Time‐dependent fading of the activation of KATP channels, induced by aprikalim and nucleotides, in excised membrane patches from cardiac myocytes

The effects of the potassium channel opener (KCO) aprikalim (RP 52891) on the nucleotide‐induced modulation of ATP‐sensitive K+ (KATP) channels in freshly dissociated ventricular myocytes of guinea‐pig heart, were studied by use of the inside‐out patch‐clamp technique. The internal surface of the excised membrane patch was initially bathed with a standard solution (Mg2+‐free with EDTA), then sequentially superfused with solutions containing nucleoside diphosphates (NDPs: 200 UM ADP and 50 UM GDP) and NDPs plus 1 mM MgCl2 (with EGTA; referred to as Mg‐NDP solution). The normalized concentration‐response (channel closing) relationship to ATP was shifted to the right when the standard solution was replaced by the Mg‐NDP solution. Hence, the internal concentration of ATP ([ATP]i) inhibiting the channel activity by half (Ki) increased from 56 UM to 180 μM, with an apparently constant slope factor (s = 2.37). NDPs in the absence of Mg2+ did not decrease the sensitivity of the channels to ATP. In standard solution, aprikalim (100 μM) activated KATP channels in the presence of a maximally inhibitory [ATP]i (500 μM). This effect was strongly enhanced when aprikalim was applied to patches exposed to Mg‐NDP solution, as demonstrated by the 9 fold increase in Kifor [ATP]i (from 180μM to 1.5 mM and s = 2.37). The ability of aprikalim to overcome the channel closing effects of ATP in Mg‐NDP solution waned rapidly. Similarly, the NDP‐induced activation of ATP‐blocked channels was also time‐dependent. Both activation processes disappeared before the channel run‐down phenomenon appeared in ATP‐free conditions. In conclusion, aprikalim is much more potent in opening KATP channels in membrane patches bathed in Mg‐NDP solution than in standard solution. However, under the former experimental conditions, the effect of aprikalim waned rapidly. It is proposed that the waning phenomenon results from changes in the intrinsic enzymatic activity of the KATP channel protein (possibly linked to the experimental conditions) which lead to the channel closure.