The Role of P2Y12 in Non-Pathological Microglial Functions during Synaptic Plasticity

Thesis (Ph.D.)--University of Rochester. School of Medicine & Dentistry. Dept. of Neurobiology and Neuroanatomy, 2016.Synaptic plasticity is critical for neurodevelopment and proper function of the adult nervous system. Studies show that microglia play critical roles in neurodevelopment, but mechanisms driving these roles are poorly understood. I explored purinergic signaling as a potential mediator between microglia and neurons during synaptic plasticity. Purinergic signaling has been implicated in microglial behavior, but studies focused on inflammatory roles. Non-inflamed microglia highly and selectively express the purinergic receptor, P2Y12, which functions in microglial chemotaxis. I posited that purinergic signaling contributes to the microglial motility underlying synapse surveillance and may be critical for microglial roles in synaptic refinement. My evidence suggests that P2Y12 disruption prevents ocular dominance shifts indicative of synaptic plasticity. P2Y12 disruption also decreases microglial process complexity, without affecting basal microglial process dynamics. In addition, microglial process dynamics appear to be regulated by arousal with increased surveillance during slow-wave sleep-like states. I find that noradrenergic signaling, contributing to arousal, is sufficient to suppress microglial process dynamics and inhibit microglial P2Y12-mediated roles in synaptic plasticity via microglial β2 adrenergic receptors. These data suggest microglia are active participants in cortical network remodeling in adolescent synaptic plasticity primarily during sleep states. These results not only describe novel neuro-immune interactions in the non-pathological brain, but provoke broader considerations of the importance of sleep in microglial roles during neurodevelopment and in neuropathology

Astrocyte glutamate uptake coordinates experience‐dependent, eye‐specific refinement in developing visual cortex

The uptake of glutamate by astrocytes actively shapes synaptic transmission, however its role in the development and plasticity of neuronal circuits remains poorly understood. The astrocytic glutamate transporter, GLT1 is the predominant source of glutamate clearance in the adult mouse cortex. Here, we examined the structural and functional development of the visual cortex in GLT1 heterozygous (HET) mice using two-photon microscopy, immunohistochemistry and slice electrophysiology. We find that though eye-specific thalamic axonal segregation is intact, binocular refinement in the primary visual cortex is disrupted. Eye-specific responses to visual stimuli in GLT1 HET mice show altered binocular matching, with abnormally high responses to ipsilateral compared to contralateral eye stimulation and a greater mismatch between preferred orientation selectivity of ipsilateral and contralateral eye responses. Furthermore, we observe an increase in dendritic spine density in the basal dendrites of layer 2/3 excitatory neurons suggesting aberrant spine pruning. Monocular deprivation induces atypical ocular dominance plasticity in GLT1 HET mice, with an unusual depression of ipsilateral open eye responses; however, this change in ipsilateral responses correlates well with an upregulation of GLT1 protein following monocular deprivation. These results demonstrate that a key function of astrocytic GLT1 function during development is the experience-dependent refinement of ipsilateral eye inputs relative to contralateral eye inputs in visual cortex

Astrocyte glutamate uptake coordinates experience‐dependent, eye‐specific refinement in developing visual cortex

The uptake of glutamate by astrocytes actively shapes synaptic transmission, however its role in the development and plasticity of neuronal circuits remains poorly understood. The astrocytic glutamate transporter, GLT1 is the predominant source of glutamate clearance in the adult mouse cortex. Here, we examined the structural and functional development of the visual cortex in GLT1 heterozygous (HET) mice using two-photon microscopy, immunohistochemistry and slice electrophysiology. We find that though eye-specific thalamic axonal segregation is intact, binocular refinement in the primary visual cortex is disrupted. Eye-specific responses to visual stimuli in GLT1 HET mice show altered binocular matching, with abnormally high responses to ipsilateral compared to contralateral eye stimulation and a greater mismatch between preferred orientation selectivity of ipsilateral and contralateral eye responses. Furthermore, we observe an increase in dendritic spine density in the basal dendrites of layer 2/3 excitatory neurons suggesting aberrant spine pruning. Monocular deprivation induces atypical ocular dominance plasticity in GLT1 HET mice, with an unusual depression of ipsilateral open eye responses; however, this change in ipsilateral responses correlates well with an upregulation of GLT1 protein following monocular deprivation. These results demonstrate that a key function of astrocytic GLT1 function during development is the experience-dependent refinement of ipsilateral eye inputs relative to contralateral eye inputs in visual cortex

Distinct prefrontal top-down circuits differentially modulate sensorimotor behavior

© 2020, The Author(s). Sensorimotor behaviors require processing of behaviorally relevant sensory cues and the ability to select appropriate responses from a vast behavioral repertoire. Modulation by the prefrontal cortex (PFC) is thought to be key for both processes, but the precise role of specific circuits remains unclear. We examined the sensorimotor function of anatomically distinct outputs from a subdivision of the mouse PFC, the anterior cingulate cortex (ACC). Using a visually guided two-choice behavioral paradigm with multiple cue-response mappings, we dissociated the sensory and motor response components of sensorimotor control. Projection-specific two-photon calcium imaging and optogenetic manipulations show that ACC outputs to the superior colliculus, a key midbrain structure for response selection, principally coordinate specific motor responses. Importantly, ACC outputs exert control by reducing the innate response bias of the superior colliculus. In contrast, ACC outputs to the visual cortex facilitate sensory processing of visual cues. Our results ascribe motor and sensory roles to ACC projections to the superior colliculus and the visual cortex and demonstrate for the first time a circuit motif for PFC function wherein anatomically non-overlapping output pathways coordinate complementary but distinct aspects of visual sensorimotor behavior

Noradrenergic signaling in the wakeful state inhibits microglial surveillance and synaptic plasticity in the mouse visual cortex

© 2019, The Author(s), under exclusive licence to Springer Nature America, Inc. Microglia are the brain’s resident innate immune cells and also have a role in synaptic plasticity. Microglial processes continuously survey the brain parenchyma, interact with synaptic elements and maintain tissue homeostasis. However, the mechanisms that control surveillance and its role in synaptic plasticity are poorly understood. Microglial dynamics in vivo have been primarily studied in anesthetized animals. Here we report that microglial surveillance and injury response are reduced in awake mice as compared to anesthetized mice, suggesting that arousal state modulates microglial function. Pharmacologic stimulation of β2-adrenergic receptors recapitulated these observations and disrupted experience-dependent plasticity, and these effects required the presence of β2-adrenergic receptors in microglia. These results indicate that microglial roles in surveillance and synaptic plasticity in the mouse brain are modulated by noradrenergic tone fluctuations between arousal states and emphasize the need to understand the effect of disruptions of adrenergic signaling in neurodevelopment and neuropathology