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A mitochondrial sulphonylurea-sensitive, ATP-sensitive K+ channel (mitoKATP) that is selectively inhibited by 5-hydroxydecanoate (5-HD) and activated by diazoxide has been implicated in ischaemic preconditioning. Here we re-evaluate the evidence for the existence of this mitoKATP by measuring changes in light scattering (A520) in parallel with direct determination of mitochondrial matrix volumes using 3H2O and [14C]sucrose. Incubation of rat liver and heart mitochondria in KCl medium containing Mg2+ and inorganic phosphate caused a decrease in light scattering over 5 min, which was accompanied by a small (15-30 %) increase in matrix volume. The presence of ATP or ADP in the buffer from the start greatly inhibited the decline in A520, whilst addition after a period of incubation (1-5 min) induced a rapid increase in A520, especially in heart mitochondria. Neither response was accompanied by a change in matrix volume, as measured isotopically. However, the effects of ATP and ADP on A520 were abolished by carboxyatractyloside and bongkrekic acid, inhibitors of the adenine nucleotide translocase (ANT) that lock the transporter in two discrete conformations and cause distinct changes in A520 in their own right. These data suggest that rather than matrix volume changes, the effects of ATP and ADP on A520 reflect changes in mitochondrial shape induced by conformational changes in the ANT. Furthermore, we were unable to demonstrate either a decrease in A520 or increase in matrix volume with a range of ATP-sensitive K+ channel openers such as diazoxide. Nor did glibencamide or 5-HD cause any reduction of matrix volume, whereas the K+ ionophore valinomycin (0.2 nM), produced a 10-20 % increase in matrix volume that was readily detectable by both techniques. Our data argue against the existence of a sulphonylurea-inhibitable mitoKATP channel.
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