat?rio no hipocampo tem revelado subtipos oscilat?rios distintos dentro da defini??o tradicional da banda gama. Mais ainda, trabalhos recentes t?m mostrado a exist?ncia de oscila??es acopladas ao ritmo teta em frequ?ncias mais altas (>100 Hz), embora haja uma diverg?ncia na literatura atual sobre at? aonde estas oscila??es de altas frequ?ncias representariam atividade oscilat?ria genu?na de redes neuronais ou se seriam derivadas de efeitos esp?rios oriundos de contamina??es por resqu?cios de potencias de a??o registrados extracelularmente. A presente tese de doutorado visa contribuir para o maior entendimento dos padr?es oscilat?rios produzidos por redes neuronais do hipocampo, com particular foco nas rela??es de acoplamento entre oscila??es de diferentes frequ?ncias. Atrav?s de dados pr?prios e compartilhados de terceiros de animais implantados cronicamente com matrizes de m?ltiplos eletrodos, obtivemos registros da atividade el?trica da regi?o CA1 de ratos durante a explora??o de ambientes familiares e per?odos de sono. Investigamos a exist?ncia conjunta de distintos padr?es oscilat?rios do hipocampo em diferentes frequ?ncias atrav?s de marcadores eletrofisiol?gicos, anat?micos e comportamentais de cada oscila??o neuronal que, quando combinados, levaram a um perfil ?nico para cada banda de frequ?ncia. Nossos resultados mostram a exist?ncia de m?ltiplas bandas de frequ?ncia moduladas pelo ritmo teta hipocampal. As modula??es s?o dotadas de diversos mecanismos separat?rios, provavelmente de forma a minimizar interfer?ncias. Demonstramos ainda que padr?es oscilat?rios esp?rios e genu?nos podem co-existir numa mesma faixa de frequ?ncia, e que, ao contr?rio de trabalhos recentes, n?o h? evid?ncia para acoplamentos do tipo fase-fase n:m no hipocampo. A capacidade de uma oscila??o neural interagir com outras oscila??es, aparentemente independentes, levanta questionamentos naturais sobre sua signific?ncia biol?gica, que, apesar de diversos avan?os na ?rea, ainda permanece um mist?rio na sua ess?ncia. / Spectral analysis of extracellular electrophysiological recordings revealed that the brain electrical activity is often organized in rhythmic patterns, known as neuronal oscillations. Recently, it was discovered that oscillations of distinct frequencies are not independent, but can interact to each other. In the last two decades, several analysis tools were developed or incorporated from other fields to study cross-frequency coupling between neural oscillations. Neural oscillations are said to be coupled if there is a dependency between their features, such as phase, amplitude or frequency. Among them, phase ? amplitude coupling is characterized by an increase in the instantaneous amplitude of one frequency band conditioned to the instantaneous phase of another frequency band, whereas n:m phase ? phase coupling is characterized by a fixed relation between m cycles of one frequency to n cycles of another one. The hippocampus is a brain region involved in memory formation and spatial navigation. As in other brain structures, hippocampal neural networks generate several oscillatory patterns, which vary according to the stage of the sleep-waking cycle. Among these patterns, theta (4 ? 12 Hz) and gamma (30 ? 100 Hz) oscillations are prominent during active waking and REM sleep. However, the study of coupling patterns in the hippocampus has revealed distinct sub-types of oscillatory activity inside the traditional gamma band. Moreover, recent studies have shown the existence of even faster oscillations coupled to theta in the hippocampus (> 100 Hz), although there is a current divergence in the literature about whether they represent genuine network activity or spurious by-products from incomplete filtering of extracellular spikes. This thesis investigates oscillatory patterns generated by hippocampal neural networks, focusing in the coupling relation among oscillations of different frequencies. Using our own data and shared third-party ones of chronically implanted animals with multisite electrodes, we recorded electrical activity in the CA1 region of rats while exploring a familiar environment and during sleep stages. We investigated the existence of simultaneous but distinct oscillatory patterns in the hippocampus separated by electrophysiological, anatomic and behavioral markers, which, once taken together, can lead to a unique profile for each frequency band. Our results point to the existence of several frequency bands coupled to the hippocampal theta rhythm. All modulations are found to be separated by mechanisms that can potentially avoid interferences. We also demonstrate that a spurious oscillatory patterns can emerge and co-exist in the same frequency band of genuine oscillations and, contrary to recent work, we show that there is lack of evidence for n:m phase ? phase coupling in the hippocampus. The capacity of neural oscillations to interact with one another raises questions about the biological significance of such phenomenon; despite recent progress in the field, however, its essence remains a mystery.

No description available.

Neuron types (e.g., pyramidal cells) within one area of the brain are often considered homogeneous, despite variability in their biophysical properties. Here we review literature demonstrating variability in the electrical activity of CA1 hippocampal pyramidal cells (PCs), including responses to somatic current injection, synaptic stimulation, and spontaneous network-related activity. In addition, we describe how responses of CA1 PCs vary with development, experience, and aging, and some of the underlying ionic currents responsible. Finally, we suggest directions that may be the most impactful in expanding this knowledge, including the use of text and data mining to systematically study cellular heterogeneity in more depth; dynamical systems theory to understand and potentially classify neuron firing patterns; and mathematical modeling to study the interaction between cellular properties and network output. Our goals are to provide a synthesis of the literature for experimentalists studying CA1 PCs, to give theorists an idea of the rich diversity of behaviors models may need to reproduce to accurately represent these cells, and to provide suggestions for future research.

Single-cell real-time quantitative RT-PCR was used to characterize the mRNA expression of rat neuronal nicotinic acetylcholine receptor (nAChR) subunits α3 and β2 in CA1 hippocampus stratum radiatum and stratum oriens interneurons. α3β2 co-expression was detected in 43% of interneurons analyzed. The nAChR subtype α3β2 was transiently expressed in cells derived from the human embryonic kidney cell line 293 at mRNA levels found in the CA1. The functional properties of α3β2 in HEK-293 cells were characterized by whole-cell patch clamping using acetylcholine (ACh) as an agonist. The kinetics of α3β2 channels were further analyzed by altering the level of α3 DNA transfected into HEK-293 cells. Varying the α3 concentration by more than 100,000 fold did not significantly alter the majority of the kinetics; the 10%-90% rise-time was the main characteristic found to be significantly different. A decrease in α3 concentration illustrated a significant increase in rise time. This and future studies will further our understanding of the extensive role neuronal nAChRs play in modulating hippocampal activity and consequently influencing cognition and memory.