Physiological role of large, Ca2+-activated K+ channels in human glomerular mesangial cells

1. Contraction assays and patch clamp methods were used to determine the role of K+ channels in the regulation of contractile tone of human mesangial cells (MC) in culture. 2. MC contraction was induced by vasoconstrictor agents, such as angiotensin II (AngII; 100 nmol/L) and glybenclamide (Glyb), but not by iberiotoxin (IbTX), a blocker of large Ca2+-activated K+ channels (BK(Ca)). These results suggest that Glyb-sensitive K+ channels, but not BK(Ca) channels, were active at rest. 3. In the presence of 100 nmol/L IbTX, contraction by AngII was slightly, but not significantly, enhanced, indicating that BK(Ca) has a minimal role as a negative feedback regulator of contraction. Nitroprusside (NP; 100 mu mol/L) a nitric oxide (NO) donor, atrial natriuretic peptide (ANP; 1.0 mu mol/L) and db-cGMP (10 mu mol/L) attenuated AngII-induced contraction in the absence, but not in the presence, of IbTX, suggesting that BK(Ca) channels were activated by cGMP. 4. In patch clamp experiments, three distinct K+-selective channels of 9, 65 and 150 pS (outward currents) were found in excised, inside-out patches. The 150 pS channel was completely inhibited by 100 nmol/L IbTX and displayed voltage- and calcium-dependent gating qualitatively similar to BK(Ca) in other cell types. 5. In cell attached (CA) patches, the response of BK(Ca) to bath AngII (100 nmol/L) was relatively minor in control solutions, but was considerably greater in the presence of db-cGMP. 6. In excised patches, Mg-ATP (1 mmol/L) plus db-cGMP (1 mu mol/L) activated BK(Ca) in the absence, but not the presence, of the non-specific kinase inhibitor, staurosporine. 7. Separate experiments showed that BK(Ca) were also activated by arachidonic acid and high ambient glucose concentrations. 8. These results indicate that: (i) resting MC tone is sensitive to glybenclamide and apamin; and (ii) the role of BK(Ca) as a negative feedback regulator of contraction is minimal under normal conditions but is markedly enhanced by cGMP-stimulating relaxants and arachidonic acid.