Figure 4

Exogenous CRT mediates a dominant inhibitory effect on primary phagocytosis by acting on microglia. (A + B) Co-cultures of cerebellar neurons and glia were incubated with LPS for 72 hours in the presence or absence of 2 μg/ml exogenous CRT prior to quantification of neuronal survival (A) and microglial activation as measured by nitrite levels in culture supernatant at 72 hour timepoint (B). (C + D) Co-cultures were incubated with Aβ peptide for 72 hours in the presence or absence of exogenous CRT prior to quantification of neuronal survival (C) and microglial density (D). (E) Exogenous CRT inhibits LPS-induced primary phagocytosis when pre-incubated with microglia but not with primed neurons. CRT was added to either neurons or microglia separately, washed out and then microglia were added to neurons for six hours prior to quantification of neuronal survival (E). (F) Co-cultures were treated with LME to remove microglia prior to addition of exogenous CRT where indicated. Pure microglia, either unactivated or LPS-activated, were then added back to neurons for six hours prior to quantification of neuronal survival. (A-E) Data represent three experiments with two replicates per experiment. (F) Data represent six experiments with two replicates per experiment. All error bars represent SEM. Aβ, amyloid-β peptide_1-42; CRT, calreticulin; LME, L-leucine methyl ester; LPS, lipopolysaccharide.