ROLE OF CA2+ CAMK-II IN CA2+-INDUCED K+ CHANNEL INHIBITION IN RAT CCDPRINCIPAL CELL/

The apical low-conductance K+ channel of rat cortical collecting duct (CCD) is inhibited by increased intracellular Ca2+ concentrations. Thi s effect has been shown to be mediated at least in part by activation of protein kinase C (PKC). In the present study, we used the patch-cla mp technique to examine the role of Ca2+/calmodulin-dependent protein kinase II (CaMK II) in mediating the Ca2+-induced inhibitory effect. I n cell-attached patches of principal cells of rat tubules, clamping of intracellular Ca2+ concentration at 400 nM by using 1 mu M ionomycin reduced channel activity to 26.5% of the control value. A further redu ction in channel activity, to 8.8% of the control value, was observed following the addition of phorbol 12-myristate 13-acetate (PMA), an ag ent known to activate PKC. Pretreatment of cells with KN-62 (CaMK II i nhibitor) or GF-109203X (PKC inhibitor) attenuated the inhibitory effe ct of Ca2+ on K+ channel activity (83.2 and 50.7% of the control value , respectively). Even in the presence of KN-62, addition of 10 mu M PM A significantly decreased channel activity to 57.2% of the control val ue. The Ca2+-induced inhibition was completely abolished by simultaneo us incubation with both KN-62 and GF-109203X. In inside-out patches, a ddition of 20 mu g/ml CaMK II in the presence of a PKC inhibitor reduc ed channel activity to 66.2% of control values. It is concluded that C aMK II is involved in mediating the Ca2+-induced inhibition of the act ivity of the apical K+ channel of rat CCD.