Department of Neurosurgery, University of Maryland School of Medicine, Baltimore, Maryland, 21201-1595.
Department of Cellular and Molecular Medicine, Laboratory of Ion Channel Research, Katholieke Universiteit Leuven, Leuven, 3000, Belgium.
Glia. 2018 Jan;66(1):108-125. doi: 10.1002/glia.23231. Epub 2017 Sep 14.
Astrocyte swelling occurs after central nervous system injury and contributes to brain swelling, which can increase mortality. Mechanisms proffered to explain astrocyte swelling emphasize the importance of either aquaporin-4 (AQP4), an astrocyte water channel, or of Na -permeable channels, which mediate cellular osmolyte influx. However, the spatio-temporal functional interactions between AQP4 and Na -permeable channels that drive swelling are poorly understood. We hypothesized that astrocyte swelling after injury is linked to an interaction between AQP4 and Na -permeable channels that are newly upregulated. Here, using co-immunoprecipitation and Förster resonance energy transfer, we report that AQP4 physically co-assembles with the sulfonylurea receptor 1-transient receptor potential melastatin 4 (SUR1-TRPM4) monovalent cation channel to form a novel heteromultimeric water/ion channel complex. In vitro cell-swelling studies using calcein fluorescence imaging of COS-7 cells expressing various combinations of AQP4, SUR1, and TRPM4 showed that the full tripartite complex, comprised of SUR1-TRPM4-AQP4, was required for fast, high-capacity transmembrane water transport that drives cell swelling, with these findings corroborated in cultured primary astrocytes. In a murine model of brain edema involving cold-injury to the cerebellum, we found that astrocytes newly upregulate SUR1-TRPM4, that AQP4 co-associates with SUR1-TRPM4, and that genetic inactivation of the solute pore of the SUR1-TRPM4-AQP4 complex blocked in vivo astrocyte swelling measured by diolistic labeling, thereby corroborating our in vitro functional studies. Together, these findings demonstrate a novel molecular mechanism involving the SUR1-TRPM4-AQP4 complex to account for bulk water influx during astrocyte swelling. These findings have broad implications for the understanding and treatment of AQP4-mediated pathological conditions.
星形胶质细胞肿胀发生在中枢神经系统损伤后，导致脑水肿，从而增加死亡率。解释星形胶质细胞肿胀的机制强调了水通道蛋白-4（AQP4），一种星形胶质细胞水通道，或介导细胞渗透溶质流入的 Na + 渗透性通道的重要性。然而，驱动肿胀的 AQP4 和 Na + 渗透性通道之间的时空功能相互作用知之甚少。我们假设损伤后星形胶质细胞肿胀与新上调的 AQP4 和 Na + 渗透性通道之间的相互作用有关。在这里，我们使用共免疫沉淀和Förster 共振能量转移，报告 AQP4 与磺酰脲受体 1-瞬时受体电位 melastatin 4（SUR1-TRPM4）单价阳离子通道物理地组装形成一种新型异源三聚体水/离子通道复合物。使用表达各种 AQP4、SUR1 和 TRPM4 组合的 COS-7 细胞的 calcein 荧光成像进行体外细胞肿胀研究表明，由 SUR1-TRPM4-AQP4 组成的完整三聚体复合物是快速、大容量跨膜水转运所必需的，该转运驱动细胞肿胀，这些发现在培养的原代星形胶质细胞中得到证实。在涉及小脑冷损伤的脑水肿小鼠模型中，我们发现星形胶质细胞新上调 SUR1-TRPM4，AQP4 与 SUR1-TRPM4 共关联，并且 SUR1-TRPM4-AQP4 复合物的溶质孔的遗传失活阻断了体内通过二羟标记测量的星形胶质细胞肿胀，从而证实了我们的体外功能研究。总之，这些发现证明了一种涉及 SUR1-TRPM4-AQP4 复合物的新型分子机制，以解释星形胶质细胞肿胀期间的大量水流入。这些发现对理解和治疗 AQP4 介导的病理状况具有广泛的意义。
