La Maladie d’Alzheimer (MA) est la première cause de démence au monde. Un fort coût de prise en charge, associé une faible efficacité des traitements actuels font de la découverte d’une thérapie efficace une priorité. Dans ce contexte les récepteurs sérotoninergiques de type 4 (5-HT4R) représentent une cible prometteuse. En effet, l’utilisation d’agonistes des 5-HT4R chez l’animal entraîne à la fois des effets pro-mnésiants et anti-amnésiants et une action sur la physiopathologie de cette maladie. Par ailleurs, l’aspect multifactoriel de la MA a conduit à faire émerger ces derniers années un consensus quant à la nécessité du développement de stratégie thérapeutique multi-cibles. Dans un premier temps, nous avons démontré l’intérêt de l’association d’un inhibiteur de l’acétylcholinestérase (IAChE) - l’un des rares médicaments disponibles, à un agoniste des 5-HT4R (RS67333) sur les performances de mémoire de travail et de référence chez la souris. Par la suite et afin de mieux comprendre les mécanismes mises en jeu dans les effets anti-amnésiant du RS67333, nous avons étudié les effets de la stimulation des 5-HT4R sur la plasticité synaptique mesurée au moyen d’une approche ex vivo au niveau de la région CA1 de l’hippocampe chez la souris saine NMRI. En effet, la plasticité synaptique, est un élément essentiel des processus d’apprentissage et de mémoire. Une première étude chez la souris saine a montré que la stimulation des 5-HT4R inhibait la potentialisation à long terme (LTP) induite par stimulation thêta-burst (TBS). Nos résultats suggèrent que l’inhibition de la LTP par l’agoniste est médiée par une modification de la neurotransmission GABAergique. La dernière partie de mes travaux a porté sur un modèle transgénique murins mimant certains des aspects de la MA (souris 5xFAD). Ainsi, si la stimulation des 5-HT4R conduisait à des résultats similaires en terme de transmission synaptique de base ou de plasticité à long-terme, elle modifie de façon importante la plasticité à court terme. L’ensemble de ces résultats obtenus ex vivo nécessiteraient d’être associé à des mesures in vivo, pour faire le lien entre les effets sur la plasticité synaptique et les performances de mémoire. Nos travaux montrent l’existence d’une association entre les 5-HT4R et la transmission GABAergique dans la modulation de la plasticité synaptique hippocampique de la région CA1. Plus généralement, ils renforcent l’intérêt de stratégies multi-cibles contenant notamment un agoniste des 5-HT4R. / Alzheimer’s disease (AD) is the first cause of dementia in the world. Due to its high cost of care combined with the lack of efficient treatment, the discovery of an effective therapy is a priority. In that regard, 5-HT4R are a promising target. Indeed, preclinical studies showed promnesic and anti-amnesic effect of 5-HT4R agonist as well as a disease-modifying effect on amyloid processing. Furthermore, given the multifactorial aspect of AD pathophysiology, there is a consensus concerning the necessity for multi-target therapy to treat effectively this disease. First, we confirmed the beneficial effect of a combined treatment with galantamine, an IAChE, and RS67333, a 5-HT4R agonist, on working and reference memory performances in a pharmacological model of scopolamine-induced amnesia. Then, for a better understanding of the mechanisms involved in 5-HT4R stimulation-induced increase in memory performances, we investigated the effects of such stimulation on CA1 area synaptic plasticity. Indeed, synaptic plasticity is a key component of learning and memory processes. Through ex-vivo electrophysiological recordings, we demonstrated that 5-HT4R activation impairs TBS-induced LTP in wild-type healthy mice. Further experiments suggested that such impairment involves a modulation of GABAergic neurotransmission. In addition, a third study on a transgenic model of AD (5xFAD mice) showed similar results. These results, obtained exclusively ex vivo, need to be associated with in vivo experiments to close the gap with behavioral experiments and allow an interpretation of memory performances through synaptic plasticity modifications. Our work shows the existence of an interplay between 5-HT4R and GABAergic transmission in the regulation of synaptic plasticity in hippocampal CA1 area. Furthermore, we strengthen the interest toward multi-target treatment involving 5-HT4R agonists in the field of AD.

