Yuan Xiaoqing, Chittajallu Ramesh, Belachew Shibeshih, Anderson Stacie, McBain Chris J, Gallo Vittorio
Laboratory of Cellular and Synaptic Neurophysiology, National Institute of Child Health and Human Development, National Institutes of Health, Bethesda, Maryland, USA.
J Neurosci Res. 2002 Nov 15;70(4):529-45. doi: 10.1002/jnr.10368.
We generated a transgenic mouse expressing the enhanced green fluorescent protein (EGFP) under the control of the 2'-3'-cyclic nucleotide 3'-phosphodiesterase (CNP) promoter. EGFP(+) cells were visualized in live tissue throughout embryonic and postnatal development. Immunohistochemical analysis in brain tissue and in sciatic nerve demonstrated that EGFP expression was restricted to cells of the oligodendrocyte and Schwann cell lineages. EGFP was also strongly expressed in "adult" oligodendrocyte progenitors (OPs) and in gray matter oligodendrocytes. Fluorescence-activated cell sorting allowed high-yield purification of EGFP(+) oligodendrocyte-lineage cells from transgenic brains. Electrophysiological patch clamp recordings of EGFP(+) cells in situ demonstrated that OP cells displayed large outward tetraethylammonium (TEA)-sensitive K(+) currents and very small inward currents, whereas mature oligodendrocytes were characterized by expression of large inward currents and small outward K(+) currents. The proliferation rate of EGFP(+) cells in developing white matter decreased with the age of the animals and was strongly inhibited by TEA. Oligodendrocyte development and physiology can be studied in live tissue of CNP-EGFP transgenic mice, which represent a source of pure EGFP(+) oligodendrocyte-lineage cells throughout development.
我们构建了一种转基因小鼠,其在2'-3'-环核苷酸3'-磷酸二酯酶(CNP)启动子的控制下表达增强型绿色荧光蛋白(EGFP)。在胚胎期和出生后的整个发育过程中,均可在活体组织中观察到EGFP(+)细胞。对脑组织和坐骨神经进行免疫组织化学分析表明,EGFP的表达仅限于少突胶质细胞系和施万细胞系的细胞。EGFP在“成年”少突胶质前体细胞(OPs)和灰质少突胶质细胞中也有强烈表达。荧光激活细胞分选可从转基因大脑中高效纯化EGFP(+)少突胶质细胞系细胞。对原位EGFP(+)细胞进行电生理膜片钳记录表明,OP细胞表现出较大的外向四乙铵(TEA)敏感钾电流和非常小的内向电流,而成熟少突胶质细胞的特征是具有较大的内向电流和较小的外向钾电流。发育中的白质中EGFP(+)细胞的增殖率随动物年龄的增长而降低,并受到TEA的强烈抑制。可在CNP-EGFP转基因小鼠的活体组织中研究少突胶质细胞的发育和生理学,该小鼠在整个发育过程中代表了纯EGFP(+)少突胶质细胞系细胞的来源。