Genetic deletion of laminin isoforms β2 and γ3 induces a reduction in Kir4.1 and aquaporin-4 expression and function in the retina.
Genetic deletion of laminin isoforms β2 and γ3 induces a reduction in Kir4.1 and aquaporin-4 expression and function in the retina.
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DOI：
10.1371/journal.pone.0016106
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发表时间：
2011-01-21
期刊：
影响因子：
3.7
通讯作者：
Hirrlinger J
中科院分区：
文献类型：
--
作者：
Hirrlinger PG;Pannicke T;Winkler U;Claudepierre T;Varshney S;Schulze C;Reichenbach A;Brunken WJ;Hirrlinger J
神经胶质细胞，如视网膜Müler神经胶质细胞，参与神经组织钾离子和水的稳态。在这些细胞中，内向整流钾(KIR)通道和AQP4水通道在空间钾缓冲和水排出过程中起着重要作用。此外，Kir4.1通道参与了负Müller细胞膜电位的维持。Kir4.1和Aquaporin-4通道的亚细胞分布似乎通过与细胞外和细胞内分子的相互作用来维持。细胞外基质中的层粘连蛋白、膜上的抗肌营养不良蛋白多糖和细胞基质中的抗肌营养不良蛋白形成一个复合体，介导Müler细胞Kir4.1和Aquaporin-4的极化表达。本研究的目的是检测含层粘连蛋白的β-2和γ-3在小鼠Müler细胞中的功能。我们使用β2和γ3层粘连蛋白基因缺失的基因敲除小鼠来测试对Kir4.1和Aquaporin-4的影响。免疫组织化学、Western Blot和定量RT-PCR分别检测蛋白质和mRNA的表达，膜片钳实验检测细胞的膜电流。我们发现，在层粘连蛋白基因敲除的小鼠中，Kir4.1的mRNA和蛋白以及水通道蛋白-4的蛋白表达下调。此外，层粘连蛋白β2和γ3基因敲除小鼠层粘连蛋白基因敲除的Müler细胞减少了Kir介导的内向电流，其膜电位比年龄匹配的野生型小鼠更具正性。这些结果表明，层粘连蛋白β2和γ3亚基对Müler细胞中两个重要的膜蛋白水通道蛋白4和Kir4.1的表达和功能都有很大的影响。
Glial cells such as retinal Müller glial cells are involved in potassium ion and water homeostasis of the neural tissue. In these cells, inwardly rectifying potassium (Kir) channels and aquaporin-4 water channels play an important role in the process of spatial potassium buffering and water drainage. Moreover, Kir4.1 channels are involved in the maintenance of the negative Müller cell membrane potential. The subcellular distribution of Kir4.1 and aquaporin-4 channels appears to be maintained by interactions with extracellular and intracellular molecules. Laminins in the extracellular matrix, dystroglycan in the membrane, and dystrophins in the cytomatrix form a complex mediating the polarized expression of Kir4.1 and aquaporin-4 in Müller cells. The aim of the present study was to test the function of the β2 and γ3 containing laminins in murine Müller cells. We used knockout mice with genetic deletion of both β2 and γ3 laminin genes to assay the effects on Kir4.1 and aquaporin-4. We studied protein and mRNA expression by immunohistochemistry, Western Blot, and quantitative RT-PCR, respectively, and membrane currents of isolated cells by patch-clamp experiments. We found a down-regulation of mRNA and protein of Kir4.1 as well as of aquaporin-4 protein in laminin knockout mice. Moreover, Müller cells from laminin β2 and γ3 knockout mice had reduced Kir-mediated inward currents and their membrane potentials were more positive than those in age-matched wild-type mice. These findings demonstrate a strong impact of laminin β2 and γ3 subunits on the expression and function of both aquaporin-4 and Kir4.1, two important membrane proteins in Müller cells.