Perillán P R, Li X, Potts E A, Chen M, Bredt D S, Simard J M
Department of Neurosurgery, University of Maryland School of Medicine, Baltimore, Maryland 21201-1595, USA.
Glia. 2000 Aug;31(2):181-92. doi: 10.1002/1098-1136(200008)31:2<181::aid-glia90>3.0.co;2-8.
Astrocytic inward rectifying K(+) channels that participate in K(+) spatial buffering in the central nervous system have been extensively investigated, but specific gene products have not been fully identified. We studied primary cultured reactive astrocytes of stellate and polygonal morphology from adult rat brains, as well as stellate astrocytes from neonatal rat brains. Single-channel recordings of cell-attached patches revealed that polygonal reactive astrocytes expressed only one hyperpolarization-activated single-channel conductance of 11-15 pS whose open probability was independent of voltage, whereas stellate reactive and stellate neonatal astrocytes exhibited two conductances, 11-15 pS and 24-27 pS. All three subtypes of astrocytes exhibited a hyperpolarization-activated macroscopic inward K(+) current that was strongly rectifying and was abrogated by 1 mM intracellular Mg(2+) introduced during conventional but not perforated patch whole-cell recording. This Mg(2+)-sensitive current comprised the total inward rectifier current in polygonal reactive astrocytes, but only a fraction of the inward rectifier current in stellate reactive and stellate neonatal astrocytes. Because a strongly rectifying, inward rectifier K(+) channel with a single-channel conductance of 11-15 pS that is voltage independent is consistent with features of Kir2.3 (IRK3), we performed immunofluorescence experiments with anti-Kir2.3 and anti-glial fibrillary acidic protein antibodies. Both antibodies co-localized to all three subtypes of astrocytes in primary culture and to reactive astrocytes in situ within brain and gelatin sponge implants. Our data indicate that astrocytes of both polygonal and stellate morphology, from both adult and neonatal rat brain, express Kir2.3 both in vivo and in vitro. Constitutive expression of Kir2.3 regardless of cell morphology or age of origin of the source tissue suggests an important functional role for this channel in astrocytes.
参与中枢神经系统钾离子空间缓冲的星形胶质细胞内向整流钾离子通道已得到广泛研究，但具体的基因产物尚未完全确定。我们研究了成年大鼠脑内原代培养的星状和多角形形态的反应性星形胶质细胞，以及新生大鼠脑内的星状星形胶质细胞。细胞贴附式膜片钳单通道记录显示，多角形反应性星形胶质细胞仅表达一种超极化激活的单通道电导，为11 - 15 pS，其开放概率与电压无关，而星状反应性星形胶质细胞和新生星状星形胶质细胞则表现出两种电导，分别为11 - 15 pS和24 - 27 pS。所有三种星形胶质细胞亚型均表现出超极化激活的宏观内向钾离子电流，该电流具有强烈的整流特性，并且在传统的而非穿孔膜片钳全细胞记录过程中引入1 mM细胞内镁离子时被消除。这种镁离子敏感电流构成了多角形反应性星形胶质细胞的总内向整流电流，但仅占星状反应性星形胶质细胞和新生星状星形胶质细胞内向整流电流的一部分。由于一种单通道电导为11 - 15 pS且与电压无关的强烈整流内向整流钾离子通道符合Kir2.3（IRK3）的特征，我们用抗Kir2.3和抗胶质纤维酸性蛋白抗体进行了免疫荧光实验。两种抗体在原代培养的所有三种星形胶质细胞亚型以及脑内和明胶海绵植入物中原位的反应性星形胶质细胞中均共定位。我们的数据表明，来自成年和新生大鼠脑的多角形和星状形态的星形胶质细胞在体内和体外均表达Kir2.3。无论细胞形态或来源组织的年龄如何都持续表达Kir2.3，这表明该通道在星形胶质细胞中具有重要的功能作用。
