Figure 1

AIMS associated mutations cause complete K_ATP channel loss-of-function. (A) Whole-cell patch clamp recordings were performed in HEK293 cells transfected with K_ir6.2 and wild-type (WT) or mutant SUR2A. Initial ambient levels of intracellular ATP means channels are inhibited immediately after membrane rupture to whole-cell configuration. Over time, ATP levels are depleted by dilution with the pipette solution. (B) Example current traces from voltage ramps for cells transfected with GFP alone, or K_ir6.2 alongside SUR2A-WT, SUR2[Arg620Ter], SUR2[Arg938Ter], SUR2[Phe1405SerfsTer8] or SUR2[Leu714SerfsTer7]. (C) Summary showing whole-cell currents at 0 mV measured at 10 min after establishing the whole-cell recording configuration. Box and whisker plot shows median as horizontal line, mean as ‘X’, and interquartile range as coloured box. P-values from Dunn’s pairwise comparisons versus SUR2A-WT following Kruskal-Wallis test shown. (D) K_ATP currents were recorded from cells transfected with K_ir6.2 and SUR2A-WT (top, grey) or SUR2[Phe1405SerfsTer8] (bottom, orange). Whole cell currents were recorded from ramp protocols as shown above with 300 µM ATP included in the patch pipette. Currents at 0 mV from sweeps recorded at 5-s intervals are shown. K_ATP channels from SUR2A-WT expressing cells displayed robust activation upon administration of 100 µM pinacidil, which was reversed by the K_ATP inhibitor glibenclamide (10 µM). Dotted line shows zero current level. (E) Summary of currents recorded prior to and after pinacidil administration in cells transfected with K_ir6.2 and SUR2A-WT or SUR2[Phe1405SerfsTer8]. P-values from Dunn’s pairwise comparisons versus SUR2A-WT currents in pinacidil following Kruskal-Wallis test shown. GFP = green fluorescent protein.