Nalmefene Prevents Alcohol-Induced Neuroinflammation and Alcohol Drinking Preference in Adolescent Female Mice: Role of TLR4

Background: We previously showed that, by activating innate immune receptors Toll-like 4 (TLR4), adolescent intermittent ethanol (EtOH) exposure causes neuroinflammation, myelin damage, and behavioral dysfunctions. Recent findings reveal that clinically used opioid antagonists naltrexone (NT) and naloxone (NX) inhibit opioid-induced TLR4 signaling and that NT, NX, and nalmefene (NF), the 6-methylene derivative of NX, are able to reduce alcohol drinking escalation. Methods: NF (0.1 mg/kg, intraperitoneally) was injected 1 hour prior to EtOH (3 g/kg, intraperitoneally) following intermittent treatment in female (PND35) adolescent mice. Inflammatory molecules, myelin proteins, and apoptotic markers were assessed in the prefrontal cortex (PFC) and striatum/nucleus accumbens (STR/NAcc). The effect of NF on alcohol drinking preference was evaluated in both the wild-type and TLR4 knockout (KO) adolescent mice. Using astroglial cells, the inhibitory potential of NT, NX, and NF on lipopolysaccharide (LPS), or the EtOH-triggered TLR4 response, was compared. Results: Our findings indicate that NF prevents the up-regulation of cytokines (IL-1b, IL-17A, TNF-alpha), chemokines (MCP-1, MIP-1, KC), and pro-inflammatory mediators (iNOS, COX-2), along with myelin damage and apoptotic events, in both PFC and STR/NAcc. NF also abolishes EtOH-induced escalation of alcohol preference/consumption, but has no effect when administered to TLR4-KO mice. In vitro experiments indicate that NX and NF inhibit TLR4 activation upon LPS or EtOH stimulation. Immunofluorescence studies and lipid rafts isolation show that NF is able to prevent TLR4 translocation to lipid rafts/caveolae in astrocytes. Conclusions: These results suggest that NF prevents neuroinflammation and brain damage by blocking the TLR4 response and also support the role of central pro-inflammatory immune signaling in the modulation of alcohol consumption/addiction.