Toll-Like Receptor 9 Is Required for Opioid-Induced Microglia Apoptosis

Opioids have been widely applied in clinics as one of the most potent pain relievers for centuries, but their abuse has deleterious physiological effects beyond addiction. However, the underlying mechanism by which microglia in response to opioids remains largely unknown. Here we show that morphine induces the expression of Toll-like receptor 9 (TLR9), a key mediator of innate immunity and inflammation. Interestingly, TLR9 deficiency significantly inhibited morphine-induced apoptosis in microglia. Similar results were obtained when endogenous TLR9 expression was suppressed by the TLR9 inhibitor CpGODN. Inhibition of p38 MAPK by its specific inhibitor SB203580 attenuated morphine-induced microglia apoptosis in wild type microglia. Morphine caused a dramatic decrease in Bcl-2 level but increase in Bax level in wild type microglia, but not in TLR9 deficient microglia. In addition, morphine treatment failed to induce an increased levels of phosphorylated p38 MAPK and MAP kinase kinase 3/6 (MKK3/6), the upstream MAPK kinase of p38 MAPK, in either TLR9 deficient or mu-opioid receptor (mu OR) deficient primary microglia, suggesting an involvement of MAPK and mu OR in morphine-mediated TLR9 signaling. Moreover, morphine-induced TLR9 expression and microglia apoptosis appears to require mu OR. Collectively, these results reveal that opioids prime microglia to undergo apoptosis through TLR9 and mu OR as well. Taken together, our data suggest that inhibition of TLR9 and/or blockage of mu OR is capable of preventing opioid-induced brain damage.