Nusser Z, Ahmad Z, Tretter V, Fuchs K, Wisden W, Sieghart W, Somogyi P
Medical Research Council Anatomical Neuropharmacology Unit, Oxford, UK.
Eur J Neurosci. 1999 May;11(5):1685-97. doi: 10.1046/j.1460-9568.1999.00581.x.
Any given subunit of the heteromultimeric type-A gamma-aminobutyric acid (GABA) GABAA receptor may be present in several receptor subtypes expressed by individual neurons. Changes in the expression of a subunit may result in differential changes in the expression of other subunits depending on the subunit composition of the receptor subtype, leading to alterations in neuronal responsiveness to GABA. We used the targeted disruption of the alpha6 subunit gene to test for changes in the expression of other GABAA receptor subunits. Immunoprecipitation and ligand binding experiments indicated that GABAA receptors were reduced by approximately 50% in the cerebellum of alpha6 -/- mice. Western blot experiments indicated that the alpha6 subunit protein completely disappeared from the cerebellum of alpha6 -/- mice, which resulted in the disappearance of the delta subunit from the plasma membrane of granule cells. The amount of beta2, beta3 and gamma2 subunits was reduced by approximately 50%, 20% and 40%, respectively, in the cerebella of alpha6 -/- mice. A comparison of the reduction in the level of alpha1, beta2, beta3, gamma2, or delta-subunit-containing receptors in alpha6 -/- cerebellum with those observed after removal of alpha6-subunit-containing receptors from the cerebella of alpha6 +/+ mice by immuno-affinity chromatography demonstrated the presence of a significantly higher than expected proportion of receptors containing beta3 subunits in alpha6 -/- mice. The receptors containing alpha1, beta2, beta3 and gamma2 subunits were present in the plasma membrane of granule cells of alpha6 -/- mice at both synaptic and extrasynaptic sites, as shown by electron microscopic immunocytochemistry. Despite the changes, the alpha1 subunit content of Golgi-cell-to-granule-cell synapses in alpha6 -/- animals remained unaltered, as did the frequency of alpha1 immunopositive synapses in the glomeruli. Furthermore, no change was apparent in the expression of the alpha1, beta2 and gamma2 subunits in Purkinje cells and interneurons of the molecular layer. These results demonstrate that in alpha6 -/- mice, the cerebellum expresses only half of the number of GABAA receptors present in wild-type animals. Since these animals have no gross motor deficits, synaptic integration in granule cells is apparently maintained by alpha1-subunit-containing receptors with an altered overall subunit composition, and/or by changes in the expression of other ligand and voltage gated channels.
异源多聚体型Aγ-氨基丁酸（GABA）GABAA受体的任何一个给定亚基可能存在于单个神经元表达的几种受体亚型中。一个亚基表达的变化可能会导致其他亚基表达的差异变化，这取决于受体亚型的亚基组成，从而导致神经元对GABA反应性的改变。我们利用α6亚基基因的靶向破坏来检测其他GABAA受体亚基表达的变化。免疫沉淀和配体结合实验表明，α6 -/-小鼠小脑中的GABAA受体减少了约50%。蛋白质印迹实验表明，α6亚基蛋白在α6 -/-小鼠的小脑中完全消失，这导致颗粒细胞膜上的δ亚基消失。在α6 -/-小鼠的小脑中，β2、β3和γ2亚基的量分别减少了约50%、20%和40%。通过免疫亲和色谱从α6 +/+小鼠小脑中去除含α6亚基的受体后，将α6 -/-小鼠小脑中含α1、β2、β3、γ2或δ亚基的受体水平的降低与观察到的情况进行比较，结果表明α6 -/-小鼠中含β3亚基的受体比例明显高于预期。如电子显微镜免疫细胞化学所示，含α1、β2、β3和γ2亚基的受体存在于α6 -/-小鼠颗粒细胞的突触和突触外膜中。尽管发生了这些变化，但α6 -/-动物中高尔基细胞到颗粒细胞突触的α1亚基含量保持不变，肾小球中α1免疫阳性突触的频率也保持不变。此外，浦肯野细胞和分子层中间神经元中α1、β2和γ2亚基的表达没有明显变化。这些结果表明，在α6 -/-小鼠中，小脑表达的GABAA受体数量仅为野生型动物的一半。由于这些动物没有明显的运动缺陷，颗粒细胞中的突触整合显然是由含α1亚基且整体亚基组成改变的受体，和/或由其他配体门控通道和电压门控通道表达的变化来维持的。
