Toll‐like receptor activation reveals developmental reorganization and unmasks responder subsets of microglia

Abstract

The sentinel and immune functions of microglia require rapid and appropriate reactions to infection and damage. Their Toll-like receptors (TLRs) sense both as threats. However, whether activated microglia mount uniform responses or whether subsets conduct selective tasks is unknown. We demonstrate that murine microglia reorganize their responses to TLR activations postnatally and that this process comes with a maturation of TLR4-organized functions. Although induction of MHCI for antigen presentation remains as a pan-populational feature, synthesis of TNFα becomes restricted to a subset, even within adult central nervous system regions. Response heterogeneity is evident ex vivo, in situ, and in vivo, but is not limited to TNFα production or to TLR-triggered functions. Also, clearance activities for myelin under physiological and pathophysiological conditions, IFNγ-enforced upregulation of MHCII, or challenged inductions of other proinflammatory factors reveal dissimilar microglial contributions. Notably, response heterogeneity is also confirmed in human brain tissue. Our findings suggest that microglia divide by constitutive and inducible capacities. Privileged production of inflammatory mediators assigns a master control to subsets. Sequestration of clearance of endogenous material versus antigen presentation in exclusive compartments can separate potentially interfering functions. Finally, subsets rather than a uniform population of microglia may assemble the reactive phenotypes in responses during infection, injury, and rebuilding, warranting consideration in experimental manipulation and therapeutic strategies.

Experimental Cortical Spreading Depression Induces NMDA Receptor Dependent Potassium Currents in Microglia

“…We never did observe any response to exposure of NMDA (n ϭ 23; Fig. 8A), whereas ATP always triggered an increase in membrane currents as previously described (Walz et al, 1993). We also performed calcium imaging with Fluo-4 loaded adult microglia in vitro with the same wash-in protocol.…”

“…Primary adult microglia cultures were prepared as described before using P49 -P56 mice (Scheffel et al, 2012). In short, the cerebellum and olfactory bulb were removed and the cerebrum was freed of blood vessels and meninges and stored in HBSS.…”

The subpopulation of microglia sensitive to neurotransmitters/neurohormones is modulated by stimulation with LPS, interferon-γ, and IL-4

“…Microglia were also cultured from adult mice (P49-56) as previously described in detail (Scheffel et al 2012). Briefly, cortical and midbrain tissue was freed of blood vessels and meninges in Hank's balanced salt solution (HBBS), mechanically dissociated into 1mm 3 pieces and trypsinized in 1% trypsin and 0.05% deoxyribonuclease for 5 min at room temperature, as described for neonatal microglia elsewhere (Giulian and Baker 1986).…”

Resistance of the Brain to Escherichia coli K1 Infection Depends on MyD88 Signaling and the Contribution of Neutrophils and Monocytes

“…Animal experiments were approved by the Animal Care Committee of the University Hospital of Göttingen and by the Niedersächsische Landesamt für Verbraucherschutz und Lebensmittelsicherheit (LAVES), Braunschweig, Lower Saxony, Germany. Meningitis was induced by injection of 3 ϫ 10 3 to 5.5 ϫ 10 3 CFU of E. coli K1 into the right frontal neocortex (18,19) under intraperitoneal anesthesia with ketamine (100 mg/kg of body weight) and xylazine (10 mg/kg of body weight). Mice (2 to 3 months old; weight, 20 to 30 g) were weighed daily and scored clinically (0, no apparent behavioral abnormality; 1, moderate lethargy; 2, severe lethargy; 3, unable to walk; 4, dead) (19).…”

“…The sensitivity of the assays for these cytokines/chemokines was 7.5 pg/ml. ELISAs were performed according to the manufacturer's instructions (12,18).…”