Life and Medical Sciences Institute, Molecular Genetics, University of Bonn, D-53115 Bonn, Germany.
J Neurosci. 2013 May 1;33(18):7985-96. doi: 10.1523/JNEUROSCI.5874-12.2013.
Panglial networks are essential for normal physiology in the CNS, and the function of distinct connexins participating in these networks is not well understood. We generated Connexin32 (Cx32)-deficient mice with additional deletion of astrocytic Cx43 to explore the role of both connexins in panglial networks. Cx43/Cx32 double knock-out (dKO) mice revealed strong microglial activation in corpus callosum and cingulum along with severe astrogliosis and scar formation. In addition, most of the fine myelinated fibers projecting from the corpus callosum into the cortex were lost. Myelin loss was caused by a strong decrease of oligodendrocytes in the cingulum of Cx43/Cx32dKO mice. Immunoblot analyses using newly generated specific Cx47 antibodies revealed that oligodendrocytic Cx47 is phosphorylated in vivo depending on astrocytic Cx43 expression. In Cx43-deficient mice, Cx47 protein levels were strongly decreased, whereas Cx47 mRNA levels were not altered. Using Cx43G138R/Cx30KO mice, we show that Cx47 expression depends on the presence of astrocytic Cx43 protein and that its gap junctional channel function is not necessary for Cx47 stabilization. In consequence, Cx43/Cx32dKO mice additionally lack Cx47 expression and therefore cannot form oligodendrocytic gap junctions, which explains the phenotypic similarities to Cx32/Cx47dKO mice. Our findings provide strong evidence that phosphorylation and stability of oligodendrocytic Cx47 proteins is dependent on astrocytic Cx43 expression. These results further unravel the complexity of panglial networks and show that results of previous studies using astrocytic Cx43-deficient mice have to be reconsidered.
神经胶质细胞连接网络对于中枢神经系统的正常生理功能至关重要，而参与这些网络的不同连接蛋白的功能尚未得到很好的理解。我们生成了Connexin32（Cx32）缺陷型小鼠，并进一步缺失星形胶质细胞的 Cx43，以探究这两种连接蛋白在神经胶质细胞连接网络中的作用。Cx43/Cx32 双重敲除（dKO）小鼠在胼胝体和扣带回中表现出强烈的小胶质细胞激活，同时伴有严重的星形胶质细胞增生和瘢痕形成。此外，来自胼胝体投射到皮质的大多数精细髓鞘纤维丢失。Cx43/Cx32 dKO 小鼠扣带回中的少突胶质细胞大量减少导致髓鞘丢失。使用新生成的特异性 Cx47 抗体进行免疫印迹分析显示，体内的少突胶质细胞 Cx47 依赖于星形胶质细胞 Cx43 的表达而被磷酸化。在 Cx43 缺陷型小鼠中，Cx47 蛋白水平显著降低，而 Cx47 mRNA 水平没有改变。使用 Cx43G138R/Cx30KO 小鼠，我们证明 Cx47 的表达依赖于星形胶质细胞 Cx43 蛋白的存在，并且其间隙连接通道功能对于 Cx47 的稳定不是必需的。因此，Cx43/Cx32 dKO 小鼠还缺乏 Cx47 的表达，因此不能形成少突胶质细胞间隙连接，这解释了它们与 Cx32/Cx47 dKO 小鼠表型相似的原因。我们的研究结果提供了强有力的证据，表明少突胶质细胞 Cx47 蛋白的磷酸化和稳定性依赖于星形胶质细胞 Cx43 的表达。这些结果进一步揭示了神经胶质细胞连接网络的复杂性，并表明之前使用星形胶质细胞 Cx43 缺陷型小鼠进行的研究结果需要重新考虑。
