The phosphorylation state of large-conductance calcium-activated potassium (BKCa) channels regulates their activity and is dynamically regulated by protein phosphatases and kinases, including protein kinase C (PKC). In this study, we showed that PKC activators up-regulate the activity of the BKCa channel alpha (alpha)-subunit, Slol, in cell-attached patches of transfected COS7 cells. In an immune complex kinase assay, BKCa channels isolated from rat brain were phosphorylated in the presence of PKC activators, without the addition of exogenous PKC, which suggests that PKC and BKCa channels functionally interact in vivo. Four different PKC isozymes, including PKC delta, phosphorylated the C-terminus of Slol and the addition of purified PKC delta-activated BKCa channels in excised patches of transfected HEK293 cells. Our results demonstrate that PKC up-regulates BKCa channels and that PKC delta may functionally interact with BKCa channel complexes in vivo. (c) 2007 Elsevier Inc. All rights reserved.