Okamoto Hiroyuki, Unno Toshihiro, Arima Daisuke, Suzuki Maki, Yan Hai-Dun, Matsuyama Hayato, Nishimura Masakazu, Komori Seiichi
United Graduate School of Veterinary Science, Gifu University, Gifu, Japan.
J Pharmacol Sci. 2004 Jun;95(2):203-13. doi: 10.1254/jphs.fp0030635.
In guinea pig single ileal smooth muscle cells held under voltage-clamp, the role of phospholipase C (PLC) in activation of the muscarinic receptor-operated cationic current (I(cat)) was studied. U73122, a PLC inhibitor, prevented the generation of I(cat) by the muscarinic agonist carbachol. The effect did not involve muscarinic receptor block since it also blocked I(cat) which was evoked by GTPgammaS applied intracellularly to activate G proteins bypassing muscarinic receptors. Also, neither cationic channel block nor other possible nonspecific actions seemed to be involved since its analogue (U73343), structurally close but deficient of the PLC-inhibiting activity, did not significantly affect carbachol- or GTPgammaS-evoked I(cat). Antibodies against the alpha subunits of G(q)/G(11) proteins (Galpha(q)/Galpha(11)-antibody) blocked only the small component of carbachol-evoked I(cat), which was associated with an increase in Ca(2+) linked to an increase in G(q/11) protein-regulated PLC activity. 1-Oleoyl-2-acetyl-sn-glycerol (OAG), an analogue of diacylglycerol (DAG) produced via PLC-catalyzed metabolism, produced no or only a small current by itself, with the carbachol-evoked I(cat) remaining unchanged. These results provide evidence for the importance of PLC in I(cat) generation, and they also strongly suggest that the activity of PLC involved in the primary activation of I(cat) is neither under regulation by G(q/11) proteins nor dependent on the action of DAG.
在电压钳制下的豚鼠单个回肠平滑肌细胞中，研究了磷脂酶C（PLC）在毒蕈碱受体介导的阳离子电流（I(cat)）激活中的作用。PLC抑制剂U73122可阻止毒蕈碱激动剂卡巴胆碱产生I(cat)。该作用不涉及毒蕈碱受体阻断），因为它也阻断了通过细胞内应用GTPγS激活G蛋白从而绕过毒蕈碱受体所诱发的I(cat)。此外，似乎既不涉及阳离子通道阻断也不涉及其他可能的非特异性作用，因为其类似物（U73343）结构相近但缺乏PLC抑制活性，对卡巴胆碱或GTPγS诱发的I(cat)没有显著影响。针对G(q)/G(11)蛋白α亚基的抗体（Gα(q)/Gα(11)抗体）仅阻断了卡巴胆碱诱发的I(cat)的小部分，这与[Ca(2+)]i的增加有关，而[Ca(2+)]i的增加与G(q/11)蛋白调节的PLC活性增加有关。1-油酰基-2-乙酰基-sn-甘油（OAG）是通过PLC催化代谢产生的二酰基甘油（DAG）的类似物，其自身产生的电流很小或几乎没有，卡巴胆碱诱发的I(cat)保持不变。这些结果证明了PLC在I(cat)产生中的重要性，并且还强烈表明参与I(cat)初始激活的PLC活性既不受G(q/11)蛋白调节，也不依赖于DAG的作用。
