Constitutive Activity of Inwardly Rectifying K+ Channel at Physiological [Ca]i Is Mediated by Ca2+/CaMK II Pathway in Opossum Kidney Proximal Tubule Cells

Abstract

Using patch-clamp technique, we studied the role of the Ca2+/calmodulin kinase II (CaMK II)-mediated phosphorylation process on the K+ channel with an inward conductance of 90 pS in opossum kidney proximal tubule cells (OKPCs). The intracellular Ca2+ concentration ([Ca]i) was measured by use of the fluorescent dye fura 2. The following results were obtained: (i) In cell-attached patches, the channel activity was inhibited by a decrease in [Ca]i induced by perfusion with low Ca2+ (10(-8) M), La3+ (100 microM), or EGTA/AM (100 microM) contained in the bath solution. The application of KN-62 (10 microM) or KN-93 (5 microM), inhibitors of CaMK II, also inhibited the channel activity. (ii) The membrane potential measured with nystatin-perforated patches was significantly decreased by the fall in [Ca]i induced by the perfusion with EGTA- or La(3+)-containing solution. Also, the application of KN-62 (10 microM) or KN-93 (5 microM) to the bath significantly decreased the membrane potential. (iii) In inside-out patches, the channel activity was significantly stimulated by the application of CaMK II (300 pM) at 10(-7) M Ca2+ in the bath. Furthermore, the application of KN-62 (10 microM) to the bath significantly decreased the channel activity. Our findings show that the constitutive activity of inwardly rectifying K+ channel at physiological [Ca]i is mediated by the Ca2+/CaMK II pathway in OKPCs.