Hippocampus is one of the main components of the limbic system. It plays an important role in the process of memory formation and recent literature shows that, like in other peripheral organs and brain structures, there is a circadian clock present in the hippocampus. The aim of the thesis was to study the hippocampal circadian clock under various conditions. The first part of the thesis examines the influence of glucocorticoid hormones on the clock gene expression in the hippocampus. Glucocorticoids are mammalian steroid hormones secreted from the adrenal glands that affect many processes in the organism. The glucocorticoid secretion is under the control of the circadian system, causing their levels to exhibit a pronounced diurnal rhythm. The hippocampus provides feedback to the hypothalamo-pituitary-adrenal (HPA) axis and is thus involved in the regulation of glucocorticoid hormone secretion. Hippocampal neurons also express glucocorticoid hormone receptors (GR). This thesis explores the effect of the absence of glucocorticoid hormones on the clock gene expression in the hippocampus. The absence of glucocorticoids was due to adrenalectomy and its effect was further compared with the effect of endogenous glucocorticoid replacement by daily injections of the GR agonist dexamethasone, emulating...

La théorie de la formation de la mémoire "en deux étapes" postule que la consolidation mnésique repose sur un dialogue pendant le sommeil entre l'hippocampe, où les traces mnésiques sont initialement formées, et le néocortex (notamment le cortex préfrontal médian, CPm), où elles seraient stockées à long terme. En particulier, la coordination entre oscillations au sein du réseau hippocampo-cortical pendant le sommeil apparaît comme un mécanisme plausible pour la consolidation. Nous avons augmenté la coordination temporelle entre ces deux structures pendant le sommeil suivant un apprentissage d'une tâche de mémoire spatiale limité en temps n'induisant pas de consolidation. Les rats ayant suivi le protocole permettant l'augmentation des intéractions entre ondulations hippocampiques, ondes delta corticales et fuseaux thalamo-corticaux par le biais d'impulsions électriques intra-corticales ont fait preuve d'un haut niveau de performance de rappel 24 h plus tard, contrairement à des rats de contrôle qui ne faisaient pas mieux que le hasard. Ainsi, cette augmentation des intéractions hippocampo-corticales pendant le sommeil a-t-elle stabilisé des traces mnésiques qui autrement n'auraient pas subsisté entre l'entraînement et le test. De plus, nous avons observé une réorganisation des réseaux neuronaux du CPm pendant le sommeil, ainsi qu'une réponse accrue du CPm à la tâche le lendemain. Ces travaux constituent une mise en évidence directe du rôle causal du dialogue hippocampo-cortical pendant le sommeil dans la consolidation mnésique, dont les mécanismes sous-jacents impliquent une coordination temporelle précise entre ondulations, ondes delta et fuseaux (Maingret et al., 2016). / The `two-stage' theory of memory posits that memory consolidation involves a dialogue during sleep between the hippocampus, where traces are initially formed, and the neocortex (notably the prefrontal cortex), where they are stored for long-term retention. Temporally coordinated oscillations in the hippocampo-cortical network could be a key mechanism for sleep-dependent memory consolidation. We dynamically manipulated the temporal coordination between the two structures during sleep following training on a spatial memory task specifically designed to trigger encoding, but not memory consolidation. Reinforcing the endogenous coordination between hippocampal sharp wave-ripples, cortical delta waves and spindles by timed electrical stimulation resulted in a high recall performance on the next day, contrary to control rats which performed at chance levels. Thus, this enhancement of hippocampo-cortical interactions during sleep stabilized memory traces that would have otherwise vanished between training and test. In addition, we observed a reorganization of prefrontal cortical networks during sleep, along with subsequent increased prefrontal responsivity to the task on the next day. These results provide direct evidence for a causal role of a hippocampo-cortical dialogue during sleep in memory consolidation, and indicate that the underlying mechanism involves a fine-tuned coordination between sharp wave-ripples, delta waves and spindles (Maingret et al., 2016).

