Donaldson J, Hill S J
Eur J Pharmacol. 1986 Sep 23;129(1-2):25-31. doi: 10.1016/0014-2999(86)90332-8.
The effect of the disulphide bond reducing agent 1,4-dithiothreitol (DTT) on the binding characteristics of the H1-selective ligand [3H]mepyramine has been examined in homogenates of guinea-pig cerebral cortex and cerebellum. DTT was found to be without effect on antagonist binding. This was evident from studies using different concentrations of [3H]mepyramine (0.5-10.0 nM), which showed no change in either the equilibrium dissociation constant (KD) or specific binding site capacity (Bmax). Furthermore, the IC50 values and slope parameters determined from the inhibition of the binding of 1 nM [3H]mepyramine by non-radioactive mepyramine were similarly insensitive to DTT in both cerebral cortex and cerebellum. In contrast, DTT shifted the inhibition curve for histamine to lower agonist concentrations and reduced the Hill coefficient in these two tissues. Analysis of these inhibition curves as double hyperbolae revealed two binding sites in the presence of DTT and only one low affinity site in the absence of DTT. Similar changes in the location (IC50) and slope (Hill coefficient) parameters were obtained with the H1-selective agonist 2-thiazolylethylamine in cerebellum and with 2-methylhistamine in both brain regions. The results of this study show that DTT affects agonist but not antagonist binding in guinea-pig cerebellum and cerebral cortex, and suggest that DTT stabilises a proportion of the agonist binding sites in a high affinity state.
已在豚鼠大脑皮层和小脑的匀浆中研究了二硫键还原剂1,4-二硫苏糖醇（DTT）对H1选择性配体[3H]美吡拉敏结合特性的影响。发现DTT对拮抗剂结合没有影响。这在使用不同浓度的[3H]美吡拉敏（0.5 - 10.0 nM）的研究中很明显，该研究表明平衡解离常数（KD）或特异性结合位点容量（Bmax）均无变化。此外，由非放射性美吡拉敏抑制1 nM [3H]美吡拉敏结合所确定的IC50值和斜率参数在大脑皮层和小脑中对DTT同样不敏感。相反，DTT使组胺的抑制曲线向较低的激动剂浓度移动，并降低了这两个组织中的希尔系数。将这些抑制曲线分析为双双曲线表明，在存在DTT的情况下有两个结合位点，而在不存在DTT的情况下只有一个低亲和力位点。在小脑中使用H1选择性激动剂2-噻唑基乙胺以及在两个脑区使用2-甲基组胺时，在位置（IC50）和斜率（希尔系数）参数上获得了类似的变化。本研究结果表明，DTT影响豚鼠小脑和大脑皮层中的激动剂结合但不影响拮抗剂结合，并表明DTT使一部分激动剂结合位点稳定在高亲和力状态。
