On the mechanism of ADP‐induced alteration of sulphonylurea sensitivity in cardiac ATP‐sensitive K + channels

To study the mechanism of regulation of sulphonylurea sensitivity in ATP‐sensitive K + (K ATP ) channels, we used the inside‐out patch clamp technique in guinea‐pig ventricular myocytes. In the absence of nucleotides, the half maximal concentration of tolbutamide inhibition of K ATP channels (IC 50 ) was 0.4 m M , and it decreased to 0.1 m M when 0.1 m M ATP was added. Increasing the ADP concentration from 0 to 0.1 and 0.3 m M in the absence of ATP shifted the IC 50 from 0.4 to 5.3 and 11.4 m M , respectively. Increasing the ADP concentration further to 1 and 3 m M conversely reduced the IC 50 to 9.5 and 4.4 m M , respectively. In the absence of Mg 2+ and ADP, the IC 50 was calculated to 16.6 m M which was found to be less, 12.3, 5.1 and 2.5 m M , respectively, when the ADP concentration was increased to 0.1, 0.3 and 1 m M . The IC 50 s for tolbutamide obtained at various concentrations of ADP in the presence of Mg 2+ were best fitted by equations reflecting a model that assumed two binding sites for ADP; one is a high affinity site that reduces the sensitivity to the sulphonylurea, while the other is a low affinity site that increases such sensitivity. Dissociation constants calculated for ADP to sites 1 and 2 were 2.6 μ M and 46.7 m M , respectively. In the absence of Mg 2+ , data were fitted by equations corresponding to a single site model (site 2); the dissociation constant for ADP was 25.0 m M . It is concluded that ADP modifies tolbutamide sensitivity by binding to two sites. The high affinity site is strongly Mg 2+ ‐dependent, whereas the low affinity site is Mg 2+ ‐independent.British Journal of Pharmacology (2000) 130 , 1411–1417; doi: 10.1038/sj.bjp.0703423