A Potassium-Selective Current Affected by Micromolar Concentrations of Anion Transport Inhibitors

Abstract

Background/Aims: In the human genome, more than 400 genes encode ion channels, which are ubiquitously expressed and often coexist and participate in almost all physiological processes. Therefore, ion channel blockers represent fundamental tools in discriminating the contribution of individual channel types to a physiological phenomenon. However, unspecific effects of these compounds may represent a confounding factor. Three commonly used chloride channel inhibitors, i.e. 4,4′-diisothiocyano-2,2′-stilbene-disulfonic acid (DIDS), 5-nitro-2-[(3-phenylpropyl) amino]benzoic acid (NPPB) and the anti-inflammatory drug niflumic acid were tested to identify the lowest concentration effective on Cl- channels and ineffective on K+ channels. Methods: The activity of the above mentioned compounds was tested by whole cell patch-clamp on the swelling-activated Cl- current ICl,swell and on the endogenous voltage-dependent, outwardly rectifying K+ selective current in human kidney cell lines (HEK 293/HEK 293 Phoenix). Results: Micromolar (1-10 µM) concentrations of DIDS and NPPB could not discriminate between the Cl- and K+ selective currents. Specifically, 1 µM DIDS only affected the K+ current and 10 µM NPPB equally affected the Cl- and K+ currents. Only relatively high (0.1-1 mM) concentrations of DIDS and prolonged (5 minutes) exposure to 0.1-1 mM NPPB preferentially suppressed the Cl- current. Niflumic acid preferentially inhibited the Cl- current, but also significantly affected the K+ current. The endogenous voltage-dependent, outwardly rectifying K+ selective current in HEK 293/HEK 293 Phoenix cells was shown to arise from the Kv 3.1 channel, which is extensively expressed in brain and is involved in neurological diseases. Conclusion: The results of the present study underscore that sensitivity of a given physiological phenomenon to the Cl- channel inhibitors NPPB, DIDS and niflumic acid may actually arise from an inhibition of Cl- channels but can also result from an inhibition of voltage-dependent K+ channels, including the Kv 3.1 channel. The use of niflumic acid as anti-inflammatory drug in patients with concomitant Kv 3.1 dysfunction may result contraindicated.