Abstract
Intermediate conductance potassium (IKCa) channels are exquisitively Ca2+ sensitive, intracellular Ca2+ regulating channel activity by complexing with calmodulin (CaM), which is bound to the cytosolic carboxyl tail. Although CaM antagonists might be expected to decrease IKCa channel activity, the effect of W-7 in human T lymphocytes are conflicting. We therefore evaluated the effect of W-7 on basolateral IKCa channels in human colonic crypt cells. Intact crypts obtained from normal human colonic biopsies by Ca2+ chelation were used for patch clamp studies of basolateral IKCa channels in the cell-attached configuration. IKCa channel activity was studied when the bath Ca2+ concentration was changed from 1.2 mmol/L to 100 μmol/L and back to 1.2 mmol/L, as well as from 100 μmol/L to 1.2 mmol/L and back to 100 μmol/L, both in the absence and presence of 25 μmol/L W-7. Decreasing bath Ca2+ from 1.2 mmol/L to 100 μmol/L decreased IKCa channel activity reversibly in the absence of W-7, whereas there was a uniformly high level of channel activity at both bath Ca2+ concentrations in the presence of W-7. In separate experiments, increasing bath Ca2+ from 100 μmol/L to 1.2 mmol/L increased IKCa channel activity reversibly in the absence of W-7, whereas there was again a uniformly high level of channel activity at both bath Ca2+ concentrations in the presence of W-7. We, therefore, propose that W-7 has a specific stimulatory effect on basolateral IKCa channel activity, despite its ability to inhibit Ca2+/CaM-mediated, IKCa channel-dependent Cl− secretion in human colonic epithelial cells.Graphic
Highlights
Intermediate conductance Ca2+-sensitive K+ channels (IKCa) dominate the K + conductance of the basolateral membrane in human colonic crypts (Al-Hazza et al 2012)
25 μmol/L W-7 had no effect on channel activity at a bath C a2+ concentration of 1.2 mmol/L, W-7 stimulated channel activity at a bath C a2+ concentration of 100 μmol/L
The initial aim of this study was to evaluate the effect of the CaM antagonist W-7 on the regulation of basolateral IKCa channels by C a2+-mediated agonists in human colonic crypt cells
Summary
Intermediate conductance Ca2+-sensitive K+ channels (IKCa) dominate the K + conductance of the basolateral membrane in human colonic crypts (Al-Hazza et al 2012). Sandle cAMP, and rapidly inhibited in a non-genomic manner by aldosterone (Sandle et al 1994; Bowley et al 2003) They are a major component of the electrogenic Cl− secretory process in human colon and readily inhibited by somatostatin and its stable synthetic analogue octreotide, which is widely used as an anti-diarrhoeal agent (Sandle et al 1999). Their inhibitory effects on human colonic IKCa channels are mediated by inhibitory G proteins and may reflect a decrease in the sensitivity of the channels to intracellular Ca2+ (Sandle et al 1999). Under these circumstances, Ca2+ binding to CaM induces a conformational change in CaM, which is transmitted to the channel subunits, leading to channel gating (Fanger et al 1999)
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