α-Subunit Tyrosine Phosphorylation Is Required for Activation of the Large Conductance Ca2+-Activated Potassium Channel in the Rabbit Sphincter of Oddi

Large conductance Ca 2+ -activated potassium (BK Ca ) channels are regulated by intracellular free Ca 2+ concentrations ([Ca 2+ ]i) and channel protein phosphorylation. In hypercholesterolemia (HC), motility impairment of the sphincter of Oddi (SO) is associated with abnormal [Ca 2+ ]i accumulation in smooth muscle cells of the rabbit SO (RSOSMCs), which is closely related to BK Ca channel activity. However, the underlying mechanisms regulating channel activity remain unclear. In this study, an HC rabbit model was generated and used to investigate BK Ca channel activity of RSOSMCs via SO muscle tone measurement in vitro and manometry in vivo , electrophysiological recording, intracellular calcium measurement, and Western blot analyses. BK Ca channel activity was decreased, which correlated with [Ca 2+ ]i overload and reduced tyrosine phosphorylation of the BK Ca α-subunit in the HC group. The abnormal [Ca 2+ ]i accumulation and decreased BK Ca channel activity were partially restored by Na 3 VO 4 pretreatment but worsened by genistein in RSOSMCs in the HC group. This study suggests that α-subunit tyrosine phosphorylation is required for [Ca 2+ ]i to activate BK Ca channels, and there is a negative feedback between the BK Ca channel and the L-type voltage-dependent Ca 2+ channel that regulates [Ca 2+ ]i. This study provides direct evidence that tyrosine phosphorylation of BK Ca α-subunits is required for [Ca 2+ ]i to activate BK Ca channels in RSOSMCs, which may be the underlying physiological and pathologic mechanism regulating the activity of BK Ca channels in SO cells.