In primary visual cortex (V1), precise spatiotemporal neuronal responses are known to underlie visual processing. Though neuronal roles in visual processing have been well studied, the role of non-neuronal cells, particularly astrocytes, in cortical synapses and circuits remains poorly understood. Cortical astrocytes contact and ensheathe nearly all excitatory synapses, creating discrete functional units consisting of a presynaptic input, a postsynaptic spine and an astrocyte process. A crucial function of astrocytes is the active uptake of glutamate from the synaptic cleft via transporters, particularly GLT1. We propose that astrocytes contribute fundamentally to V1 circuits via GLT1 activity, actively shaping synaptic and neuronal response profiles. Focal Ca2+ transients potentially related to synaptic glutamate uptake have recently been demonstrated within astrocyte processes, and synaptic transmission shown to actively recruit astrocytic GLT1 to sites of synaptic activity. Novel high- resolution imaging techniques, together with cell-specific markers, new optical probes, and genetically engineered mice with specific temporal and spatial control of protein expression, enable us to analyze the crosstalk between astrocyte and neuronal activity at unprecedented resolution in awake mice in vivo.
We aim to take advantage of the exquisite organization of visual inputs to V1 neurons to examine the interaction of Ca2+ microdomains, mitochondria and glutamate transporters in astrocyte processes, the functional contribution of astrocyte transporters to neuronal synapses and circuits during visual processing, and the impact of altered glutamate transport on the development and plasticity of V1 circuits.
In Aim 1, we will examine astrocyte microdomain Ca2+ responses to visual stimuli, including orientation-specific gratings and complex natural images, their relationship to mitochondria, and how genetic or pharmacological reduction of GLT1 impacts the specificity and reliability of astrocyte and cell-specific neuronal responses.
In Aim 2, we will determine the functional relationship between single dendritic spines and adjacent astrocytic processes using simultaneous dual-wavelength imaging of astrocytes and neurons during visual stimulation. We will also determine how GLT1 reduction affects astrocytic process and neuronal spine responses and structures.
In Aim 3, we will determine the role of GLT1 in the development and plasticity of astrocyte responses and visual cortex circuits. We will examine how germline reduction of GLT1 alters neuronal and astrocyte microdomain responses during normal development and following monocular deprivation, along with the sharpening of orientation selectivity and the binocular matching of orientation preference. Our overarching goal is to critically examine the hypothesis that astrocytes and their transporters are integral functional partners with neurons in the function and development of cortical circuits. As such, an understanding of normal and abnormal function in a host of neurodevelopmental and neurodegenerative disorders will require incorporating the role of astrocytes.
Neuronal networks are the focus of most research into cortical function; however, astrocytes comprise a significant proportion of cells in the cerebral cortex and their role is information processing remains poorly understood. In this project, we will use high-resolution imaging methods in visual cortex of mice to characterize visually evoked Ca2+ responses within astrocyte processes and correlate them with neuronal responses; additionally, we will examine how these responses are modulated by glutamate uptake through astrocyte transporters, their relationship to mitochondria, and how glutamate transporter function impacts the development of visual cortex circuits. These studies will provide critical information for understanding the role of astrocytes in normal function and brain disorders.