Regulation of Na+-K+-2Cl- cotransport in turkey red cells: The role of oxygen tension and protein phosphorylation

M. C. Muzyamba, A. R. Cossins, J. S. Gibson*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

52 Citations (Scopus)


1. Na+-K+-2Cl- cotransport (NKCC) was studied in turkey red cells using Na+ dependence or bumetanide sensitivity of 86Rb+ influx to monitor activity of the transporter. 2. Deoxygenation was the major physiological stimulus for NKCC activity: oxygen tensions (P(O2) over the physiological range modulated the transporter, with a P(O2) for half-maximal activation of about 41 mmHg (n = 3). In air, activity of NKCC was also stimulated by shrinkage and isoproteronol (isoprenaline, 5 μM). By contrast, in deoxygenated cells, although the transporter activity was markedly elevated, it was no longer sensitive to volume or β-adrenergic stimulation. 3. Calyculin A, a protein phosphatase inhibitor, stimulated cotransport with a lag of about 5 min. N-Ethylmaleimide (NEM) inhibited cotransport and also blocked the stimulatory effect of calyculin A if administered before calyculin A. Stimulation by calyculin A and deoxygenation were not additive. Staurosporine (2 μM) inhibited deoxygenated-stimulated KC influxes, but not those stimulated by calyculin A. NEM added during calyculin A stimulation, i.e. during the 5 min lag, caused transport activity to be clamped at levels intermediate between maximal (calyculin A alone) and control. Cells treated with calyculin A alone or with calyculin A followed by NEM were no longer sensitive to volume, isoproteronol or P(O2). 4. The results have characterized the interaction between deoxygenation and other stimuli of NKCC activity. They have also shown that it is possible to manipulate the transporter in a reciprocal way to that shown previously for K+-Cl- cotransport.

Original languageEnglish
Pages (from-to)421-429
Number of pages9
JournalJournal of Physiology
Issue number2
Publication statusPublished - 1 Jun 1999
Externally publishedYes


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