蛋白激酶A激活和磷脂酶A_2抑制参与腺苷对大鼠肾髓袢升支粗段管周膜50 pS钾通道的易化效应(英文)

Involvement of protein kinase A activation and phospholipase A_2 inhibition in the adenosine-activated basolateral 50 pS K^+ channels in the thick ascending limb of the rat kidney

本文旨在研究蛋白激酶A (protein kinase A, PKA)和磷脂酶A2(phospholipase A2,PLA2)在腺苷对髓袢升支粗段管周膜50 pS钾通道刺激过程中的作用。解剖显微镜下分离髓袢升支粗段,膜片钳技术记录50 pS钾通道电流,免疫印迹技术测定PKA及PLA2磷酸化水平的蛋白表达情况。结果显示,环己基腺苷(cyclohexyladenosine, CHA,一种腺苷类似物)可增加50 pS钾通道的活性(P<0.05)。在PKA阻断剂H8的存在下,CHA不再增加50 pS钾通道的活性。在PLA2阻断剂AACOCF3存在下,CHA不再增加50 pS钾通道的活性。CHA可以增加髓袢升支粗段中PKA的磷酸化水平,而抑制PLA2的磷酸化水平(P<0.01)。在AACOCF3存在下,CHA依旧可以增加PKA的磷酸化水平(P<0.01);在H8存在下,CHA不再抑制PLA2的磷酸化水平(P>0.05)。以上结果提示,腺苷对髓袢升支粗段管周膜50 pS钾通道的刺激作用是通过先激活PKA,而后再抑制PLA2途径而实现的。

The present study was designed to investigate the role of protein kinase A （PKA） and phospholipase A2 （PLA2） in the stimulating effect of adenosine on the basolateral 50 pS K＋ channels in the thick ascending limb （TAL） of the rat kidney. Under the anatomic microscope, the TAL was dissected. The current of 50 PS K^＋ channels were recorded by patch clamp technology. The protein expression of phosphorylated PKA and phosphorylated PLA2 were examined by Western blot. The results showed that cyclohexyladenosine （CHA）, an analog of adenosine, increased the 50 pS K^＋ channel activity （P 〈 0.05）. In the presence of H8, an antagonist of PKA, CHA did not affect the 50 pS K^＋ channel activity. In the presence ;ofAACOCF3 （an antagonist of PLA2）, CHA did not further increase the 50 pS K^＋ channel activity. CHA increased phosphorylation level of PKA, whereas inhibited phosphorylation of PLA2 in the TAL of the rat kidney （P 〈 0.01）. Furthermore, after blocking the PLA2 with AACOCF3, CHA still increased the expression ofphosphorylated PKA. On the contrary, CHA did not obviously change the expression of phosphorylated PLA2 after H8 pretreatment. The results suggest that the stimulation ofbasolateral 50 pS K^＋ channels by CHA is mediated by the activation of PKA followed by the inhibition of PLA2 in the TAL of the rat kidney.