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  • Title: Regulation of Na(+)-K(+)-Cl(-) cotransporter in primary astrocytes by dibutyryl cAMP and high [K(+)](o).
    Author: Su G, Haworth RA, Dempsey RJ, Sun D.
    Journal: Am J Physiol Cell Physiol; 2000 Dec; 279(6):C1710-21. PubMed ID: 11078685.
    Abstract:
    In this study, we examined the Na(+)-K(+)-Cl(-) cotransporter activity and expression in rat cortical astrocyte differentiation. Astrocyte differentiation was induced by dibutyryl cAMP (DBcAMP, 0. 25 mM) for 7 days, and cells changed from a polygonal to process-bearing morphology. Basal activity of the cotransporter was significantly increased in DBcAMP-treated astrocytes (P < 0.05). Expression of an approximately 161-kDa cotransporter protein was increased by 91% in the DBcAMP-treated astrocytes. Moreover, the specific [(3)H]bumetanide binding was increased by 67% in the DBcAMP-treated astrocytes. Inhibition of protein synthesis by cyclohexamide (2-3 microgram/ml) significantly attenuated the DBcAMP-mediated upregulation of the cotransporter activity and expression. The Na(+)-K(+)-Cl(-) cotransporter in astrocytes has been suggested to play a role in K(+) uptake. In 75 mM extracellular K(+) concentration, the cotransporter-mediated K(+) influx was stimulated by 147% in nontreated cells and 79% in DBcAMP-treated cells (P < 0.05). To study whether this high K(+)-induced stimulation of the cotransporter is attributed to membrane depolarization and Ca(2+) influx, the role of the L-type voltage-dependent Ca(2+) channel was investigated. The high-K(+)-mediated stimulation of the cotransporter activity was abolished in the presence of either 0.5 or 1.0 microM of the L-type channel blocker nifedipine or Ca(2+)-free HEPES buffer. A rise in intracellular free Ca(2+) in astrocytes was observed in high K(+). These results provide the first evidence that the Na(+)-K(+)-Cl(-) cotransporter protein expression can be regulated selectively when intracellular cAMP is elevated. The study also demonstrates that the cotransporter in astrocytes is stimulated by high K(+) in a Ca(2+)-dependent manner.
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