McHugh J, Mok W M, Wang G K, Strichartz G
Anesthesia Research Laboratories, Brigham and Women's Hospital, Harvard Medical School, Boston, Massachusetts 02115, USA.
J Gen Physiol. 1995 Feb;105(2):267-87. doi: 10.1085/jgp.105.2.267.
We have synthesized a model local anesthetic (LA), N-(2-di-N-butyl-aminoethyl)-4-azidobenzamide (DNB-AB), containing the photoactivatable aryl azido moiety, which is known to form a covalent bond to adjacent molecules when exposed to UV light (Fleet, G.W., J.R. Knowles, and R.R. Porter. 1972. Biochemical Journal. 128:499-508. Ji, T.H. 1979. Biochimica et Biophysica Acta. 559:39-69). We studied the effects of DNB-AB on the sodium current (INa) under whole-cell voltage clamp in clonal mammalian GH3 cells and on 3[H]-BTX-B binding to sheep brain synaptoneurosomes. In the absence of UV illumination, DNB-AB behaved similarly to known LAs, producing both reversible block of peak INa (IC50 = 26 microM, 20 degrees C) and reversible inhibition of 3[H]-BTX-B (50 nM in the presence of 0.12 microgram/liter Leiurus quinquestriatus scorpion venom) binding (IC50 = 3.3 microM, 37 degrees C), implying a noncovalent association between DNB-AB and its receptor(s). After exposure to UV light, both block of INa and inhibition of 3[H]-BTX-B binding were only partially reversible (INa = 42% of control; 3[H]-BTX-B binding = 23% of control) showing evidence of a light-dependent, covalent association between DNB-AB and its receptor(s). In the absence of drug, UV light had less effect on INa (post exposure INa = 96% of control) or on 3[H]-BTX-B binding (post exposure binding = 70% of control). The irreversible block of INa was partially protected by coincubation of DNB-AB with 1 mM bupivacaine (IC50 = 45 microM, for INa inhibition at 20 degrees C, Wang, G.K., and S.Y. Wang. 1992. Journal of General Physiology. 100:1003-1020), (post exposure INa = 73% of control). The irreversible inhibition of 3[H]-BTX-B binding also was partially protected by coincubation with bupivacaine (500 microM, 37 degrees C) (post exposure binding = 51% of control), suggesting that the site of irreversible inhibition of both INa and 3[H]-BTX-B binding is shared with the clinical LA bupivacaine.
我们合成了一种模型局部麻醉药（LA），即N-(2-二正丁基氨基乙基)-4-叠氮基苯甲酰胺（DNB-AB），它含有可光活化的芳基叠氮部分，已知该部分在紫外光照射下会与相邻分子形成共价键（Fleet, G.W., J.R. Knowles, and R.R. Porter. 1972. 《生物化学杂志学报》. 128:499 - 508. Ji, T.H. 1979. 《生物化学与生物物理学报》. 559:39 - 69）。我们研究了DNB-AB在克隆的哺乳动物GH3细胞的全细胞膜片钳记录模式下对钠电流（INa）的影响，以及对3[H]-BTX-B与羊脑突触神经小体结合的影响。在没有紫外线照射的情况下，DNB-AB的表现与已知的局部麻醉药相似，能使峰INa产生可逆性阻断（IC50 = 26 microM，20摄氏度），并对3[H]-BTX-B（在存在0.12微克/升的以色列金蝎毒液时为50 nM）的结合产生可逆性抑制（IC50 = 3.3 microM，37摄氏度），这意味着DNB-AB与其受体之间存在非共价结合。紫外线照射后，INa的阻断和3[H]-BTX-B结合的抑制仅部分可逆（INa = 对照的42%；3[H]-BTX-B结合 = 对照的23%），表明DNB-AB与其受体之间存在光依赖性的共价结合。在没有药物的情况下，紫外线对INa（照射后INa = 对照的96%）或3[H]-BTX-B结合（照射后结合 = 对照的70%）的影响较小。将DNB-AB与1 mM布比卡因共同孵育可部分保护INa的不可逆阻断（IC50 = 45 microM，用于20摄氏度时的INa抑制，Wang, G.K., and S.Y. Wang. 1992. 《普通生理学杂志》. 100:1003 - 1020），（照射后INa = 对照的73%）。将3[H]-BTX-B结合的不可逆抑制与布比卡因（500 microM，37摄氏度）共同孵育也可部分受到保护（照射后结合 = 对照的51%），这表明INa和3[H]-BTX-B结合的不可逆抑制位点与临床局部麻醉药布比卡因相同。
