The neural circuitry responsible for memory consists of complex components with dynamic interactions. In hippocampal area CA1, interactions between excitatory pyramidal cells and inhibitory interneurons shape ensemble activity which encodes sequential experience. An extremely diverse set of inhibitory interneurons, with variation in gene expression, synaptic targeting, state-dependent activity, and connectivity, contribute substantially to circuit activity, such as theta and sharp wave-ripple oscillations. The precise roles of each interneuron group is not well understood, though characterization of their activity reveals mechanisms underlying hippocampal circuit computation. In this dissertation, I aim to identify and disrupt interactions between pyramidal cells and local interneurons to clarify their role in shaping cell assembly activity. We characterized axo-axonic cell activity in sharp wave-ripples, and compared their control of pyramidal cell activity and ripple events to parvalbumin expressing neurons. We identified pyramidal cell-interneuron interactions during ripples, suggesting they serve as lateral inhibitors between cell assemblies. We additionally developed and implemented a novel neural device to explore the role of cannabinoid disruption of hippocampal oscillations and organization of assemblies in vivo in awake animals. We demonstrate that cannabinoid receptor type 1 within CA1 is responsible for suppression of theta and SPW-Rs. We also found that cannabinoid activation within CA1 circuitry, regardless of muted input from CA3, was sufficient to disrupt sharp wave-ripples, likely through interference of pyramidal cell-interneuron interactions. The work in this dissertation provides insight suggesting that interneuron activity must be studied at the spiking timescale to characterize their control over cell assembly activity. / Doctor of Philosophy / Understanding how the brain creates memory remains one of the greatest questions in the field of neuroscience. Coordinated brain activity serves to build communication on large and small scales, across brain regions and within circuits consisting of small groups of neurons. Precise coordination of activity and communication across neurons and regions is thought to build salient experience, which is achieved through the timing of neuron action potentials, or spikes. Neurons receive thousands of inputs that control their spiking activity. "Go and stop" signals from excitatory and inhibitory interneurons act to conduct synchronized activity, which is required for proper circuit function. Importantly, coordinated spiking across large groups of neurons is responsible for observed "brain waves", or oscillations, which reflect organized activity. In CA1 of the hippocampus, there are >20 subtypes of interneurons that all make distinct contributions to memory function, and the roles of these interneurons have not been fully studied within behaving animals. As engineers develop new tools, new methods become available to study and classify how unique groups of interneurons play a part in circuit activity. Thus, we sought to characterize the role of axo-axonic cells, a specialized interneuron with strong control over spiking activity, in hippocampal oscillations that are responsible for memory encoding and consolidation. We identified a new role for axo-axonic cells in the regulation of pyramidal cell spiking in sharp wave-ripple oscillations. Additionally, we developed a novel neural device that allowed us to investigate the mechanisms that underlie cannabinoids, molecules found in Cannabis sativa, and memory dysfunction. We leveraged the multifunctionality of our T-DOpE probe to focally deliver synthetic cannabinoid into the hippocampus in combination with optical control of circuits, with simultaneous recording of activity. We found that cannabinoids acting within CA1 sufficiently disrupt hippocampal oscillations, likely through hindering pyramidal cell-interneuron interactions. Together, these findings suggest that the spatial and temporal resolution required to study diverse roles of interneurons is high, and experiments designed to explore interneuron activity should especially emphasize fine time-scales.

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Komplexe Berechnungen im Gehirn werden durch das Zusammenspiel von exzitatorischen und hemmenden Neuronen in lokalen Netzwerken ermöglicht. In kortikalen Netzwerken, wird davon ausgegangen, dass hemmende Neurone, besonders Parvalbumin positive Korbzellen, ein „blanket of inhibition” generieren. Dieser Sichtpunkt wurde vor kurzem durch Befunde strukturierter Inhibition infrage gestellt, jedoch ist die Organisation solcher Konnektivität noch unklar. In dieser Dissertation, präsentiere ich die Ergebnisse unserer Studie Parvabumin positiver Korbzellen, in Schichten II / III des entorhinalen Kortexes und Präsubiculums der Ratte. Im entorhinalen Kortex haben wir dorsale und ventrale Korbzellen beschrieben und festgestellt, dass diese morphologisch und physiologisch ähnlich, jedoch in ihrer Konnektivität zu Prinzipalzellen dorsal stärker als ventral verbunden sind. Dieser Unterschied korreliert mit Veränderungen der Gitterzellenphysiologie. Ähnlich zeige ich im Präsubiculum, dass inhibitorische Konnektivität eine essenzielle Rolle im lokalen Netzwerk spielt. Hemmung im Präsubiculum ist deutlich spärlicher ist als im entorhinalen Kortex, was ein unterschiedliches Prinzip der Netzwerkorganisation suggeriert. Um diesen Unterschied zu studieren, haben wir Morphologie und Netzwerkeigenschaften Präsubiculärer Korbzellen