The hippocampus is a complex brain structure that has been studied extensively and is subject to abnormal structural change in various neuropsychiatric disorders. The highest definition in vivo method of visualizing the anatomy of this structure is structural Magnetic Resonance Imaging (MRI). Gross structure can be assessed by the naked eye inspection of MRI scans but measurement is required to compare scans from individuals within normal ranges, and to assess change over time in individuals. The gold standard of such measurement is manual tracing of the boundaries of the hippocampus on scans. This is known as a Region Of Interest (ROI) approach. ROI is laborious and there are difficulties with test-retest and inter-rater reliability. These difficulties are primarily due to uncertainty in designation of the hippocampus boundary. An improved, less labour intensive and more reliable method is clearly desirable. This thesis describes a fully automated hybrid methodology that is able to first locate and then extract hippocampal volumes from 3D 1.5T MRI T1 brain scans automatically. The hybrid algorithm uses brain atlas mappings and fuzzy inference to locate hippocampal areas and create initial hippocampal boundaries. This initial location is used to seed a deformable manifold algorithm. Rule based deformations are then applied to refine the estimate of the hippocampus locations. Finally, the hippocampus boundaries are corrected through an inference process that assures adherence to an expected hippocampus volume. The ICC values of this methodology when compared to the manual segmentation of the same hippocampi result in a 0.73 for the left and 0.81 for the right hippocampi. These values both fall within the range of reliability testing according to the manual ‘gold standard’ technique. Thus, this thesis describes the development and validation of a genuinely automated approach to hippocampal volume extraction of potential utility in studies of a range of neuropsychiatric disorders and could eventually find clinical applications.

