Inhibitory Effect Of Naloxone On Lipopolysaccharide(LPS)-Induced Retinal Microglial Activation And The Underlying Mechanism

Purpose:To investigate the effects of naloxone on lipopolysaccharide (LPS)-induced retinal microglial activation and the underlying mechanism.Methods:Primary cultured microglial cells were isolated from the retinas of Sprague-Dawley rats and activated LPS; in the experimental groups the cells were treated with various concentrations of naloxone (0.5μM,1.0μM and2.0μM)30min before addition of LPS. The culture supernatants were collected12h after LPS treatment to determine the levels of tumour necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) by ELISA assays. Western blot analysis and double immunofluorescence (0X42and P-p38) were used to examine the activation of the p38Mitogen-Activated Protein Kinase (p38MAPK) Signaling Pathway. The cultured microglial cells were also pretreated with2.5μM and5.0μM of SB203580, a selective inhibitor of the p38MAPK signalling pathway,12h before treatments with naloxone and/or LPS. Then, ELISA analysis was used to measure concentrations of TNF-α and IL-1β again.Results:LPS triggered a robust increase of proinflammatory and neurotoxic factors (IL-1β and TNF-a) in the supernatants. Naloxone inhibited LPS-induced microglial production of IL-1β and TNF-α. The activation of the p38MAPK pathway was inhibited by naloxone in a dose-dependent pattern, which was further supported by the fact that naloxone failed to change TNF-α and IL-1β production in microglia pretreated with the p38MAPK inhibitor SB203580. However, naloxone showed no effects on the levels of extracellular signal-regulated kinases (ERK) and c-Jun amino-terminal kinases (JNK).Conclusions:Naloxone might have the potential of neuro-protection by suppressing microglial activation in the retina, and this suppression occurs partly through the p38MAPK signalling pathway.