Aguilar J S, Fonseca M I, Lunt G G
Department of Biochemistry, University of Bath, U.K.
Neurochem Res. 1989 Aug;14(8):765-70. doi: 10.1007/BF00964956.
We have compared the effect of ethanol, a membrane perturbant, on the muscarinic binding sites in neural membranes from a vertebrate (rat) and an insect (locust). The binding of the muscarinic antagonist [3H]quinuclidinyl benzilate ([3H]QNB) to both rat and locust neural membranes was inhibited by ethanol at 10-500 mM concentrations; but this inhibition was greater in the locust. Ethanol (500 mM) increased the apparent dissociation constant (KD') of [3H]QNB binding to rat membranes from 0.13 +/- 0.01 nM in control to 0.20 +/- 0.02 nM; there was also an small but significant reduction in the number of binding sites Bmax. In locust, 500 mM ethanol reduced the Bmax of [3H]QNB binding from 590 +/- 30 in control to 320 +/- 40 pmol/g protein; no significant alteration in the KD was detected. The dissociation rate constant (koff) of [3H]QNB increased from 0.020 +/- 0.003 in controls to 0.031 +/- 0.004 (min-1) in the presence of 500 mM ethanol, the association rate constant (Kon) did not change significantly. In locust, 500 mM ethanol did not affect either Kon or Koff. Competition experiments revealed that the binding affinities of both the agonist carbamylcholine and the antagonist atropine to the rat membranes were reduced in the presence of ethanol. In contrast, ethanol caused no alteration in the binding affinities of these ligands to the locust membranes. This differential effect of ethanol on rat and locust muscarinic binding suggests a difference in the hydrophobic domains and/or the membrane interactions of the muscarinic receptors in the two species.
我们比较了膜扰动剂乙醇对脊椎动物（大鼠）和昆虫（蝗虫）神经膜中毒蕈碱结合位点的影响。毒蕈碱拮抗剂[³H]喹核醇基苯甲酸酯（[³H]QNB）与大鼠和蝗虫神经膜的结合在10 - 500 mM浓度的乙醇作用下受到抑制；但这种抑制在蝗虫中更明显。乙醇（500 mM）使[³H]QNB与大鼠膜结合的表观解离常数（KD'）从对照中的0.13±0.01 nM增加到0.20±0.02 nM；结合位点数量Bmax也有小幅但显著的减少。在蝗虫中，500 mM乙醇使[³H]QNB结合的Bmax从对照中的590±30降低到320±40 pmol/g蛋白质；未检测到KD有显著变化。[³H]QNB的解离速率常数（koff）从对照中的0.020±0.003增加到500 mM乙醇存在时的0.031±0.004（min⁻¹），结合速率常数（Kon）没有显著变化。在蝗虫中，500 mM乙醇对Kon和koff均无影响。竞争实验表明，在乙醇存在下，激动剂氨甲酰胆碱和拮抗剂阿托品与大鼠膜的结合亲和力均降低。相反，乙醇对这些配体与蝗虫膜的结合亲和力没有影响。乙醇对大鼠和蝗虫毒蕈碱结合的这种差异效应表明这两个物种中毒蕈碱受体的疏水结构域和/或膜相互作用存在差异。