Το ενδογενές σύστημα των κανναβινοειδών αποτελεί ένα πολύπλοκο ενδογενές σύστημα μεταγωγής σήματος το οποίο επηρεάζει ένα σημαντικό αριθμό φυσιολογικών διεργασιών και μεταβολικών μονοπατιών του οργανισμού (Cota and Woods, 2005). Απαρτίζεται από τους διαμεμβρανικούς υποδοχείς των κανναβινοειδών (CBR), τα ενδοκανναβινοειδή και τις πρωτεΐνες που είναι υπεύθυνες για την βιοσύνθεση και την αποικοδόμηση των δεύτερων. (Petrocellis et al., 2004). Τα κανναβινοειδή παρουσιάζουν ποικιλία επιπτώσεων, όπως δυσλειτουργία στη μάθηση και μνήμη, διαφοροποίηση των συναισθηματικών καταστάσεων, μείωση κινητικού ελέγχου και αναλγησία. Αναστέλλουν τη συναπτική διαβίβαση σε διάφορες περιοχές του εγκεφάλου όπως ο ιππόκαμπος, ο επικλινής πυρήνας και ο προμετωπιαίος φλοιός κυρίως μέσω προσυναπτικών μηχανισμών. Το γλουταμινικό οξύ είναι ο κύριος διεγερτικός νευροδιαβιβαστής στο Κεντρικό Νευρικό Σύστημα των θηλαστικών. Οι γλουταμινεργικοί νευρώνες διανέμονται ευρέως στο κεντρικό νευρικό σύστημα και παίζουν ρόλο σε πολλές βιολογικές διεργασίες, συμπεριλαμβανομένης της κωδικοποίησης των πληροφοριών, το σχηματισμό και την ανάκτηση των μνημών, τη χωρική αναγνώριση και τη διατήρηση της προσοχής (McEntee και Crook, 1993). Λόγω του ρόλου του στη συναπτική πλαστικότητα, το γλουταμινικό εμπλέκεται σε γνωσιακές λειτουργίες του εγκεφάλου, όπως η μάθηση και η μνήμη (McEntee και Crook, 1993). Το γλουταμινικό δρα στα μετασυναπτικά κύτταρα σε τρεις οικογένειες ιοντοτροπικών υποδοχέων τους NMDA, τους AMPA και τους καϊνικούς υποδοχείς, ενώ υπάρχουν, επίσης, και γλουταμινικοί μεταβολοτροπικοί υποδοχείς που συνδέονται με G πρωτεΐνες. Η αμυγδαλή εκτελεί ένα πρωταρχικό ρόλο στην επεξεργασία των συναισθηματικών αντιδράσεων και τη μνήμη, ενώ ο ιππόκαμπος έχει σημαντικό ρόλο στην εδραίωση των πληροφοριών από βραχυχρόνια μνήμη σε μακροχρόνια μνήμη και στη χωρική πλοήγηση. Οι δύο αυτές περιοχές είναι πλούσιες σε CB1 και NMDA υποδοχείς, ενώ τα τελευταία χρόνια, πληθώρα ερευνών υποδεικνύει ότι η έκθεση πειραματοζώων σε κανναβινοειδή επιφέρει σημαντικές αλλαγές σ’ αυτές τις δύο περιοχές. Στόχος της ερευνητικής εργασίας ήταν η μελέτη της επίδρασης των κανναβινοειδών στο γλουταμινεργικό σύστημα, στην αμυγδαλή και τον ιππόκαμπο επίμυων. Η μελέτη επικεντώθηκε στους NMDA υποδοχείς του γλουταμινικού οξέος με τη χρήση των μεθόδων της in situ υβριδοποίησης για το mRNA των NR1, NR2A και NR2B υπομονάδων του υποδοχέα και της ποσοτικής αυτοραδιογραφίας υποδοχέων, σε επίμυς που τους χορηγήθηκε WIN55212-2, ένας συνθετικός αγωνιστής του CB1 κανναβινοειδικού υποδοχέα, ο οποίος εμφανίζει παρόμοια δράση με την Δ9-τετραϋδροκανναβινόλη (THC), το φυσικό συστατικό του φυτού Cannabis sativa. Μελετήθηκαν τέσσερις ομάδες επίμυων όπου σε δύο από αυτές χορηγήθηκε WIN και στις υπόλοιπες δύο χορηγήθηκε ο διαλύτης του WIN, Vehicle. Στις δύο ομάδες που χορηγήθηκε WIN55212- 2 1mg/kg για 20 ημέρες, οι επίμυς της μίας ομάδας (ομάδα WIN) θανατώθηκαν 2 ώρες μετά από την τελευταία δόση ενώ οι επίμυς της άλλης ομάδας (ομάδα WIN+WITHD) θανατώθηκαν 7 ημέρες μετά την τελευταία δόση. Κατά το διάστημα αυτό δεν έγινε καμία χορήγηση ώστε να μελετηθεί αν οι επιπτώσεις της χρόνιας χορήγησης του WIN ήταν μόνιμες. Τα αποτελέσματά μας έδειξαν ότι μετά τη χρόνια χορήγηση του WIN τα επίπεδα έκφρασης των υπομονάδων του NMDA υποδοχέα μειώθηκαν στο ιππόκαμπο και την αμυγδαλή των επίμυων, ενώ μετά την διακοπή της χορήγησης του WIN, τα επίπεδα έκφρασης επανέχρονται. Είναι πιθανό ότι οι αλλαγές στην έκφραση και λειτουργία των υποδοχέων του γλουταμινικού που παρουσιάζονται ως προσαρμοστικές αλλαγές στο νευρικό σύστημα, να είναι ένας κοινός μηχανισμός με τον οποίο τα εθιστικά ναρκωτικά επηρεάζουν την νευρική λειτουργία. / The endocannabinoid system is a complicated endogenous signaling system that affects a variety of physiological processes and metabolic routes in human body (Cota and Woods, 2005). The endocannabinoid system includes the transmembrane cannabinoid receptors (CBR), the endocannabinoids and the enzymes that synthesize and degrade the endocannabinoids (Petrocellis et al., 2004). Cannabinoids have a variety of effects, such as impairment in learning and memory, modulation of emotional states, reduced motor control and analgesia. Cannabinoids inhibit synaptic transmission in several brain regions such as the hippocampus, nucleus accumbens and the prefrontal cortex mainly via presynaptic mechanisms. Glutamate is the most abundant excitatory neurotransmitter in the mammalian central nervous system. The glutamatergic neurons are widely distributed in the central nervous system and play a role in many biological processes, including the coding of information, the formation and recovery of memories, the spatial recognition and maintaining the attention (McEntee και Crook, 1993). Because of its role in synaptic plasticity, glutamate is involved in cognitive functions like learning and memory in the brain. (McEntee και Crook, 1993). Glutamate activates three families of ionotropic receptors in postsynaptic cells, those are NMDA, AMPA and kainate receptors, while there are also metabotropic G proteins coupled glutamate receptors. Amygdala has a primary role in the processing of emotional reactions and memory, whereas hippocampus has an important role in the consolidation of informations from short term memory into long term memory and spatial navigation. These two brain regions contain a large number of CB1 and NMDA receptors, while recently, many studies suggest that animals treated with cannabinoids display significant changes in these two areas. The aim of this research was to study the changes in glutamatergic system in the amygdala and hippocampus of rats treated with cannabinoids. We focused on NMDA glutamate receptors, using in situ hybridization for studying the expression of NR1, NR2A and NR2B subunits and quantitative receptor autoradiography, in rats treated with WIN55212-2, a synthetic agonist of the CB1 cannabinoid receptor, which shows similar effects with delta-9-tetrahydrocannabinol (THC), a natural component of the plant Cannabis sativa. Adult rats were injected with WIN55212-2 (1mg/kg) and Vehicle. Animals received repeated administrations of WIN55212-2 1mg/kg once a day for 20 days. Animals in group WIN were sacrificed 2 hours after the last administration whereas in group WIN + WITHD were sacrificed 7 days after the last administration. During this time there was no administration of WIN so we could study whether the effects of chronic exposure were permanent. Our results demonstrate that chronic exposure to WIN55212-2 produced significant decreases in the expression of NMDA receptor's subunits in hippocampus and amygdala. These changes were reversed one week after abstinence. These adaptive synaptic changes may share common mechanisms with addictive drugs in modifying neural circuitry.