Alzheimer’s disease (AD) is a neurodegenerative disorder resulting in the decline of cognitive function, memory formation and retrieval, and abrupt changes in personality. Damage to brain networks occur during prodromal stages of AD, prior to the development of clinical symptoms of dementia. Further characterising this state and identifying reliable biomarkers for early detection are priorities in AD research. I characterised neuronal changes within the dorsal CA1 and subiculum regions of the hippocampal formation (HF) in the well-characterised 3xTgAD mouse model of AD. These regions are well-established sites for early neurodegeneration in both AD patients and AD animal models. We inserted multi-electrode recording arrays into CA1 and subiculum of urethane anaesthetised 3xTgAD mice and recorded spontaneous local field potential activity. Using traditional and novel information theoretic approaches, I determined the information carrying capacity of the CA1- subiculum network during different network rhythms, and how this altered with age and AD-like pathology. A bipolar stimulating electrode was inserted into CA1, allowing the assessment of synaptic integrity between CA1 and subiculum. Results showed that synaptic and network changes occur in CA1 and subiculum during the early stages of AD-like pathology and correlates with the development of intracellular beta-amyloid. There is a progressive breakdown in synaptic facilitation as early as 3 months in the 3xTgAD mouse. These data support an advanced ageing-like phenotype in AD model mice, with an enhanced age/pathology-dependent breakdown in neuronal communication compared to age-matched controls. In agreement with other studies, 3xTgAD mice demonstrate evidence of pathology-related changes in the network rhythms of the HF. 3xTgAD mice show an increase in the power of alpha and beta rhythms, and a concurrent reduction in the power of delta oscillations. Application of novel information theoretic techniques results in a breakdown in the information carrying capacity of the hippocampal system. This deficit manifests as a reduction in information flow during delta-dominant periods of EEG rhythms, with a specific reduction during slow-wave ripple activity. This change in neuronal communication correlates with the onset of memory-retention/consolidation deficits. These network changes are complex, with alterations in the information carrying capacity of the system during theta rhythms at 6 months, and during slow-wave components by 9 months in the 3xTgAD mouse. This study provides the first evidence of an early and progressive decline in neuronal connectivity and communication that correlates with changes in cognition in the 3xTgAD mouse. Application of novel analytical techniques to multi-site EEG recording revealed early and measureable changes in information processing during the onset of AD-like pathology. These are important new biomarkers for early AD characterisation.

Synaptic transmission occurs as a result of either a spontaneous release of presynaptic vesicles or a batch release of presynaptic vesicles driven by action potentials. The physiological consequence of synaptic transmission driven by different patterns and frequencies of presynaptic stimulation has been extensively investigated. However, the physiological nature, mechanism as well as relevance of prolonged presynaptic stimulation have been poorly characterized. In this dissertation, I present three projects in which prolonged stimulation of synaptic transmission in different forms and different brain regions was studied for its effect on synaptic transmission, mechanisms and physiological relevance. In the first project, prolonged electrical stimulation (100 sec) at high frequency induced a deep synaptic depression in acute hippocampal slices, followed by a recovery of synaptic transmission after ~15 min. The deep synaptic depression was attributed to a complete depletion of presynaptic vesicle pools. In the second project, attempts were made to characterize the mechanism of nuclear activation of gene transcription induced by prolonged electrical stimulation (100 sec). Our results demonstrated that reduced inactivation of non-L-type calcium channels failed to provide calcium required for gene transcription, leaving the activation of gene transcription a selective function for L-type calcium channels. In the third project, we sought to study the physiological relevance of enhanced miniature events of inhibitory synapses induced by prolonged chemical stimulation. We showed that prolonged application (2 min) of nicotine to the striatal slice enhanced the frequency of miniature inhibitory currents that was accompanied with a reduction in the amplitude of evoked response. This reduction in the amplitude of evoked responses was ascribed to a compromised action potential invasion of presynaptic terminals possibly due to inactivation of sodium channels resulting from nicotine-induced depolarization. To summarize, prolonged stimulation of presynaptic vesicle release imposes significant influence upon neuron-to-neuron communication, with distinct mechanisms in different brain regions. / Medicine, Faculty of / Graduate

