Project Summary: Project 2 - Linking Fast Timescale Neuron-Astrocyte Communication to Neural Circuit Function and Behavior A fundamental yet unresolved question in neuroscience is how non-neuronal cells communicate with the surrounding neurons, influence their function, and potentially affect animal behavior. Astrocytes are in a unique position to modulate neural circuit function. They are ubiquitous in all CNS regions, express receptors for neurotransmitters, neuromodulators, and neuropeptides, extend highly ramified processes that interact with synapses and other CNS elements, and can operate as a syncytium partly due to their gap junctional coupling. These structural and functional properties enable them to modulate synaptic plasticity and neuronal excitability. Indeed, experimental evidence from multiple species and CNS regions now suggests that astrocytes modulate neural circuit function and behavior on both slow and fast timescales. Nevertheless, precisely how astrocytes respond to the composite molecular signals in their environment and how their intricate excitation patterns influence neural circuit function on fast timescales (sub-seconds to minutes) remains unclear. This Project will test the hypothesis that the heterogeneity of astrocyte transients can be understood by the temporal integration of the time-varying molecular signals in their environment. Previous studies have also suggested that astrocytes operate in at least two different modes: 1) Individually, and 2) as a syncytium. Yet, the relevance of these various forms of chemical excitation for neural circuit function remains a mystery. This Project's second hypothesis is that the different activity modes serve distinct physiological roles, enabling astrocytes to influence neural circuits and behavior on different timescales. This Project proposes four major Aims to tackle these issues as part of a team initiative. Aim 1 will determine how molecular signaling by local neurons relates to astrocyte excitation. Aim 2 focuses on elucidating how neuromodulator signaling by projection neurons influences astrocyte activity. Aim 3 will determine how targeted manipulation of astrocyte function (e.g., their ability to detect, temporally integrate, communicate, or respond to extracellular signals) modulates their excitation patterns, neural circuit function, and behavior. Aim 4 will generate a multilayer, multilevel atlas of the investigated neuron-astrocyte circuits. These data will be acquired from a common set of mouse cortical regions involved in sensorimotor processing using a reward-based quantitative behavioral assay. Computational analyses and modeling of this data will be used to identify variables controlling astrocyte excitation, cell-intrinsic parameters constraining this activity, distinct activity modes, and neuronal properties affected by these astrocytic features. Together, the functional and anatomical studies of this Project will a) provide foundational information about how astrocytes (individually or as a syncytium) respond to, integrate, and modulate neural circuit function (Projects 1 and 2); b) guide the development of novel genetically encoded indicators and interventional tools to interrogate neuron-astrocyte circuits in vivo (Projects 2, 3, and 4); c) inform, test, and refine predictive neuron-astrocyte circuit models of sensorimotor processing (Projects 1, 2, and Data Science Resource Core).