Borden L A, Gibbs T T
Department of Anatomy and Cell Biology, State University of New York, Brooklyn 11203.
Eur J Pharmacol. 1990 Jun 12;188(6):391-7. doi: 10.1016/0922-4106(90)90199-8.
The benzodiazepine drugs modulate gamma-aminobutyric acid (GABA)-mediated synaptic transmission via a high-affinity binding site that is part of the GABA(A) receptor complex, but which is distinct from the GABA binding site. Ro15-4513 is a benzodiazepine negative modulator of GABA action that displays unique anti-ethanol properties both in vivo and in vitro. Ro15-4513 has been reported to photoaffinity label nearly 100% of the benzodiazepine binding sites in rat brain homogenates. In contrast, the benzodiazepine positive modulator flunitrazepam photoaffinity labels only 25% of the sites. Here, we have examined the reversible binding of [3H]Ro15-4513, [3H]flumazenil (Ro15-1788), and [3H]flunitrazepam to embryonic chick brain membranes, and to membranes that have been photoaffinity labeled with nonradioactive flunitrazepam. Photoaffinity labeling with flunitrazepam decreased the subsequent reversible binding of [3H]flunitrazepam and [3H]flumazenil, but increased the binding of [3H]Ro15-4513. The increase in [3H]Ro15-4513 binding after flunitrazepam photoaffinity labeling was due to a decrease in the apparent Kd, with no change in Bmax. Following photoaffinity labeling, negative modulation of [3H]Ro15-4513 binding by GABA was lost, whereas positive modulation of residual [3H]flunitrazepam binding was retained. We conclude that the site photoaffinity labeled by flunitrazepam is distinct from the site responsible for reversible binding of [3H]Ro15-4513.
苯二氮䓬类药物通过作为GABA(A)受体复合物一部分的高亲和力结合位点来调节γ-氨基丁酸（GABA）介导的突触传递，但该位点与GABA结合位点不同。Ro15 - 4513是一种GABA作用的苯二氮䓬类负性调节剂，在体内和体外均表现出独特的抗乙醇特性。据报道，Ro15 - 4513能对大鼠脑匀浆中近100%的苯二氮䓬结合位点进行光亲和标记。相比之下，苯二氮䓬类正性调节剂氟硝西泮仅能对25%的位点进行光亲和标记。在此，我们研究了[3H]Ro15 - 4513、[3H]氟马西尼（Ro15 - 1788）和[3H]氟硝西泮与胚胎鸡脑膜以及用非放射性氟硝西泮进行光亲和标记的膜的可逆结合。用氟硝西泮进行光亲和标记会降低随后[3H]氟硝西泮和[3H]氟马西尼的可逆结合，但会增加[3H]Ro15 - 4513的结合。氟硝西泮光亲和标记后[3H]Ro15 - 4513结合的增加是由于表观解离常数（Kd）降低，最大结合量（Bmax）不变。光亲和标记后，GABA对[3H]Ro15 - 4513结合的负性调节作用丧失，而对残留[3H]氟硝西泮结合的正性调节作用得以保留。我们得出结论，氟硝西泮光亲和标记的位点与负责[3H]Ro15 - 4513可逆结合的位点不同。
