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Updated: Jun 3, 2026

Study of the Functions and Activities of Neuronal K-Cl Co-Transporter KCC2 Using Western Blotting
Published on: December 9, 2022
A key role for KCl cotransport in cell volume regulation in human erythroleukemia cells
Chiun-Chien Huang1, Poh-Hong Lim, Andrew C Hall
1Department of Physiology, Chung Shan Medical University, Taichung, Taiwan. huanglim@ms71.hinet.net
Aims:
KCl cotransport is believed to be involved in volume regulation in various erythroid cells of vertebrates, although the mechanism of activation and the role of the signaling elements involved remain uncertain. In this study, we characterized KCl cotransport activated by hypo-osmotic stress, and clarified several signaling elements involved in the regulation of this pathway within the human erythroleukemia cell line K562.
Main Methods:
The Cl(-)-dependent K(+) efflux (measured using (86)Rb(+)) and regulatory volume decrease (RVD) from pre-loaded K562 cells subjected to hypo-osmotic challenge were measured in cells treated with/without KCl cotransport inhibitors [(dihydroindenyl)oxy]alkanoic acid (DIOA) and Ba(2+). This Cl(-)-dependent K(+) efflux has also been measured in cells treated with the phorbol 12-myristate 13-acetate (protein kinase C (PKC) activator), RO 31-8220 or calphostin C (PKC inhibitor), genistein (protein tyrosine kinase (PTK) inhibitor), PP2 (Src kinase inhibitor), AG18 or AG1478 (epidermal growth factor receptor (EGFR) kinase inhibitor), wortmannin or LY294002 (phosphatatidylinositol 3-kinase (PI 3-kinase) inhibitor), or PD98059 (mitogen-activated protein (MAP) kinase inhibitor).
Key Findings:
Cl(-)-dependent K(+) efflux was strongly stimulated by hypo-osmotic challenge and this increased K(+) efflux was mediated by the DIOA- and Ba(2+)-sensitive KCl cotransport. RO 31-8220, calphostin C, genistein, PP2, AG18, AG1478, wortmannin, LY294002 and PD98059 were shown to significantly inhibit or stimulate the activity of this pathway.
Significance:
Our results suggest that the hypo-osmotically-activated KCl cotransport is an important regulator of K562 cell volume, and the activity of this pathway is modulated by PKC, PTK, PI 3-kinase and/or MAP kinases.
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