The Mitochondrial K Channel as a Receptor for Potassium Channel Openers

Abstract

Plasma membrane K ATP channels are highly sensitive to the family of drugs known as K ؉ channel openers, raising the question whether mitochondrial K ATP channels are similarly sensitive to these agents. We addressed this question by measuring K ؉ flux in intact rat liver mitochondria and in liposomes containing K ATP channels purified from rat liver and beef heart mitochondria. K ؉ channel openers completely reversed ATP inhibition of K ؉ flux in both systems. In liposomes, ATP-inhibited K ؉ flux was restored by diazoxide (K1 ⁄2 ‫؍‬ 0.4 M), cromakalim (K1 ⁄2 ‫؍‬ 1 M), and two developmental cromakalim analogues, EMD60480 and EMD57970 (K1 ⁄2 ‫؍‬ 6 nM). Similar K1 ⁄2 values were observed in intact mitochondria. These potencies are well within the range observed with plasma membrane K ATP channels. We also compared the potencies of these K ؉ channel openers on the plasma membrane K ATP channel purified from beef heart myocytes. The K ATP channel from cardiac mitochondria is 2000-fold more sensitive to diazoxide than the channel from cardiac sarcolemma, indicating that two distinct receptor subtypes coexist within the myocyte. We suggest that the mitochondrial K ATP channel is an important intracellular receptor that should be taken into account in considering the pharmacology of K ؉ channel openers.K ϩ channel openers (KCOs) 1 activate ATP-inhibited K ATP channels. As described in several excellent reviews (1-3), members of this drug family exhibit a rich and clinically important pharmacology. Thus, cell membrane K ATP channels (cellK ATP ) in different tissues are considered to mediate the hypotensive and diabetogenic effects of diazoxide (4) and the cardioprotective effects of cromakalim and its derivatives (5). It is important to determine whether these drugs also act on mitochondrial K ATP channels (mitoK ATP ) in their therapeutic range.In the first reports of KCO actions in mitochondria, Belyaeva et al. (6) and Szewczyk et al. (7) observed stimulation of K ϩ uptake by KCOs in respiring mitochondria. RP66471 was the most potent KCO studied (K1 ⁄2 ϭ 50 M), whereas P1060 and diazoxide were only weakly active at 700 M. Because these concentrations are much higher than K1 ⁄2 values observed with cellK ATP (1), these results appear to imply that mitochondrial actions of KCOs are not pharmacologically important.We now report that diazoxide, cromakalim, and two experimental benzopyran derivatives are very potent activators of K ϩ flux through ATP-inhibited mitoK ATP , with K1 ⁄2 values similar to those observed with cellK ATP . KCO activation of K ϩ flux was observed in both intact mitochondria and proteoliposomes containing reconstituted mitoK ATP . No effect was observed on uninhibited K ϩ flux, which likely explains the low potencies observed by previous workers (6, 7) in assays that did not include Mg 2ϩ and ATP. We also found that mitoK ATP and cellK ATP from beef heart differed strongly in their sensitivity to diazoxide, indicating distinct receptor subtypes among K ATP channels from the same cell. Our result...

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