published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

published_or_final_version / Biological Sciences / Doctoral / Doctor of Philosophy

This thesis explores the role of path integration on the firing of hippocampal place cells and medial entorhinal grid cells. Grid cells fire at equidistant locations in an environment, indicating that they keep track of the distance and direction an animal has moved in an environment. One class of model of path integration uses a continuous attractor network to update position information. The first part of this thesis showed that such a network can generate a "look-ahead" of neural activity that sweeps through the positions just visited and about to be visited, on the short time scale that is observed<italic>in vivo</italic>. Adding intrinsic currents to the neurons in the network model allowed this look-ahead to recur every theta cycle, and generate grid fields of a size comparable to data. Grid cells are a major input the hippocampus, and are hypothesized to be the source of the place specificity of place cells. When an animal explores an open environment, place cells are active in a particular location regardless of the direction in which the animal travels through it. While performing a specific task, such as visiting specific locations in the environment in sequence, however, most place cells are active only in one direction. The second part of this thesis studied the development of this directionality. It was determined that upon the initial appearance of place fields in a novel environment, place cells fired in all directions, supporting the hypothesis that the path integration is the primary determinant of place specificity. The directionality of place fields developed gradually, possibly as a result of learning. Ideas about how this directionality could develop are explored.

The present study examined social status and adult neurogenesis in the naked mole rat. These animals live in large colonies with a strict reproductive dominance hierarchy; one female and 1-3 males breed, while other members are subordinate and reproductively suppressed. We examined whether social status affects doublecortin (DCX; a marker for immature neurons) immunoreactivity in the dentate gyrus, piriform cortex (PCx), and basolateral amygdala (BLA) by comparing breeders to subordinates. We also examined subordinates removed from their colony and paired with opposite- or same-sex conspecifics for 6 months. Breeders had reduced DCX immunoreactivity in all areas, with BLA effects confined to females. Effects of housing condition were region-specific, with higher PCx DCX immunoreactivity observed in opposite- than same-sex paired subordinates regardless of gonadal status. The opposite pattern was observed in the BLA. Future work will clarify whether findings are attributable to status differences in stress, behavioural plasticity, or life stage.

CA1 is the main source of afferents from the hippocampus, but the function of CA1 and its perforant path (PP) input remains unclear. In this thesis, Marr’s model of the hippocampus is used to investigate previously hypothesized functions, and also to investigate some of Marr’s unexplored theoretical ideas. The last part of the thesis explains the excitatory responses to PP activity in vivo, despite inhibitory responses in vitro. Quantitative support for the idea of CA1 as a relay of information from CA3 to the neocortex and subiculum is provided by constraining Marr’s model to experimental data. Using the same approach, the much smaller capacity of the PP input by comparison implies it is not a one-shot learning network. In turn, it is argued that the entorhinal-CA1 connections cannot operate as a short-term memory network through reverberating activity. The PP input to CA1 has been hypothesized to control the activity of CA1 pyramidal cells. Marr suggested an algorithm for self-organising the output activity during pattern storage. Analytic calculations show a greater capacity for self-organised patterns than random patterns for low connectivities and high loads, confirmed in simulations over a broader parameter range. This superior performance is maintained in the absence of complex thresholding mechanisms, normally required to maintain performance levels in the sparsely connected networks. These results provide computational motivation for CA3 to establish patterns of CA1 activity without involvement from the PP input. The recent report of CA1 place cell activity with CA3 lesioned (Brun et al., 2002. Science, 296(5576):2243-6) is investigated using an integrate-and-fire neuron model of the entorhinal-CA1 network. CA1 place field activity is learnt, despite a completely inhibitory response to the stimulation of entorhinal afferents. In the model, this is achieved using N-methyl-D-asparate receptors to mediate a significant proportion of the excitatory response. Place field learning occurs over a broad parameter space. It is proposed that differences between similar contexts are slowly learnt in the PP and as a result are amplified in CA1. This would provide improved spatial memory in similar but different contexts.

