Peter Proks

Abstract:

Potassium channels selectively conduct K⁺ ions across cell membranes and have key roles in cell excitability. Their opening and closing can be spontaneous or controlled by membrane voltage or ligand binding. We used Ba²⁺ as a probe to determine the location of the ligand-sensitive gate in an inwardly rectifying K⁺ channel (Kir6.2). To a K⁺ channel, Ba²⁺ and K⁺ are of similar sizes, but Ba²⁺ blocks the pore by binding within the selectivity filter. We found that internal Ba²⁺ could still access its binding site when the channel was shut, which indicates that the ligand-sensitive gate lies above the Ba²⁺-block site, and thus within or above the selectivity filter. This is in marked contrast to the voltage-dependent gate of KV channels, which is located at the intracellular mouth of the pore.

Abstract:

Potassium channels selectively conduct K(+) ions across cell membranes and have key roles in cell excitability. Their opening and closing can be spontaneous or controlled by membrane voltage or ligand binding. We used Ba(2+) as a probe to determine the location of the ligand-sensitive gate in an inwardly rectifying K(+) channel (Kir6.2). To a K(+) channel, Ba(2+) and K(+) are of similar sizes, but Ba(2+) blocks the pore by binding within the selectivity filter. We found that internal Ba(2+) could still access its binding site when the channel was shut, which indicates that the ligand-sensitive gate lies above the Ba(2+)-block site, and thus within or above the selectivity filter. This is in marked contrast to the voltage-dependent gate of K(V) channels, which is located at the intracellular mouth of the pore.

Abstract:

Sulfonylureas are widely used to treat type 2 diabetes because they stimulate insulin secretion from pancreatic beta-cells. They primarily act by binding to the SUR subunit of the ATP-sensitive potassium (K(ATP)) channel and inducing channel closure. However, the channel is still able to open to a limited extent when the drug is bound, so that high-affinity sulfonylurea inhibition is not complete, even at saturating drug concentrations. K(ATP) channels are also found in cardiac, skeletal, and smooth muscle, but in these tissues are composed of different SUR subunits that confer different drug sensitivities. Thus tolbutamide and gliclazide block channels containing SUR1 (beta-cell type), but not SUR2 (cardiac, smooth muscle types), whereas glibenclamide, glimepiride, repaglinide, and meglitinide block both types of channels. This difference has been exploited to determine residues contributing to the sulfonylurea-binding site. Sulfonylurea block is decreased by mutations or agents (e.g., phosphatidylinositol bisphosphate) that increase K(ATP) channel open probability. We now propose a kinetic model that explains this effect in terms of changes in the channel open probability and in the transduction between the drug-binding site and the channel gate. We also clarify the mechanism by which MgADP produces an apparent increase of sulfonylurea efficacy on channels containing SUR1 (but not SUR2).

Abstract:

Most imidazolines inhibit ATP-sensitive K(+) (K(ATP)) channels. Since these drugs are potentially clinically relevant insulin secretagogues, it is important to know whether extrapancreatic K(ATP) channels are targeted. We examined the effects of three imidazoline-derived antidiabetic drugs on the cloned K(ATP) channel, expressed in Xenopus laevis oocytes, and their specificity for interaction with the pore-forming Kir6.2 or the sulphonylurea receptor (SUR) 1 subunit. Midaglizole, LY397364 and LY389382 blocked Kir6.2deltaC currents with IC(50) of 3.8, 6.1 and 0.7 microM, respectively. The block of Kir6.2/SUR1 currents by LY397364 and LY389382 was best fit by a two-site model, suggesting that these drugs also interact with SUR1. However, since all three drugs interact with the Kir6.2 subunit, and Kir6.2 forms the pore of extrapancreatic K(ATP) channels, these drugs are unlikely to be specific for the beta-cell.