This thesis investigated the contribution of the nucleus accumbens core (AcbC) and the hippocampus (H) to choice and learning involving reinforcement that was delayed or unlikely. Animals must frequently act to influence the world even when the reinforcing outcomes of their actions are delayed. Learning with action-outcome delays is a complex problem, and little is known of the neural mechanisms that bridge such delays. Impulsive choice, one aspect of impulsivity, is characterized by an abnormally high preference for small, immediate rewards over larger delayed rewards, and is a feature of attention-deficit/hyperactivity disorder (ADHD), addiction, mania, and certain personality disorders. Furthermore, when animals choose between alternative courses of action, seeking to maximize the benefit obtained, they must also evaluate the likelihood of the available outcomes. Little is known of the neural basis of this process, or what might predispose individuals to be overly conservative or to take risks excessively (avoiding or preferring uncertainty, respectively), but risk taking is another aspect of the personality trait of impulsivity and is a feature of a number of psychiatric disorders, including pathological gambling and some personality disorders. The AcbC, part of the ventral striatum, is required for normal preference for a large, delayed reward over a small, immediate reward (self-controlled choice) in rats, but the reason for this is unclear. Chapter 3 investigated the role of the AcbC in learning a free-operant instrumental response using delayed reinforcement, performance of a previously learned response for delayed reinforcement, and assessment of the relative magnitudes of two different rewards. Groups of rats with excitotoxic or sham lesions of the AcbC acquired an instrumental response with different delays (0, 10, or 20 s) between the lever-press response and reinforcer delivery. A second (inactive) lever was also present, but responding on it was never reinforced. The delays retarded learning in normal rats. AcbC lesions did not hinder learning in the absence of delays, but AcbC-lesioned rats were impaired in learning when there was a delay, relative to sham-operated controls. Rats were subsequently trained to discriminate reinforcers of different magnitudes. AcbC-lesioned rats were more sensitive to differences in reinforcer magnitude than sham-operated controls, suggesting that the deficit in self-controlled choice previously observed in such rats was a consequence of reduced preference for delayed rewards relative to immediate rewards, not of reduced preference for large rewards relative to small rewards. AcbC lesions also impaired the performance of a previously learned instrumental response in a delay-dependent fashion. These results demonstrate that the AcbC contributes to instrumental learning and performance by bridging delays between subjects' actions and the ensuing outcomes that reinforce behaviour. When outcomes are delayed, they may be attributed to the action that caused them, or mistakenly attributed to other stimuli, such as the environmental context. Consequently, animals that are poor at forming context-outcome associations might learn action-outcome associations better with delayed reinforcement than normal animals. The hippocampus contributes to the representation of environmental context, being required for aspects of contextual conditioning. It was therefore hypothesized that animals with H lesions would be better than normal animals at learning to act on the basis of delayed reinforcement. Chapter 4 tested the ability of H-lesioned rats to learn a free-operant instrumental response using delayed reinforcement, and their ability to exhibit self-controlled choice. Rats with sham or excitotoxic H lesions acquired an instrumental response with different delays (0, 10, or 20 s) between the response and reinforcer delivery. H-lesioned rats responded slightly less than sham-operated controls in the absence of delays, but they became better at learning (relative to shams) as the delays increased; delays impaired learning less in H-lesioned rats than in shams. In contrast, lesioned rats exhibited impulsive choice, preferring an immediate, small