Minturn J E, Black J A, Angelides K J, Waxman S G
Department of Neurology, Yale University School of Medicine, New Haven, Connecticut 06510.
Glia. 1990;3(5):358-67. doi: 10.1002/glia.440030507.
Astrocytes cultured from neonatal rat optic nerve can be classified into two subtypes, distinguished by their morphology (stellate or fibroblast-like) and their ability to bind monoclonal antibody A2B5. The presence of sodium channels in astrocytes cultured from rat optic nerve was demonstrated by indirect immunofluorescence with polyclonal antibody 7493, which is directed against purified rat brain sodium channel protein. Astrocytes cultured from postnatal day 7 (P7) rat optic nerves exhibited sodium channel immunostaining on both A2B5+ and A2B5- astrocytes up to 6 days in vitro (DIV). Staining was distributed throughout the cytoplasm and cell processes, with areas of greater intensity in the perinuclear region. At 6 DIV, the A2B5-/GFAP+ cells exhibited a loss of sodium channel immunostaining, while the A2B5+/GFAP+ cells continued to display 7493 immunoreactivity. This sodium channel staining pattern persisted for up to 28 DIV (the longest time point examined). Astrocyte cultures derived from PO rat optic nerves exhibited sodium channel immunoreactivity during the first 6 DIV. The P0 astrocyte cultures, in which, the vast majority of cells are A2B5-/GFAP+, displayed a similar staining pattern to those astrocytes with corresponding phenotype derived from P7 optic nerves. P0-derived A2B5- astrocytes showed loss of 7493 immunostaining at 6 DIV, while the rare (less than 1% of cells) A2B5+/GFAP+ cells continued to express sodium channels reactive to 7493. The reduction of sodium channel immunoreactivity in A2B5- but not A2B5+, astrocytes from both P0 and P7 optic nerves after a similar latency (approximately 6 DIV) suggests that the loss of immunostaining may result from the absence of neuronal associations in the culture environment, rather than an intrinsic biologically timed change in astrocytic expression of sodium channels.
从新生大鼠视神经培养的星形胶质细胞可分为两种亚型，可通过其形态（星状或成纤维细胞样）以及结合单克隆抗体A2B5的能力来区分。用针对纯化大鼠脑钠通道蛋白的多克隆抗体7493进行间接免疫荧光，证实了从大鼠视神经培养的星形胶质细胞中存在钠通道。从出生后第7天（P7）大鼠视神经培养的星形胶质细胞在体外培养6天（DIV）内，A2B5 +和A2B5 -星形胶质细胞均表现出钠通道免疫染色。染色分布在整个细胞质和细胞突起中，核周区域强度更大。在6 DIV时，A2B5 - / GFAP +细胞的钠通道免疫染色消失，而A2B5 + / GFAP +细胞继续显示7493免疫反应性。这种钠通道染色模式持续长达28 DIV（检查的最长时间点）。来自P0大鼠视神经的星形胶质细胞培养物在最初6 DIV期间表现出钠通道免疫反应性。P0星形胶质细胞培养物中绝大多数细胞为A2B5 - / GFAP +，其染色模式与来自P7视神经的具有相应表型的星形胶质细胞相似。P0来源的A2B5 -星形胶质细胞在6 DIV时显示7493免疫染色消失，而罕见的（少于1%的细胞）A2B5 + / GFAP +细胞继续表达对7493有反应的钠通道。P0和P7视神经的A2B5 -而非A2B5 +星形胶质细胞在相似的潜伏期（约6 DIV）后钠通道免疫反应性降低，这表明免疫染色的丧失可能是由于培养环境中缺乏神经元联系，而不是星形胶质细胞钠通道表达的内在生物学定时变化。
