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 μM) induced Ca²⁺-activated K⁺ current [I_K(ca)] in a concentration-dependent manner with an EC₅₀ value of 4 μM. Squamocin-stimulated I_K(ca) was suppressed by iberiotoxin (200 nM), paxilline (1 μM), or tetraethylammonium chloride (5 mM), yet not by apamin (200 nM) or glibenclamide (10 μM). 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 μM) 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 μM) for 5 h did not alter the magnitude of squamocin-induced I_K(ca). However, squamocin (30 μM) 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⁺ (BK_Ca) channels without altering single-channel conductance. Conversely, in inside-out patches, squamocin applied to the intracellular surface had no effect on BK_Ca 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.