Role of P2X receptors in synthesis and release of IL-1β during oxygen-glucose deprivation in rat hippocampus

Abstract

Objective To evaluate the role of P2X receptors in the synthesis and release of IL-1β during oxygen-glucose deprivation (OGD) in rat hippocampus. Methods Male SD rats weighing 150-200 g were anesthetized with ether and decapitated. The hippocampi were removed and sagittally sliced (400 μm thick) and placed in artificial cerebrospinal fluid (aCSF) aerated with 95% O2-5% CO2. One hundred and sixty hippocampal slices were randomly divided into 4 groups ( n = 40 each): Ⅰ control group (group C); Ⅱ OGD group; Ⅲ OGD +BBG group; Ⅳ OGD + anti-P2X4 group (group OP). In group C, the hippocampal slices were continously incubated with aCSF aerated with 95% O2-5% CO2 . In group OGD, the hippocampal slices were incubated with glucose-free aCSF and aerated with 95% N2-5%CO2 . In group OGD + BBG, the hippocampal slices were incubated with aCSF containning P2X7 receptor-specific antagonist G (BBG, final concentration 1 μmol/L) and aerated with 95% O2-5% CO2 for 20 min, then exposed to OGD, and BBG (final concentration 1 μ mol/L) was added in glucose-free aCSF. In group OP, the hippocampal slices were incubated with aCSF containning P2X4 receptor antibody (final concentration 1.5 μg/ml) and aerated with 95% O2-5% CO2 for 60 min, then exposed to OGD, and P2X4 receptor antibody (final concentration 1.5 μg/ml) was added in glucose-free aCSF. LDH and IL-1β release was detected before OGD and at 20, 40 and 60 min of OGD. Histological changes were observed using HE staining.Intracellular pro-IL-1β precursor protein expression was detected by Western blot at 60 min of OGD. Results LDH and IL-1β release and expression of intracellular pro-IL-1β precursor protein were signifcantly higher in the other groups than in group C ( P ＜ 0.05 ). Compared with group OGD, LDH and IL- 1 β release was signifcantly decreased, while the expression of intracellular pro-IL-1β precursor protein un-regulated in group OGD + BBG ( P ＜0.05), but no signifcant difference was found in the prarameters mentioned above in group OP ( P ＞ 0.05). Conclusion P2X7 receptor mediates the synthesis and release of IL-1β during OGD in rat hippocampus, but P2X4 receptor does not. Key words: Receptors, purinergic P2; Interleukin-1 beta; Hypoxia-ischemia,brain; Hippocampus