目的:通过建立星形胶质细胞氨中毒的模型,探讨瞬时受体电位M2 (TRPM2)阳离子通道在其中发挥的作用.方法:分离并培养小鼠原代星形胶质细胞.实验分为5组:对照组、氯化铵处理组、氯化铵+3-氨基苯甲酰胺(3-AB)处理组、氯化铵+PJ-34处理组和TRPM2基因敲除小鼠+氧化铵处理组.测定细胞的活性、caspase-3活性、细胞坏死和体积大小,以此来衡量氨中毒的程度.用全细胞膜片钳记录TRPM2通道电流变化.结果:氯化铵引起细胞肿胀伴随着细胞坏死.聚腺苷二磷酸核糖聚合酶(PARP)抑制剂3-AB和PJ-34抑制了氯化铵引起的阳离子电流,并减轻了氯化铵引起的相应细胞伤害.TRPM2基因敲除小鼠组细胞的伤害明显减轻(P＜0.01).结论:TRMP2通道的激活是细胞暴露于氯化铵后发生肿胀、坏死的必要步骤；氯化铵诱导的星形胶质细胞肿胀与坏死密切相关.%AIM:To study the effects of transient receptor potential melastatin 2 (TRPM2) cation channel on ammonia intoxication in astrocytes.METHODS:Primary astrocytes were isolated,cultured and divided into 5 groups:control group,ammonium chloride (NH4Cl) treatment group,NH4Cl + 3-aminobenzamide (3-AB) treatment group,NH4Cl + PJ-34 treatment group,and TRPM2 knockout + NH4Cl treatment group.Cell viability,caspase-3 activity,cell necrosis and cell volume were measured to assess the extent of ammonia toxicity.Whole-cell patch-clamp was performed to record the currents via TRPM2 channels.RESULTS:NH4Cl caused cell swelling accompanied with cell necrosis.The poly(ADP-ribose) polymerase (PARP) inhibitors,3-AB and PJ-34,inhibited the cation currents activated by NH4Cl,and attenuated NH4 Cl-induced cell damage.In TRPM2-deficient astrocytes,decreased cell damage was observed.CONCLUSION:TRPM2 activation is essential for cell swelling and necrosis in astrocytes exposed to NH4Cl.NH4 Cl-triggered astrocyte swelling is closely correlated with necrosis.
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