Regulation of stably expressed and native BK channels from human myometrium by cGMP- and cAMP-dependent protein kinase

Abstract

 The cloned BK channel α subunit from human myometrium was stably expressed in Chinese hamster ovary cells, either alone (CHOα cells) or in combination with the auxiliary β subunit (CHOα+β cells). We studied basic channel properties and the effects of cGMP- and cAMP-dependent protein kinases on the BK channel activity. Coexpression of α and β subunits enhanced the Ca²⁺ and voltage sensitivity of the BK channel, and decreased the inhibitory potency of iberiotoxin. Blocking and stimulating effects on BK channel activity by charybdotoxin and nitric oxide, respectively, were independent of the β subunit. The cGMP kinase Iα and cAMP kinase failed to affect BK channel activity in CHOα and CHOα+β cells at different [Ca²⁺]_i and voltages. In contrast, BK channels in freshly isolated myometrial cells from postmenopausal women responded to cAMP kinase and cGMP kinase with a fourfold and twofold decrease in their open probability (NP _o), respectively. These effects could be reversed by alkaline phosphatase and remained unaffected by the phosphatase inhibitor okadaic acid (100 nM). In 28% of myometrial cells, however, cAMP and cGMP kinases increased NP _o 2-fold and 3.5-fold, respectively. This stimulation was enhanced rather than reversed by alkaline phosphatase and was abolished by 100 nM okadaic acid. The results suggest that in stably transfected CHO cells the expressed BK channel is not regulated by cAMP kinase and cGMP kinase. However, in native myometrial cells stimulatory and inhibitory regulation of BK channels by cAMP kinase and cGMP kinase was observed, suggesting that channel regulation by the protein kinases requires factors that are not provided by CHO cells. Alternatively, failure of regulation may have been due to the primary structure of the myometrial BK channel protein used in this study.