reward to a delayed, larger reward, even though they preferred the large reward when it was not delayed. These results support the view that the H hinders action-outcome learning with delayed outcomes, perhaps because it promotes the formation of context-outcome associations instead. However, although lesioned rats were better at learning with delayed reinforcement, they were worse at choosing it, suggesting that self-controlled choice and learning with delayed reinforcement tax different psychological processes. Chapter 5 examined the effects of excitotoxic lesions of the AcbC on probabilistic choice in rats. Rats chose between a single food pellet delivered with certainty (probability p = 1) and four food pellets delivered with varying degrees of uncertainty (p = 1, 0.5, 0.25, 0.125, and 0.0625) in a discrete-trial task, with the large-reinforcer probability decreasing or increasing across the session. Subjects were trained on this task and then received excitotoxic or sham lesions of the AcbC before being retested. After a transient period during which AcbC-lesioned rats exhibited relative indifference between the two alternatives compared to controls, AcbC-lesioned rats came to exhibit risk-averse choice, choosing the large reinforcer less often than controls when it was uncertain, to the extent that they obtained less food as a result. Rats behaved as if indifferent between a single certain pellet and four pellets at p = 0.32 (sham-operated) or at p = 0.70 (AcbC-lesioned) by the end of testing. When the probabilities did not vary across the session, AcbC-lesioned rats and controls strongly preferred the large reinforcer when it was certain, and strongly preferred the small reinforcer when the large reinforcer was very unlikely (p = 0.0625), with no differences between AcbC-lesioned and sham-operated groups. These results suggest that the AcbC contributes to action selection by promoting the choice of uncertain, as well as delayed, reward.

At the author’s request, the abstract has been removed due to the confidential nature of the thesis. It will be added once the embargo period has passed.

This study assessed the in vivo effects of various serotonin (5-HT) receptor modulators on 5-HT neurotransmission in the rat hippocampus. Vortioxetine, humanized-vortioxetine, and escitalopram blocked the 5-HT transporter, but similar to ipsapirone did not dampen the sensitivity of postsynaptic 5-HT1A receptors. Long-term administration of all treatments increased the tonic activation of postsynaptic 5-HT1A heteroreceptors, an effect common to all antidepressants. Vortioxetine decreased the function of the terminal 5-HT1B autoreceptor under high but not a low degree of activation, thus showing that its partial agonism led to increased 5-HT release and that long-term administration results in the desensitization of terminal 5-HT1B autoreceptors. Vortioxetine overcame the effects of 5-HT1B and 5-HT3 receptor agonists. This study was unable to determine the involvement of 5-HT7 receptor antagonism exerted by vortioxetine affects 5-HT neurotransmission. Therefore, vortioxetine would appear to exert different actions, via transporter and receptor activity, on the serotonergic system in the hippocampus, consistent with its unique pharmacological profile.

Les astrocytes sont des partenaires clés des neurones. Dans l’hippocampe, et tout particulièrement au niveau des synapses CA3-CA1, en libérant la D-sérine, ces cellules gliales régulent l’activité des récepteurs glutamatergiques de type N-methyl-D-aspartate (NMDA) et de ce fait la mémoire synaptique, aussi connue sous le nom de plasticité synaptique à long terme. Cependant, le signal synaptique à l’origine de la libération de la D-sérine par les astrocytes reste à ce jour méconnu. De par des données rapportées dans la littérature nous nous sommes tout particulièrement intéressés aux récepteurs astrocytaires aux ephrins de type B3 (EphB3) et aux endocannabinoïdes de type 1 (CB1). Pour ce faire nous avons principalement utilisé une approche électrophysiologique sur des tranches aiguës d’hippocampe de souris adulte. Dans une première étude, nos données indiquent que l’activation des récepteurs EphB3 augmente la présence de D-sérine synaptique et en conséquence l’activité des récepteurs NMDA synaptiques. A l’inverse, leur inhibition diminue à la fois l’activité des récepteurs NMDA synaptiques et la potentialisation à long-terme qui en dépend (LTP ; une forme de plasticité synaptique à long terme). L’interaction EphB3-ephrinB3 contrôle donc la LTP en contrôlant la disponibilité en D-sérine synaptique. Dans une seconde étude, nous avons utilisé un modèle transgénique permettant d’inhiber l’expression des récepteurs CB1 