Ca²⁺/Calmodulin kinase II is required to activate inwardly rectifying K⁺ channel in opossum kidney proximal tubule cells

Abstract

We have previously reported that the K⁺ channel with an inward conductance of 90 pS in opossum kidney proximal tubule (OKP) cells was inactivated by protein kinase C-mediated phosphorylation. In this study, we further examined the role of Ca²⁺/calmodulin kinase II (CaMK II)-mediated phosphorylation process on this channel activity. 1) In cell-attached patches, the channel activity was inhibited by a decrease in cytosolic Ca²⁺ by the perfusion of low Ca²⁺ (10⁻⁹ M) or 100 μM La³⁺ containing solution, or by the addition of 100 μM EGTA/AM to the bath. The application of KN-62 (10 μM) or KN-93 (5 μM), inhibitors of CaMK II, inhibited the channel activity, whereas L-NAME (5 μM) or L-NNA (10 μM), cNOS inhibitors, failed to inhibit the channel activity. These results suggest that Ca²⁺/CaMK II-mediated phosphorylation process is involved in the regulation of channel activity. 2) Membrane potential measured with nystatin-perforated patches was significantly decreased by a fall in cytosolic Ca²⁺ with low Ca²⁺ (10⁻⁹ M), La³⁺ (100 μM), or EGTA/AM (100 μM) containing solution in the bath. In addition, the application of KN-62 (10 μM) or KN-93 (5 μM) to the bath significantly decreased the membrane potential. 3) In inside-out patches with 10^−6.5 M Ca²⁺ in the bath, the channel activity was dramatically stimulated by the application of CaMK II (300 pM). In conclusion, Ca²⁺/CaMK II-mediated phosphorylation process is involved in maintaining the K⁺ channel activity in OKP cells. [Jpn J Physiol 54 Suppl:S115 (2004)]