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The version posted must include the following notice on the first page: This is the peer reviewed version of the following article: http://onlinelibrary.wiley.com/doi/10.1002/cne.23477/abstract, which has been published in final form at DOI: 10.1002/cne.23477. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving. / 京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第19610号 / 医博第4117号 / 新制||医||1015(附属図書館) / 32646 / 京都大学大学院医学研究科医学専攻 / (主査)教授 渡邉 大, 教授 髙橋 良輔, 教授 宮本 享 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

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The research discussed here examines how normal healthy aging affects spatial suppressive mechanisms in a variety of visual tasks using both static and dynamic stimuli. Prior research has suggested that younger adults demonstrate a center-surround antagonistic pattern in which they show spatial summation at low contrast and spatial suppression at high contrast in brief motion direction discrimination tasks. Older adults have been shown to have reduced spatial suppression at high contrast and this is thought to be related to reduced GABAergic inhibition in the visual cortex. The results obtained from this program of research suggest that age-related changes in optical and neural visual mechanisms do not affect spatiotemporal mechanisms for static stimuli when the target is presented with the mask (embedded masking). However, when the mask appears immediately before (forward masking) or after (backward masking) the target, older adults require more contrast to detect the target (Chapter 2). In addition, spatial suppression is not reduced for older adults in a task with moving stimuli presented at long durations, even with increasing speed (Chapter 3). In Chapter 4, we used static stimuli presented at brief durations to induce a sudden motion onset and found that although there was no significant age difference in spatial suppression, there was a trend showing reduced levels of spatial suppression in older adults. These results taken together suggest that inhibitory neural mechanisms in the visual cortex may mediate spatial suppression for briefly presented stimuli only. / Thesis / Doctor of Philosophy (PhD)

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No description available.

