Wu G, Miller K W
Department of Anesthesia, Massachusetts General Hospital, Boston 02114.
Biochemistry. 1994 Aug 9;33(31):9085-91. doi: 10.1021/bi00197a009.
The reversible decline of the nicotinic acetylcholine receptor's response to acetylcholine during prolonged exposure to acetylcholine is known as desensitization. Here, we studied ethanol's modulation of fast agonist-induced desensitization of the nicotinic acetylcholine receptor in postsynaptic membrane vesicles from Torpedo using a fast kinetic technique: pulsed quenched flow. Preincubation of the vesicles with various concentrations of acetylcholine at 4 degrees C for times ranging from 80 ms to 1.5 s caused fast desensitization, which was revealed as a decreased 86Rb+ influx when the vesicles were subsequently briefly exposed to a saturating concentration of acetylcholine in 86RbCl. Acetylcholine-induced fast desensitization had a maximum observed rate, kdmax, of 6.8 s-1, a half-effect concentration, KD, of 157 microM, and a Hill coefficient of 1.4. Increasing the ethanol concentration up to 1.0 M causes a linear increase in kdmax, such that 1.0 M ethanol doubles the rate. Ethanol (1 M) also decreased KD 10-fold without changing the Hill coefficient. We consider a modified sequential model to interpret our data. Two acetylcholine molecules bind sequentially to the receptor's resting state to form a pre-open (closed) state, which then opens and, at very high acetylcholine concentrations, is inhibited. A priori fast desensitization might occur from any of these acetylcholine-occupied states. If we assume fast desensitization to occur solely from the pre-open state, our data predict an excessively large action of ethanol on the fast desensitization rate constant (> 200-fold increase in the desensitization rate constant at 1 M ethanol). When we assume fast desensitization to occur from all states, ethanol is seen to have two actions.(ABSTRACT TRUNCATED AT 250 WORDS)
在长时间暴露于乙酰胆碱期间，烟碱型乙酰胆碱受体对乙酰胆碱的反应出现可逆性下降，这一现象被称为脱敏。在此，我们使用一种快速动力学技术——脉冲淬灭流，研究了乙醇对来自电鳐突触后膜囊泡中烟碱型乙酰胆碱受体快速激动剂诱导的脱敏的调节作用。在4℃下，将囊泡与不同浓度的乙酰胆碱预孵育80毫秒至1.5秒，会导致快速脱敏，这表现为随后将囊泡短暂暴露于86RbCl中饱和浓度的乙酰胆碱时，86Rb+内流减少。乙酰胆碱诱导的快速脱敏观察到的最大速率kdmax为6.8 s-1，半效应浓度KD为157 microM，希尔系数为1.4。将乙醇浓度增加至1.0 M会导致kdmax呈线性增加，使得1.0 M乙醇使速率翻倍。乙醇（1 M）还使KD降低了10倍，而希尔系数不变。我们考虑用一个修正的顺序模型来解释我们的数据。两个乙酰胆碱分子依次与受体的静息状态结合，形成一个预开放（关闭）状态，然后该状态开放，并且在非常高的乙酰胆碱浓度下受到抑制。先验地，快速脱敏可能发生在这些乙酰胆碱占据的任何状态。如果我们假设快速脱敏仅发生在预开放状态，我们的数据预测乙醇对快速脱敏速率常数的作用过大（在1 M乙醇时脱敏速率常数增加超过200倍）。当我们假设快速脱敏发生在所有状态时，乙醇表现出两种作用。（摘要截短于250字）