analysiert. Prinzipalzellen werden über ein vorherrschendes reziprokes Motif gehemmt die durch die polarisierte Struktur der Korbzellaxone ermöglicht wird. Unsere Netzwerksimulationen zeigen, dass eine polarisierte Inhibition Kopfrichtungs-Tuning verbessert. Insgesamt zeigen diese Ergebnisse, dass inhibitorische Konnektivität, funktioneller Anforderungen der lokalen Netzwerke zur Folge, unterschiedlich strukturiert sein kann. Letztlich stelle ich die Hypothese auf, dass für lokale inhibitorische Konnektivität eine Abweichung von „blanket of inhibition― zur „maßgeschneiderten― Inhibition zur Lösung spezifischer computationeller Probleme vorteilhaft sein kann. / Local microcircuits in the brain mediate complex computations through the interplay of excitatory and inhibitory neurons. It is generally assumed that fast-spiking parvalbumin basket cells, mediate a non-selective -blanket of inhibition-. This view has been recently challenged by reports structured inhibitory connectivity, but it’s precise organization and relevance remain unresolved. In this thesis, I present the results of our studies examining the properties of fast-spiking parvalbumin basket cells in the superficial medial entorhinal cortex and presubiculum of the rat. Characterizing these interneurons in the dorsal and ventral medial entorhinal cortex, we found basket cells of the two subregions are more likely to be connected to principal cells in the dorsal compared to the ventral region. This difference is correlated with changes in grid physiology. Our findings further indicated that inhibitory connectivity is essential for local computation in the presubiculum. Interestingly though, we found that in this region, local inhibition is lower than in the medial entorhinal cortex, suggesting a different microcircuit organizational principle. To study this difference, we analyzed the properties of fast-spiking basket cells in the presubiculum and found a characteristic spatially organized connectivity principle, facilitated by the polarized axons of the presubicular fast-spiking basket cells. Our network simulations showed that such polarized inhibition can improve head direction tuning of principal cells. Overall, our results show that inhibitory connectivity is differently organized in the medial entorhinal cortex and the presubiculum, likely due to functional requirements of the local microcircuit. As a conclusion to the studies presented in this thesis, I hypothesize that a deviation from the blanket of inhibition, towards a region-specific, tailored inhibition can provide solutions to distinct computational problems.

Dans le cortex visuel des mammifères, une cellule à panier (BC) qui représente un sous-type majoritaire d’interneurones GABAergiques, innerve une centaine de neurones par une multitude de synapses localisées sur le soma et sur les dendrites proximales de chacune de ses cibles. De plus, ces cellules sont importantes pour la génération des rythmes gammas, qui régulent de nombreuses fonctions cognitives, et pour la régulation de la plasticité corticale. Bien que la fonction des BC au sein des réseaux corticaux est à l'étude, les mécanismes qui contrôlent le développement de leur arborisation complexe ainsi que de leurs nombreux contacts synaptiques n’ont pas été entièrement déterminés. En utilisant les récepteurs allatostatines couplés aux protéines G de la drosophile (AlstR), nous démontrons in vitro que la réduction de l'excitation ainsi que la réduction de la libération des neurotransmetteurs par les BCs corticales individuelles des souris, diminuent le nombre de cellules innervées sans modifier le patron d'innervation périsomatique, durant et après la phase de prolifération des synapses périsomatiques. Inversement, lors de la suppression complète de la libération des neurotransmetteurs par les BCs individuelles avec l’utilisation de la chaîne légère de la toxine tétanus, nous observons des effets contraires selon le stade de développement. Les BCs exprimant TeNT-Lc pendant la phase de prolifération sont caractérisées par des arborisations axonales plus denses et un nombre accru de petits boutons homogènes autour des somas innervés. Toutefois, les cellules transfectées avec TeNT-Lc après la phase de la prolifération forment une innervation périsomatique avec moins de branchements terminaux d’axones et un nombre réduit de boutons avec une taille irrégulière autour des somas innervés. Nos résultats révèlent le rôle spécifique des niveaux de l’activité neuronale et de la neurotransmission dans l'établissement du territoire synaptique des cellules GABAergiques corticaux. Le facteur neurotrophique dérivé du cerveau (BDNF) est un modulateur puissant de la maturation activité-dépendante des synapses GABAergiques. Grâce à l'activation et à la signalisation de son récepteur tyrosine kinase B (TrkB), la liaison de mBDNF module fortement la prolifération des synapses périsomatiques GABAergiques formés par les BCs. Par contre, le rôle du récepteur neurotrophique de faible affinité, p75NTR, dans le développement du territoire synaptique des cellules reste encore inconnu. Dans ce projet, nous démontrons que la suppression de p75NTR au niveau des BCs individuelles in vitro provenant de souris p75NTRlox induit la formation d'une innervation périsomatique exubérante. BDNF est synthétisé sous une forme précurseur, proBDNF, qui est par la suite clivée par des enzymes, y compris la plasmine activée par tPA, pour produire une forme mature de BDNF (m)BDNF. mBDNF et proBDNF se lient avec une forte affinité à TrkB et p75NTR, respectivement. Nos résultats