Astrocytes are highly complex cells with hundreds of thousands of fine processes that contact and ensheathe neuronal synapses. These perisynaptic astrocyte processes actively participate in synaptic development and function by regulating neurotransmitter release, maintaining ion homeostasis, and modulating synaptic connectivity. Despite the importance of astrocytes in synaptic development and function, we know very little about the molecular and cellular mechanisms that control the establishment of complex astrocyte morphology and astrocyte-synapse interactions. In our preliminary experiments, we found that in the mouse visual cortex the establishment of the complex astrocyte morphology is a developmentally regulated process that occurs during a period of extensive synapse formation. Manipulation of visual experience, i.e. dark rearing mice during first three weeks of postnatal development, strongly stunts cortical astrocyte development, indicating that experience- dependent changes in synaptic connectivity can alter morphological maturation of astrocytes. How is the complex astrocyte morphology attained and remodeled? To mechanistically address this question, we developed a primary cortical neuron and astrocyte co-culture system that takes advantage of the following basic observation: Astrocytes cultured by themselves have a simple fibroblast-like morphology; however, contact with neurons, even for a short period of time, is sufficient to trigger extensive elaboration of the astrocytes. This morphological shift in astrocytes is primarily driven by direct neuronal contact, but not by soluble secreted factors. Using this system, we conducted a candidate-based genetic screen and identified that the astrocytic expression of neuroligin (NL) family cell-adhesion molecules (CAMs), NL1, NL2 and NL3 control neuronal adherence and morphological maturation of astrocytes in vitro and in vivo. Based on these findings, our objective here is to determine the functions of NLs in astrocyte development and astrocyte- synapse interactions. To do so, we will test three hypotheses: 1) Astrocytic NLs control astrocyte morphological complexity by mediating astrocyte-synapse association. 2) NLs perform these functions via their extracellular interactions with axonal/presynaptic neurexins and 3) via their critical intracellular domains that control cytoskeletal dynamics within astrocytes. These studies have the potential to significantly advance our understanding of the molecular basis of astrocyte development and astrocyte-synapse interactions. Moreover, the results obtained here will provide critical new avenues for studying the formation of the tripartite synapses and reveal a new paradigm through which astrocytic NLs control brain development, a process that may be critically impaired in neurological disorders.!

Astrocytes are the most abundant cells in our brains. Their functional and structural interactions with neuronal synapses are required for proper brain physiology. This study aims to uncover how neuroligin family cell adhesion molecules regulate the complex morphology and synapse association of astrocytes. The roles of neuroligins in astrocytes are largely unexplored and neglected. Because mutations in Neuroligins are linked to many neurological disorders, it is critical to understand the full functions of these proteins in all brain cells that can alter brain connectivity. !