谷氨酸损伤大鼠皮层神经元及抑制钙/钙调蛋白依赖性蛋白激酶Ⅱ活性

Inhibitory effect of glutamate on calcium/calmodulin- dependent protein kinase Ⅱ and damaging effect on cultured rat cortical neurons

利用原代培养的大鼠胎鼠皮层神经元 ,采用乳酸脱氢酶 (LDH)测定和32 P参入方法 ,研究谷氨酸 (Glu)对皮层神经元损伤 ,钙 /钙调蛋白依赖性蛋白激酶 (Ca MK )活性的影响及其机理 .Glu(50- 1 0 0 0μmol· L-1)作用 1 0 min,导致皮层神经元Ca MK 活性立即明显下降 ,神经元形态及 LDH释放早期无明显改变 ,2 4 h时 LDH释放显著增加 ,神经元损伤 .N-甲基 - D-天冬氨酸 (NMDA)受体拮抗剂地佐环平 (MK- 80 1 )明显降低 LDH释放 ,保护Ca MK 活性 ,而非 NMDA受体拮抗剂 6,7-二硝基喹口恶啉二酮 (DNQX)无保护作用 .去除胞外 Ca2 +可明显保护 Ca MK 活性 .Ca MK 抑制剂 KN- 62可明显拮抗 Ca MK 活性下降 ,部分保护神经元损伤 .结果提示 ,Glu兴奋毒引起皮层神经元迟发性损伤和 Ca MK 活性早期明显下降 ,主要由 NMDA受体介导和胞外 Ca2 +内流增加有关 .

In cultured rat cortical neurons, the damaging effects and mechanisms of glutamate (Glu) on neurons and calcium/calmodulin-dependent protein kinase Ⅱ (CaMKⅡ) activty were studied. The activity of CaMKⅡ, measured by 32P incorporation, decreased immediately after exposure to Glu (50-1000 μmol·L -1, 10 min), while the morphology and lactate dehydrogenase(LDH) release did not change at 1 h until at 24 h. MK-801 significantly reduced the LDH release, protected the activity of CaMKⅡ, while 6,7-dinitroquinoxaline-2,3(1H,4H)-dione(DNQX) had no effect. Removing extracellular Ca 2+ could protect the activity of CaMKⅡsignificantly. KN-62〔1-(N,O-bis[5-isoquinolinesulfonyl]-N-methyl-L-tyrosyl)-4-phenylpiperazine〕 also protected the activity of CaMKⅡand had partial protection on LDH releasing. The results suggest that the NMDA receptor play an important role in the excitotoxicity of Glu causing delayed neuronal death and the decreased CaMKⅡactivity.