McCabe R T, Olsen R W, Yezuita J P, Wamsley J K
Department of Psychiatry, University of Utah School of Medicine, Salt Lake City.
J Pharmacol Exp Ther. 1988 Apr;245(1):342-9.
Inasmuch as the presence of endogenous gamma-aminobutyric acid (GABA) may affect benzodiazepine binding to tissue sections in autoradiographic studies, a protocol designed to check for this influence has been investigated. [3H]Flunitrazepam (1 nM) was used to label benzodiazepine receptors for autoradiographic localization. Bicuculline was added to the incubation medium of an additional set of tissue sections to antagonize any potential effect of endogenous GABA. Binding in these sections was compared to that occurring in another set in which excess GABA was added to "create" further GABA enhancement. Binding also was compared to adjacent sections which were treated similarly but also preincubated in distilled-deionized water to burst the cells by osmotic shock and eliminate endogenous GABA, thereby preventing any effect on benzodiazepine binding. The results indicated that endogenous GABA is indeed present in the slide-mounted tissue sections and is affecting benzodiazepine receptor binding differentially in various regions of the brain depending on the density of GABAergic innervation. Scatchard analysis of saturation data demonstrated that the alteration in benzodiazepine binding due to GABA was a result of a change in the affinity rather than number of receptors present. These experiments have been compared to the binding of the imidazodiazepine, [3H] Ro15-1788. We also show that the treatments affect endogenous GABA and not the receptors themselves. This suggests strongly that, when using a single nonsaturating concentration of radiolabeled benzodiazepine antagonist, autoradiographic studies to date may have been subject to erroneous interpretation due to the differential effects of endogenous GABA on benzodiazepine binding (increased affinity).(ABSTRACT TRUNCATED AT 250 WORDS)
鉴于内源性γ-氨基丁酸（GABA）的存在可能会在放射自显影研究中影响苯二氮䓬与组织切片的结合，因此对一项旨在检查这种影响的方案进行了研究。使用[³H]氟硝西泮（1 nM）标记苯二氮䓬受体以进行放射自显影定位。在另一组组织切片的孵育培养基中加入荷包牡丹碱，以拮抗内源性GABA的任何潜在影响。将这些切片中的结合与另一组切片中的结合进行比较，在另一组切片中加入过量的GABA以“产生”进一步的GABA增强作用。还将结合与相邻切片进行比较，这些相邻切片经过类似处理，但也在蒸馏去离子水中预孵育，以通过渗透压休克使细胞破裂并消除内源性GABA，从而防止对苯二氮䓬结合产生任何影响。结果表明，内源性GABA确实存在于载玻片上的组织切片中，并且根据GABA能神经支配的密度，在大脑的各个区域对苯二氮䓬受体结合产生不同的影响。对饱和数据的Scatchard分析表明，由于GABA导致的苯二氮䓬结合改变是亲和力变化而非存在的受体数量变化的结果。这些实验已与咪唑并二氮䓬[³H]Ro15-1788的结合进行了比较。我们还表明，这些处理影响内源性GABA而非受体本身。这强烈表明，当使用单一非饱和浓度的放射性标记苯二氮䓬拮抗剂时，由于内源性GABA对苯二氮䓬结合的不同影响（亲和力增加），迄今为止的放射自显影研究可能受到了错误的解释。（摘要截断于250字）
