Goldman-Wohl D S, Chan E, Baird D, Heintz N
Howard Hughes Medical Institute, Rockefeller University, New York, New York 10021.
J Neurosci. 1994 Feb;14(2):511-22. doi: 10.1523/JNEUROSCI.14-02-00511.1994.
A two-step hybridization/subtraction procedure was employed to isolate markers for the later stages of Purkinje cell differentiation. From this screen, a novel Shaw potassium channel cDNA (Kv3.3b) was identified that is developmentally regulated. Expression of this channel is highly enriched in the brain, particularly in the cerebellum, where its expression is confined to Purkinje cells and deep cerebellar nuclei. Sequence analysis revealed that it is an alternatively spliced form of the mouse Kv3.3 gene, and that the previously reported Kv3.3 mRNA (Ghanshani et al., 1992) is not expressed in cerebellum. Expression of the Kv3.3b mRNA begins in cerebellar Purkinje cells between postnatal day 8 (P8) and P10 and continues through adulthood, coinciding with elaboration of the mature Purkinje cell dendritic arbor. The timing of expression of Kv3.3b mRNA is maintained in mixed, dissociated primary cerebellar cell culture. These results suggest that the Kv3.3b K+ channel function is restricted to terminally differentiated Purkinje cells, and that analysis of the mechanisms governing its expression in vivo and in vitro can reveal molecular mechanisms governing Purkinje cell differentiation.
采用两步杂交/扣除法来分离浦肯野细胞分化后期的标记物。通过该筛选,鉴定出一种新的肖氏钾通道cDNA(Kv3.3b),其表达受发育调控。该通道的表达在脑中高度富集,尤其是在小脑,其表达局限于浦肯野细胞和小脑深部核团。序列分析表明,它是小鼠Kv3.3基因的一种选择性剪接形式,并且先前报道的Kv3.3 mRNA(Ghanshani等人,1992年)在小脑中不表达。Kv3.3b mRNA的表达在出生后第8天(P8)至第10天之间开始于小脑浦肯野细胞,并持续至成年期,这与成熟浦肯野细胞树突分支的形成相一致。Kv3.3b mRNA的表达时间在混合的、解离的原代小脑细胞培养中得以维持。这些结果表明,Kv3.3b钾通道功能仅限于终末分化的浦肯野细胞,并且对其体内和体外表达调控机制的分析可以揭示浦肯野细胞分化的分子机制。