Benzodiazepines modulate GABAA receptor surface levels and synaptic inhibition

说明（由申请人提供）：苯二氮卓类药物广泛用于治疗焦虑、失眠和癫痫。此外，它们是精神分裂症、抑郁症和酒精解毒的关键辅助治疗方法。然而，由于耐受性的发展，这些安全有效的化合物的使用受到严重限制。苯二氮卓类药物增强了3-氨基丁酸（GABAA）受体的活性，GABAA是中枢神经系统中主要的抑制性神经递质受体。耐受性产生的分子机制尚未确定。越来越明显的是，苯二氮卓类药物优先增强突触部位的GABAA受体亚型的活性，这些突触部位主要由？1 - 3,然后呢?2单元。相比之下，大多数介导强直抑制的突触外GABAA受体（GABAARs）的结构不同于它们的突触对应物，对苯二氮卓类药物的功能调节不敏感。然而，苯二氮卓类药物治疗神经元与GABAAR亚型表面水平变化和神经元抑制效果之间的联系机制尚未得到证实。这些现象与我们的初步研究一起产生了驱动本提案中描述的实验的中心假设：将神经元暴露于苯二氮卓类药物会促进？2个亚基GABAARs从质膜中分离出来，并随后降解，导致抑制性突触大小和数量减少，突触抑制效果下降。我们的建议围绕三个具体目标：(1)我们将表征苯二氮卓类药物（BZ）处理对GABAAR膜运输和降解的影响；(2)我们将测量BZ治疗对突触抑制的影响，并确定改变突触效能的机制；(3)我们将测量BZ处理对野生型小鼠和？2H101R突变小鼠表达？含2的对BZ不敏感的GABAAR。

DESCRIPTION (provided by applicant): Benzodiazepines are widely used to treat anxiety, insomnia and seizure disorders. Furthermore they are key adjunct treatments in schizophrenia, depression and alcohol detoxification. However, the use of these safe and efficacious compounds is severely limited due to the development of tolerance. Benzodiazepines potentiate the activity of 3-aminobutyric acid (GABAA) receptors, the major inhibitory neurotransmitter receptors in the central nervous system. The molecular mechanism underlying the development of tolerance has not yet been determined. It is increasingly evident that benzodiazepines preferentially enhance the activity of GABAA receptor subtypes present at synaptic sites that are largely composed of ?1-3, ? and ?2 subunits. In contrast, the majority of extrasynaptic GABAA receptors (GABAARs), which mediate tonic inhibition, have structures different from those of their synaptic counterparts and are insensitive to functional modulation by benzodiazepines. However, a mechanism linking benzodiazepine treatment of neurons to changes in surface levels of GABAAR subtypes and the efficacy of neuronal inhibition has not been demonstrated. These phenomena together with our preliminary studies generated the central hypothesis driving the experiments described in this proposal: Exposure of neurons to benzodiazepines promotes the removal of ?2 subunit-containing GABAARs from the plasma membrane and their subsequent degradation, leading to a reduction in inhibitory synapse size and number along with a decrease in the efficacy of synaptic inhibition. Our proposal centers on three specific aims: (1) We will characterize the effects of benzodiazepine (BZ) treatment on GABAAR membrane trafficking and degradation; (2) We will measure the effects of BZ treatment on synaptic inhibition and identify the mechanism altering synaptic efficacy; and (3) We will measure the ability of BZ treatment to modulate cell surface accumulation of GABAARs and synaptic inhibition in wild type mice and ?2H101R mutant mice that express ?2-containing GABAAR that are BZ insensitive. PUBLIC HEALTH RELEVANCE: Benzodiazepines are widely used to treat anxiety, insomnia and seizure disorders, but their clinical use is severely limited due to the development of tolerance. The therapeutic actions of benzodiazepines are primarily exerted by potentiating the activity of ?-aminobutyric acid (GABA) type A receptors, the major inhibitory neurotransmitter receptors in the central nervous system. We will investigate the effects of benzodiazepines on GABA type A receptors to define a molecular mechanism underlying benzodiazepine tolerance, facilitating the development of new therapeutic agents to improve patient quality of life.