démontrent qu’un traitement des cultures organotypiques avec la forme résistante au clivage de proBDNF (mut-proBDNF) réduit fortement le territoire synaptique des BCs. Les cultures traitées avec le peptide PPACK, qui inactive tPA, ou avec tPA altèrent et favorisent respectivement la maturation de l’innervation synaptique des BCs. Nous démontrons aussi que l’innervation exubérante formée par les BCs p75NTR-/- n’est pas affectée par un traitement avec mut-proBDNF. L’ensemble de ces résultats suggère que l'activation de p75NTR via proBDNF régule négativement le territoire synaptique des BCs corticaux. Nous avons ensuite examiné si mut-proBDNF affecte l’innervation périsomatique formée par les BCs in vivo, chez la souris adulte. Nous avons constaté que les boutons GABAergiques périsomatiques sont significativement diminués dans le cortex infusé avec mut-proBDNF par rapport à l’hémisphère non-infusé ou traité avec de la saline. En outre, la plasticité de la dominance oculaire (OD) est rétablie par ce traitement chez la souris adulte. Enfin, en utilisant des souris qui ne possèdent pas le récepteur p75NTR dans leurs BCs spécifiquement, nous avons démontré que l'activation de p75NTR via proBDNF est nécessaire pour induire la plasticité de la OD chez les souris adultes. L’ensemble de ces résultats démontre un rôle critique de l'activation de p75NTR dans la régulation et le maintien de la connectivité des circuits GABAergiques, qui commencent lors du développement postnatal précoce jusqu’à l'âge adulte. De plus, nous suggérons que l'activation contrôlée de p75NTR pourrait être un outil utile pour restaurer la plasticité dans le cortex adulte. / Cortical GABAergic basket cells (BC) innervate hundreds of postsynaptic targets with synapses clustered around the soma and proximal dendrites. They are important for gamma oscillation generation, which in turn regulate many cognitive functions, and for the regulation of developmental cortical plasticity. Although the function of BC within cortical networks is being explored, the mechanisms that control the development of their extensive arborisation and synaptic contacts have not been entirely resolved. By using the Drosophila allatostatin G-protein-coupled receptors (AlstR), we show that reducing excitation, and thus neurotransmitter release, in mouse cortical single BC in slice cultures decreases the number of innervated cells without changing the pattern of perisomatic innervation, both at the peak and after the proliferation phase of perisomatic synapse formation. Conversely, suppressing neurotransmitter release in single BCs by using the tetanus toxin light-chain can have completely opposite effects depending on the developmental stage. Basket cells expressing TeNT-Lc during the peak of the proliferation were characterized by denser axonal arbors and an increased number of smaller, homogenous boutons around the innervated somatas compared with control cells. However, after the peak of the synapse proliferation, TeNT-Lc transfected BCs formed perisomatic innervation with fewer terminal axon branches and fewer irregular-sized boutons around innervated somatas. Our results reveal a remarkably specific and age-dependent role of neural activity and neurotransmission levels in the establishment of the synaptic territory of cortical GABAergic cells. Brain derived neurotrophic factor (BDNF) has been shown to be a strong modulator of activity-dependent-maturation of GABAergic synapses. Through the activation and signaling of their receptor Tropomyosin-related kinase B (TrkB), mBDNF binding strongly modulates the proliferation of GABAergic perisomatic synapses formed by BCs. Whether the low-affinity neurotrophin-receptor p75NTR also play a role in the development of basket cell synaptic territory is unknown. Here, we show that single-cell deletion of p75NTR in BCs in cortical organotypic cultures from p75NTRlox mice induce the formation of exuberant perisomatic innervations by the mutant basket cells, in a cell-autonomous fashion. BDNF is synthesized as a precursor, proBDNF, which is cleaved by enzymes, including tPA-activated plasmin, to produce mature (m)BDNF. mBDNF and proBDNF bind with high-affinity to TrkB and p75NTR, respectively. Our results show that treating organotypic cultures with cleavage-resistant proBDNF (mut-proBDNF) strongly reduces the synaptic territory of BCs. Treating cultures with the tPA-inactivating peptide PPACK or with tPA impairs and promotes the maturation of BC synaptic innervations, respectively. We further show that the exuberant innervations formed by p75NTR-/- basket cells are not affected by mut-proBDNF treatment. All together, these results suggest that proBDNF-mediated p75NTR activation negatively regulates the synaptic territory of BCs. We next examined if mut-proBDNF affects perisomatic innervation formed by BCs in vivo, in the adult mouse. We found that perisomatic GABAergic boutons are significantly decreased in the cortex infused with mut-proBDNF as compared to non-infused or saline-treated hemispheres. Further, ocular dominance (OD) plasticity is restored by this treatment in adult mice. Finally, we found that proBDNF-mediated activation of p75NTR is necessary to induce OD plasticity in the adult mice, by using mice that lack p75NTR specifically in BCs. All together, these results demonstrate a critical role of p75NTR activation in regulating and maintaining GABAeric circuit connectivity from early postnatal development to adulthood. Further, we suggest that controlled activation of p75NTR could be a useful tool to restore plasticity in adult cortex.