Stimulatory effects of squamocin, an Annonaceous acetogenin, on Ca2+-activated K+ current in cultured smooth muscle cells of human coronary artery

The patch-clamp recording technique was used to investigate the effect of squamocin, an Annonaceous acetogenin, on ion currents in cultured smooth muscle cells of human coronary artery. In whole-cell configuration, squamocin (0.3-100 muM) induced Ca2+-activated K+ current [I-K(Ca)] in a concentration-dependent manner with an EC50 value of 4 muM. Squamocin-stimulated I-K(Ca) was suppressed by iberiotoxin (200 nM), paxilline (1 muM), or tetraethylammonium chloride (5 mM), yet not by apamin (200 nM) or glibenclamide (10 muM). In cells dialyzed with 10 mM EGTA, this compound had little effect on I-K(Ca). When cells were exposed to Ca2+-free solution, squamocin (3 muM) induced a transient increase in I-K(Ca). In continued presence of squamocin, an additional increase in extracellular Ca2+ (1 mM) caused a significant increase in I-K(Ca). Pretreatment with carbonyl cyanide m-chlorophenyl hydrazone (CCCP; 3 muM) for 5 h did not alter the magnitude of squamocin-induced I-K(Ca). However, squamocin (30 muM) suppressed the amplitude of voltage-dependent L-type Ca2+ Current. In cell-attached configuration of single-channel recordings, squamocin applied to the bath increased the activity of large-conductance Ca2+-activated K+ (BKCa) channels without altering single-channel conductance. Conversely, in inside-out patches, squamocin applied to the intracellular surface had no effect on BKCa channel activity, whereas niflumic acid increased it effectively. These findings provide the evidence that squamocin can activate I-K(Ca) in coronary arterial smooth muscle cells. Initial transient activation of I-K(Ca) may reflect the squamocin-induced Ca2+ release from intracellular Ca2+ stores, whereas the sustained activation of I-K(Ca) may arise from the squamocin-induced Ca2+ influx across the cell membrane. The stimulatory effect of squamocin on these channels should affect the functional activity of vascular smooth muscle cells.