京都大学 / 新制・課程博士 / 博士(医学) / 甲第24088号 / 医博第4864号 / 新制||医||1059(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 林 康紀, 教授 渡邉 大, 教授 髙橋 良輔 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Tinnitus, ringing in the ears, is a phantom sound percept affecting roughly 10-20% of the total world population. Tinnitus severely impacts the quality of life of 10% of tinnitus sufferers, affecting their sleep, concentration, emotion, social enjoyment, and sometimes leading to depression and suicidal tendencies. In humans, most forms of tinnitus are associated with noise-exposure, leading to compensatory maladaptive plasticity of central auditory neurons. Human and animal studies suggest that tinnitus alters normal adult attentional resources. Human studies by McKenna, Hallam and Surlock 1996, suggested tinnitus-related impairment in sustained attention, vigilance, visual conceptualization and visuo-motor memory. Additionally, tinnitus may impact aspects of selective or divided attention as well as working and long-term memory. The involvement of primary auditory cortex and nicotinic signaling in selective attention, working and long-term memory has been well established. Neuronal nicotinic acetylcholine receptors (nAChRs) are present on presynaptic and postsynaptic inputs that innervate neurons across layers of primary auditory cortex (A1). Layer 5 pyramidal neurons (PNs) in the A1 are major output neurons, conveying auditory information to corticocortical and subcortical nuclei. The excitation of PNs is regulated by a complex microcircuitory of inhibitory neurons with vasointestinal peptide positive (VIP) neurons playing a key role in regulating the excitation. The focus of present studies was to: 1) Characterize tinnitus-related changes in the physiology and nAChR signaling of layer 5 PNs and VIP neurons in the A1 and 2) Determine the ability of nAChR partial/desensitizing agonists to ameliorate tinnitus pathology in subcellular studies. Wild-type, ChAT-Cre and VIP-Cre:Rosa26-loxP-stop-loxP-tdTomato (VIP-Cre:Rosa-tdTomato Long-Evans rats were used in the present study. CHAT-Cre rats allowed us to selectively express cre-inducible AAV-EF1a-DIO-hChR2(H134R)-EYFP and stimulate the cholinergic neurons of basal forebrain (BF). VIP-Cre:Rosa-tdTomato express fluorescent tdTomato protein in the VIP positive neurons allowing us to identify them under fluorescence microscopy using 550 nm wavelength. An established noise-exposure (one hour of 116 dB narrowband noise centered at 16 kHz) was used to induce behavioral tinnitus in a rat model. Approximately 50-60% noise-exposed animals (53/92) exhibited behavioral evidence of tinnitus with significant shifts in hearing threshold contiguous to the exposure frequency. Animals were classified as control, exposed tinnitus and non-tinnitus. In vitro whole-cell patch clamp recordings were performed in control and tinnitus animals. Results: Numerous tinnitus-related changes in the physiology of layer 5 PNs and VIP neurons, and changes in the activity of excitatory and inhibitory input neurons were observed. The resting membrane potential of A1 layer 5 PNs from tinnitus animals was significantly depolarized compared to PNs from unexposed controls. PNs from the A1 of animals with behavioral evidence of tinnitus showed increases in the frequency of excitatory and decreases in frequency of inhibitory spontaneous postsynaptic currents, which directly correlated with the rat’s tinnitus score. Optical stimulation of thalamocortical terminals from PNs in tinnitus animals evoked significantly larger excitatory/inward currents than in currents evoked in PNs from controls. A1 layer 5 PNs showed tinnitus-related decreases in postsynaptic gamma-amino butyric acid (GABA) signaling suggestive of GABA-A receptors (GABA-ARs) subunit switches or loss of GABA-ARs. VIP neurons favoring excitation of layer 5 PNs via disinhibition, were depolarized with significantly lower current to evoke action potentials (rheobase current). The excitability of VIP neurons was significantly increased, with this increase being strongly correlated to the rat’s tinnitus score. Tinnitus-related changes in nAChR signaling were then tested in layer 5 PNs and VIP neurons. Both PNs and VIP neurons receive cholinergic input from basal forebrain and were highly sensitive to nicotinic stimulation. Optical stimulation of basal forebrain (BF) terminals evoked a depolarizing current from VIP neurons. In tinnitus animals, layer 5 PNs showed a significant loss of nAChR signaling, while, VIP neurons showed tinnitus-related increase in responses to nicotinic stimulation. Most of the nAChR responses in auditory cortex are believed to be mediated via volume transmission of acetylcholine (ACh). Continuous voltage clamped recordings were used to examine the activity of excitatory and inhibitory neurons impacting PNs in the presence of bath applied ACh. We observed significant tinnitus-related changes in nAChR signaling with layer 5 PNs showing significantly larger GABAergic input after prolonged bath application of ACh. This led us to hypothesize that desensitization of nAChRs could increase/normalize the activity of GABAergic input neurons. To test this hypothesis, nAChR partial desensitizing agonists sazetidine-A and varenicline were used in cellular and behavioral studies. Immediately after bath application of sazetidine-A or varenicline, a dramatic increase in the activity of inhibitory input neurons onto PNs was observed. In a behavioral tinnitus test, both sazetidine-A and varenicline were effective in lowering the tinnitus-like behavior. In conclusion, we identified a significant tinnitus-related disruption in intrinsic physiology of layer 5 PNs and VIP neurons, with strong evidence of dysregulated cholinergic signaling. Partial/desensitizing agonists sazetidine-A and varenicline increased the activity of inhibitory input neurons, showing therapeutic potential in both subcellular and behavioral studies.

The activity of networks subserving memory and learning in the hippocampus is under the control of GABAergic interneurons. In order to test the contribution of distinct cell types, I have recorded extracellularly, labelled, and identified different types of interneuron in area CA3 of the hippocampus, a region implicated in the generation of gamma and theta oscillations, and the initiation of sharp-waves. I present here a detailed analysis of the spike timing of parvalbumin-positive (PV) basket and physiologically identified pyramidal cells in area CA3, relative to various network states recorded in area CA3 and CA1 simultaneously. Additionally, I have shown by detailed analysis that five classes of previously recorded and identified CA1 interneuron fired with cell type specific firing patterns relative to local gamma oscillations. In CA3, PV basket cells fired phase locked to theta and gamma oscillations recorded in CA1 as well as in CA3, and increased their firing rates during CA1 sharp-waves. Pyramidal cells in CA3 were also phase-locked, but fired at phases different from basket cells. During theta oscillations, CA3 pyramidal and PV basket cells were phase locked to both CA1 and CA3 theta equally, suggesting a wide coherence of these oscillations; in contrast, cells fired more strongly phase-locked to gamma oscillations in CA3 than in CA1, suggesting a specific role for CA3 in the generation of this rhythm. In contrast to theta and gamma oscillations, CA3 basket cells were phase-locked to ripples in area CA3 but not in CA1. Overall, my results show that the spike timing of several types of interneuron in CA1, and PV basket cells in CA3, is correlated in a cell- and area-specific manner with the generation of particular states of synchronous activity.