CNSC-63. UNRAVELING MICROGLIAL CONTRIBUTIONS TO NEURON–GLIOMA SYNAPTIC INTERACTIONS IN GLIOBLASTOMA

Abstract Glioblastoma (GBM) is the most aggressive primary brain tumor, with a dismal median survival of 12–15 months. Recent evidence shows that neurons form functional synapses with GBM cells, promoting invasion and growth. Microglia, the brain’s resident immune cells, can modulate synaptic connections; however, their role in neuron-glioma synaptic interactions remains poorly understood. Here, we co-cultured patient-derived GBM cells with either human iPSC-derived microglia-like cells or HMC3 microglia in five-month-old iPSC-derived cortical organoids, assessing GBM calcium activity over 30 minutes using GCaMP6s-based imaging. Compared to GBM-only controls, cells grown with microglia displayed elevated mean ΔF/F0 (t-test, p = 6.536×10^-6,****) and higher calcium peak counts (p = 0.0002181 by t-test, ***). Tumor region- specific single-cell transcriptomic analysis of the Darmanis et. al dataset, containing GBM samples from four patients revealed eight distinct immune subclusters and highlighted a spatial dichotomy in immune function, with upregulation of complement and phagocytic signals in the tumor core, and unique ligand–receptor interactions in the periphery. CellChat analysis identified SEMA4D–PLXNB1 as a candidate ligand–receptor pair enriched between immune and neoplastic cells at the tumor periphery. Resident microglia exhibited significantly higher SEMA4D expression, while neoplastic cells showed elevated PLXNB1. These data suggest that microglia intensify GBM calcium signaling and engage in SEMA4D–PLXNB1 signaling. Future studies will determine the impact of these observations on neuron-gioma synapses and whether genetic ablation of PLXNB1 disrupts microglia-driven calcium signaling and slows tumor progression, offering mechanistic insight into the interplay of immune cells, neurons, and GBM.