Sulfonylurea binding to a low-affinity site inhibits the Na/K-ATPase and the K-ATP channel in insulin-secreting cells

We have used hamster insulinoma tumor (HIT) cells, an insulin-secreting tumor cell line, to investigate modulation of the Na/K-ATPase and of the ATP-sensitive K channel (K-ATP) by the sulfonylurea glyburide. Membrane proteins from cells cultured in RPMI with 11 mM glucose have at least two glyburide receptor populations, as evidenced by high and low binding affinity constants, (K-d = 0.90 and 91 nM, respectively). In these cells K-ATP channel activity was blocked by low glyburide concentrations, IC50 = 5.4 nM. At 12.5 nM glyburide the inhibition developed slowly, tau = 380 s, and caused reduction of channel activity by 75%. At higher concentrations, however, inhibition occur-red at a fast rate, tau = 42 s at 100 nM, and was almost complete. Na/K-ATPase activity measured enzymatically and electrophsiologically was also suppressed by glyburide, but higher concentrations were needed, IC50 = 20-40 nM. Inhibition occurred rapidly, tau = 30 s at 50 nM, when maximum, activity was reduced by 40%. By contrast, cells cultured in RPMI supplemented with 25 mM glucose exhibit a single receptor population binding glyburide with low affinity, K-d = 68 nM. In these cells inhibition of the Na/K-ATPase by the sulfonylurea was similar to that observed in cells cultured in 11 mM glucose, but K-ATP channel inhibition was markedly altered. Inhibition occurred only at high concentrations of glyburide and at a fast rate; maximum inhibition was observed at similar to 100 nM. Based on these data, we propose that glyburide binding to the high affinity site affects primarily K-ATP channel activity, while inter action with the low affinity site inhibits both Na/K-ATPase and K-ATP channel activities. The latter observation suggests possible functional interactions between the Na/K-ATPase and the K-ATP channel.