Harris R A, Valenzuela C F, Brozowski S, Chuang L, Hadingham K, Whiting P J
Denver VA Medical Center, Colorado, USA.
J Pharmacol Exp Ther. 1998 Jan;284(1):180-8.
We studied the adaptation of gamma-aminobutyric acid type A (GABAA) receptor function to chronic ethanol exposure in cells stably transfected with the following GABAA receptor subunits: alpha-1 beta-2 gamma-2L, alpha-1 beta-2 gamma-2S, alpha-1 beta-3 gamma-2S, alpha-1 beta-1, alpha-5 beta-3 gamma-3 and alpha-6 beta-3 gamma-2S. Chronic exposure to ethanol resulted in a decrease in muscimol-stimulated 36Cl- flux and a decrease in modulation of that flux by ethanol, flunitrazepam, methyl-6,7-4-dimethoxy-4-ethyl-beta-carboline-3-carboxylate and pregnanolone without any change in the modulation by pentobarbital or zinc. Direct activation of the GABAA receptor by pentobarbital was enhanced by chronic ethanol treatment. Reduction of the action of muscimol, ethanol and flunitrazepam differed in the duration and amount of ethanol required to see an effect. Reduction of the action of ethanol of alpha-1 beta-2 gamma-2L cells occurred within 15 min and was near-maximal for 25 mM ethanol, whereas reduction of the actions of muscimol and flunitrazepam actions required hours of exposure and higher concentrations of ethanol. Chronic ethanol exposure produced a reduction in the Emax value for the action of muscimol for all six subunit combinations, but quantification of surface receptors by immunolabeling showed no change in GABAA receptor density. The differences in alcohol sensitivity and time courses for different effects of ethanol indicate multiple mechanisms of adaptation of GABAA receptors. Use of stably transfected cells rules out "subunit substitution" as a mechanism for these changes and points to post-translational changes (e.g., phosphorylation, receptor assembly) as the most likely mechanisms. These in vitro findings are compared with results from in vivo studies.
我们研究了在稳定转染以下γ-氨基丁酸A型（GABAA）受体亚基的细胞中，GABAA受体功能对慢性乙醇暴露的适应性：α-1β-2γ-2L、α-1β-2γ-2S、α-1β-3γ-2S、α-1β-1、α-5β-3γ-3和α-6β-3γ-2S。慢性乙醇暴露导致蝇蕈醇刺激的36Cl-通量降低，以及乙醇、氟硝西泮、甲基-6,7-4-二甲氧基-4-乙基-β-咔啉-3-羧酸酯和孕烷醇酮对该通量的调节作用降低，而戊巴比妥或锌的调节作用没有任何变化。慢性乙醇处理增强了戊巴比妥对GABAA受体的直接激活作用。蝇蕈醇、乙醇和氟硝西泮作用的降低在所需乙醇的持续时间和量方面有所不同。α-1β-2γ-2L细胞中乙醇作用的降低在15分钟内发生，对于25 mM乙醇接近最大值，而蝇蕈醇和氟硝西泮作用的降低需要数小时的暴露和更高浓度的乙醇。慢性乙醇暴露导致所有六种亚基组合的蝇蕈醇作用的Emax值降低，但通过免疫标记对表面受体进行定量显示GABAA受体密度没有变化。乙醇不同作用的酒精敏感性和时间进程差异表明GABAA受体存在多种适应机制。使用稳定转染的细胞排除了“亚基替代”作为这些变化的机制，并指出翻译后变化（例如，磷酸化、受体组装）是最可能的机制。将这些体外研究结果与体内研究结果进行了比较。
