Copper disrupts S-nitrosothiol signaling in activated BV2 microglia

• Copper inhibits GSNOR activity in activated BV2 microglia.
• Cu(I) reduces nitrite release and SNO proteins in LPS-treated BV2 microglia.
• SNO-formation is inhibited by 100 μM Cu(I) in activated BV2 microglia.

Microglia, the primary resident immune cells of the central nervous system (CNS), responds rapidly to pathogens and injury by secreting immune mediators including nitric oxide (NO). The reaction of NO with the anti-oxidant glutathione forms S-nitrosoglutathione (GSNO), the major pool of biologic NO in the body. GSNO is degraded by GSNO reductase (GSNOR). Recently, we have shown that copper (Cu(I)) inhibits the release of NO in lipopolysaccharide (LPS)-stimulated BV2 microglia and induces BV2 microglia to acquire a mixed a profile with both pro- and anti-inflammatory characteristics. Since GSNOR is the critical enzyme in GSNO metabolism, we sought to determine whether Cu(I) affects GSNOR activity and S-nitrosothiol (SNO) accumulation in activated BV2 microglia. Our results show that GSNOR protein expression is reduced by Cu(I) treatment in LPS-stimulated BV2 microglia. Our results also show a decrease in S-nitrosothiol content despite a reduced GSNOR expression. This effect is most likely due to Cu(I) reacting with the central thiol of the SNO bond resulting in the degradation of SNO. A dose of 1 μM Cu(I) did not affect SNO protein accumulation in LPS-stimulated BV2 microglia, however, a dose of 100 μM Cu(I) inhibited SNO protein in accordance with inhibition of S-nitrosothiols. These data provide direct evidence that Cu(I) disrupts S-nitrosothiol homeostasis and NO metabolism, and, thus, provide new insights into the mechanisms involved in microglia-mediated-CNS disorders.