Astrocyte Network Facilitation of Neuroplasticity

/Abstract Astrocyte gap junctions are necessary for memory formation, synaptic plasticity, coordination of neuronal signaling, and closing the visual and motor critical periods; we also know signaling through astrocyte gap junctions can drive changes between behavioral states. Studies on networks of gap-junction-connected astrocytes have been limited to in vitro methods, slice electrophysiology, and pan-astrocyte knockouts. However, these methods cannot deeply examine the complexity and function of distinct, 3D, in vivo astrocyte networks, and largely limit mechanistic work to calcium dynamics. However, studies in other model systems have found that gap junctions flux many molecules known to facilitate neuroplasticity – especially connexin 43 (cx43), which has the largest pore limit and is the predominant astrocyte connexin. To address this gap in neuroscience, I designed and built a vector that marks functional astrocyte networks by biotinylating molecules fluxed by an infected astrocyte's gap junctions. Using a tissue-clearing method I optimized to preserve astrocyte morphology, I was able to map functional astrocyte network communication across whole, cleared brains. I found that there are many independent astrocyte networks across the brain, each connecting specific brain regions rather than diffusing indiscriminately. Through mass spectrometry, I also identified over 200 biotinylated metabolites, antioxidants, and peptides that traverse astrocyte networks. I will now build on these findings to test a central hypothesis: astrocyte networks circulate pools of antioxidants that buffer oxidatively stressed local environments and facilitate neuroplasticity. I will study neuroplasticity in adult mouse barrel cortex induced by whisker trim, a robust model of neuronal remodeling. Using my vector, I found that astrocyte networks usually connect several brain regions. However, barrel cortex astrocyte networks are almost exclusively limited to barrel cortex itself. This makes barrel cortex an optimal system to establish the fundamental organizational and mechanistic properties of astrocyte networks. Here, I first use state-of-the-art spatial transcriptomics and mass spectrometry imaging to establish the fundamental organizational properties of astrocyte networks. I then use chemogenetics in neurons to challenge broad astrocyte networks to functionally adapt to extremes in local environments and determine mechanisms by which astrocyte networks support the varied needs of the multiple brain regions they traverse. Finally, the first experiments in my independent laboratory will manipulate astrocyte network wide antioxidant pools to determine if these pools can rescue neuronal remodeling during periods of oxidative stress, then eliminate astrocyte networks in the brain to determine if antioxidant support is the sole way astrocyte networks support remodeling neurons. Together, these experiments provide a foundational understanding of astrocytic networks while exposing mechanisms necessary for neuronal plasticity to occur. Project Number: 1K99NS139313-01A1 | Fiscal Year: 2025 | NIH Institute/Center: National Institute of Neurological Disorders and Stroke (NINDS) | Principal Investigator: Melissa Cooper | Institution: NEW YORK UNIVERSITY SCHOOL OF MEDICINE, NEW YORK, NY | Award Amount: $141,102 | Activity Code: K99 | Study Section: Special Emphasis Panel[ZMH1 ERB-G (01)] View on NIH RePORTER: https://reporter.nih.gov/project-details/11138214