<p>Humans and animals have remarkable capabilities in keeping time and using time as a guide to orient their learning and decision making. Psychophysical models of timing and time perception have been proposed for decades and have received behavioral, anatomical and pharmacological data support. However, despite numerous studies that aimed at delineating the neural underpinnings of interval timing, a complete picture of the neurobiological network of timing in the seconds-to-minutes range remains elusive. Based on classical interval timing protocols and proposing a Timing, Immersive Memory and Emotional Regulation (TIMER) test battery, the author investigates the contributions of the dorsal and ventral hippocampus as well as the dorsolateral and the dorsomedial striatum to interval timing by comparing timing performances in mice after they received cytotoxic lesions in the corresponding brain regions. On the other hand, a timing-based theoretical framework for the emergence of conscious experience that is closely related to the function of the claustrum is proposed so as to serve both biological guidance and the research and evolution of “strong” artificial intelligence. Finally, a new “Double Saturation Model of Interval Timing” that integrates the direct- and indirect- pathways of striatum is proposed to explain the set of empirical findings.</p> / Dissertation

Stress is known to impair spatial navigation in rat models of declarative memory, and declarative memory in humans, but the effects on spatial navigation in humans are unclear. At least four models have been proposed to account for the cognitive effects of stress, based on the two different physiological stress response systems (the sympathetic-adrenal-medullary (SAM) and the hypothalamic-pituitary-adrenal (HPA) systems) and the effects of these responses on the hippocampus and (sometimes) other subcortical structures. In this dissertation, I examined the effects of an acute (experimental) stressor on human spatial navigation in three variations of virtual Morris water mazes designed to dissociate between hippocampus-dependent (allocentric) and hippocampus-independent (egocentric) forms of navigation. Results were considered in the light of all 4 models. Experiment 1 used a dual-strategy Morris water maze to test whether acute stress influences navigational strategy selection and whether this effect is mediated by the activation of the HPA or the SAM system. Surprisingly, stress increased hippocampus-based strategy selection, and did so in the presence of SAM but not HPA activation. Experiment 2 used new dual-strategy and place mazes to examine the effects of acute stress on both strategy selection and allocentric navigational performance. It also attempted to contrast the effects of stress at a short delay, which would favour mediation by the SAM system, and a longer, 30 minute delay (from stressor onset), which would favour mediation by the HPA system. Contrary to expectations, results revealed no effect of stress when tested immediately and sex-dependent impairments of performance (in females) and allocentric strategy selection (in males) at the delay. Experiment 3 used the same mazes as Experiment 2, plus a new cue maze to examine the effects of acute stress on strategy selection and both allocentric and egocentric navigational performance after a 30 minute delay. Results confirmed that stress reduces allocentric strategy selection and impairs allocentric performance, but also has sex-dependent effects on egocentric performance: in females, stress enhanced navigation (as expected) but in males, stress impaired it. None of the 4 models provided a good explanation for these results, suggesting that current accounts of the cognitive effects of stress may be inadequate. / Graduate

Functional neuroimaging studies of autobiographical memory have grown dramatically in recent years. These studies are important because they can investigate the neural correlates of processes that are difficult to study using laboratory stimuli, including: (i) complex constructive processes, (ii) recollective qualities of emotion and vividness, and (iii) remote memory retrieval. Constructing autobiographical memories involves search, monitoring and self-referential processes that are associated with activity in separable prefrontal regions. The contributions of emotion and vividness have been linked to the amygdala and visual cortex respectively. Finally, there is evidence that recent and remote autobiographical memories might activate the hippocampus equally, which has implications for memory-consolidation theories. The rapid development of innovative methods for eliciting personal memories in the scanner provides the opportunity to delve into the functional neuroanatomy of our personal past. / Dissertation