Disentangling neuroimmune landscapes across peripheral activation paradigms resolves divergent glial state programs

Lipopolysaccharide (LPS), a gram-negative bacterial cell-wall component, is a well-characterized immunostimulant acting via Toll-like receptor 4 (TLR4) and is widely used to model systemic inflammation-to-brain immune signaling. A single intraperitoneal high dose evokes a robust peripheral inflammatory state that is rapidly relayed to the central nervous system (CNS), resulting in profound neuroinflammation. By contrast, repeated low-dose LPS engages innate immune memory and has been associated with neuroprotective effects. Here we sought to comprehensively characterize the CNS-specific effects of the repeated LPS regimen relative to a neurotoxic single high-dose challenge. High-dose LPS induced robust forebrain inflammatory cytokine expression, disrupted homeostatic microglial and astrocytic marker programs, and produced transcriptomic signatures enriched for NF-κB signaling and apoptosis. In striking contrast, repeated low-dose LPS preserved homeostatic glial marker expression while increasing IBA1/F4/80-positive microglial signal across forebrain regions without a parallel increase in inflammatory cytokine transcripts, demonstrating a dissociation between gliosis marker intensity and inflammatory pathway activation. Whole-forebrain RNA-seq demonstrated selective enrichment of phagocytosis-related pathways under the repeated regimen in the absence of pro-inflammatory transcriptional amplification. Flow cytometry revealed an expansion of CD45^high CD11b⁺ myeloid cells expressing the phagocytic marker CD206 following repeated low-dose LPS. Cell-type-resolved transcriptional profiling showed that this CD45^high CD11b⁺ subset preferentially upregulated phagocytic programs while lacking prominent pro-inflammatory and apoptotic pathway activation. In parallel, astrocytes maintained homeostatic gene expression without enrichment of neurotoxic inflammatory signatures. Together, these findings delineate how distinct systemic LPS dosing paradigms differentially shape glial transcriptional and phenotypic responses in the CNS.