astrocytaires (souris GFAP-CB1-KO). Nous avons découvert que la suppression de ces récepteurs diminue la disponibilité en D-sérine synaptique. De plus, nos travaux montrent que les récepteurs CB1 astrocytaires sont nécessaires à l’induction de la LTP via la D-serine. En conclusion, ces travaux de Thèse révèlent que les récepteurs astrocytaires EphB3 et CB1 régulent les fonctions dépendantes des récepteurs NMDA via le contrôle qu’ils exercent sur la disponibilité en D-sérine. / Astrocytes are key partners of neurons. In the hippocampus, and more particularly at CA3-CA1 synapses, by releasing D-serine, these glial cells regulate the activity of synaptic Nmethyl-D-aspartate (NMDA) receptors and thus synaptic memory, also known as long-term synaptic plasticity. Yet, the synaptic signal inducing D-serine release by astrocytes is still unknown. Based on interesting data from the literature we have investigated the role of the astrocytic receptors for ephrinB3 (EphB3) and endocannabinoids (CB1). To this end we used electrophysiological approaches on acute hippocampal slices of adult mice. In a first study, our data indicate on one hand that the activation of EphB3 receptors increases synaptic D-serine availability and in consequences the activity of synaptic NMDA receptor activity. On the other hand, inhibition of EphB3 receptors induces a decrease of synaptic NMDA receptor activity as well as the induction of the long-term potentiation (LTP; a form of long-term plasticity). Thus, EphB3-ephrinB3 interaction controls LTP induction through the availability of synaptic D-serine. In a second study, we used a transgenic model allowing the inhibition of CB1 receptors expression in astrocytes (GFAP-CB1-KO mice). We discovered that their deletion reduced synaptic D-serine availability. Our work shows that astrocytic CB1 receptors are necessary for LTP induction via this D-serine. All together, this PhD work reveals that astrocytic EphB3 and CB1 receptors regulate synaptic NMDA receptor functions through the control of D-serine availability.

The hippocampus is critical for the encoding and retrieval of episodic memories. During ongoing experience, the hippocampus exhibits activity patterns related to the current spatiotemporal context. How hippocampal firing patterns relate to the representation of mental maps important for behavioral and cognitive processes is still an open question. Here a series of experiments aimed to test how the hippocampus represents the spatiotemporal context of ongoing experience. Extracellular recordings from the dorsal CA1 region of the hippocampus were collected from rats engaged in a blocked serial reversal object-association task. Behaviorally, rats did not utilize the temporal segregation between task blocks as a way to correctly match object valence and rather treated each block of trials as separate episodes. This lack of an alternating context was further uncovered in the neural coding of the rat’s hippocampal firing patterns. Furthermore, gradual drift in the hippocampal ensemble representation of experience was discovered, correlating with the temporal duration of the task and not the blocked organization of the behavioral paradigm. In the next two experiments, extracellular recordings from dorsal CA1 were collected from rats traversing a linear track environment, with different environmental manipulations. During variable starting location recording sessions, it was found that positional coding by the hippocampal population was relative to starting location and that place field allocation was biased towards the reference frame at the start of the journey, demonstrating that hippocampal place fields are not uniformly distributed and express compressed activity patterns referenced to the beginning point of trajectories. During blocked manipulation of lighting condition, individual units showed preference to specific lighting conditions and the hippocampal population rapidly remapped between lights ‘ON’ and lights ‘OFF’ blocks of trials, suggesting that hippocampal maps of space are not solely governed by internal dynamics and that alterations in sensory input can modify hippocampal motifs of ongoing experience. Overall, the findings of the three experiments further our understanding of how the hippocampus represents ongoing experience, highlighting the role of temporal drift as well as demonstrating how both external and internal stimuli and frames of reference coalesce into a comprehensive cognitive map of experience.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第22312号 / 医博第4553号 / 新制||医||1040(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 伊佐 正, 教授 林 康紀, 教授 高橋 淳 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM