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Updated: Oct 9, 2026
Stab-Wound Mouse Model for Studying Hemorrhage and Inflammation in Traumatic Brain Injury
Published on: February 21, 2025
Microglia at brain stab wounds express connexin 43 and in vitro form functional gap junctions after treatment with
E A Eugenín1, D Eckardt, M Theis
1Departamento de Ciencias Fisiológicas, Facultad de Ciencias Biológicas, Pontificia Universidad Católica de Chile, Santiago 340, Chile.
Abstract:
Gap junctional communication between microglia was investigated at rat brain stab wounds and in primary cultures of rat and mouse cells. Under resting conditions, rat microglia (FITC-isolectin-B4-reactive cells) were sparsely distributed in the neocortex, and most (95%) were not immunoreactive for Cx43, a gap junction protein subunit. At brain stab wounds, microglia progressively accumulated over several days and formed aggregates that frequently showed Cx43 immunoreactivity at interfaces between cells. In primary culture, microglia showed low levels of Cx43 determined by Western blotting, diffuse intracellular Cx43 immunoreactivity, and a low incidence of dye coupling. Treatment with the immunostimulant bacterial lipopolysaccharide (LPS) or the cytokines interferon-gamma (INF-gamma) or tumor necrosis factor-alpha (TNF-alpha) one at a time did not increase the incidence of dye coupling. However, microglia treated with INF-gamma plus LPS showed a dramatic increase in dye coupling that was prevented by coapplication of an anti-TNF-alpha antibody, suggesting the release and autocrine action of TNF-alpha. Treatment with INF-gamma plus TNF-alpha also greatly increased the incidence of dye coupling and the Cx43 levels with translocation of Cx43 to cell-cell contacts. The cytokine-induced dye coupling was reversibly inhibited by 18 alpha-glycyrrhetinic acid, a gap junction blocker. Cultured mouse microglia also expressed Cx43 and developed dye coupling upon treatment with cytokines, but microglia from homozygous Cx43-deficient mice did not develop significant dye coupling after treatment with either INF-gamma plus LPS or INF-gamma plus TNF-alpha. This report demonstrates that microglia can communicate with each other through gap junctions that are induced by inflammatory cytokines, a process that may be important in the elaboration of the inflammatory response.
Insights
Microglia communicate via gap junctions, especially during brain inflammation. Inflammatory cytokines induce connexin 43 (Cx43) expression and enhance microglial communication, crucial for immune responses.
Area of Science:
- Neuroscience
- Immunology
- Cell Biology
Background:
- Microglia are the primary immune cells of the central nervous system.
- Gap junctional communication (GJC) is essential for cell-to-cell signaling.
- The role of GJC in microglial inflammatory responses is not well understood.
Purpose of the Study:
- To investigate GJC in microglia under physiological and pathological conditions.
- To determine the role of inflammatory cytokines in regulating microglial GJC.
- To elucidate the involvement of connexin 43 (Cx43) in microglial communication.
Main Methods:
- Examined Cx43 expression and dye coupling in rat and mouse microglia in vivo and in primary cultures.
- Utilized Western blotting and immunofluorescence for Cx43 detection.
- Assessed GJC using dye transfer assays and manipulated cytokine/LPS treatments and Cx43 deficiency.
Main Results:
- Microglia exhibit low basal Cx43 expression and GJC.
- Brain injury induces microglial aggregation and increased Cx43 at cell interfaces.
- Inflammatory cytokines (IFN-γ, TNF-α) combined with LPS or with each other significantly enhance microglial GJC and Cx43 levels.
- Cytokine-induced GJC is dependent on TNF-α and Cx43, and is inhibited by a gap junction blocker.
- Cx43-deficient microglia show impaired GJC induction.
Conclusions:
- Microglia communicate through gap junctions.
- Inflammatory cytokines induce and enhance microglial GJC via Cx43.
- This cytokine-mediated GJC is a critical component of the microglial inflammatory response in the brain.
