Hirota K, Lambert D G
University Department of Anaesthesia and Pain Management, Leicester Royal Infirmary, UK.
Can J Anaesth. 2000 May;47(5):467-70. doi: 10.1007/BF03018979.
The cellular target site(s) for anesthetic action remain controversial. In this study we have examined any interaction of i.v. anesthetics (thiopental, pentobarbital, ketamine, etomidate, propofol, alphaxalone), local anesthetics (lidocaine, prilocaine, procaine and tetracaine), and the non anesthetic barbiturate, barbituric acid with the omega-conotoxin MVII(A) binding site on N-type voltage sensitive Ca2+ channels in rat cerebrocortical membranes.
[125I] omega-conotoxin MVII(A) binding assays were performed in 0.5 ml volumes of Tris.HCl buffer containing BSA 0.1% for 30 min at 20 degrees C using fresh cerebrocortical membranes (5 microg of protein). Non-specific binding was defined in the presence of excess (10(-8) M) omega-conotoxin MVII(A). The interaction of i.v. (alphaxolone, etomidate, propofol, pentobarbitone, ketamine and thiopentone), local (lidocaine, prilocaine, procaine and tetracaine) anesthetics and barbituric acid was determined by displacement of [125I] omega-conotoxin MVII(A) (approximately 1 pM).
The binding of [125I] omega-conotoxin was concentration-dependent and saturable with Bmax and Kd of 223 +/- 15 fmol/mg protein and 2.13 +/- 0.14 pM, respectively. Unlabelled omega-conotoxin MVII(A) displaced [125I] omega-conotoxin MVII(A) yielding a pKd of 11.04 +/- 0.04 (9.2 pM). All i.v. and local anesthetics at clinically relevant concentrations did not show any interaction with the omega-conotoxin MVII(A) binding site.
The present study suggests that omega-conotoxin MVII(A) binding site on N-type voltage sensitive Ca2+ channels may not be a target for i.v. and local anesthetic agents.
麻醉作用的细胞靶点仍存在争议。在本研究中，我们检测了静脉麻醉药（硫喷妥钠、戊巴比妥、氯胺酮、依托咪酯、丙泊酚、alphaxalone）、局部麻醉药（利多卡因、丙胺卡因、普鲁卡因和丁卡因）以及非麻醉性巴比妥酸盐巴比妥酸与大鼠大脑皮质膜上N型电压敏感性Ca2+通道上的ω-芋螺毒素MVII(A)结合位点之间的相互作用。
在含有0.1%牛血清白蛋白的Tris.HCl缓冲液中，使用新鲜大脑皮质膜（5微克蛋白质），于20℃下进行30分钟的[125I]ω-芋螺毒素MVII(A)结合试验，反应体积为0.5毫升。非特异性结合通过在过量（10(-8)M）ω-芋螺毒素MVII(A)存在的情况下进行测定。静脉麻醉药（alphaxolone、依托咪酯、丙泊酚、戊巴比妥、氯胺酮和硫喷妥钠）、局部麻醉药（利多卡因、丙胺卡因、普鲁卡因和丁卡因）以及巴比妥酸的相互作用通过[125I]ω-芋螺毒素MVII(A)（约1皮摩尔）的置换来确定。
[125I]ω-芋螺毒素的结合呈浓度依赖性且可饱和，Bmax和Kd分别为223±15飞摩尔/毫克蛋白质和2.13±0.14皮摩尔。未标记的ω-芋螺毒素MVII(A)置换[125I]ω-芋螺毒素MVII(A)，产生的pKd为11.04±0.04（9.2皮摩尔）。所有临床相关浓度的静脉麻醉药和局部麻醉药均未显示与ω-芋螺毒素MVII(A)结合位点有任何相互作用。
本研究表明，N型电压敏感性Ca2+通道上的ω-芋螺毒素MVII(A)结合位点可能不是静脉麻醉药和局部麻醉药的作用靶点。
