Joo Jinhyeong, Kim Ki Jung, Lim Jiwoon, Choi Sun Yeong, Koh Wuhyun, Lee C Justin
Center for Cognition and Sociality, Institute for Basic Science (IBS), Daejeon 34126, Korea.
IBS School, Korea University of Science and Technology (UST), Daejeon 34113, Korea.
Exp Neurobiol. 2024 Aug 31;33(4):180-192. doi: 10.5607/en24019.
Bestrophin-1 (BEST1) is a Ca-activated anion channel known for its role in astrocytes. Best1 is permeable to gliotransmitters, including GABA, to contribute to tonic GABA inhibition and modulate synaptic transmission in neighboring neurons. Despite the crucial functions of astrocytic BEST1, there is an absence of genetically engineered cell-type specific conditional mouse models addressing these roles. In this study, we developed an astrocyte-specific BEST1 conditional knock-out (BEST1 aKO) mouse line. Using the embryonic stem cell (ES cell) targeting method, we developed floxed mice (C57BL/6JCya-/Cya), which have exon 3, 4, 5, and 6 of flanked by two loxP sites. By crossing with hGFAP-CreER mice, we generated floxed/hGFAP-CreER mice, which allowed for the tamoxifen-inducible deletion of under the human GFAP promoter. We characterized its features across various brain regions, including the striatum, hippocampal dentate gyrus (HpDG), and Parafascicular thalamic nucleus (Pf). Compared to the Cre-negative control, we observed significantly reduced BEST1 protein expression in immunohistochemistry (IHC) and tonic GABA inhibition in patch clamp recordings. The reduction in tonic GABA inhibition was 66.7% in the striatum, 46.4% in the HpDG, and 49.6% in the Pf. Our findings demonstrate that the BEST1 channel in astrocytes significantly contributes to tonic inhibition in the local brain areas. These mice will be valuable for future studies not only on tonic GABA release but also on tonic release of gliotransmitters mediated by astrocytic BEST1.
最佳rophin-1（BEST1）是一种钙激活阴离子通道，因其在星形胶质细胞中的作用而闻名。Best1对包括GABA在内的神经胶质递质具有通透性，有助于维持GABA的张力性抑制并调节邻近神经元的突触传递。尽管星形胶质细胞BEST1具有关键功能，但目前缺乏针对这些作用的基因工程细胞类型特异性条件性小鼠模型。在本研究中，我们开发了一种星形胶质细胞特异性BEST1条件性敲除（BEST1 aKO）小鼠品系。利用胚胎干细胞（ES细胞）靶向方法，我们培育出了floxed小鼠（C57BL/6JCya-/Cya），其第3、4、5和6外显子两侧有两个loxP位点。通过与hGFAP-CreER小鼠杂交，我们得到了floxed/hGFAP-CreER小鼠，该小鼠允许在人GFAP启动子下通过他莫昔芬诱导缺失Best1。我们对其在包括纹状体、海马齿状回（HpDG）和束旁丘脑核（Pf）在内的各个脑区的特征进行了表征。与Cre阴性对照相比，我们在免疫组织化学（IHC）中观察到BEST1蛋白表达显著降低，在膜片钳记录中观察到GABA张力性抑制降低。纹状体中GABA张力性抑制的降低为66.7%，HpDG中为46.4%，Pf中为49.6%。我们的研究结果表明，星形胶质细胞中的BEST1通道对局部脑区的张力性抑制有显著贡献。这些小鼠不仅对未来关于GABA张力性释放的研究有价值，而且对由星形胶质细胞BEST1介导的神经胶质递质张力性释放的研究也有价值。
